M. Peter Marinkovich, MD

All Publications

• Trial of Beremagene Geperpavec (B-VEC) for Dystrophic Epidermolysis Bullosa. The New England journal of medicine Guide, S. V., Gonzalez, M. E., Bagci, I. S., Agostini, B., Chen, H., Feeney, G., Steimer, M., Kapadia, B., Sridhar, K., Quesada Sanchez, L., Gonzalez, F., Van Ligten, M., Parry, T. J., Chitra, S., Kammerman, L. A., Krishnan, S., Marinkovich, M. P. 2022; 387 (24): 2211-2219

  Abstract

  BACKGROUND: Dystrophic epidermolysis bullosa is a rare genetic blistering skin disease caused by mutations in COL7A1, which encodes type VII collagen (C7). Beremagene geperpavec (B-VEC) is a topical investigational herpes simplex virus type 1 (HSV-1)-based gene therapy designed to restore C7 protein by delivering COL7A1.METHODS: We conducted a phase 3, double-blind, intrapatient randomized, placebo-controlled trial involving patients 6 months of age or older with genetically confirmed dystrophic epidermolysis bullosa. For each patient, a primary wound pair was selected, with the wounds matched according to size, region, and appearance. The wounds within each pair were randomly assigned in a 1:1 ratio to receive weekly application of either B-VEC or placebo for 26 weeks. The primary end point was complete wound healing of treated as compared with untreated wounds at 6 months. Secondary end points included complete wound healing at 3 months and the change from baseline to weeks 22, 24, and 26 in pain severity during changes in wound dressing, assessed with the use of a visual analogue scale (scores range from 0 to 10, with higher scores indicating greater pain).RESULTS: Primary wound pairs were exposed to B-VEC and placebo in 31 patients. At 6 months, complete wound healing occurred in 67% of the wounds exposed to B-VEC as compared with 22% of those exposed to placebo (difference, 46 percentage points; 95% confidence interval [CI], 24 to 68; P=0.002). Complete wound healing at 3 months occurred in 71% of the wounds exposed to B-VEC as compared with 20% of those exposed to placebo (difference, 51 percentage points; 95% CI, 29 to 73; P<0.001). The mean change from baseline to week 22 in pain severity during wound-dressing changes was -0.88 with B-VEC and -0.71 with placebo (adjusted least-squares mean difference, -0.61; 95% CI, -1.10 to -0.13); similar mean changes were observed at weeks 24 and 26. Adverse events with B-VEC and placebo included pruritus and chills.CONCLUSIONS: Complete wound healing at 3 and 6 months in patients with dystrophic epidermolysis bullosa was more likely with topical administration of B-VEC than with placebo. Pruritus and mild systemic side effects were observed in patients treated with B-VEC. Longer and larger trials are warranted to determine the durability and side effects of B-VEC for this disease. (Funded by Krystal Biotech; GEM-3 ClinicalTrials.gov number, NCT04491604.).

  View details for DOI 10.1056/NEJMoa2206663

  View details for PubMedID 36516090

• In vivo topical gene therapy for recessive dystrophic epidermolysis bullosa: a phase 1 and 2 trial. Nature medicine Gurevich, I., Agarwal, P., Zhang, P., Dolorito, J. A., Oliver, S., Liu, H., Reitze, N., Sarma, N., Bagci, I. S., Sridhar, K., Kakarla, V., Yenamandra, V. K., O'Malley, M., Prisco, M., Tufa, S. F., Keene, D. R., South, A. P., Krishnan, S. M., Marinkovich, M. P. 2022

  Abstract

  Recessive dystrophic epidermolysis bullosa (RDEB) is a lifelong genodermatosis associated with blistering, wounding, and scarring caused by mutations in COL7A1, the gene encoding the anchoring fibril component, collagen VII (C7). Here, we evaluated beremagene geperpavec (B-VEC), an engineered, non-replicating COL7A1 containing herpes simplex virus type 1 (HSV-1) vector, to treat RDEB skin. B-VEC restored C7 expression in RDEB keratinocytes, fibroblasts, RDEB mice and human RDEB xenografts. Subsequently, a randomized, placebo-controlled, phase 1 and 2 clinical trial (NCT03536143) evaluated matched wounds from nine RDEB patients receiving topical B-VEC or placebo repeatedly over 12weeks. No grade 2 or above B-VEC-related adverse events or vector shedding or tissue-bound skin immunoreactants were noted. HSV-1 and C7 antibodies sometimes presented at baseline or increased after B-VEC treatment without an apparent impact on safety or efficacy. Primary and secondary objectives of C7 expression, anchoring fibril assembly, wound surface area reduction, duration of wound closure, and time to wound closure following B-VEC treatment were met. A patient-reported pain-severity secondary outcome was not assessed given the small proportion of wounds treated. A global assessment secondary endpoint was not pursued due to redundancy with regard to other endpoints. These studies show that B-VEC is an easily administered, safely tolerated, topical molecular corrective therapy promoting wound healing in patients with RDEB.

  View details for DOI 10.1038/s41591-022-01737-y

  View details for PubMedID 35347281

• Epidermolysis bullosa. Nature reviews. Disease primers Bardhan, A., Bruckner-Tuderman, L., Chapple, I. L., Fine, J., Harper, N., Has, C., Magin, T. M., Marinkovich, M. P., Marshall, J. F., McGrath, J. A., Mellerio, J. E., Polson, R., Heagerty, A. H. 2020; 6 (1): 78

  Abstract

  Epidermolysis bullosa (EB) is an inherited, heterogeneous group of rare genetic dermatoses characterized by mucocutaneous fragility and blister formation, inducible by often minimal trauma. A broad phenotypic spectrum has been described, with potentially severe extracutaneous manifestations, morbidity and mortality. Over 30 subtypes are recognized, grouped into four major categories, based predominantly on the plane of cleavage within the skin and reflecting the underlying molecular abnormality: EB simplex, junctional EB, dystrophic EB and Kindler EB. The study of EB has led to seminal advances in our understanding of cutaneous biology. To date, pathogenetic mutations in 16 distinct genes have been implicated in EB, encoding proteins influencing cellular integrity and adhesion. Precise diagnosis is reliant on correlating clinical, electron microscopic and immunohistological features with mutational analyses. In the absence of curative treatment, multidisciplinary care is targeted towards minimizing the risk of blister formation, wound care, symptom relief and specific complications, the most feared of which - and also the leading cause of mortality - is squamous cell carcinoma. Preclinical advances in cell-based, protein replacement and gene therapies are paving the way for clinical successes with gene correction, raising hopes amongst patients and clinicians worldwide.

  View details for DOI 10.1038/s41572-020-0210-0

  View details for PubMedID 32973163

• Safety and Wound Outcomes Following Genetically Corrected Autologous Epidermal Grafts in Patients With Recessive Dystrophic Epidermolysis Bullosa. JAMA Siprashvili, Z., Nguyen, N. T., Gorell, E. S., Loutit, K., Khuu, P., Furukawa, L. K., Lorenz, H. P., Leung, T. H., Keene, D. R., Rieger, K. E., Khavari, P., Lane, A. T., Tang, J. Y., Marinkovich, M. P. 2016; 316 (17): 1808-1817

  Abstract

  Recessive dystrophic epidermolysis bullosa (RDEB) is a devastating, often fatal, inherited blistering disorder caused by mutations in the COL7A1 gene encoding type VII collagen. Support and palliation are the only current therapies.To evaluate the safety and wound outcomes following genetically corrected autologous epidermal grafts in patients with RDEB.Single-center phase 1 clinical trial conducted in the United States of 4 patients with severe RDEB with a measured area of wounds suitable for grafting of at least 100 cm2. Patients with undetectable type VII collagen keratinocyte expression were excluded.Autologous keratinocytes isolated from biopsy samples collected from 4 patients with RDEB were transduced with good manufacturing practice-grade retrovirus carrying full-length human COL7A1 and assembled into epidermal sheet grafts. Type VII collagen gene-corrected grafts (approximately 35 cm2) were transplanted onto 6 wounds in each of the patients (n = 24 grafts).The primary safety outcomes were recombination competent retrovirus, cancer, and autoimmune reaction. Molecular correction was assessed as type VII collagen expression measured by immunofluorescence and immunoelectron microscopy. Wound healing was assessed using serial photographs taken at 3, 6, and 12 months after grafting.The 4 patients (mean age, 23 years [range, 18-32 years]) were all male with an estimated body surface area affected with RDEB of 4% to 30%. All 24 grafts were well tolerated without serious adverse events. Type VII collagen expression at the dermal-epidermal junction was demonstrated on the graft sites by immunofluorescence microscopy in 9 of 10 biopsy samples (90%) at 3 months, in 8 of 12 samples (66%) at 6 months, and in 5 of 12 samples (42%) at 12 months, including correct type VII collagen localization to anchoring fibrils. Wounds with recombinant type VII collagen graft sites displayed 75% or greater healing at 3 months (21 intact graft sites of 24 wound sites; 87%), 6 months (16/24; 67%), and 12 months (12/24; 50%) compared with baseline wound sites.In this preliminary study of 4 patients with RDEB, there was wound healing in some type VII collagen gene-corrected grafts, but the response was variable among patients and among grafted sites and generally declined over 1 year. Long-term follow-up is necessary for these patients, and controlled trials are needed with a broader range of patients to better understand the potential long-term efficacy of genetically corrected autologous epidermal grafts.clinicaltrials.gov Identifier: NCT01263379.

  View details for DOI 10.1001/jama.2016.15588

  View details for PubMedID 27802546

• Tumour microenvironment: laminin 332 in squamous-cell carcinoma. Nature reviews. Cancer Marinkovich, M. P. 2007; 7 (5): 370-380

  Abstract

  Basement membranes can be a barrier to tumour growth, but basement membrane molecules, including laminins, are also important autocrine factors produced by cancers to promote tumorigenesis. Many studies have shown the importance of laminin 332 (previously known as laminin 5) in this process, especially in squamous cell carcinoma. Through interactions with several cell-surface receptors (including alpha6beta4 and alpha3beta1 integrins, epidermal growth factor receptor and syndecan 1) and other basement membrane components (including type VII collagen), laminin 332 drives tumorigenesis through phosphatidylinositol-3 kinase (PI3K) and RAC1 activation, promoting tumour invasion and cell survival. The extracellular interactions of laminin 332 appear amenable to antibody-mediated therapies.

  View details for PubMedID 17457303

• THE DERMAL EPIDERMAL JUNCTION OF HUMAN SKIN CONTAINS A NOVEL LAMININ VARIANT JOURNAL OF CELL BIOLOGY Marinkovich, M. P., Lunstrum, G. P., Keene, D. R., Burgeson, R. E. 1992; 119 (3): 695-703

  Abstract

  We report the identification of a novel laminin variant that appears to be unique to a subset of epithelial basement membranes. The variant contains two chains electrophoretically and immunologically identical to the B1 and B2 chains. Epitopes contained in the laminin A chain are absent from the molecule, and a 190-kD chain substitutes for the A chain. V8 protease analysis and Western blotting studies indicate that the variant 190-kD chain shows structural and immunological similarity to the 200-kD chain of kalinin. Rotary shadowing analysis indicates that the 190-kD chain contributes a large globular structure to the variant long arm, but lacks the short arm contributed to laminin by the A chain. The variant is produced by cultured skin explants, human keratinocytes and a squamous cell carcinoma line, and is present in human amniotic fluid. Polyclonal antibodies raised to kalinin, a recently characterized novel component of anchoring filaments, and mAb BM165 which recognizes a subunit of kalinin (Rousselle et al., 1991) cross react with the variant under nonreducing conditions. Immunohistological surveys of human tissues using the crossreacting antikalinin antiserum indicate that the distribution of this laminin variant is at least restricted to anchoring filament containing basement membranes. We propose the name K-laminin for this variant.

  View details for Web of Science ID A1992JV26900019

  View details for PubMedID 1383241

  View details for PubMedCentralID PMC2289679

• Repurposing an epithelial sodium channel inhibitor as a therapy for murine and human skin inflammation. Science translational medicine Winge, M. C., Nasrallah, M., Jackrazi, L. V., Guo, K. Q., Fuhriman, J. M., Szafran, R., Ramanathan, M., Gurevich, I., Nguyen, N. T., Siprashvili, Z., Inayathullah, M., Rajadas, J., Porter, D. F., Khavari, P. A., Butte, A. J., Marinkovich, M. P. 2024; 16 (777): eade5915

  Abstract

  Inflammatory skin disease is characterized by a pathologic interplay between skin cells and immunocytes and can result in disfiguring cutaneous lesions and systemic inflammation. Immunosuppression is commonly used to target the inflammatory component; however, these drugs are often expensive and associated with side effects. To identify previously unidentified targets, we carried out a nonbiased informatics screen to identify drug compounds with an inverse transcriptional signature to keratinocyte inflammatory signals. Using psoriasis, a prototypic inflammatory skin disease, as a model, we used pharmacologic, transcriptomic, and proteomic characterization to find that benzamil, the benzyl derivative of the US Food and Drug Administration-approved diuretic amiloride, effectively reversed keratinocyte-driven inflammatory signaling. Through three independent mouse models of skin inflammation (Rac1G12V transgenic mice, topical imiquimod, and human skin xenografts from patients with psoriasis), we found that benzamil disrupted pathogenic interactions between the small GTPase Rac1 and its adaptor NCK1. This reduced STAT3 and NF-κB signaling and downstream cytokine production in keratinocytes. Genetic knockdown of sodium channels or pharmacological inhibition by benzamil prevented excess Rac1-NCK1 binding and limited proinflammatory signaling pathway activation in patient-derived keratinocytes without systemic immunosuppression. Both systemic and topical applications of benzamil were efficacious, suggesting that it may be a potential therapeutic avenue for treating skin inflammation.

  View details for DOI 10.1126/scitranslmed.ade5915

  View details for PubMedID 39661704

• Practical considerations relevant to treatment with the gene therapy beremagene geperpavec-svdt for dystrophic epidermolysis bullosa. The Journal of dermatological treatment Paller, A. S., Guide, S. V., Ayala, D., Gonzalez, M. E., Lucky, A. W., Bagci, I. S., Marinkovich, M. P. 2024; 35 (1): 2350232

  Abstract

  Dystrophic epidermolysis bullosa (DEB), a rare genetic skin disease caused by loss-of-function mutations in COL7A1, the gene encoding type VII collagen (COL7), is characterized by skin blistering, scarring, and extracutaneous manifestations that markedly reduce patient quality-of-life. Beremagene geperpavec-svdt ('B-VEC') is a gene therapy employing a non-integrating, replication-defective herpes simplex virus type 1 (HSV-1)-based vector encoding two copies of full-length human COL7A1 to restore COL7 protein after topical administration to DEB wounds. B-VEC was approved in the United States in 2023 as the first topical gene therapy and the first approved treatment for DEB. However, few providers have experience with use of this gene therapy.Data was obtained through literature review and the experience of providers who participated in the B-VEC clinical study or initiated treatment after B-VEC approval.This review discusses the burden of disease, describes the clinical trial outcomes of B-VEC, and provides physician and patient/caregiver recommendations as a practical guide for the real-world use of B-VEC, which can be administered in-office or at the patient's home.By continuing to optimize the practical aspects of B-VEC administration, the focus will continue to shift to patient-centric considerations and improved patient outcomes.

  View details for DOI 10.1080/09546634.2024.2350232

  View details for PubMedID 38724041

• A tyrosine residue on β4 integrin endodomain plays a key role in epidermal tumorigenesis Tripathi, P., Marinkovich, M., Kariya, Y. ELSEVIER SCIENCE INC. 2024: S149

  View details for Web of Science ID 001276879002240

• Functional genotype classification groups distinguish disease severity in recessive dystrophic epidermolysis bullosa Eid, E., Pathmarajah, P., Nazaroff, J., So, J., Mittal, V., Gaona, R., Harris, N., Li, S., Marinkovich, M., Oro, A., Tang, J. Y. ELSEVIER SCIENCE INC. 2024: S74

  View details for Web of Science ID 001276879001194

• Immune reactions in dystrophic epidermolysis bullosa patients following gene therapy Bagci, I., Tripathi, P., Sun, A., Dolorito, J., Sridhar, K., Momin, N., Marinkovich, M. ELSEVIER SCIENCE INC. 2024: S74

  View details for Web of Science ID 001276879001193

• 11-year safety profile of genetically engineered autologous cell therapy in recessive dystrophic epidermolysis bullosa Gaona, R., Gorell, E. S., Marinkovich, M., Chiou, A., Wiss, K., Eid, E., Reusch, D., Truesdale, A., Abdelwahab, S., Tang, J. Y. ELSEVIER SCIENCE INC. 2024: S171

  View details for Web of Science ID 001276879002356

• Lateral association of anchoring fibrils in the absence of anti-parallel dimer formation in two dystrophic epidermolysis bullosa patients Momin, N., Bagci, I., Dolorito, J., Tufa, S., Tripathi, P., Sridhar, K., Keene, D., Marinkovich, M. ELSEVIER SCIENCE INC. 2024: S74

  View details for Web of Science ID 001276879001191

• Collagen VII's Dual Mesenchymal and Epithelial Origins: Implications for Molecular Corrective Therapies. The Journal of investigative dermatology Bagci, I. S., Gurevich, I., Dolorito, J. A., Tripathi, P., Momin, N. S., Sun, A., La, T., Sridhar, K., Marinkovich, M. P. 2024

  View details for DOI 10.1016/j.jid.2024.01.040

  View details for PubMedID 38980231

• Wound behavior in dystrophic epidermolysis bullosa: A retrospective natural history case series. Journal of the American Academy of Dermatology La, T., Chen, H., Chen, S., Bagci, I. S., Sridhar, K., Momin, N., Krishnan, S., Marinkovich, M. P. 2024

  View details for DOI 10.1016/j.jaad.2024.06.060

  View details for PubMedID 38971181

• Functional Genotype-Phenotype Associations in Recessive Dystrophic Epidermolysis Bullosa. Journal of the American Academy of Dermatology So, J. Y., Nazaroff, J., Yenamandra, V. K., Gorell, E. S., Harris, N., Fulchand, S., Eid, E., Dolorito, J. A., Marinkovich, M. P., Tang, J. Y. 2024

  Abstract

  Genotype-phenotype associations in recessive dystrophic epidermolysis bullosa (RDEB) have been difficult to elucidate.To investigate RDEB genotype-phenotype associations and explore a functional approach to genotype classification.Clinical examination and genetic testing of RDEB subjects, including assessment of clinical disease by RDEB subtype and extent of blistering. Genotypes were evaluated according to each variant's effect on type VII collagen (C7) function per updated literature, and subsequently categorized by degree of impact on C7 function as low-impact (splice/missense, missense/missense), medium-impact (premature termination codon [PTC]/missense, splice/splice), and high-impact (PTC/PTC, PTC/splice). Genotype-phenotype associations were investigated using Kruskal-Wallis and Fisher's exact tests, and age-adjusted regressions.83 participants were included. High-impact variants were associated with worse RDEB subtype and clinical disease, including increased prevalence of generalized blistering (55.6% for low-impact vs. 72.7% medium-impact vs. 90.4% high-impact variants, p=0.002). In age-adjusted regressions, participants with high-impact variants had 40.8-fold greater odds of squamous cell carcinoma compared to low-impact variants (p=0.02), and 5.7-fold greater odds of death compared to medium-impact variants (p=0.05).Cross-sectional design.Functional genotype categories may stratify RDEB severity; high-impact variants correlated with worse clinical outcomes. Further validation in larger cohorts is needed.

  View details for DOI 10.1016/j.jaad.2024.04.073

  View details for PubMedID 38735484

• Cell volume expansion and local contractility drive collective invasion of the basement membrane in breast cancer. Nature materials Chang, J., Saraswathibhatla, A., Song, Z., Varma, S., Sanchez, C., Alyafei, N. H., Indana, D., Slyman, R., Srivastava, S., Liu, K., Bassik, M. C., Marinkovich, M. P., Hodgson, L., Shenoy, V., West, R. B., Chaudhuri, O. 2023

  Abstract

  Breast cancer becomes invasive when carcinoma cells invade through the basement membrane (BM)-a nanoporous layer of matrix that physically separates the primary tumour from the stroma. Single cells can invade through nanoporous three-dimensional matrices due to protease-mediated degradation or force-mediated widening of pores via invadopodial protrusions. However, how multiple cells collectively invade through the physiological BM, as they do during breast cancer progression, remains unclear. Here we developed a three-dimensional in vitro model of collective invasion of the BM during breast cancer. We show that cells utilize both proteases and forces-but not invadopodia-to breach the BM. Forces are generated from a combination of global cell volume expansion, which stretches the BM, and local contractile forces that act in the plane of the BM to breach it, allowing invasion. These results uncover a mechanism by which cells collectively interact to overcome a critical barrier to metastasis.

  View details for DOI 10.1038/s41563-023-01716-9

  View details for PubMedID 37957268

• Treatment With Ataluren for Wound Healing and Health Complications in a Patient With Junctional Epidermolysis Bullosa. JAMA dermatology Orlowski, G. M., Amano, S. U., Flanagan, K. E., Rieger, K. E., Marinkovich, M. P., Wiss, K. 2023

  View details for DOI 10.1001/jamadermatol.2023.2077

  View details for PubMedID 37585209

• Anti-NC16A IgA from linear IgA bullous dermatosis patients induce neutrophil-dependent subepidermal blistering in mice. The Journal of investigative dermatology Jing, K., Jordan, T. J., Li, N., Burette, S., Yang, B., Marinkovich, M. P., Diaz, L. A., Googe, P., Thomas, N. E., Feng, S., Liu, Z. 2023

  Abstract

  Linear IgA bullous dermatosis (LABD) is an acquired autoimmune subepidermal blistering skin disease characterized by circulating and tissue-bound IgA autoantibodies that recognize epitopes within the hemidesmosomal protein BP180, including its NC16A domain. Histologically, LABD has long been defined by neutrophil infiltration and dermal-epidermal separation. However, the pathogenic roles of anti-NC16A IgA and neutrophils in LABD, as well as their interactions, have not been thoroughly studied. We show that passive transfer of patient-derived anti-NC16A IgA induce clinical and histologic LABD pathology in humanized NC16A mice that are reconstituted locally or systemically with human neutrophils. The lesional skin of mice exhibits significantly elevated levels of the neutrophil chemoattractants CXCL-1 and CXCL-2. Furthermore, we show significantly increased levels of the neutrophil chemoattractant IL-8 in blister fluids of LABD patients. This study provides direct evidence that anti-NC16A IgA in LABD patients are pathogenic and interact with neutrophils to mediate tissue injury and subepidermal blister formation. This study further corroborates the importance of neutrophil-mediated tissue injury in LABD disease physiology and establishes a clinically relevant in vivo model system that can be used to systematically dissect the immunopathogenesis of LABD.

  View details for DOI 10.1016/j.jid.2023.05.027

  View details for PubMedID 37437774

• Mapping the burden of severe forms of epidermolysis bullosa - Implications for patient management. JAAD international Mellerio, J. E., Kiritsi, D., Marinkovich, M. P., Haro, N. R., Badger, K., Arora, M., Dziasko, M. A., Vithlani, M., Martinez, A. E. 2023; 11: 224-232

  Abstract

  Background: The pathophysiological processes underlying the phenotypic spectrum of severe forms of epidermolysis bullosa (EB) are complex and poorly understood.Objective: To use burden mapping to explore relationships between primary pathomechanisms and secondary clinical manifestations in severe forms of EB (junctional and dystrophic EB [JEB/DEB]) and highlight strengths and weaknesses in evidence regarding the contribution of different pathways.Methods: Literature searches were performed to identify evidence regarding the pathophysiological and clinical aspects of JEB/DEB. Identified publications and clinical experience were used to construct burden maps to visually communicate plausible connections and their relative importance by subtype.Results: Our findings suggest that most of the clinical consequences of JEB/DEB may result from an abnormal state and/or faulty skin remodeling driven by a vicious cycle of delayed wound healing, predominantly mediated through inflammation. The quantity and quality of evidence varies by individual manifestations and disease subtype.Limitations: The burden maps are provisional hypotheses requiring further validation and are limited by the published evidence base and subjectivity in clinical opinion.Conclusions: Delayed wound healing appears to be a key driver of the burden of JEB/DEB. Further studies are warranted to understand the role of inflammatory mediators and accelerated wound healing in patient management.

  View details for DOI 10.1016/j.jdin.2023.02.016

  View details for PubMedID 37179539

• Functional genotype-phenotype associations in recessive dystrophic epidermolysis bullosa Pathmarajah, P., So, J., Nazaroff, J., Harris, N., Marinkovich, M. P., Tang, J. Y. ELSEVIER SCIENCE INC. 2023: S149

  View details for Web of Science ID 001037960301115

• Long term use of topical beremagene geperpavec (B-VEC) in two patients with dystrophic epidermolysis bullosa Bagci, I., Momin, N., Agostini, B., Chen, H., Feeney, G., Steimer, M., Sridhar, K., Kapadia, B., Krishnan, S., Marinkovich, M. P. ELSEVIER SCIENCE INC. 2023: S155

  View details for Web of Science ID 001037960301152

• Neutrophil elastase is critical in linear IgA bullous dermatosis in mice Li, N., Burette, S., Yang, B., Marinkovich, M. P., Diaz, L., Googe, P., Thomas, N., Liu, Z. ELSEVIER SCIENCE INC. 2023: S30

  View details for Web of Science ID 001037960300178

• Results from VIITAL: A phase 3, randomized, intrapatient-controlled trial of an investigational collagen type VII gene-corrected autologous cell therapy, EB-101, for the treatment of recessive dystrophic epidermolysis bullosa (RDEB) Tang, J. Y., Marinkovich, M. P., Wiss, K., McCarthy, D., Truesdale, A., Chiou, A. S., McIntyre, J. K., Moore, A., Grachev, I. ELSEVIER SCIENCE INC. 2023: S138

  View details for Web of Science ID 001037960301053

• Mixed IgM- and IgA-mediated epidermolysis bullosa acquisita associated with IgM-λ paraproteinemia in an 81-year-old woman. JAAD case reports Chau, T., Wu, J., Kahn, B., Elco, C., Marinkovich, M. P., Rieger, K. E., Robinson-Bostom, L., Firoz, E. F. 2023; 34: 7-9

  View details for DOI 10.1016/j.jdcr.2023.01.024

  View details for PubMedID 36923393

  View details for PubMedCentralID PMC10009333

• Characterization of DSG3-CAART cells prior to & following adoptive transfer in mucosal Pemphigus Vulgaris Basu, S., Volkov, J., Nunez, D., Fouch, M., Stadanlick, J., Binder, G., Chang, D., Hoffman, K., Porter, D., Abedi, M., Weng, W. K., Micheletti, R., Maverakis, E., Marinkovich, M. P., Milone, M. C., Payne, A. S. MARY ANN LIEBERT, INC. 2022: A123

  View details for Web of Science ID 000899950600388

• Long-term safety and efficacy of gene-corrected autologous keratinocyte grafts for recessive dystrophic epidermolysis bullosa. Orphanet journal of rare diseases So, J. Y., Nazaroff, J., Iwummadu, C. V., Harris, N., Gorell, E. S., Fulchand, S., Bailey, I., McCarthy, D., Siprashvili, Z., Marinkovich, M. P., Tang, J. Y., Chiou, A. S. 2022; 17 (1): 377

  Abstract

  BACKGROUND: Recessive dystrophic epidermolysis bullosa (RDEB) is a rare, devastating blistering genodermatosis caused by mutations in the COL7A1 gene, which encodes for type VII collagen and is necessary for dermal-epidermal adhesion and integrity. Disease manifestations include severe and debilitating wounds, aggressive squamous cell carcinomas, and premature death; however, there are currently no approved therapies. This Phase 1/2a, open-label study evaluated the long-term efficacy and safety of gene-corrected autologous keratinocyte grafts (EB-101) for chronic RDEB wounds.METHODS: Autologous keratinocytes were harvested from participants with severe RDEB, transduced with a retrovirus containing the full-length COL7A1 gene, and grown into 5*7cm (35 cm2) sheets. Gene-corrected keratinocyte sheets were then transplanted onto chronic RDEB wounds present for≥12weeks.RESULTS: Seven adult participants with severe RDEB were grafted with six sheets each (42 total sheets) onto wounds and followed for a mean of 5.9years (range 4-8years). Long-term improvements in wound healing and symptoms were observed. At year five, 70% (21/30) of treated sites demonstrated≥50% wound healing compared to baseline by investigator global assessment. No sites with≥50% wound healing were painful or pruritic, compared to 67% (6/9) of sites with<50% wound healing (p<0.001) at year five. Grafts were well-tolerated throughout long-term follow-up. No serious adverse events related to treatment were reported over a mean of 5.9years of follow-up. No persistent systemic autoimmunity against type VII collagen or replication-competent retrovirus infections were identified, and no participants developed squamous cell carcinomas related to treatment during long-term follow-up.CONCLUSIONS: Treatment with EB-101 appears safe and efficacious, and produces long-term improvements in wound healing, pain, and itch for RDEB patients. Results from the Phase 3 randomized controlled trial are forthcoming.TRIAL REGISTRATION: ClinicalTrials.gov, NCT01263379. Registered December 15, 2010. https://clinicaltrials.gov/ct2/show/NCT01263379.

  View details for DOI 10.1186/s13023-022-02546-9

  View details for PubMedID 36253825

• Localized CO2 laser treatment of a recalcitrant oral ulceration in pemphigus vulgaris CLINICAL ADVANCES IN PERIODONTICS Chainani-Wu, N., Gopal-Murthy, V., Wu, A., Marinkovich, M. 2022

  Abstract

  Recalcitrant oral lesions of pemphigus vulgaris (PV), an autoimmune blistering disease, can result in significant discomfort, difficulty in eating, and maintaining oral hygiene. Increasing the dosage of systemic medications to control such localized lesions results in an increased risk of adverse effects.We describe a male patient diagnosed at age 51 with PV by oral biopsy that included a direct immunofluorescence examination. After further baseline laboratory testing, he was started on prednisone and mycophenolate. These medications were slowly tapered with adjustments guided by clinical signs. Mycophenolate was replaced with intravenous immunoglobulin monthly infusions due to adverse effects about 2 years after initiation. During the 4.5-year follow-up period after diagnosis, his oral and skin lesions were well-controlled apart from minor transient flares. However, a painful ulcerated lesion on the facial gingiva between #11 and 12 was nonresponsive, even with the use of topical clobetasol in trays. A carbon dioxide (CO2 ) laser was used to vaporize the recalcitrant lesion under local anesthesia. The procedure resulted in complete healing of ulceration with no recurrence until the most recent examination, 2 years postlaser surgery.Adjunctive procedures that can facilitate a decrease in the cumulative dosage of corticosteroids and immunosuppressants have great value in the management of PV. CO2 laser vaporization is safe, with minimal morbidity and no long-term side effects. It should be considered an adjunctive treatment option for the management of recalcitrant lesions in patients with oral PV.Why is this case new information? To our knowledge, this is the second report on the use of a CO2 laser in the treatment of recalcitrant oral lesions of PV and the first report with a documented long-term resolution of the treated lesions. What are the keys to the successful management of this case? A localized recalcitrant lesion was treated with this approach. All other mucosal and cutaneous sites were well controlled on the patient's systemic medication regimen. What are the primary limitations to success in this case? This approach is only relevant for the management of recalcitrant lesions in patients whose disease activity is otherwise well controlled. The availability of specialized equipment and trained clinicians is necessary.

  View details for DOI 10.1002/cap.10210

  View details for Web of Science ID 000844531200001

  View details for PubMedID 35649436

• Genotype-phenotype associations in recessive dystrophic epidermolysis bullosa (RDEB) So, J., Harris, N., Fulchand, S., Gorell, E., Nazaroff, J., Yenamandra, V., Marinkovich, M., Tang, J. ELSEVIER SCIENCE INC. 2022: S77

  View details for Web of Science ID 000829693001011

• A phase 1 trial of DSG3-CAART cells in mucosal-dominant pemphigus vulgaris (mPV) patients: Preliminary data Chang, D. J., Basu, S., Micheletti, R., Maverakis, E., Marinkovich, M., Porter, D. L., Abedi, M., Weng, W., Hoffman, K., Volkov, J., Nunez, D., Milone, M. C., Binder, G. K., Payne, A. S. ELSEVIER SCIENCE INC. 2022: B18

  View details for Web of Science ID 000829693000085

• GEM-3: phase 3 safety and immunogenicity results of beremagene geperpavec (B-VEC), an investigational, topical gene therapy for dystrophic epidermolysis bullosa (DEB) Marinkovich, M., Gonzalez, M., Guide, S., Bagci, I. S., Chitra, S., Agostini, B., Chen, H., Parry, T., Krishnan, S. ELSEVIER SCIENCE INC. 2022: S79

  View details for Web of Science ID 000829693001020

• Characterization of DSG3-CAART Cells Prior to & Following Adoptive Transfer in Mucosal Pemphigus Vulgaris Basu, S., Volkov, J. S., Chang, D., Nunez, D., Hoffman, K., Manfredo-Vieira, S., Porter, D., Abedi, M., Weng, W., Micheletti, R., Maverakis, E., Marinkovich, M., Milone, M., Payne, A. CELL PRESS. 2022: 329-330

  View details for Web of Science ID 000794043701300

• A Phase 1 Trial of Targeted DSG3-CAART Cell Therapy in Mucosal-Dominant Pemphigus Vulgaris (mPV) Patients: Early Cohort Data Chang, D. J., Basu, S., Porter, D., Abedi, M., Weng, W., Micheletti, R., Maverakis, E., Marinkovich, M., Bryer, J., Downing, L., Bagci, I., Hoffman, K., Volkov, J., Nunez, D., Milone, M., Payne, A. S. CELL PRESS. 2022: 373

  View details for Web of Science ID 000794043701395

• The Treatment of Wounds Associated with Recessive Dystrophic Epidermolysis Bullosa with Local Injections of Gene-Corrected, Collagen VII-Expressing Autologous Human Dermal Fibroblasts Marinkovich, M., Sridhar, K. J., Bagci, I., Dolorito, J. M., Keene, D. R., Yonchek, M., Blumenthal, R. L., Spellman, M. C. CELL PRESS. 2022: 376

  View details for Web of Science ID 000794043701401

• Patient-reported outcomes and quality of life in dominant dystrophic epidermolysis bullosa: A global cross-sectional survey. Pediatric dermatology Fulchand, S., Harris, N., Li, S., Barriga, M., Gorell, E., De Souza, M., Murrell, D., Marinkovich, P., Krishna Yenamandra, V., Tang, J. Y. 2021

  Abstract

  INTRODUCTION: Dystrophic epidermolysis bullosa is a debilitating skin condition, without curative treatment. Previous research has focused on the recessive variant, which is known to cause severe disease. Limited work focusing on the clinical manifestations and outcomes of dominant dystrophic epidermolysis bullosa is found (DDEB).METHODS: Analysis of an online survey of 42 DDEB patients.RESULTS: Self-reported severity of disease did not correlate with size of the wound or number of dressing changes, but did correlate with severity of pain reported in the last 12months (3.4 mild vs 6.8 severe disease, P=0.0002). Patients with severe DDEB also reported more severe internal disease symptoms, such as difficulty swallowing (62.5%, P=0.01) and greater analgesic use during dressing changes (4.4% mild vs 81.3% severe, P=<0.001).DISCUSSION: Patient perception of disease severity in DDEB appears to be most impacted by pain, presence of chronic open wounds, difficulty swallowing, difficulty walking, and anal strictures. As research on DDEB increases, future studies focused on these symptoms might be the most impactful for DDEB patients. However, distinguishing DDEB from other subtypes remains a challenge.

  View details for DOI 10.1111/pde.14802

  View details for PubMedID 34515355

• Measurement of skin adhesion in recessive dystrophic epidermolysis bullosa patients JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Nazaroff, J., Manoukian, M., Barriga, M., Lane, A., Marinkovich, M., Tang, J. Y. 2021; 85 (2): 491-492

  View details for DOI 10.1016/j.jaad.2018.07.034

  View details for Web of Science ID 000687299800034

• Patient reported outcomes following EB-101 treatment of recessive dystrophic epidermolysis bullosa (rdeb) wounds showed durable wound healing and reduction in disease burden Tang, J., Marinkovich, M., Barriga, M., Bailey, I., Harris, N., Rudin, D. ELSEVIER SCIENCE INC. 2021: S31

  View details for Web of Science ID 000641872800173

• Assessment of safety in repeat dosing of an in vivo topical gene therapy for the treatment of recessive dystrophic epidermolysis bullosa (RDEB) in a phase I/II trial Marinkovich, M., Forte, S., Oliver, S., Dolorito, J., Sridhar, K., Liu, H., Reitze, N., Sarma, N., Krishnan, S. ELSEVIER SCIENCE INC. 2021: S28

  View details for Web of Science ID 000641872800155

• A systematic literature review of the disease burden in patients with recessive dystrophic epidermolysis bullosa. Orphanet journal of rare diseases Tang, J. Y., Marinkovich, M. P., Lucas, E., Gorell, E., Chiou, A., Lu, Y., Gillon, J., Patel, D., Rudin, D. 2021; 16 (1): 175

  Abstract

  BACKGROUND/OBJECTIVE: Recessive dystrophic epidermolysis bullosa (RDEB) is a genetic collagen disorder characterized by skin fragility leading to blistering, wounds, and scarring. There are currently no approved curative therapies. The objective of this manuscript is to provide a comprehensive literature review of the disease burden caused by RDEB.METHODS: A systematic literature review was conducted in MEDLINE and Embase in accordance with PRISMA guidelines. Observational and interventional studies on the economic, clinical, or humanistic burden of RDEB were included.RESULTS: Sixty-five studies were included in the review. Patients had considerable wound burden, with 60% reporting wounds covering more than 30% of their body. Increases in pain and itch were seen with larger wound size. Chronic wounds were larger and more painful than recurrent wounds. Commonly reported symptoms and complications included lesions and blistering, anemia, nail dystrophy and loss, milia, infections, musculoskeletal contractures, strictures or stenoses, constipation, malnutrition/nutritional problems, pseudosyndactyly, ocular manifestations, and dental caries. Many patients underwent esophageal dilation (29-74%; median dilations, 2-6) and gastrostomy tube placement (8-58%). In the severely affected population, risk of squamous cell carcinoma (SCC) was 76% and mortality from SCC reached 84% by age 40. Patients with RDEB experienced worsened quality of life (QOL), decreased functioning and social activities, and increased pain and itch when compared to other EB subtypes, other skin diseases, and the general population. Families of patients reported experiencing high rates of burden including financial burden (50-54%) and negative impact on private life (79%). Direct medical costs were high, though reported in few studies; annual payer-borne total medical costs in Ireland were $84,534 and annual patient-borne medical costs in Korea were $7392. Estimated annual US costs for wound dressings ranged from $4000 to $245,000. Patients spent considerable time changing dressings: often daily (13-54% of patients) with up to three hours per change (15-40%).CONCLUSION: Patients with RDEB and their families/caregivers experience significant economic, humanistic, and clinical burden. Further research is needed to better understand the costs of disease, how the burden of disease changes over the patient lifetime and to better characterize QOL impact, and how RDEB compares with other chronic, debilitating disorders.

  View details for DOI 10.1186/s13023-021-01811-7

  View details for PubMedID 33849616

• Clinical characteristics associated with increased wound size in patients with recessive dystrophic epidermolysis bullosa. Pediatric dermatology Solis, D. C., Gorell, E. S., Teng, C., Barriga, M., Nazaroff, J., Li, S., Subica, A., Lu, Y., Marinkovich, M. P., Tang, J. Y. 2021

  Abstract

  As more therapeutic clinical trials focus on treatment of individual wounds in patients with recessive dystrophic epidermolysis bullosa, it has become crucial to understand the baseline clinical characteristics of these wounds. To investigate these features, we administered an RDEB-specific wound survey. Forty participants reported on location, size, pain, infection frequency, wound type, and duration of 189 wounds; a subset of 22 participants reported on pruritus in 63 wounds. Increased wound size was significantly associated with increased pain, increased pruritus, longer wound duration, increased infection frequency, and patients with mutations resulting in truncated type VII collagen.

  View details for DOI 10.1111/pde.14576

  View details for PubMedID 33749033

• QR-313, an antisense oligonucleotide, shows therapeutic efficacy for treatment of dominant and recessive dystrophic epidermolysis bullosa: a preclinical study. The Journal of investigative dermatology Bornert, O., Hogervorst, M., Nauroy, P., Bischof, J., Swildens, J., Athanasiou, I., Tufa, S. F., Keene, D. R., Kiritsi, D., Hainzl, S., Murauer, E. M., Marinkovich, M. P., Platenburg, G., Hausser, I., Wally, V., Ritsema, T., Koller, U., Haisma, E. M., Nystrom, A. 2020

  Abstract

  Dystrophic epidermolysis bullosa (DEB) is a blistering skin disease caused by mutations in the gene COL7A1 encoding collagen VII. DEB can be inherited in a recessive (RDEB) or dominant (DDEB) manner and is associated with a high wound burden. Perpetual cycles of wounding and healing drive fibrosis in DDEB and RDEB, as well as the formation of a tumor-permissive microenvironment. Prolonging wound free episodes by improving the quality of wound healing would therefore confer substantial benefit for individuals with DEB. The collagenous domain of collagen VII is encoded by 82 in-frame exons, which makes splice-modulation therapies attractive for DEB. Indeed, antisense oligonucleotide (AON)-based exon skipping has shown promise for RDEB. However, the suitability of AONs for treatment of DDEB remains unexplored. Here, we developed QR-313 - a clinically applicable, potent AON specifically targeting exon73. We show the feasibility of topical delivery of QR-313 in a carbomer-composed gel for treatment of wounds to restore collagen VII abundance in human RDEB skin. Importantly, our data reveal that QR-313 also shows direct benefit for DDEB caused by exon73 mutations. Thus, the same topically applied therapeutic could be used to improve the wound healing quality in RDEB and DDEB.

  View details for DOI 10.1016/j.jid.2020.08.018

  View details for PubMedID 32946877

• Classification of Two Distinct Wound Types in Recessive Dystrophic Epidermolysis Bullosa: A Retrospective and Cohort Natural History Study. Journal of the American Academy of Dermatology Solis, D. C., Teng, C., Gorell, E. S., Barriga, M., Nazaroff, J., Li, S., Lu, Y., Bruckner, A., Marinkovich, M. P., Tang, J. Y. 2020

  View details for DOI 10.1016/j.jaad.2020.08.118

  View details for PubMedID 32896597

• Multidisciplinary Care of Epidermolysis Bullosa during the COVID-19 Pandemic - Consensus: Recommendations by an International Panel of Experts. Journal of the American Academy of Dermatology Murrell, D. F., Lucky, A. W., Salas-Alanis, J. C., Woodley, D. T., Palisson, F., Natsuga, K., Nikolic, M., Ramirez-Quizon, M., Paller, A. S., Lara-Corrales, I., Barzegar, M. A., Sprecher, E., Has, C., Laimer, M., Bruckner, A. L., Bilgic, A., Nanda, A., Purvis, D., Hovnanian, A., Murat-Susic, S., Bauer, J., Kern, J. S., Bodemer, C., Martin, L. K., Mellerio, J., Kowaleski, C., Robertson, S. J., Bruckner-Tuderman, L., Pope, E., Marinkovich, M. P., Tang, J. Y., Su, J., Uitto, J., Eichenfield, L. F., Teng, J., Aan Koh, M. J., Lee, S. E., Khuu, P., Rishel, H. I., Sommerlund, M., Wiss, K., Hsu, C., Chiu, T. W., Martinez, A. E. 2020

  View details for DOI 10.1016/j.jaad.2020.06.1023

  View details for PubMedID 32682031

• Neutrophils are critical in linear IgA bullous dermatosis in mice Li, N., Burette, S., Jing, K., Mulligan, E., Yanik, J., Yang, B., Marinkovich, M. P., Diaz, L., Feng, S., Liu, Z. ELSEVIER SCIENCE INC. 2020: S10

  View details for Web of Science ID 000554564400117

• Larger wounds in recessive dystrophic epidermolysis bullosa patients associated with worse quality of life: Results of a global cross-sectional survey Gorell, E., Eng, V., Solis, D., Choi, S., Nazaroff, J., Li, S., de Souza, M., Murrell, D., Marinkovich, M. P., Tang, J. ELSEVIER SCIENCE INC. 2020: S34

  View details for Web of Science ID 000554564400256

• In vivo correction of recessive dystrophic epidermolysis bullosa (RDEB) by direct cutaneous COL7A1 gene replacement: Results of a phase 1-2 trial Marinkovich, M. P., Vinzant, S., Karkala, V., Sridhar, K., Gurevitch, I., Dolorito, J., Agarwal, P., Krishnan, S. ELSEVIER SCIENCE INC. 2020: S37

  View details for Web of Science ID 000554564400274

• Topical QR-313, an Antisense Oligonucleotide, in the Treatment of Dystrophic Epidermolysis Bullosa Marinkovich, M. P., Sridhar, K., Karkala, V., Yenamandra, V. K., Gurevitch, I., Dolorito, J., Bagci, I. S., O'Mara, C., Ramsdell, D., Landy, H. ELSEVIER SCIENCE INC. 2020: S37

  View details for Web of Science ID 000554564400277

• Premature thymic involution and oropharyngeal blistering cause early lethality in generalized severe junctional epidermolysis bullosa Yenamandra, V. K., Dolorito, J., Gurevitch, I., Godoy, E., Casey, K., Marinkovich, M. P. ELSEVIER SCIENCE INC. 2020: S35

  View details for Web of Science ID 000554564400263

• Type VII collagen NC2 domain expression differentiates severe from milder recessive dystrophic epidermolysis bullosa subtypes Ponakala, A., Yenamandra, V. K., Teng, C. E., Barriga, M., Dolorito, J., Gorell, E., Nguyen, N., Tufa, S., Rieger, K., Keene, D., Tang, J., Marinkovich, M. P. ELSEVIER SCIENCE INC. 2020: S37

  View details for Web of Science ID 000554564400276

• Diagnosis and management of pemphigus: Recommendations of an international panel of experts JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Murrell, D. F., Pena, S., Joly, P., Marinovic, B., Hashimoto, T., Diaz, L. A., Sinha, A. A., Payne, A. S., Daneshpazhooh, M., Eming, R., Jonkman, M. F., Mimouni, D., Borradori, L., Kim, S., Yamagami, J., Lehman, J. S., Saleh, M., Culton, D. A., Czernik, A., Zone, J. J., Fivenson, D., Ujiie, H., Wozniak, K., Akman-Karakas, A., Bernard, P., Korman, N. J., Caux, F., Drenovska, K., Prost-Squarcioni, C., Vassileva, S., Feldman, R. J., Cardones, A., Bauer, J., Ioannides, D., Jedlickova, H., Palisson, F., Patsatsi, A., Uzun, S., Yayli, S., Zillikens, D., Amagai, M., Hertl, M., Schmidt, E., Aoki, V., Grando, S. A., Shimizu, H., Baum, S., Cianchini, G., Feliciani, C., Iranzo, P., Mascaro Jr, J. M., Kowalewski, C., Hall, R., Groves, R., Harman, K. E., Marinkovich, M., Maverakis, E., Werth, V. P. 2020; 82 (3): 575-+

  View details for DOI 10.1016/j.jaad.2018.02.021

  View details for Web of Science ID 000513949800028

• Consensus reclassification of inherited epidermolysis bullosa and other disorders with skin fragility. The British journal of dermatology Has, C., Bauer, J. W., Bodemer, C., Bolling, M. C., Bruckner-Tuderman, L., Diem, A., Fine, J., Heagerty, A., Hovnanian, A., Marinkovich, M. P., Martinez, A. E., McGrath, J. A., Moss, C., Murrell, D. F., Palisson, F., Schwieger-Briel, A., Sprecher, E., Tamai, K., Uitto, J., Woodley, D. T., Zambruno, G., Mellerio, J. E. 2020

  Abstract

  BACKGROUND: Several new genes and clinical subtypes have been identified since the publication in 2014 of the report of the last International Consensus Meeting on Epidermolysis Bullosa (EB).OBJECTIVES: We sought to reclassify disorders with skin fragility, with a focus on EB, based on new clinical and molecular data.METHODS: This was a consensus expert review.RESULTS: In this latest consensus report, we introduce the concept of genetic disorders with skin fragility, of which classical EB represents the prototype. Other disorders with skin fragility, where blisters are a minor part of the clinical picture or are not seen because skin cleavage is very superficial, are classified as separate categories. These include peeling skin disorders, erosive disorders, hyperkeratotic disorders, and connective tissue disorders with skin fragility. Because of the common manifestation of skin fragility, these 'EB-related' disorders should be considered under the EB umbrella in terms of medical and socioeconomic provision of care.CONCLUSIONS: The proposed classification scheme should be of value both to clinicians and researchers, emphasizing both clinical and genetic features of EB. What is already known about this topic? Epidermolysis bullosa (EB) is a group of genetic disorders with skin blistering. The last updated recommendations on diagnosis and classification were published in 2014. What does this study add? We introduce the concept of genetic disorders with skin fragility, of which classical EB represents the prototype. Clinical and genetic aspects, genotype-phenotype correlations, disease-modifying factors and natural history of EB are reviewed. Other disorders with skin fragility, e.g. peeling skin disorders, erosive disorders, hyperkeratotic disorders, and connective tissue disorders with skin fragility are classified as separate categories; these 'EB-related' disorders should be considered under the EB umbrella in terms of medical and socioeconomic provision of care.

  View details for DOI 10.1111/bjd.18921

  View details for PubMedID 32017015

• RESEARCH UPDATE (GENE AND PROTEIN) Marinkovich, M. ACTA DERMATO-VENEREOLOGICA. 2020: 23–24

  View details for Web of Science ID 000568327600066

• Cells from discarded dressings differentiate chronic from acute wounds in patients with Epidermolysis Bullosa. Scientific reports Fuentes, I. n., Guttmann-Gruber, C. n., Tockner, B. n., Diem, A. n., Klausegger, A. n., Cofré-Araneda, G. n., Figuera, O. n., Hidalgo, Y. n., Morandé, P. n., Palisson, F. n., Rebolledo-Jaramillo, B. n., Yubero, M. J., Cho, R. J., Rishel, H. I., Marinkovich, M. P., Teng, J. M., Webster, T. G., Prisco, M. n., Eraso, L. H., Piñon Hofbauer, J. n., South, A. P. 2020; 10 (1): 15064

  Abstract

  Impaired wound healing complicates a wide range of diseases and represents a major cost to healthcare systems. Here we describe the use of discarded wound dressings as a novel, cost effective, accessible, and non-invasive method of isolating viable human cells present at the site of skin wounds. By analyzing 133 discarded wound dressings from 51 patients with the inherited skin-blistering disease epidermolysis bullosa (EB), we show that large numbers of cells, often in excess of 100 million per day, continually infiltrate wound dressings. We show, that the method is able to differentiate chronic from acute wounds, identifying significant increases in granulocytes in chronic wounds, and we show that patients with the junctional form of EB have significantly more cells infiltrating their wounds compared with patients with recessive dystrophic EB. Finally, we identify subsets of granulocytes and T lymphocytes present in all wounds paving the way for single cell profiling of innate and adaptive immune cells with relevance to wound pathologies. In summary, our study delineates findings in EB that have potential relevance for all chronic wounds, and presents a method of cellular isolation that has wide reaching clinical application.

  View details for DOI 10.1038/s41598-020-71794-1

  View details for PubMedID 32934247

• Patient Reported Outcomes and Quality of Life in Recessive Dystrophic Epidermolysis Bullosa: A Global Cross-sectional Survey. Journal of the American Academy of Dermatology Eng, V. A., Solis, D. C., Gorell, E. S., Choi, S. n., Nazaroff, J. n., Li, S. n., de Souza, M. P., Murrell, D. F., Marinkovich, M. P., Tang, J. Y. 2020

  Abstract

  A spectrum of skin disease severity exists in patients with recessive dystrophic epidermolysis bullosa (RDEB).To characterize the patient reported outcomes and quality of life (QOL) in RDEB patients.A cross-sectional study of RDEB patients surveyed through the global EBCare Registry. Patient reported outcomes included skin disease severity, wound characteristics, pain, itch, extra-cutaneous symptoms, and medications. QOL was measured using the validated Quality of Life in Epidermolysis Bullosa (QOLEB) instrument.85 RDEB patients reported on 1,226 wounds (937 recurrent wounds and 289 chronic open wounds). Overall skin disease severity was self-reported as mild (26%, 22/83), moderate (48%, 40/83), or severe (25%, 21/83). Worsening skin disease severity was significantly associated with larger wounds, increased opiate use, anemia, gastrostomy tube use, infections, osteoporosis, and squamous cell carcinoma. Larger wound size was associated with worse quality of life scores.All data were self-reported from an online EB patient registry.This study shows a significant correlation between larger wound size with worsening skin disease severity and quality of life in RDEB participants. Worsening skin disease severity significantly correlated with key clinical manifestations. These results demonstrate that RDEB patients are able to self-report their skin disease severity and wounds.

  View details for DOI 10.1016/j.jaad.2020.03.028

  View details for PubMedID 32199895

• RECOMBINANT HUMAN COLLAGEN VII DECREASES MARKERS OF FIBROSIS AFTER CORNEAL ABRASION IN MICE WITH EPIDERMOLYSIS BULLOSA Chen, V. M., Shelke, R., Gipson, I., Kumar-Singh, R., Panjwani, N., Cao, Z., Ramadan, A., Marinkovich, M. ACTA DERMATO-VENEREOLOGICA. 2020: 70

  View details for Web of Science ID 000568327600187

• RELATIONSHIPS BETWEEN WOUND SIZE, CLINICAL MANIFESTATIONS, AND QUALITY OF LIFE IN RECESSIVE DYSTROPHIC EPIDERMOLYSIS BULLOSA: A GLOBAL CROSS-SECTIONAL SURVEY Gorell, E. S., Eng, V., Solis, D., Choi, S., Nazaroff, J., de Souza, M., Murrell, D., Marinkovich, M. P., Tang, J. Y. ACTA DERMATO-VENEREOLOGICA. 2020: 39–40

  View details for Web of Science ID 000568327600108

• COSTS AND ACCESSIBILITY Marinkovich, M. ACTA DERMATO-VENEREOLOGICA. 2020: 17

  View details for Web of Science ID 000568327600042

• CLASSIFICATION OF TWO DISTINCT WOUND TYPES IN RECESSIVE DYSTROPHIC EPIDERMOLYSIS BULLOSA: A NATURAL HISTORY STUDY Teng, C., Solis, D. C., Barriga, M., Li, S., Marinkovich, M., Tang, J. Y. ACTA DERMATO-VENEREOLOGICA. 2020: 39

  View details for Web of Science ID 000568327600107

• IMMUNE CELL PROFILING OF WOUNDS FROM EPIDERMOLYSIS BULLOSA PATIENTS Fuentes, I., Guttmann-Gruber, C., Tockner, B., Diem, A., Klausegger, A., Cofre-Araneda, G., Figuera, O., Hidalgo, Y., Morande, P., Palisson, F., Rebolledo-Jaramillo, B., Yubero, M., Cho, R. J., Rishel, H. I., Marinkovich, M., Teng, J., Webster, T. G., Prisco, M., Eraso, L. H., Hofbauer, J., South, A. P. ACTA DERMATO-VENEREOLOGICA. 2020: 46

  View details for Web of Science ID 000568327600126

• GENE THERAPY FOR RDEB (EX VIVO VS IN VIVO) Marinkovich, M. ACTA DERMATO-VENEREOLOGICA. 2020: 13

  View details for Web of Science ID 000568327600027

• PHASE 1/2A CLINICAL TRIAL OF GENE-CORRECTED AUTOLOGOUS CELL THERAPY FOR RECESSIVE DYSTROPHIC EPIDERMOLYSIS BULLOSA Gorell, E., Eichstadt, S., Barriga, M., Ponakala, A., Teng, C., Nguyen, N., Siprashvili, Z., Nazaroff, J., Chiou, A., Taylor, L., Khuu, P., Keene, D., Rieger, K., Khosla, R., Lorenz, H. P., Furukawa, L., Marinkovich, M. P., Tang, J. Y. ACTA DERMATO-VENEREOLOGICA. 2020: 75

  View details for Web of Science ID 000568327600201

• RESULTS FROM A PHASE I/II STUDY OF A TOPICAL GENE THERAPY (BERCOLAGENE TELSERPAVEC, B-VEC) IN PATIENTS WITH RECESSIVE DYSTROPHIC EPIDERMOLYSIS BULLOSA (RDEB) Marinkovich, M. P., Vinzant, S., Agarwal, P., Krishna, S. ACTA DERMATO-VENEREOLOGICA. 2020: 48

  View details for Web of Science ID 000568327600130

• UNDERSTANDING OCULAR DISEASE IN THE DEB MOUSE MODEL: CHALLENGES OF ASYMMETRY AND SURVIVAL Chen, V. M., Richey, L., Esmail, M., Shelke, R., Cao, Z., Panjwani, N., Marinkovich, M. ACTA DERMATO-VENEREOLOGICA. 2020: 69

  View details for Web of Science ID 000568327600185

• A PHASE 1/2 STUDY OF GENETICALLY-CORRECTED, COLLAGEN VII EXPRESSING AUTOLOGOUS HUMAN DERMAL FIBROBLASTS INJECTED INTO THE SKIN OF PATIENTS WITH RECESSIVE DYSTROPHIC EPIDERMOLYSIS BULLOSA (RDEB) Marinkovich, M. P., Lane, A., Sridhar, K., Keene, D., Malyala, A., Spellman, M., Maslowski, J. ACTA DERMATO-VENEREOLOGICA. 2020: 75

  View details for Web of Science ID 000568327600200

• From Clinical Phenotype to Genotypic Modelling: Incidence and Prevalence of Recessive Dystrophic Epidermolysis Bullosa (RDEB). Clinical, cosmetic and investigational dermatology Eichstadt, S., Tang, J. Y., Solis, D. C., Siprashvili, Z., Marinkovich, M. P., Whitehead, N., Schu, M., Fang, F., Erickson, S. W., Ritchey, M. E., Colao, M., Spratt, K., Shaygan, A., Ahn, M. J., Sarin, K. Y. 2019; 12: 933-942

  Abstract

  Recessive dystrophic epidermolysis bullosa (RDEB) is an inherited genetic disorder characterized by recurrent and chronic open wounds with significant morbidity, impaired quality of life, and early mortality. RDEB patients demonstrate reduction or structural alteration type VII collagen (C7) owing to mutations in the gene COL7A1, the main component of anchoring fibrils (AF) necessary to maintain epidermal-dermal cohesion. While over 700 alterations in COL7A1 have been reported to cause dystrophic epidermolysis bullosa (DEB), which may be inherited in an autosomal dominant (DDEB) or autosomal recessive pattern (RDEB), the incidence and prevalence of RDEB is not well defined. To date, the widely estimated incidence (0.2-6.65 per million births) and prevalence (3.5-20.4 per million people) of RDEB has been primarily characterized by limited analyses of clinical databases or registries.Using a genetic modelling approach, we use whole exome and genome sequencing data to estimate the allele frequency of pathogenic variants. Through the ClinVar and NCBI database of human genome variants and phenotypes, DEB Register, and analyzing premature COL7A1 termination variants we built a model to predict the pathogenicity of previously unclassified variants. We applied the model to publicly available sequences from the Exome Aggregation Consortium (ExAC) and Genome Aggregation Database (gnomAD) and identified variants which were classified as pathogenic for RDEB from which we estimate disease incidence and prevalence.Genetic modelling applied to the whole exome and genome sequencing data resulted in the identification of predicted RDEB pathogenic alleles, from which our estimate of the incidence of RDEB is 95 per million live births, 30 times the 3.05 per million live birth incidence estimated by the National Epidermolysis Bullosa Registry (NEBR). Using a simulation approach, we estimate a mean of approximately 3,850 patients in the US who may benefit from COL7A1-mediated treatments in the US.We conclude that genetic allele frequency estimation may enhance the underdiagnosis of rare genetic diseases generally, and RDEB specifically, which may improve incidence and prevalence estimates of patients who may benefit from treatment.

  View details for DOI 10.2147/CCID.S232547

  View details for PubMedID 31920360

  View details for PubMedCentralID PMC6935313

• Epidermolysis bullosa with pyloric atresia consistently demonstrates concurrent low intra-basal epidermal and lamina lucida cleavage planes: a survey of six cases. Journal of the European Academy of Dermatology and Venereology : JEADV Wang, J. Y., Marinkovich, M. P., Rieger, K. E. 2019

  Abstract

  Epidermolysis bullosa with pyloric atresia (EB-PA) is a rare, autosomal recessive form of epidermolysis bullosa characterized by mucocutaneous fragility, intestinal obstruction, and frequent urologic and renal abnormalities. The clinical course is typically lethal in the first few weeks of life, with an overall mortality of nearly 80%.1 Mutations in ITGB4, ITGA6, and PLEC1, which encode hemidesmosome components beta4 integrin, alpha6 integrin, and plectin, respectively, are most commonly implicated.

  View details for DOI 10.1111/jdv.16153

  View details for PubMedID 31851393

• Phase 1/2a clinical trial of gene-corrected autologous cell therapy for recessive dystrophic epidermolysis bullosa. JCI insight Eichstadt, S., Barriga, M., Ponakala, A., Teng, C., Nguyen, N. T., Siprashvili, Z., Nazaroff, J., Gorell, E. S., Chiou, A. S., Taylor, L., Khuu, P., Keene, D. R., Rieger, K., Khosla, R. K., Furukawa, L. K., Lorenz, H. P., Marinkovich, M. P., Tang, J. Y. 2019; 4 (19)

  Abstract

  BACKGROUNDRecessive dystrophic epidermolysis bullosa (RDEB) patients have mutations in the COL7A1 gene and thus lack functional type VII collagen (C7) protein; they have marked skin fragility and blistering. This single-center phase 1/2a open-label study evaluated the long-term efficacy, safety, and patient-reported outcomes in RDEB patients treated with gene-corrected autologous cell therapy.METHODSAutologous keratinocytes were isolated from participant skin biopsies. Epidermal sheets were prepared from cells transduced with a retrovirus carrying the full-length human COL7A1 gene. These gene-corrected autologous epidermal sheets measured 5 * 7 cm (35 cm2) and were transplanted onto 6 wound sites in each of 7 adult participants (n = 42 sites total) from 2013 to 2017. Participants were followed for 2 to 5 years.RESULTSNo participants experienced any serious related adverse events. Wound healing of 50% or greater by Investigator Global Assessment was present in 95% (36 of 38) of treated wounds versus 0% (0 of 6) of untreated control wounds at 6 months (P < 0.0001). At year 1, 68% (26 of 38) of treated wounds had 50% or greater healing compared with 17% (1 of 6) of control wounds (P = 0.025). At year 2, 71% (27 of 38) of treated wounds had 50% or greater healing compared with 17% (1 of 6) of control wounds (P = 0.019).CONCLUSIONC7 expression persisted up to 2 years after treatment in 2 participants. Treated wounds with 50% or greater healing demonstrated improvement in patient-reported pain, itch, and wound durability. This study provides additional data to support the clinically meaningful benefit of treating chronic RDEB wounds with ex vivo, C7 gene-corrected autologous cell therapy. This approach was safe and promoted wound healing that was associated with improved patient-reported outcomes.TRIAL REGISTRATIONClinicaltrials.gov identifier: NCT01263379.FUNDINGEpidermolysis Bullosa Research Partnership, Epidermolysis Bullosa Medical Research Foundation, NIH R01 AR055914, Office of Research and Development at the Palo Alto Veteran's Affairs Medical Center, and the Dermatology Foundation.

  View details for DOI 10.1172/jci.insight.130554

  View details for PubMedID 31578311

• Gene Therapy for Epidermolysis Bullosa JOURNAL OF INVESTIGATIVE DERMATOLOGY Marinkovich, M., Tang, J. Y. 2019; 139 (6): 1221–26

  View details for DOI 10.1016/j.jid.2018.11.036

  View details for Web of Science ID 000468701800015

• Genetically corrected autologous keratinocyte epidermal grafts improve wound healing and patient reported outcomes in patients with recessive dystrophic epidermolysis bullosa Eichstadt, S., Barriga, M., Teng, C., Nguyen, N. T., Gorell, E., Siprashvili, Z., Loutit, K., Dutt-Singkh, Y., Nazaroff, J., Marinkovich, M. P., Tang, J. ELSEVIER SCIENCE INC. 2019: S64

  View details for Web of Science ID 000465561503087

• First in human use of a novel in vivo gene therapy to successfully correct recessive dystrophic epidermolysis bullosa (RDEB) skin: Results of a phase 1/2 placebo controlled trial Marinkovich, M. P., Sridhar, K., Gurevich, I., Ponakala, A., Boddu, S., Keene, D., Vinzant, S., Agarwal, P., Krishnan, S. ELSEVIER SCIENCE INC. 2019: S66

  View details for Web of Science ID 000465561503101

• The use of human skin equivalents to evaluate the effectivity of QR-313, an antisense oligonucleotide, in gel formulation Hogervorst, M., van Berkel, M., Oort, C., Marinkovich, M. P., Keene, D., Ritsema, T., Swildens, J., Haisma, I. ELSEVIER SCIENCE INC. 2019: S64

  View details for Web of Science ID 000465561503088

• Natural history of wounds in patients with recessive dystrophic epidermolysis bullosa Teng, C., Solis, D., Tang, J., Barriga, M., Marinkovich, M. P. ELSEVIER SCIENCE INC. 2019: S43

  View details for Web of Science ID 000465561502251

• From Clinical Phenotype to Genotypic Modelling: Incidence and Prevalence of Recessive Dystrophic Epidermolysis Bullosa (RDEB) CLINICAL COSMETIC AND INVESTIGATIONAL DERMATOLOGY Eichstadt, S., Tang, J. Y., Solis, D. C., Siprashvili, Z., Marinkovich, M., Whitehead, N., Schu, M., Fang, F., Erickson, S. W., Ritchey, M. E., Colao, M., Spratt, K., Shaygan, A., Ahn, M. J., Sarin, K. Y. 2019; 12: 933–42

  View details for DOI 10.2147/CCID.S232547

  View details for Web of Science ID 000503975800001

• Gene Therapy for Epidermolysis Bullosa. The Journal of investigative dermatology Marinkovich, M. P., Tang, J. Y. 2019

  Abstract

  Epidermolysis bullosa is a family of diseases characterized by blistering and fragility of the skin in response to mechanical trauma. Advances in our understanding of epidermolysis bullosa pathophysiology have provided the necessary foundation for the first clinical trials of gene therapy for junctional and dystrophic epidermolysis bullosa. These therapies show that gene therapy is both safe and effective, with the potential to correct the molecular and clinical phenotype of patients with epidermolysis bullosa. Improvements in gene delivery and in preventing immune reactions will be among the challenges that lie ahead during further therapeutic development.

  View details for PubMedID 31068252

• Measurement of Skin Adhesion in Recessive Dystrophic Epidermolysis Bullosa Patients. Journal of the American Academy of Dermatology Nazaroff, J., Manoukian, M., Barriga, M., Lane, A., Marinkovich, M. P., Tang, J. Y. 2018

  View details for PubMedID 30081112

• A new deletion mutation sheds light on laminin-332 biology. The British journal of dermatology Marinkovich, M. P. 2018; 178 (6): 1245

  View details for PubMedID 29897122

• A new deletion mutation sheds light on laminin-332 biology BRITISH JOURNAL OF DERMATOLOGY Marinkovich, M. P. 2018; 178 (6): 1245

  View details for DOI 10.1111/bjd.16595

  View details for Web of Science ID 000435078600107

• CyTOF analysis allows characterization of Axl-Expressing Dendritic cells in healthy human donors Leylek, R., Alcantara-Hernandez, M., Wagar, L. E., Engleman, E. G., Marinkovich, M. P., Davis, M. M., Nolan, G. P., Idoyaga, J. WILEY. 2018: 29

  View details for Web of Science ID 000434106700041

• Phase I / II Clinical Trial for Recessive Dystrophic Epidermolysis Bullosa Using EB-101 (COL7A1 Gene-Corrected Autologous Keratinocytes) Tang, J. Y., Marinkovich, M. P., Siprashvili, Z., Nguyen, N. T., Gorell, E. S., Loutit, K., Dutt-Singkh, Y., Barriga, M., Solis, D., Khuu, P., Furukawa, L., Lorenz, H. P., Leung, T. H., Keene, D. R., Rieger, K. E., Khavari, P. A., Lane, A. T. CELL PRESS. 2018: 158

  View details for Web of Science ID 000435342202011

• A phase 1/2 study of genetically-corrected, collagen VII expressing autologous human dermal fibroblasts injected into the skin of patients with recessive dystrophic epidermolysis bullosa (RDEB) Marinkovich, M., Lane, A., Sridhar, K., Keene, D., Malyala, A., Maslowski, J. ELSEVIER SCIENCE INC. 2018: S100

  View details for Web of Science ID 000431188500592

• 50% wound healing correlates with RDEB patient reported outcomes in pain, itch and skin durability Dutt-Singkh, Y., Barriga, M., Nazaroff, J., Solis, D., Li, S., Marinkovich, M., Tang, J. ELSEVIER SCIENCE INC. 2018: S56

  View details for Web of Science ID 000431188500331

• Measurement of skin adherence in recessive dystrophic epidermolysis bullosa patients Nazaroff, J., Li, S., Lane, A., Marinkovich, M., Tang, J. ELSEVIER SCIENCE INC. 2018: S102

  View details for Web of Science ID 000431188500603

• Defining chronic wound types in recessive dystrophic epidermolysis bullosa patients for clinical outcome assessment Solis, D., Nazaroff, J., Dutt-Singkh, Y., Choi, S., Barriga, M., Li, S., Marinkovich, M., Tang, J. ELSEVIER SCIENCE INC. 2018: S97

  View details for Web of Science ID 000431188500571

• Successful in vivo COL7A1 gene delivery and correction of recessive dystrophic epidermolysis bullosa (RDEB) skin using an off the shelf HSV-1 vector (KB103) Gurevich, I., Agarwal, P., Dolorito, J., Prisco, M., O'Malley, M., Regula, L., Wittmer, L., Coghlan, S., Fuentes, I., South, A. P., Krishnan, S., Marinkovich, M. ELSEVIER SCIENCE INC. 2018: S129

  View details for Web of Science ID 000431498600021

• Targeting pathogenic interactions between Rac1 and NCK1 in psoriasis Winge, M. G., Nasrallah, M., Fuhriman, J. M., Ramanathan, M., Azameera, A., Nguyen, N., Inayathullah, M., Rajadas, J., Khavari, P., Butte, A., Marinkovich, M. ELSEVIER SCIENCE INC. 2018: S161

  View details for Web of Science ID 000431498600212

• A high-dimensional phenotypic map of human Dendritic cells paves the way for therapeutics Alcantara-Hernandez, M., Leylek, R., Wagar, L. E., Engleman, E. G., Keler, T., Marinkovich, M. P., Davis, M. M., Nolan, G. P., Idoyaga, J. WILEY. 2018: 25–26

  View details for Web of Science ID 000434106700035

• ITK and RLK Inhibitor PRN694 Improves Skin Disease in Two Mouse Models of Psoriasis. The Journal of investigative dermatology Fuhriman, J. M., Winge, M. C., Haberstock-Debic, H., Funk, J. O., Bradshaw, J. M., Marinkovich, M. P. 2018; 138 (4): 864–71

  Abstract

  The chronic and highly prevalent skin disorder psoriasis vulgaris is characterized by a hyperproliferative epidermis and aberrant immune activity. Many studies have highlighted the role of differentiated T lymphocytes in psoriasis progression. Several biologics are currently available that target proinflammatory cytokines produced by T lymphocytes, but the need for improved therapies persists. The small molecule PRN694 covalently binds ITK and RLK, two Tec kinases activated downstream of T-lymphocyte activation, both of which are up-regulated in psoriatic skin. These Tec kinases are involved in signaling cascades mediating T-lymphocyte proliferation, differentiation, and migration and proinflammatory cytokine production. Invitro analysis showed that PRN694 effectively inhibited IL-17A production from murine T helper type 17-differentiated T lymphocytes. Additionally, PRN694 effectively reduced the psoriasis-like phenotype severity and reduced epidermal proliferation and thickness in both the Rac1V12 and imiquimod mouse models of psoriasis. PRN694 also inhibited CD3+ T-cell and gammadelta T-cell infiltration into skin regions. Inhibition of ITK and RLK attenuated psoriasis-associated signaling pathways, indicating that PRN694 is an effective psoriasis therapeutic.

  View details for PubMedID 29129599

• ITK and RLK Inhibitor PRN694 Improves Skin Disease in Two Mouse Models of Psoriasis JOURNAL OF INVESTIGATIVE DERMATOLOGY Fuhriman, J. M., Winge, M. G., Haberstock-Debic, H., Funk, J., Bradshaw, J., Marinkovich, M. 2018; 138 (4): 864–71

  View details for DOI 10.1016/j.jid.2017.10.029

  View details for Web of Science ID 000428135400024

• Diagnosis and Management of Pemphigus: recommendations by an International Panel of Experts. Journal of the American Academy of Dermatology Murrell, D. F., Pena, S., Joly, P., Marinovic, B., Hashimoto, T., Diaz, L. A., Sinha, A. A., Payne, A. S., Daneshpazhooh, M., Eming, R., Jonkman, M. F., Mimouni, D., Borradori, L., Kim, S., Yamagami, J., Lehman, J. S., Saleh, M. A., Culton, D. A., Czernik, A., Zone, J. J., Fivenson, D., Ujiie, H., Wozniak, K., Akman-Karakas, A., Bernard, P., Korman, N. J., Caux, F., Drenovska, K., Prost-Squarcioni, C., Vassileva, S., Feldman, R. J., Cardones, A. R., Bauer, J., Ioannides, D., Jedlickova, H., Palisson, F., Patsatsi, A., Uzun, S., Yayli, S., Zillikens, D., Amagai, M., Hertl, M., Schmidt, E., Aoki, V., Grando, S. A., Shimizu, H., Baum, S., Cianchini, G., Feliciani, C., Iranzo, P., Mascaro, J. M., Kowalewski, C., Hall, R., Groves, R., Harman, K. E., Marinkovich, M. P., Maverakis, E., Werth, V. P. 2018

  Abstract

  BACKGROUND: Several European countries recently developed international diagnostic and management guidelines for pemphigus, which have been instrumental in the standardization of pemphigus management, OBJECTIVE: We now present results from a subsequent Delphi consensus to broaden the generalizability of recommendations.METHODS: A preliminary survey, based on the European Dermatology Forum (EDF) and the European Academy of Dermatology and Venereology (EADV) guidelines, was sent to a panel of international experts to determine the level of consensus. The results were discussed at the International Bullous Diseases Consensus Group in March 2016 during the annual American Academy of Dermatology (AAD) conference. A second survey was sent following the meeting to more experts to achieve greater international consensus.RESULTS: The 39 experts participated in the first round of the Delphi-survey while 54 from 21 countries completed the second round. The number of statements in the survey was reduced from 175 topics in Delphi I to 24 topics in Delphi II based on Delphi results and meeting discussion.LIMITATIONS: Each recommendation represents the majority opinion and therefore may not reflect all possible treatment options available.CONCLUSIONS: We present here the recommendations resulting from this Delphi process. This international consensus includes intravenous CD20 inhibitors as a first line therapy option for moderate to severe pemphigus.

  View details for PubMedID 29438767

• Chronic skin inflammation accelerates macrophage cholesterol crystal formation and atherosclerosis. JCI insight Baumer, Y., Ng, Q., Sanda, G. E., Dey, A. K., Teague, H. L., Sorokin, A. V., Dagur, P. K., Silverman, J. I., Harrington, C. L., Rodante, J. A., Rose, S. M., Varghese, N. J., Belur, A. D., Goyal, A., Gelfand, J. M., Springer, D. A., Bleck, C. K., Thomas, C. L., Yu, Z. X., Winge, M. C., Kruth, H. S., Marinkovich, M. P., Joshi, A. A., Playford, M. P., Mehta, N. N. 2018; 3 (1)

  Abstract

  Inflammation is critical to atherogenesis. Psoriasis is a chronic inflammatory skin disease that accelerates atherosclerosis in humans and provides a compelling model to understand potential pathways linking these diseases. A murine model capturing the vascular and metabolic diseases in psoriasis would accelerate our understanding and provide a platform to test emerging therapies. We aimed to characterize a new murine model of skin inflammation (Rac1V12) from a cardiovascular standpoint to identify novel atherosclerotic signaling pathways modulated in chronic skin inflammation. The RacV12 psoriasis mouse resembled the human disease state, including presence of systemic inflammation, dyslipidemia, and cardiometabolic dysfunction. Psoriasis macrophages had a proatherosclerotic phenotype with increased lipid uptake and foam cell formation, and also showed a 6-fold increase in cholesterol crystal formation. We generated a triple-genetic K14-RacV12-/+/Srb1-/-/ApoER61H/H mouse and confirmed psoriasis accelerates atherogenesis (~7-fold increase). Finally, we noted a 60% reduction in superoxide dismutase 2 (SOD2) expression in human psoriasis macrophages. When SOD2 activity was restored in macrophages, their proatherogenic phenotype reversed. We demonstrate that the K14-RacV12 murine model captures the cardiometabolic dysfunction and accelerates vascular disease observed in chronic inflammation and that skin inflammation induces a proatherosclerotic macrophage phenotype with impaired SOD2 function, which associated with accelerated atherogenesis.

  View details for DOI 10.1172/jci.insight.97179

  View details for PubMedID 29321372

  View details for PubMedCentralID PMC5821196

• Chronic skin inflammation accelerates macrophage cholesterol crystal formation and atherosclerosis Chronic skin inflammation accelerates macrophage cholesterol crystal formation and atherosclerosis Marinkovich, M. P., et al 2018

  Abstract

  Inflammation is critical to atherogenesis. Psoriasis is a chronic inflammatory skin disease that accelerates atherosclerosis in humans and provides a compelling model to understand potential pathways linking these diseases. A murine model capturing the vascular and metabolic diseases in psoriasis would accelerate our understanding and provide a platform to test emerging therapies. We aimed to characterize a new murine model of skin inflammation (Rac1V12) from a cardiovascular standpoint to identify novel atherosclerotic signaling pathways modulated in chronic skin inflammation. The RacV12 psoriasis mouse resembled the human disease state, including presence of systemic inflammation, dyslipidemia, and cardiometabolic dysfunction. Psoriasis macrophages had a proatherosclerotic phenotype with increased lipid uptake and foam cell formation, and also showed a 6-fold increase in cholesterol crystal formation. We generated a triple-genetic K14-RacV12-/+/Srb1-/-/ApoER61H/H mouse and confirmed psoriasis accelerates atherogenesis (~7-fold increase). Finally, we noted a 60% reduction in superoxide dismutase 2 (SOD2) expression in human psoriasis macrophages. When SOD2 activity was restored in macrophages, their proatherogenic phenotype reversed. We demonstrate that the K14-RacV12 murine model captures the cardiometabolic dysfunction and accelerates vascular disease observed in chronic inflammation and that skin inflammation induces a proatherosclerotic macrophage phenotype with impaired SOD2 function, which associated with accelerated atherogenesis.

  View details for DOI 10.1172/jci.insight.97179

  View details for PubMedCentralID PMC5821196

• Phase I/IIa clinical trial for recessive dystrophic epidermolysis bullosa using EB-101 (COL7A1 gene-corrected autologous keratinocytes) Siprashvili, Z., Nguyen, N. T., Gorell, E. S., Loutit, K., Dutt-Singkh, Y., Nazaroff, J., Khuu, P., Furukawa, L., Lorenz, H. P., Leung, T. H., Keene, D. R., Rieger, K. E., Khavari, P. A., Lane, A. T., Tang, J. Y., Marinkovich, M. P. MARY ANN LIEBERT, INC. 2017: A10

  View details for Web of Science ID 000418410700026

• Validity and Accuracy of a Mobile Phone Application for the Assessment of Wounds in Recessive Dystrophic Epidermolysis Bullosa. Journal of the American Academy of Dermatology Nazaroff, J., Solis, D., Barriga, M., Dutt-Singkh, Y., Shufeng, L., Marinkovich, M. P., Tang, J. Y. 2017

  View details for PubMedID 29146128

• Intraepidermal Type VII Collagen by Immunofluorescence Mapping: A Specific Finding for Bullous Dermolysis of the Newborn. Pediatric dermatology Heinecke, G., Marinkovich, M. P., Rieger, K. E. 2017; 34 (3): 308-314

  Abstract

  Bullous dermolysis of the newborn (BDN) is a subtype of dystrophic epidermolysis bullosa (DEB) characterized by skin fragility and blister formation at birth that typically resolves within the first year of life. Abnormal intraepidermal retention of type VII collagen (C7) has been reported as a characteristic feature of BDN, but few studies have investigated the specificity of this finding.We retrospectively reviewed pathology reports of patients diagnosed with DEB using immunofluorescence mapping from January 2001 to January 2015. For cases describing intraepidermal accumulation of C7, we collected information on patient characteristics, including genetic testing results, clinical outcome, and concurrent electron microscopy findings, where available.Of the 143 cases of DEB with immunofluorescence mapping, eight patients had intracytoplasmic epidermal retention of C7. Of these eight patients, two were lost to follow-up, four had complete resolution of bullae, and two had marked improvement with rare residual bullae. Concurrent electron microscopic findings available for three patients were consistent with BDN.Our review of immunofluorescence mapping findings in patients with DEB found that 5.6% had abnormal intracytoplasmic epidermal retention of C7, a finding previously reported in BDN. All such patients with clinical outcomes available had resolution or marked improvement of bullae, consistent with clinical outcomes expected in BDN.

  View details for DOI 10.1111/pde.13132

  View details for PubMedID 28523885

• ITK and RLK inhibitor improves skin disease in a psoriatic mouse model Fuhriman, J. M., Winge, M. G., Haberstock-Debic, H., Funk, J., Bradshaw, M., Marinkovich, M. ELSEVIER SCIENCE INC. 2017: S120

  View details for DOI 10.1016/j.jid.2017.02.723

  View details for Web of Science ID 000406862400694

• Attenuated netrin-1 receptor mediated regulation of tiam1 is required for rac1 mutant melanoma progression Winge, M. G., Kovalski, J., Nguyen, N. T., Wu, D., Zehnder, A., Khavari, P. A., Marinkovich, M. ELSEVIER SCIENCE INC. 2017: S139

  View details for DOI 10.1016/j.jid.2017.02.836

  View details for Web of Science ID 000406862400805

• Natural history of chronic wounds in patients with recessive dystrophic epidermolysis bullosa Solis, D., Nazaroff, J., Dutt-Singkh, Y., Choi, S., Barriga, M., Bailey-Healy, I., Marinkovich, M., Tang, J. Y. ELSEVIER SCIENCE INC. 2017: S37

  View details for DOI 10.1016/j.jid.2017.02.234

  View details for Web of Science ID 000406862400219

• Quality of life in recessive dystrophic epidermolysis bullosa: The AltaVoice patient registry, 2012-2015 Choi, S., Solis, D., Nazaroff, J., Bailey-Healy, I., Barriga, M., Dutt-Singkh, Y., Li, S., Marinkovich, M., Rangel-Miller, V., Tang, J. Y. ELSEVIER SCIENCE INC. 2017: S38

  View details for DOI 10.1016/j.jid.2017.02.239

  View details for Web of Science ID 000406862400224

• Phase I/IIa clinical trial for recessive dystrophic epidermolysis bullosa using genetically corrected autologous keratinocytes Siprashvili, Z., Nguyen, N., Gorell, E., Loutit, K., Dutt-Singkh, Y., Nazaroff, J., Khuu, P., Furukawa, L., Lorenz, H., Leung, T., Keene, D., Rieger, K., Khavari, P. A., Lane, A., Tang, J. Y., Marinkovich, M. ELSEVIER SCIENCE INC. 2017: S89

  View details for DOI 10.1016/j.jid.2017.02.539

  View details for Web of Science ID 000406862400513

• Type VII collagen (C7) expression and chimerism after bone marrow/cord blood transplantation (BMCBT) for severe generalized recessive dystrophic epidermolysis bullosa (RDEB) Tolar, J., McGrath, J., Osborn, M., Keene, D., Riddle, M., Hook, K., Hordinsky, M., Marinkovich, M., Woodley, D., Chen, M., Tryon, R., DeFor, T., Ebens, C., Tamai, K., Hovnanian, A., Blazar, B., Wagner, J. ELSEVIER SCIENCE INC. 2017: S65

  View details for DOI 10.1016/j.jid.2017.02.394

  View details for Web of Science ID 000406862400376

• Validity and accuracy of a mobile phone application for the assessment of chronic wounds in recessive dystrophic epidermolysis bullosa Nazaroff, J., Solis, D., Bailey-Healy, I., Barriga, M., Choi, S., Dutt-Singkh, Y., Marinkovich, M., Tang, J. Y. ELSEVIER SCIENCE INC. 2017: S36

  View details for DOI 10.1016/j.jid.2017.02.230

  View details for Web of Science ID 000406862400215

• Bone marrow/cord blood transplantation (BMCBT) ameliorates symptoms in some, but not all, subtypes of severe generalized junctional epidermolysis bullosa (JEB) Hook, K., Tolar, J., McGrath, J., Osborn, M., Keene, D., Riddle, M., Hordinsky, M., Marinkovich, M., Tryon, R., DeFor, T., Ebens, C., Tamai, K., Hovnanian, A., Blazar, B., Wagner, J. ELSEVIER SCIENCE INC. 2017: S52

  View details for DOI 10.1016/j.jid.2017.02.324

  View details for Web of Science ID 000406862400307

• Microtubules acquire resistance from mechanical breakage through intralumenal acetylation SCIENCE Xu, Z., Schaedel, L., Portran, D., Aguilar, A., Gaillard, J., Marinkovich, M. P., Thery, M., Nachury, M. V. 2017; 356 (6335): 328-332

  Abstract

  Eukaryotic cells rely on long-lived microtubules for intracellular transport and as compression-bearing elements. We considered that long-lived microtubules are acetylated inside their lumen and that microtubule acetylation may modify microtubule mechanics. Here, we found that tubulin acetylation is required for the mechanical stabilization of long-lived microtubules in cells. Depletion of the tubulin acetyltransferase TAT1 led to a significant increase in the frequency of microtubule breakage. Nocodazole-resistant microtubules lost upon removal of acetylation were largely restored by either pharmacological or physical removal of compressive forces. In in vitro reconstitution experiments, acetylation was sufficient to protect microtubules from mechanical breakage. Thus, acetylation increases mechanical resilience to ensure the persistence of long-lived microtubules.

  View details for DOI 10.1126/science.aai8764

  View details for PubMedID 28428427

• Unique mouse monoclonal antibodies reactive with maturation-related epitopes on type VII collagen. Experimental dermatology Hayakawa, T., Hirako, Y., Teye, K., Tsuchisaka, A., Koga, H., Ishii, N., Karashima, T., Kaneda, M., Oyu, Y., Tateishi, C., Sugawara, K., Yonamine, A., Shinkuma, S., Shimizu, H., Fukano, H., Shimozato, K., Nguyen, N. T., Marinkovich, M. P., Tsuruta, D., Hashimoto, T. 2017

  Abstract

  In this study, we generated a new set of monoclonal antibodies (mAbs) to bovine and human type VII collagen (COL7) by immunizing mice with bovine cornea-derived basement membrane zone (BMZ) fraction. The four mAbs, tentatively named as COL7-like mAbs, showed speckled subepidermal staining in addition to linear BMZ staining of normal human skin and bovine cornea, a characteristic immunofluorescence feature of COL7, but showed no reactivity with COL7 by in vitro biochemical analyses. Taking advantage of the phenomenon that COL7-like mAbs did not react with mouse BMZ, we compared immunofluorescence reactivity between wild-type and COL7-rescued humanized mice and found that COL7-like mAbs reacted with BMZ of COL7-rescued humanized mice. In ELISAs, COL7-like mAbs reacted with intact triple-helical mammalian recombinant protein (RP) of COL7 but not with bacterial RP. Furthermore, COL7-like mAbs did not react with COL7 within either cultured DJM-1 cells or basal cells of skin of a bullous dermolysis of the newborn patient. These results confirmed that COL7-like mAbs reacted with human and bovine COL7. The epitopes for COL7-like mAbs were considered to be present only on mature COL7 after secretion from keratinocytes and deposition to BMZ and to be easily destroyed during immunoblotting procedure. Additional studies indicated association of the speckled subepidermal staining with both type IV collagen and elastin. These unique anti-COL7 mAbs should be useful in studies of both normal and diseased conditions, particularly dystrophic epidermolysis bullosa, which produces only immature COL7.

  View details for DOI 10.1111/exd.13306

  View details for PubMedID 28111846

• Epidermal activation of the small GTPase Rac1 in psoriasis pathogenesis. Small GTPases Winge, M. C., Marinkovich, M. P. 2017: 1-6

  Abstract

  The small GTPase Ras-related C3 botulinum toxin substrate 1 (RAC1) plays a central role in skin homeostasis, including barrier function, wound healing and inflammatory responses. Psoriasis is a common skin disease characterized by deregulation of these functions, and affected skin exhibit keratinocyte hyperproliferation, inflammation and immune cell infiltration. Although psoriasis is often triggered by environmental stimulus, there is a strong genetic association with genes expressed in both immune cells and keratinocytes, of which several are linked to Rac1 signaling. Rac1 is highly active in human psoriatic lesional skin and keratinocytes, and keratinocyte-specific overexpression of an activated mutant of Rac1, Rac1(V12), in a transgenic mouse model closely mimics the presentation of human psoriasis. Both Rac1 activation in keratinocytes and immune derived stimulus are required to drive psoriasiform signaling in transgenic mouse and human xenograft models of psoriasis. Therefore, understanding how increased Rac1 activation in psoriatic epidermis is regulated is central to understanding how the abnormal crosstalk between keratinocytes and immune cells is maintained.

  View details for DOI 10.1080/21541248.2016.1273861

  View details for PubMedID 28055293

• Gentamicin induces functional type VII collagen in recessive dystrophic epidermolysis bullosa patients. The Journal of clinical investigation Woodley, D. T., Cogan, J. n., Hou, Y. n., Lyu, C. n., Marinkovich, M. P., Keene, D. n., Chen, M. n. 2017; 127 (8): 3028–38

  Abstract

  Recessive dystrophic epidermolysis bullosa (RDEB) is an incurable disease caused by mutations in the gene encoding type VII collagen, the major component of anchoring fibrils (AF). We previously demonstrated that gentamicin produced functional type VII collagen in RDEB cells harboring nonsense mutations. Herein, we determined whether topical or intradermal gentamicin administration induces type VII collagen and AFs in RDEB patients.A double-blind, placebo-controlled pilot trial assessed safety and efficacy of topical and intradermal gentamicin in 5 RDEB patients with nonsense mutations. The topical arm tested 0.1% gentamicin ointment or placebo application 3 times daily at 2 open erosion sites for 2 weeks. The intradermal arm tested daily intradermal injection of gentamicin solution (8 mg) or placebo into 2 intact skin sites for 2 days in 4 of 5 patients. Primary outcomes were induction of type VII collagen and AFs at the test sites and safety assessment. A secondary outcome assessed wound closure of topically treated erosions.Both topical and intradermal gentamicin administration induced type VII collagen and AFs at the dermal-epidermal junction of treatment sites. Newly created type VII collagen varied from 20% to 165% of that expressed in normal human skin and persisted for 3 months. Topical gentamicin corrected dermal-epidermal separation, improved wound closure, and reduced blister formation. There were no untoward side effects from gentamicin treatments. Type VII collagen induction did not generate anti-type VII collagen autoantibodies in patients' blood or skin.Topical and intradermal gentamicin suppresses nonsense mutations and induces type VII collagen and AFs in RDEB patients. Gentamicin therapy may provide a readily available treatment for RDEB patients with nonsense mutations.ClinicalTrials.gov NCT02698735.Epidermolysis Bullosa Research Partnership, Epidermolysis Bullosa Medical Research Foundation, NIH, and VA Merit Award.

  View details for PubMedID 28691931

• High-Dimensional Phenotypic Mapping of Human Dendritic Cells Reveals Interindividual Variation and Tissue Specialization. Immunity Alcántara-Hernández, M. n., Leylek, R. n., Wagar, L. E., Engleman, E. G., Keler, T. n., Marinkovich, M. P., Davis, M. M., Nolan, G. P., Idoyaga, J. n. 2017

  Abstract

  Given the limited efficacy of clinical approaches that rely on ex vivo generated dendritic cells (DCs), it is imperative to design strategies that harness specialized DC subsets in situ. This requires delineating the expression of surface markers by DC subsets among individuals and tissues. Here, we performed a multiparametric phenotypic characterization and unbiased analysis of human DC subsets in blood, tonsil, spleen, and skin. We uncovered previously unreported phenotypic heterogeneity of human cDC2s among individuals, including variable expression of functional receptors such as CD172a. We found marked differences in DC subsets localized in blood and lymphoid tissues versus skin, and a striking absence of the newly discovered Axl+ DCs in the skin. Finally, we evaluated the capacity of anti-receptor monoclonal antibodies to deliver vaccine components to skin DC subsets. These results offer a promising path for developing DC subset-specific immunotherapies that cannot be provided by transcriptomic analysis alone.

  View details for PubMedID 29221729

• BMP1-like proteinases are essential to the structure and wound healing of skin MATRIX BIOLOGY Muir, A. M., Massoudi, D., Ngon Nguyen, N., Keene, D. R., Lee, S., Birk, D. E., Davidson, J. M., Marinkovich, M. P., Greenspan, D. S. 2016; 56: 114-131

  Abstract

  Closely related extracellular metalloproteinases bone morphogenetic protein 1 (BMP1) and mammalian Tolloid-like 1 (mTLL1) are co-expressed in various tissues and have been suggested to have overlapping roles in the biosynthetic processing of extracellular matrix components. Early lethality of mice null for the BMP1 gene Bmp1 or the mTLL1 gene Tll1 has impaired in vivo studies of these proteinases. To overcome issues of early lethality and functional redundancy we developed the novel BT(KO) mouse strain, with floxed Bmp1 and Tll1 alleles, for induction of postnatal, simultaneous ablation of the two genes. We previously showed these mice to have a skeletal phenotype that includes elements of osteogenesis imperfecta (OI), osteomalacia, and deficient osteocyte maturation, observations validated by the finding of BMP1 mutations in a subset of human patients with OI-like phenotypes. However, the roles of BMP1-like proteinase in non-skeletal tissues have yet to be explored, despite the supposed importance of putative substrates of these proteinases in such tissues. Here, we employ BT(KO) mice to investigate potential roles for these proteinases in skin. Loss of BMP1-like proteinase activity is shown to result in markedly thinned and fragile skin with unusually densely packed collagen fibrils and delayed wound healing. We demonstrate deficits in the processing of collagens I and III, decorin, biglycan, and laminin 332 in skin, which indicate mechanisms whereby BMP1-like proteinases affect the biology of this tissue. In contrast, lack of effects on collagen VII processing or deposition indicates this putative substrate to be biosynthetically processed by non-BMP1-like proteinases.

  View details for DOI 10.1016/j.matbio.2016.06.004

  View details for PubMedID 27363389

• Safety and efficacy of the JAK inhibitor tofacitinib citrate in patients with alopecia areata. JCI insight Kennedy Crispin, M., Ko, J. M., Craiglow, B. G., Li, S., Shankar, G., Urban, J. R., Chen, J. C., Cerise, J. E., Jabbari, A., Winge, M. C., Marinkovich, M. P., Christiano, A. M., Oro, A. E., King, B. A. 2016; 1 (15)

  Abstract

  Alopecia areata (AA) is an autoimmune disease characterized by hair loss mediated by CD8(+) T cells. There are no reliably effective therapies for AA. Based on recent developments in the understanding of the pathomechanism of AA, JAK inhibitors appear to be a therapeutic option; however, their efficacy for the treatment of AA has not been systematically examined.This was a 2-center, open-label, single-arm trial using the pan-JAK inhibitor, tofacitinib citrate, for AA with >50% scalp hair loss, alopecia totalis (AT), and alopecia universalis (AU). Tofacitinib (5 mg) was given twice daily for 3 months. Endpoints included regrowth of scalp hair, as assessed by the severity of alopecia tool (SALT), duration of hair growth after completion of therapy, and disease transcriptome.Of 66 subjects treated, 32% experienced 50% or greater improvement in SALT score. AA and ophiasis subtypes were more responsive than AT and AU subtypes. Shorter duration of disease and histological peribulbar inflammation on pretreatment scalp biopsies were associated with improvement in SALT score. Drug cessation resulted in disease relapse in 8.5 weeks. Adverse events were limited to grade I and II infections. An AA responsiveness to JAK/STAT inhibitors score was developed to segregate responders and nonresponders, and the previously developed AA disease activity index score tracked response to treatment.At the dose and duration studied, tofacitinib is a safe and effective treatment for severe AA, though it does not result in a durable response. Transcriptome changes reveal unexpected molecular complexity within the disease.ClinicalTrials.gov NCT02197455 and NCT02312882.This work was supported by the US Department of Veterans Affairs Office of Research and Development, National Institute of Arthritis and Musculoskeletal and Skin Diseases National Institutes of Health grant R01 AR47223 and U01 AR67173, the National Psoriasis Foundation, the Swedish Society of Medicine, the Fernström Foundation, the Locks of Love Foundation, the National Alopecia Areata Foundation, and the Ranjini and Ajay Poddar Resource Fund for Dermatologic Diseases Research.

  View details for PubMedID 27699252

• Safety and efficacy of the JAK inhibitor tofacitinib citrate in patients with alopecia areata JCI INSIGHT Crispin, M., Ko, J. M., Craiglow, B. G., Li, S., Shankar, G., Urban, J. R., Chen, J. C., Cerise, J. E., Jabbari, A., Winge, M. G., Marinkovich, M., Christiano, A. M., Oro, A. E., King, B. A. 2016; 1 (15)

  View details for DOI 10.1172/jci.insight.89776

  View details for Web of Science ID 000387123200013

• Factors That May Promote an Effective Local Research Environment JOURNAL OF INVESTIGATIVE DERMATOLOGY Wang, K., Lee, C. S., Marinkovich, M., Chang, H. Y., Oro, A. E., Khavari, P. A. 2016; 136 (8): 1529–31

  View details for PubMedID 27450496

• RAC1 activation drives pathologic interactions between the epidermis and immune cells JOURNAL OF CLINICAL INVESTIGATION Winge, M. C., Ohyama, B., Dey, C. N., Boxer, L. M., Li, W., Ehsani-Chimeh, N., Truong, A. K., Wu, D., Armstrong, A. W., Makino, T., Davidson, M., Starcevic, D., Kislat, A., Nguyen, N. T., Hashimoto, T., Homey, B., Khavari, P. A., Bradley, M., Waterman, E. A., Marinkovich, M. P. 2016; 126 (7): 2661-2677

  Abstract

  Interactions between the epidermis and the immune system govern epidermal tissue homeostasis. These epidermis-immune interactions are altered in the inflammatory disease psoriasis; however, the pathways that underlie this aberrant immune response are not well understood. Here, we determined that Ras-related C3 botulinum toxin substrate 1 (RAC1) is a key mediator of epidermal dysfunction. RAC1 activation was consistently elevated in psoriatic epidermis and primary psoriatic human keratinocytes (PHKCs) exposed to psoriasis-related stimuli, but not in skin from patients with basal or squamous cell carcinoma. Expression of a constitutively active form of RAC1 (RACV12) in mice resulted in the development of lesions similar to those of human psoriasis that required the presence of an intact immune system. RAC1V12-expressing mice and human psoriatic skin showed similar RAC1-dependent signaling as well as transcriptional overlap of differentially expressed epidermal and immune pathways. Coculture of PHKCs with immunocytes resulted in the upregulation of RAC1-dependent proinflammatory cytokines, an effect that was reproduced by overexpressing RAC1 in normal human keratinocytes. In keratinocytes, modulating RAC1 activity altered differentiation, proliferation, and inflammatory pathways, including STAT3, NFκB, and zinc finger protein 750 (ZNF750). Finally, RAC1 inhibition in xenografts composed of human PHKCs and immunocytes abolished psoriasiform hyperplasia and inflammation in vivo. These studies implicate RAC1 as a potential therapeutic target for psoriasis and as a key orchestrator of pathologic epidermis-immune interactions.

  View details for DOI 10.1172/JCI85738

  View details for Web of Science ID 000379094800028

  View details for PubMedID 27294528

  View details for PubMedCentralID PMC4922704

• Practice and Educational Gaps in Blistering Disease DERMATOLOGIC CLINICS Ehsani-Chimeh, N., Marinkovich, M. P. 2016; 34 (3): 251-?

  Abstract

  Treatment of autoimmune patients can be challenging and rewarding. These patients often remain undiagnosed for prolonged periods of time or underdiagnosed without immunologic confirmation, resulting in significant morbidity. The most important principle in management of autoimmune bullous disease is to halt blistering activity while minimizing side effects of medications, especially those caused by corticosteroids. Judicious use of systemic steroids and steroid-sparing agents are essential tools in the management of these patients. Rituximab and intravenous immunoglobulin are playing increasingly important and earlier roles in management. Understanding of and surveillance for drug side effects are critical in long-term management.

  View details for DOI 10.1016/j.det.2016.02.001

  View details for PubMedID 27363880

• CLINICAL AND PRECLINICAL ASSESSMENT OF THE ANTI-MCAM MONOCLONAL ANTIBODY PRX003, A POTENTIAL NOVEL TREATMENT FOR TH17-MEDIATED INFLAMMATORY DISEASES Koller, M., Flanagan, K., Skov, M., Goldblum, R., Griffith, S. G., Barbour, R. M., Ehsani-Chimeh, N., Marinkovich, M. P., Zago, W., Yednock, T. A., Kinney, G. G., Ness, D. BMJ PUBLISHING GROUP. 2016: 134

  View details for DOI 10.1136/annrheumdis-2016-eular.3570

  View details for Web of Science ID 000401523100385

• Transdermal Delivery of Functional Collagen Via Polyvinylpyrrolidone Microneedles ANNALS OF BIOMEDICAL ENGINEERING Sun, W., Inayathullah, M., Manoukian, M. A., Malkovskiy, A. V., Manickam, S., Marinkovich, M. P., Lane, A. T., Tayebi, L., Seifalian, A. M., Rajadas, J. 2015; 43 (12): 2978-2990

  Abstract

  Collagen makes up a large proportion of the human body, particularly the skin. As the body ages, collagen content decreases, resulting in wrinkled skin and decreased wound healing capabilities. This paper presents a method of delivering type I collagen into porcine and human skin utilizing a polyvinylpyrrolidone microneedle delivery system. The microneedle patches were made with concentrations of 1, 2, 4, and 8% type I collagen (w/w). Microneedle structures and the distribution of collagen were characterized using scanning electron microscopy and confocal microscopy. Patches were then applied on the porcine and human skin, and their effectiveness was examined using fluorescence microscopy. The results illustrate that this microneedle delivery system is effective in delivering collagen I into the epidermis and dermis of porcine and human skin. Since the technique presented in this paper is quick, safe, effective and easy, it can be considered as a new collagen delivery method for cosmetic and therapeutic applications.

  View details for DOI 10.1007/s10439-015-1353-0

  View details for PubMedID 26066056

• Characterization of patients with dystrophic epidermolysis bullosa for collagen VII therapy BRITISH JOURNAL OF DERMATOLOGY Gorell, E. S., Nguyen, N., Siprashvili, Z., Marinkovich, M. P., Lane, A. T. 2015; 173 (3): 821–23

  View details for DOI 10.1111/bjd.13737

  View details for Web of Science ID 000362176800031

  View details for PubMedID 25703736

  View details for PubMedCentralID PMC4545744

• Phase I clinical trial for recessive dystrophic epidermolysis bullosa using genetically corrected autologous keratinocytes Siprashvili, Z., Nguyen, N. T., Gorell, E., Loutit, K., Khuu, P., Furukawa, L. K., Lorenz, H. P., Leung, T. H., Keene, D. R., Khavari, P., Lane, A., Tang, J. Y., Marinkovich, M. NATURE PUBLISHING GROUP. 2015: S72

  View details for Web of Science ID 000352783200418

• A novel therapeutic inhibits Rac1 mediated invasion and metastasis in a newly described in vivo model of human melanoma Winge, M. C., Kovalski, J., Nguyen, N. T., Wu, D., Zehnder, A., Khavari, P., Marinkovich, M. NATURE PUBLISHING GROUP. 2015: S71

  View details for Web of Science ID 000352783200412

• Type VII Collagen Is the Major Autoantigen for Sub lamina Densa-Type Linear IgA Bullous Dermatosis JOURNAL OF INVESTIGATIVE DERMATOLOGY Tsuchisaka, A., Ohara, K., Ishii, N., Nguyen, N. T., Marinkovich, M., Hashimoto, T. 2015; 135 (2): 626–29

  View details for PubMedID 25207819

• Definitions and outcome measures for mucous membrane pemphigoid: Recommendations of an international panel of experts JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Murrell, D. F., Marinovic, B., Caux, F., Prost, C., Ahmed, R., Wozniak, K., Amagai, M., Bauer, J., Beissert, S., Borradori, L., Culton, D., Fairley, J. A., Fivenson, D., Jonkman, M. F., Marinkovich, M. P., Woodley, D., Zone, J., Aoki, V., Bernard, P., Bruckner-Tuderman, L., Cianchini, G., Venning, V., Diaz, L., Eming, R., Grando, S. A., Hall, R. P., Hashimoto, T., Herrero-Gonzalez, J. E., Hertl, M., Joly, P., Karpati, S., Kim, J., Kim, S. C., Korman, N. J., Kowalewski, C., Lee, S. E., Rubenstein, D. R., Sprecher, E., Yancey, K., Zambruno, G., Zillikens, D., Doan, S., Daniel, B. S., Werth, V. P. 2015; 72 (1): 168-174

  Abstract

  Mucous membrane pemphigoid encompasses a group of autoimmune bullous diseases with a similar phenotype characterized by subepithelial blisters, erosions, and scarring of mucous membranes, skin, or both. Although knowledge about autoimmune bullous disease is increasing, there is often a lack of clear definitions of disease, outcome measures, and therapeutic end points. With clearer definitions and outcome measures, it is possible to directly compare the results and data from various studies using meta-analyses. This consensus statement provides accurate and reproducible definitions for disease extent, activity, outcome measures, end points, and therapeutic response for mucous membrane pemphigoid and proposes a disease extent score, the Mucous Membrane Pemphigoid Disease Area Index.

  View details for DOI 10.1016/j.jaad.2014.08.024

  View details for PubMedID 25443626

• Inherited epidermolysis bullosa: Updated recommendations on diagnosis and classification JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Fine, J., Bruckner-Tuderman, L., Eady, R. A., Bauer, E. A., Bauer, J. W., Has, C., Heagerty, A., Hintner, H., Hovnanian, A., Jonkman, M. F., Leigh, I., Marinkovich, M. P., Martinez, A. E., McGrath, J. A., Mellerio, J. E., Moss, C., Murrell, D. F., Shimizu, H., Uitto, J., Woodley, D., Zambruno, G. 2014; 70 (6): 1103-1126

  Abstract

  Several new targeted genes and clinical subtypes have been identified since publication in 2008 of the report of the last international consensus meeting on diagnosis and classification of epidermolysis bullosa (EB). As a correlate, new clinical manifestations have been seen in several subtypes previously described.We sought to arrive at an updated consensus on the classification of EB subtypes, based on newer data, both clinical and molecular.In this latest consensus report, we introduce a new approach to classification ("onion skinning") that takes into account sequentially the major EB type present (based on identification of the level of skin cleavage), phenotypic characteristics (distribution and severity of disease activity; specific extracutaneous features; other), mode of inheritance, targeted protein and its relative expression in skin, gene involved and type(s) of mutation present, and--when possible--specific mutation(s) and their location(s).This classification scheme critically takes into account all published data through June 2013. Further modifications are likely in the future, as more is learned about this group of diseases.The proposed classification scheme should be of value both to clinicians and researchers, emphasizing both clinical and molecular features of each EB subtype, and has sufficient flexibility incorporated in its structure to permit further modifications in the future.

  View details for DOI 10.1016/j.jaad.2014.01.903

  View details for Web of Science ID 000336030400032

• Inherited epidermolysis bullosa: updated recommendations on diagnosis and classification. Journal of the American Academy of Dermatology Fine, J., Bruckner-Tuderman, L., Eady, R. A., Bauer, E. A., Bauer, J. W., Has, C., Heagerty, A., Hintner, H., Hovnanian, A., Jonkman, M. F., Leigh, I., Marinkovich, M. P., Martinez, A. E., McGrath, J. A., Mellerio, J. E., Moss, C., Murrell, D. F., Shimizu, H., Uitto, J., Woodley, D., Zambruno, G. 2014; 70 (6): 1103-1126

  Abstract

  Several new targeted genes and clinical subtypes have been identified since publication in 2008 of the report of the last international consensus meeting on diagnosis and classification of epidermolysis bullosa (EB). As a correlate, new clinical manifestations have been seen in several subtypes previously described.We sought to arrive at an updated consensus on the classification of EB subtypes, based on newer data, both clinical and molecular.In this latest consensus report, we introduce a new approach to classification ("onion skinning") that takes into account sequentially the major EB type present (based on identification of the level of skin cleavage), phenotypic characteristics (distribution and severity of disease activity; specific extracutaneous features; other), mode of inheritance, targeted protein and its relative expression in skin, gene involved and type(s) of mutation present, and--when possible--specific mutation(s) and their location(s).This classification scheme critically takes into account all published data through June 2013. Further modifications are likely in the future, as more is learned about this group of diseases.The proposed classification scheme should be of value both to clinicians and researchers, emphasizing both clinical and molecular features of each EB subtype, and has sufficient flexibility incorporated in its structure to permit further modifications in the future.

  View details for DOI 10.1016/j.jaad.2014.01.903

  View details for PubMedID 24690439

• Epidermal Rac1 hyperactivation is a key feature of human psoriasis Winge, M. C., Ohyama, B., Waterman, E. A., Dei, C., Ehsani-Chimeh, N., Li, W., Truong, A., Wu, D., Makino, T., Davidson, M., Starcevic, D., Nguyen, N., Kislat, A., Homey, B., Armstrong, A. W., Hashimoto, T., Engleman, E., Bradley, M., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2014: S18

  View details for Web of Science ID 000334560400104

• Loss of the laminin-332 alpha 3IIIa domain impairs keratinocyte migration in vitro Wang, J. Y., Lakshmireddy, H., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2014: S87

  View details for Web of Science ID 000334560400499

• Phase 1 clinical trial of genetically corrected autologous epidermal keratinocytes for recessive dystrophic epidermolysis bullosa Siprashvili, Z., Nguyen, N. T., Gorell, E., Khuu, P., Furukawa, L., Lorenz, H. P., Leung, T. H., Keene, D. R., Khavari, P., Marinkovich, M., Lane, A. T. NATURE PUBLISHING GROUP. 2014: S75

  View details for Web of Science ID 000334560400429

• Somatic Correction of Junctional Epidermolysis Bullosa by a Highly Recombinogenic AAV Variant. Molecular therapy : the journal of the American Society of Gene Therapy Melo, S. P., Lisowski, L., Bashkirova, E., Zhen, H. H., Chu, K., Keene, D. R., Marinkovich, M. P., Kay, M. A., Oro, A. E. 2014; 22 (4): 725-733

  Abstract

  Definitive correction of disease causing mutations in somatic cells by homologous recombination (HR) is an attractive therapeutic approach for the treatment of genetic diseases. However, HR-based somatic gene therapy is limited by the low efficiency of gene targeting in mammalian cells and replicative senescence of primary cells ex vivo, forcing investigators to explore alternative strategies such as retro- and lentiviral gene transfer, or genome editing in induced pluripotent stem cells. Here, we report correction of mutations at the LAMA3 locus in primary keratinocytes derived from a patient affected by recessive inherited Herlitz junctional epidermolysis bullosa (H-JEB) disorder using recombinant adenoassociated virus (rAAV)-mediated HR. We identified a highly recombinogenic AAV serotype, AAV-DJ, that mediates efficient gene targeting in keratinocytes at clinically relevant frequencies with a low rate of random integration. Targeted H-JEB patient cells were selected based on restoration of adhesion phenotype, which eliminated the need for foreign sequences in repaired cells, enhancing the clinical use and safety profile of our approach. Corrected pools of primary cells assembled functional laminin-332 heterotrimer and fully reversed the blistering phenotype both in vitro and in skin grafts. The efficient targeting of the LAMA3 locus by AAV-DJ using phenotypic selection, together with the observed low frequency of off-target events, makes AAV-DJ based somatic cell targeting a promising strategy for ex vivo therapy for this severe and often lethal epithelial disorder.

  View details for DOI 10.1038/mt.2013.290

  View details for PubMedID 24390279

• Aberrant expression of laminin-332 promotes cell proliferation and cyst growth in ARPKD AMERICAN JOURNAL OF PHYSIOLOGY-RENAL PHYSIOLOGY Vijayakumar, S., Dang, S., Marinkovich, M. P., Lazarova, Z., Yoder, B., Torres, V. E., Wallace, D. P. 2014; 306 (6): F640-F654

  Abstract

  Basement membrane abnormalities have often been observed in kidney cysts of polycystic kidney disease (PKD) patients and animal models. There is an abnormal deposition of extracellular matrix molecules, including laminin-α3,β3,γ2 (laminin-332), in human autosomal dominant PKD (ADPKD). Knockdown of PKD1 paralogs in zebrafish leads to dysregulated synthesis of the extracellular matrix, suggesting that altered basement membrane assembly may be a primary defect in ADPKD. In this study, we demonstrate that laminin-332 is aberrantly expressed in cysts and precystic tubules of human autosomal recessive PKD (ARPKD) kidneys as well as in the kidneys of PCK rats, an orthologous ARPKD model. There was aberrant expression of laminin-γ2 as early as postnatal day 2 and elevated laminin-332 protein in postnatal day 30, coinciding with the formation and early growth of renal cysts in PCK rat kidneys. We also show that a kidney cell line derived from Oak Ridge polycystic kidney mice, another model of ARPKD, exhibited abnormal lumen-deficient and multilumen structures in Matrigel culture. These cells had increased proliferation rates and altered expression levels of laminin-332 compared with their rescued counterparts. A function-blocking polyclonal antibody to laminin-332 significantly inhibited their abnormal proliferation rates and rescued their aberrant phenotype in Matrigel culture. Furthermore, abnormal laminin-332 expression in cysts originating from collecting ducts and proximal tubules as well as in precystic tubules was observed in a human end-stage ADPKD kidney. Our results suggest that abnormal expression of laminin-332 contributes to the aberrant proliferation of cyst epithelial cells and cyst growth in genetic forms of PKD.

  View details for DOI 10.1152/ajprenal.00104.2013

  View details for Web of Science ID 000333334700009

  View details for PubMedID 24370592

• A phase II randomized vehicle-controlled trial of intradermal allogeneic fibroblasts for recessive dystrophic epidermolysis bullosa JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Venugopal, S. S., Yan, W., Frew, J. W., Cohn, H. I., Rhodes, L. M., Kim Tran, K., Melbourne, W., Nelson, J. A., Sturm, M., Fogarty, J., Marinkovich, M. P., Igawa, S., Ishida-Yamamoto, A., Murrell, D. F. 2013; 69 (6): 898-915

  Abstract

  Chronic wounds are a major source of morbidity and mortality in generalized severe recessive dystrophic epidermolysis bullosa (RDEB-GS).This was a phase II double-blinded randomized controlled trial of intralesional allogeneic cultured fibroblasts in suspension solution versus suspension solution alone for wound healing in RDEB-GS.Adult patients with RDEB-GS were screened for chronic ulcers and reduced collagen VII expression. Up to 6 pairs of symmetric wounds were measured and biopsied at baseline, then randomized to cultured allogeneic fibroblasts in a crystalloid suspension solution with 2% albumin or suspension solution alone. Ulcer size, collagen VII protein and messenger RNA expression, anchoring fibril numbers, morphology, and inflammatory markers were measured at 2 weeks and at 3, 6, and 12 months.All wounds healed significantly more rapidly with fibroblasts and vehicle injections, with an area decrease of 50% by 12 weeks, compared with noninjected wounds. Collagen VII expression increased to a similar degree in both study arms in wounds from 3 of 5 patients.The number of patients with RDEB-GS who met inclusion criteria was a limitation, as was 1 trial center rather than multicenter.The injection of both allogeneic fibroblasts and suspension solution alone improved wound healing in chronic nonhealing RDEB-GS wounds independently of collagen VII regeneration. This may provide feasible therapy for wound healing in patients with RDEB-GS.

  View details for DOI 10.1016/j.jaad.2013.08.014

  View details for Web of Science ID 000327736900023

  View details for PubMedID 24075228

• Patterns of oral mucosa lesions in patients with epidermolysis bullosa: comparison and agreement between oral medicine and dermatology JOURNAL OF ORAL PATHOLOGY & MEDICINE Fortuna, G., Lozada-Nur, F., Pollio, A., Aria, M., Cepeda-Valdes, R., Marinkovich, M. P., Bruckner, A. L., Cesar Salas-Alanis, J. 2013; 42 (10): 733-740

  Abstract

  The oral mucosa in patients with epidermolysis bullosa (EB) can be affected with different lesions and degrees of severity. However, patterns of oral lesions in distinct types of EB are still unclear.The purpose of this study was to determine the frequency and distribution of four types of lesions (erythema, erosion, atrophy, and blister) for each oral site and to calculate the interobserver reliability for each type of lesion in each site.Ninety-two patients with different EB types were assessed independently by an oral medicine specialist and a dermatologist. The degree of agreement was calculated by the intraclass correlation coefficient (ICC).The most affected oral site was the tongue, with the most frequent lesion being erythema and atrophy [54(58.7%) patients] for the oral medicine specialist and erosion [54(58.7%) patients] for the dermatologist. Patients with recessive dystrophic EB-severe generalized (RDEB-sev gen) showed the highest mean of sites involved by each lesion for both oral medicine and dermatology. The interobserver reliability on the total of lesions was excellent on only 3 sites: lower lip (ICC: 0.89; 95%CI:0.83-0.92), hard palate (ICC:0.85; 95%CI:0.72-0.91), and tongue (ICC:0.89; 95%CI:0.84-0.92), whereas the interobserver reliability calculated for each single oral lesion showed a lower agreement.Total distribution of sites involved by four types of lesions was higher in RDEB-sev gen than in the rest of EB types, with a predominance of erythema followed by erosion. The agreement on the type of lesion was found to be poor-moderate for many oral sites.

  View details for DOI 10.1111/jop.12094

  View details for Web of Science ID 000326815200003

• Patterns of oral mucosa lesions in patients with epidermolysis bullosa: comparison and agreement between oral medicine and dermatology. Journal of oral pathology & medicine Fortuna, G., Lozada-Nur, F., Pollio, A., Aria, M., Cepeda-Valdes, R., Marinkovich, M. P., Bruckner, A. L., Salas-Alanís, J. C. 2013; 42 (10): 733-740

  Abstract

  The oral mucosa in patients with epidermolysis bullosa (EB) can be affected with different lesions and degrees of severity. However, patterns of oral lesions in distinct types of EB are still unclear.The purpose of this study was to determine the frequency and distribution of four types of lesions (erythema, erosion, atrophy, and blister) for each oral site and to calculate the interobserver reliability for each type of lesion in each site.Ninety-two patients with different EB types were assessed independently by an oral medicine specialist and a dermatologist. The degree of agreement was calculated by the intraclass correlation coefficient (ICC).The most affected oral site was the tongue, with the most frequent lesion being erythema and atrophy [54(58.7%) patients] for the oral medicine specialist and erosion [54(58.7%) patients] for the dermatologist. Patients with recessive dystrophic EB-severe generalized (RDEB-sev gen) showed the highest mean of sites involved by each lesion for both oral medicine and dermatology. The interobserver reliability on the total of lesions was excellent on only 3 sites: lower lip (ICC: 0.89; 95%CI:0.83-0.92), hard palate (ICC:0.85; 95%CI:0.72-0.91), and tongue (ICC:0.89; 95%CI:0.84-0.92), whereas the interobserver reliability calculated for each single oral lesion showed a lower agreement.Total distribution of sites involved by four types of lesions was higher in RDEB-sev gen than in the rest of EB types, with a predominance of erythema followed by erosion. The agreement on the type of lesion was found to be poor-moderate for many oral sites.

  View details for DOI 10.1111/jop.12094

  View details for PubMedID 23772832

• Polyvinylpyrrolidone microneedles enable delivery of intact proteins for diagnostic and therapeutic applications ACTA BIOMATERIALIA Sun, W., Araci, Z., Inayathullah, M., Manickam, S., Zhang, X., Bruce, M. A., Marinkovich, M. P., Lane, A. T., Milla, C., Rajadas, J., Butte, M. J. 2013; 9 (8): 7767-7774

  Abstract

  We present a method of fabricating microneedles from polyvinylpyrrolidone (PVP) that enables delivery of intact proteins (or peptides) to the dermal layers of the skin. PVP is known to self-assemble into branched hollow fibers in aqueous and alcoholic solutions; we utilized this property to develop dissolvable patches of microneedles. Proteins were dissolved in concentrated PVP solution in both alcohol and water, poured into polydimethylsiloxane templates shaped as microneedles and, upon evaporation of solvent, formed into concentric, fibrous, layered structures. This approach of making PVP microneedles overcomes problems in dosage, uniform delivery and stability of protein formulation as compared to protein-coated metallic microneedles or photopolymerized PVP microneedles. Here we characterize the PVP microneedles and measure the delivery of proteins into skin. We show that our method of fabrication preserves the protein conformation. These microneedles can serve as a broadly useful platform for delivering protein antigens and therapeutic proteins to the skin, for example for allergen skin testing or immunotherapy.

  View details for DOI 10.1016/j.actbio.2013.04.045

  View details for PubMedID 23648574

• Development of a mouse model for linear IgA bullous dermatosis Geng, P., Lin, L., Park, M., Li, N., Shumate, K., Marinkovich, M. P., Hall, R., Diaz, L., Liu, Z. NATURE PUBLISHING GROUP. 2013: S39

  View details for Web of Science ID 000317698900234

• Diagnosing Epidermolysis Bullosa Type and Subtype in Infancy Using Immunofluorescence Microscopy: The Stanford Experience PEDIATRIC DERMATOLOGY Berk, D. R., Jazayeri, L., Marinkovich, M. P., Sundram, U. N., Bruckner, A. L. 2013; 30 (2): 226-233

  Abstract

  The natural history of inherited epidermolysis bullosa (EB) varies significantly across subtypes. When confronted with an infant suspected to have EB, rapidly determining the type and subtype is critical in counselling families accurately about the infant's diagnosis and prognosis. Although transmission electron microscopy (TEM) has been considered the criterion standard for EB diagnosis, immunofluorescence microscopy (IFM) using monoclonal antibodies (mAbs) to EB-specific basement membrane zone proteins has several advantages, but few studies have evaluated the diagnostic utility of IFM. We sought to evaluate the clinical utility of IFM using an expanded panel of EB-specific mAbs. This was a retrospective review of pathology reports from infants younger < 1 year old with suspected EB primarily analyzed with IFM by the Stanford Dermatopathology service. Seventy-seven cases were identified for analysis, of which 20 were suboptimal for IFM analysis. Fifty-five cases were diagnosed with EB and classified as follows: EB simplex (n = 5), junctional EB (n = 31), dystrophic EB (n = 19). TEM was available in 36 of 55 cases (65%). IFM with an expanded panel of EB-specific mAbs should be considered the first-line diagnostic test to evaluate infants with clinically suspected EB.

  View details for DOI 10.1111/j.1525-1470.2012.01880.x

  View details for Web of Science ID 000315961500009

  View details for PubMedID 23461686

• Keratinocytes from Induced Pluripotent Stem Cells in Junctional Epidermolysis Bullosa JOURNAL OF INVESTIGATIVE DERMATOLOGY Tolar, J., Xia, L., Lees, C. J., Riddle, M., McElroy, A., Keene, D. R., Lund, T. C., Osborn, M. J., Marinkovich, M. P., Blazar, B. R., Wagner, J. E. 2013; 133 (2): 562-565

  View details for DOI 10.1038/jid.2012.278

  View details for Web of Science ID 000313668000036

  View details for PubMedID 22931927

  View details for PubMedCentralID PMC3514565

• Epidermolysis Bullosa Oropharyngeal Severity (EBOS) score: a multicenter development and reliability assessment. Journal of the American Academy of Dermatology Fortuna, G., Chainani-Wu, N., Lozada-Nur, F., Aria, M., Cepeda-Valdes, R., Pollio, A., Marinkovich, M. P., Martinez-Salazar, A. E., Mignogna, M. D., Bruckner, A. L., Salas-Alanís, J. C. 2013; 68 (1): 83-92

  Abstract

  Epidermolysis bullosa (EB) is a genetic mucocutaneous disorder characterized by blister formation upon mild trauma. All 4 EB types may show oropharyngeal lesions involving either hard or soft tissues. Currently, there are very few data on EB scoring that include the oropharyngeal cavity.We sought to develop an oropharyngeal severity score that was objective, valid, reliable, reproducible, easy to perform, and appropriate for all EB types.In this study, oral medicine specialists developed a new score, the EB Oropharyngeal Severity (EBOS) score. This measured oropharyngeal disease activity (erythema, atrophy, blisters, erosion/ulceration) and structural damage (microstomia, ankyloglossia, scarring phenotype beyond microstomia and ankyloglossia, enamel hypoplasia). It was tested on 92 patients with different types/subtypes of EB, and interobserver and intraobserver reliability were assessed.The EBOS mean total score was 12.9 ± 10.9 (range: 0-34). Both interobserver and intraobserver reliability for total score on all patients with EB were considered excellent (intraclass correlation coefficient 0.94; 95% confidence interval 0.90-0.96 and intraclass correlation coefficient 0.90; 95% confidence interval 0.84-0.94, respectively). Even analyzing each single parameter of the disease activity and structural damage, a substantial to excellent correlation was found in the interobserver (except for 4 sites) and intraobserver reliability. A significant correlation was found between EB types/subtypes and the EBOS median score (P < .001), but not between age and the EBOS mean total score in each group.The sample size was small and the number of EB subtypes was limited.The EBOS score seems to represent an instrument capable of truly quantifying the oropharyngeal severity in different types/subtypes of EB, demonstrating excellent interobserver and intraobserver reliability.

  View details for DOI 10.1016/j.jaad.2012.04.009

  View details for PubMedID 22575158

• Epidermolysis Bullosa Oropharyngeal Severity (EBOS) score: A multicenter development and reliability assessment JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Fortuna, G., Chainani-Wu, N., Lozada-Nur, F., Aria, M., Cepeda-Valdes, R., Pollio, A., Marinkovich, M. P., Martinez-Salazar, A. E., Mignogna, M. D., Bruckner, A. L., Cesar Salas-Alanis, J. 2013; 68 (1): 83-92

  Abstract

  Epidermolysis bullosa (EB) is a genetic mucocutaneous disorder characterized by blister formation upon mild trauma. All 4 EB types may show oropharyngeal lesions involving either hard or soft tissues. Currently, there are very few data on EB scoring that include the oropharyngeal cavity.We sought to develop an oropharyngeal severity score that was objective, valid, reliable, reproducible, easy to perform, and appropriate for all EB types.In this study, oral medicine specialists developed a new score, the EB Oropharyngeal Severity (EBOS) score. This measured oropharyngeal disease activity (erythema, atrophy, blisters, erosion/ulceration) and structural damage (microstomia, ankyloglossia, scarring phenotype beyond microstomia and ankyloglossia, enamel hypoplasia). It was tested on 92 patients with different types/subtypes of EB, and interobserver and intraobserver reliability were assessed.The EBOS mean total score was 12.9 ± 10.9 (range: 0-34). Both interobserver and intraobserver reliability for total score on all patients with EB were considered excellent (intraclass correlation coefficient 0.94; 95% confidence interval 0.90-0.96 and intraclass correlation coefficient 0.90; 95% confidence interval 0.84-0.94, respectively). Even analyzing each single parameter of the disease activity and structural damage, a substantial to excellent correlation was found in the interobserver (except for 4 sites) and intraobserver reliability. A significant correlation was found between EB types/subtypes and the EBOS median score (P < .001), but not between age and the EBOS mean total score in each group.The sample size was small and the number of EB subtypes was limited.The EBOS score seems to represent an instrument capable of truly quantifying the oropharyngeal severity in different types/subtypes of EB, demonstrating excellent interobserver and intraobserver reliability.

  View details for DOI 10.1016/j.jaad.2012.04.009

  View details for Web of Science ID 000312270900015

  View details for PubMedID 22575158

• Keratinocyte-Targeted Expression of Human Laminin gamma 2 Rescues Skin Blistering and Early Lethality of Laminin gamma 2 Deficient Mice PLOS ONE Adair-Kirk, T. L., Griffin, G. L., Meyer, M. J., Kelley, D. G., Miner, J. H., Keene, D. R., Marinkovich, M. P., Ruppert, J. M., Uitto, J., Senior, R. M. 2012; 7 (9)

  Abstract

  Laminin-332 is a heterotrimeric basement membrane component comprised of the α3, ß3, and γ2 laminin chains. Laminin-332 modulates epithelial cell processes, such as adhesion, migration, and differentiation and is prominent in many embryonic and adult tissues. In skin, laminin-332 is secreted by keratinocytes and is a key component of hemidesmosomes connecting the keratinocytes to the underlying dermis. In mice, lack of expression of any of the three Laminin-332 chains result in impaired anchorage and detachment of the epidermis, similar to that seen in human junctional epidermolysis bullosa, and death occurs within a few days after birth. To bypass the early lethality of laminin-332 deficiency caused by the knockout of the mouse laminin γ2 chain, we expressed a dox-controllable human laminin γ2 transgene under a keratinocyte-specific promoter on the laminin γ2 (Lamc2) knockout background. These mice appear similar to their wild-type littermates, do not develop skin blisters, are fertile, and survive >1.5 years. Immunofluorescence analyses of the skin showed that human laminin γ2 colocalized with mouse laminin α3 and ß3 in the basement membrane zone underlying the epidermis. Furthermore, the presence of "humanized" laminin-332 in the epidermal basement membrane zone rescued the alterations in the deposition of hemidesmosomal components, such as plectin, collagen type XVII/BP180, and integrin α6 and ß4 chains, seen in conventional Lamc2 knockout mice, leading to restored formation of hemidesmosomes. These mice will be a valuable tool for studies of organs deficient in laminin-332 and the role of laminin-332 in skin, including wound healing.

  View details for DOI 10.1371/journal.pone.0045546

  View details for Web of Science ID 000311313900155

  View details for PubMedCentralID PMC3445496

• A critical reappraisal of the current data on drug-induced linear immunoglobulin A bullous dermatosis: A real and separate nosological entity? JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Fortuna, G., Cesar Salas-Alanis, J., Guidetti, E., Marinkovich, M. P. 2012; 66 (6): 988-994

  Abstract

  Linear IgA disease (LAD) has been associated with a variety of drugs over the past 30 years.To review current literature on all available cases of drug-induced LAD, in order to ascertain whether a close relationship is justified, so that it constitutes a real and separate nosological entity.The PubMed database was searched for all articles written in English related to drug-induced LAD published between January 1980 and December 2010.The literature review shows that at least 84 articles were published, describing a total of 103 patients. Of these articles, only 46, from 13 countries, were included in this analysis, with a total of 52 patients: 24 (46.2%) were believed to be induced by vancomycin and 28 (53.8%) by drugs other than vancomycin. Challenge-dechallenge-rechallenge testing protocol was performed on only 6 (11.5%) of 52 patients, of which only 5 showed a positive result, while the Naranjo algorithm was performed on only 2 of them (0.3%).The evidence of this review analysis is based only on case reports. No study on large samples of drug-induced LAD is currently available.The literature analysis reveals no strong scientific evidence to support the notion that some drugs have induced LAD; therefore in many reviewed cases, we must question whether drug-induced LAD is really the underlying entity. Further and thorough investigations using one of the available algorithms for adverse drug reaction are warranted.

  View details for DOI 10.1016/j.jaad.2011.09.018

  View details for PubMedID 22169257

• Keratinocyte-targeted expression of human laminin ?2 rescues skin blistering and early lethality of laminin ?2 deficient mice. PloS one Adair-Kirk, T. L., Griffin, G. L., Meyer, M. J., Kelley, D. G., Miner, J. H., Keene, D. R., Marinkovich, M. P., Ruppert, J. M., Uitto, J., Senior, R. M. 2012; 7 (9)

  Abstract

  Laminin-332 is a heterotrimeric basement membrane component comprised of the α3, ß3, and γ2 laminin chains. Laminin-332 modulates epithelial cell processes, such as adhesion, migration, and differentiation and is prominent in many embryonic and adult tissues. In skin, laminin-332 is secreted by keratinocytes and is a key component of hemidesmosomes connecting the keratinocytes to the underlying dermis. In mice, lack of expression of any of the three Laminin-332 chains result in impaired anchorage and detachment of the epidermis, similar to that seen in human junctional epidermolysis bullosa, and death occurs within a few days after birth. To bypass the early lethality of laminin-332 deficiency caused by the knockout of the mouse laminin γ2 chain, we expressed a dox-controllable human laminin γ2 transgene under a keratinocyte-specific promoter on the laminin γ2 (Lamc2) knockout background. These mice appear similar to their wild-type littermates, do not develop skin blisters, are fertile, and survive >1.5 years. Immunofluorescence analyses of the skin showed that human laminin γ2 colocalized with mouse laminin α3 and ß3 in the basement membrane zone underlying the epidermis. Furthermore, the presence of "humanized" laminin-332 in the epidermal basement membrane zone rescued the alterations in the deposition of hemidesmosomal components, such as plectin, collagen type XVII/BP180, and integrin α6 and ß4 chains, seen in conventional Lamc2 knockout mice, leading to restored formation of hemidesmosomes. These mice will be a valuable tool for studies of organs deficient in laminin-332 and the role of laminin-332 in skin, including wound healing.

  View details for DOI 10.1371/journal.pone.0045546

  View details for PubMedID 23029085

• Linear immunoglobulin A bullous dermatosis CLINICS IN DERMATOLOGY Fortuna, G., Marinkovich, M. P. 2012; 30 (1): 38-50

  Abstract

  Linear immunoglobulin A (IgA) bullous dermatosis, also known as linear IgA disease, is an autoimmune mucocutaneous disorder characterized by subepithelial bullae, with IgA autoantibodies directed against several different antigens in the basement membrane zone. Its immunopathologic characteristic resides in the presence of a continuous linear IgA deposit along the basement membrane zone, which is clearly visible on direct immunofluorescence. This disorder shows different clinical features and distribution when adult-onset of linear IgA disease is compared with childhood-onset. Diagnosis is achieved via clinical, histopathologic, and immunopathologic examinations. Two common therapies are dapsone and sulfapyridine, which reduce the inflammatory response and achieve disease remission in a variable period of time.

  View details for DOI 10.1016/j.clindermatol.2011.03.008

  View details for PubMedID 22137225

• Molecular organization of the basement membrane zone CLINICS IN DERMATOLOGY Hashmi, S., Marinkovich, M. P. 2011; 29 (4): 398-411

  Abstract

  The dermal-epidermal basement membrane is a complex assembly of proteins that provide adhesion and regulate many important processes such as development, wound healing, and cancer progression. This contribution focuses on the structure and function of individual components of the basement membrane, how they assemble together, and how they participate in human tissues and diseases, with an emphasis on skin involvement. Understanding the composition and structure of the basement membrane provides insight into the pathophysiology of inherited blistering disorders, such as epidermolysis bullosa, and acquired bullous diseases, such as the pemphigoid group of autoimmune diseases and epidermolysis bullosa acquisita.

  View details for DOI 10.1016/j.clindermatol.2011.01.009

  View details for PubMedID 21679867

• Laminin-511 and integrin beta-1 in hair follicle development and basal cell carcinoma formation BMC DEVELOPMENTAL BIOLOGY DeRouen, M. C., Zhen, H., Tan, S. H., Williams, S., Marinkovich, M. P., Oro, A. E. 2010; 10

  Abstract

  Initiation of the hair follicle placode and its subsequent growth, maturation and cycling in post-natal skin requires signaling interactions between epithelial cells and adjacent dermal cells and involves Shh signaling via the primary cilium. Previous reports have implicated laminins in hair follicle epithelial invagination.Here we use a human BCC model system and mouse mutants to re-evaluate the role of laminin-511 in epithelial invagination in the skin. Blocking laminin 511 and 332 in BCCs maintains primary cilia and Shh signalling, but prevents invagination. Similarly, in laminin-511 and dermal beta-1 integrin mutants, dermal papilla development and primary cilia formation are normal. Dermal beta-1 integrin mutants have normal hair follicle development.Our data provides support for a primary role of laminin-511 promoting hair follicle epithelial downgrowth without affecting dermal primary cilia and Shh target gene induction.

  View details for DOI 10.1186/1471-213X-10-112

  View details for Web of Science ID 000284878300001

  View details for PubMedID 21067603

  View details for PubMedCentralID PMC2995472

• Long-Term Type VII Collagen Restoration to Human Epidermolysis Bullosa Skin Tissue HUMAN GENE THERAPY Siprashvili, Z., Nguyen, N. T., Bezchinsky, M. Y., Marinkovich, M. P., Lane, A. T., Khavari, P. A. 2010; 21 (10): 1299-1310

  Abstract

  In spite of advances in the molecular diagnosis of recessive dystrophic epidermolysis bullosa (RDEB), an inherited blistering disease due to a deficiency of type VII collagen at the basement membrane zone (BMZ) of stratified epithelium, current therapy is limited to supportive palliation. Gene delivery has shown promise in short-term experiments; however, its long-term sustainability through multiple turnover cycles in human tissue has awaited confirmation. To characterize approaches for long-term genetic correction, retroviral vectors were constructed containing long terminal repeat-driven full-length and epitope-tagged COL7A1 cDNA and evaluated for durability of type VII collagen expression and function in RDEB skin tissue regenerated on immune-deficient mice. Type VII collagen expression was maintained for 1 year in vivo, or over 12 epidermal turnover cycles, with no abnormalities in skin morphology or self-renewal. Type VII collagen restoration led to correction of RDEB disease features, including reestablishment of anchoring fibrils at the BMZ. This approach confirms durably corrective and noninjurious gene delivery to long-lived epidermal progenitors and provides the foundation for a human clinical trial of ex vivo gene delivery in RDEB.

  View details for DOI 10.1089/hum.2010.023

  View details for Web of Science ID 000282955500008

  View details for PubMedID 20497034

  View details for PubMedCentralID PMC2957245

• Clinical and immunological heterogeneity of canine subepidermal blistering dermatoses with anti-laminin-332 (laminin-5) auto-antibodies VETERINARY DERMATOLOGY Olivry, T., Bizikova, P., Dunston, S. M., Bond, R., Halliwell, R., Loeffler, A., Pucheu-Haston, C. M., Chen, M., Marinkovich, M. P. 2010; 21 (4): 345-357

  Abstract

  Laminin-332 (laminin-5) is a basement membrane heterotrimeric protein composed of alpha-3, beta-3 and gamma-2 laminin chains. Laminin-332 polypeptides are targeted by auto-antibodies in human patients with mucous membrane (cicatricial) pemphigoid or, more rarely, subepidermal vesicular diseases that resemble epidermolysis bullosa acquisita (EBA) or bullous pemphigoid (BP). The objectives of this report were to characterize the clinical, histopathological and immunological characteristics of nine dogs with auto-antibodies targeting laminin-332. Immunological investigations consisted of direct immunofluorescence (IF), indirect IF with intact and salt-split canine gingival, and salt-split normal or laminin-332-deficient human skin, immunoblotting with purified human laminin-332 and immunoblotting with recombinant NC1 domain of human collagen VII. All dogs exhibited varying degrees of skin blistering and ulceration associated with microscopic subepidermal vesiculation with or without inflammatory cells. Indirect IF established that circulating IgG auto-antibodies bound the dermal side of salt-split canine lip and human skin. In five dogs, IgG variably recognized the basement membrane of laminin-332-deficient human skin (three dogs negative, two dogs positive). In all nine dogs, IgG auto-antibodies detected purified human laminin-332 by immunoblotting. In two dogs, additional targeting of collagen VII-NC1 was present. These observations establish laminin-332 as a novel basement membrane antigen in dogs with autoimmune blistering diseases with variable clinical phenotypes. The names 'acquired junctional epidermolysis bullosa', 'anti-laminin-332 mucous membrane pemphigoid (MMP)' and 'mixed auto-immune subepidermal blistering dermatosis' are proposed for dogs with clinical signs reminiscent of EBA, MMP or BP respectively.

  View details for DOI 10.1111/j.1365-3164.2010.00870.x

  View details for Web of Science ID 000279406100004

  View details for PubMedID 20456722

• Observations of Skin Grafts Derived from Keratinocytes Expressing Selectively Engineered Mutant Laminin-332 Molecules JOURNAL OF INVESTIGATIVE DERMATOLOGY Sakai, N., Waterman, E. A., Nguyen, N. T., Keene, D. R., Marinkovich, M. P. 2010; 130 (8): 2147-2150

  View details for DOI 10.1038/jid.2010.85

  View details for Web of Science ID 000279984000029

  View details for PubMedID 20393483

  View details for PubMedCentralID PMC2904829

• Deletion of dermal integrin beta-1 leads to adhesion, but not hair follicle morphogenesis, defects DeRouen, M. C., Marinkovich, M. P., Oro, A. E. NATURE PUBLISHING GROUP. 2010: S104

  View details for Web of Science ID 000276455100618

• Role of Dermal-Epidermal Basement Membrane Zone in Skin, Cancer, and Developmental Disorders DERMATOLOGIC CLINICS Ko, M. S., Marinkovich, M. P. 2010; 28 (1): 1-?

  Abstract

  The dermal-epidermal basement membrane zone is an important epithelial and stromal interface, consisting of an intricately organized collection of intracellular, transmembrane, and extracellular matrix proteins. The basement membrane zone has several main functions including acting as a permeability barrier, forming an adhesive interface between epithelial cells and the underlying matrix, and controlling cellular organization and differentiation. This article identifies key molecular players of the dermal-epidermal membrane zone, and highlights recent research studies that have identified structural and functional roles of these components in the context of various blistering, neoplastic, and developmental syndromes.

  View details for DOI 10.1016/j.det.2009.10.001

  View details for PubMedID 19945611

• Loss of the Desmosomal Protein Perp Enhances the Phenotypic Effects of Pemphigus Vulgaris Autoantibodies JOURNAL OF INVESTIGATIVE DERMATOLOGY Nguyen, B., Dusek, R. L., Beaudry, V. G., Marinkovich, M. P., Attardi, L. D. 2009; 129 (7): 1710-1718

  Abstract

  Pemphigus vulgaris (PV) is an autoimmune bullous disease in which autoantibodies against proteins of the desmosomal adhesion complex perturb desmosomal function, leading to intercellular adhesion defects in the oral mucosa and skin. Previous studies have demonstrated a central role for downregulation of the desmosomal cadherin desmoglein 3 (DSG3) in the pathogenesis of PV. However, the effects of non-cadherin desmosomal proteins in modulating the cellular manifestations of PV remain poorly understood. Here, we characterize the expression and functional importance of Perp, a newly discovered tetraspan desmosomal protein, in PV. Our data demonstrate that PV autoantibodies disrupt Perp expression at the membrane and trigger its internalization along with DSG3 into the endosomal pathway, where it is ultimately targeted to the lysosome for degradation. We further show that Perp deficiency exacerbates the pathogenic effects of PV autoantibodies on keratinocytes by enhancing both the depletion of desmosomal DSG3 and intercellular adhesion defects. Together, our findings highlight the importance of non-cadherin desmosomal proteins in modulating PV phenotypes and provide new insight into Perp's role in the desmosome.

  View details for DOI 10.1038/jid.2008.419

  View details for PubMedID 19158843

• Subepidermal blistering induced by human autoantibodies to BP180 requires innate immune players in a humanized bullous pemphigoid mouse model JOURNAL OF AUTOIMMUNITY Liu, Z., Sui, W., Zhao, M., Li, Z., Li, N., Thresher, R., Giudice, G. J., Fairley, J. A., Sitaru, C., Zillikens, D., Ning, G., Marinkovich, M. P., Diaz, L. A. 2008; 31 (4): 331-338

  Abstract

  Bullous pemphigoid (BP) is a cutaneous autoimmune inflammatory disease associated with subepidermal blistering and autoantibodies against BP180, a transmembrane collagen and major component of the hemidesmosome. Numerous inflammatory cells infiltrate the upper dermis in BP. IgG autoantibodies in BP fix complement and target multiple BP180 epitopes that are highly clustered within a non-collagen linker domain, termed NC16A. Anti-BP180 antibodies induce BP in mice. In this study, we generated a humanized mouse strain, in which the murine BP180NC14A is replaced with the homologous human BP180NC16A epitope cluster region. We show that the humanized NC16A (NC16A+/+) mice injected with anti-BP180NC16A autoantibodies develop BP-like subepidermal blisters. The F(ab')(2) fragments of pathogenic IgG fail to activate the complement cascade and are no longer pathogenic. The NC16A+/+ mice pretreated with mast cell activation blocker or depleted of complement or neutrophils become resistant to BP. These findings suggest that the humoral response in BP critically depends on innate immune system players.

  View details for DOI 10.1016/j.jaut.2008.08.009

  View details for Web of Science ID 000261839400003

  View details for PubMedID 18922680

  View details for PubMedCentralID PMC2642586

• Laminin-511 is an epithelial message promoting dermal papilla development and function during early hair morphogenesis GENES & DEVELOPMENT Gao, J., DeRouen, M. C., Chen, C., Nguyen, M., Nguyen, N. T., Ido, H., Harada, K., Sekiguchi, K., Morgan, B. A., Miner, J. H., Oro, A. E., Marinkovich, M. P. 2008; 22 (15): 2111-2124

  Abstract

  Hair morphogenesis takes place through reciprocal epithelial and mesenchymal signaling; however, the mechanisms controlling signal exchange are poorly understood. Laminins are extracellular proteins that play critical roles in adhesion and signaling. Here we demonstrate the mechanism of how laminin-511 controls hair morphogenesis. Dermal papilla (DP) from laminin-511 mutants showed developmental defects by E16.5, including a failure to maintain expression of the key morphogen noggin. This maintenance was critical as exogenous introduction of noggin or sonic hedgehog (Shh) produced downstream from noggin was sufficient to restore hair follicle development in lama5(-/-) (laminin-511-null) skin. Hair development required the beta1 integrin binding but not the heparin binding domain of laminin-511. Previous studies demonstrated that Shh signaling requires primary cilia, microtubule-based signaling organelles. Laminin-511 mutant DP showed decreased length and structure of primary cilia in vitro and in vivo. Laminin-511, but not laminin-111, restored primary cilia formation in lama5(-/-) mesenchyme and triggered noggin expression in an Shh- and PDGF-dependent manner. Inhibition of laminin-511 receptor beta1 integrin disrupted DP primary cilia formation as well as hair development. These studies show that epithelial-derived laminin-511 is a critical early signal that directs ciliary function and DP maintenance as a requirement for hair follicle downgrowth.

  View details for DOI 10.1101/gad.1689908

  View details for PubMedID 18676816

• Targeting a tumor-specific laminin domain critical for human carcinogenesis CANCER RESEARCH Tran, M., Rousselle, P., Nokelainen, P., Tallapragada, S., Nguyen, N. T., Fincher, E. F., Marinkovich, M. P. 2008; 68 (8): 2885-2894

  Abstract

  Laminin-332 is critical for squamous cell carcinoma (SCC) tumorigenesis, but targeting it for cancer therapy has been unachievable due to key role of laminin-332 in promoting tissue integrity. Here, we show that a portion of laminin-332, termed G45, which is proteolytically removed and absent in normal tissues, is prominently expressed in most human SCC tumors and plays an important role in human SCC tumorigenesis. Primary human keratinocytes lacking G45 (DeltaG45) showed alterations of basal receptor organization, impaired matrix deposition, and increased migration. After SCC transformation, the absence of G45 domain in DeltaG45 cells was associated with deficient extracellular signal-regulated kinase and phosphotidylinositol 3-kinase (PI3K) pathway activation, impaired invasion, deficient metalloproteinase activity, and absent tumorgenicity in vivo. Expression of G45 or activated PI3K subunit in DeltaG45 cells reversed these abnormalities. G45 antibody treatment induced SCC tumor apoptosis, decreased SCC tumor proliferation, and markedly impaired human SCC tumorigenesis in vivo without affecting normal tissue adhesion. These results show a remarkable selectivity of expression and function for laminin-332 G45 in human SCC tumorigenesis and implicate it as a specific target for anticancer therapy.

  View details for DOI 10.1158/0008-5472.CAN-07-6160

  View details for PubMedID 18413757

• Observations of epidermal grafts derived from keratinocytes expressing selectively engineered mutant laminin-332 molecules International Investigative Dermatology Meeting Sakai, N., Waterman, E., Nguyen, N., Keene, D., Kawana, S., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2008: S43–S43

  View details for Web of Science ID 000254353800257

• Targeting a tumor specific laminin domain critical for human carcinogenesis International Investigative Dermatology Meeting Tran, M. M., Rousselle, P., Tallapragracku, S., Nguyen, N., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2008: S27–S27

  View details for Web of Science ID 000254353800157

• Discovery of Basement Membrane Components. journal of investigative dermatology Peter Marinkovich, M. 2008; 128: E3-4

  View details for DOI 10.1038/skinbio.6250012

  View details for PubMedID 26794066

• Bridging structure with function: Structural, regulatory, and developmental role of laminins INTERNATIONAL JOURNAL OF BIOCHEMISTRY & CELL BIOLOGY Tzu, J., Marinkovich, M. P. 2008; 40 (2): 199-214

  Abstract

  The basement membrane is a highly intricate and organized portion of the extracellular matrix that interfaces with a variety of cell types including epithelial, endothelial, muscle, nerve, and fat cells. The laminin family of glycoproteins is a major constituent of the basement membrane. The 16 known laminin isoforms are formed from combinations of alpha, beta, and gamma chains, with each chain containing specific domains capable of interacting with cellular receptors such as integrins and other extracellular ligands. In addition to its role in the assembly and architectural integrity of the basement membrane, laminins interact with cells to influence proliferation, differentiation, adhesion, and migration, processes activated in normal and pathologic states. In vitro these functions are regulated by the post-translational modifications of the individual laminin chains. In vivo laminin knockout mouse studies have been particularly instructive in defining the function of specific laminins in mammalian development and have also highlighted its role as a key component of the basement membrane. In this review, we will define how laminin structure complements function and explore its role in both normal and pathologic processes.

  View details for DOI 10.1016/j.biocel.2007.07.015

  View details for PubMedID 17855154

• Laminin/Integrin expression profile in malignant melanoma Wong, K., Marinkovich, M. P., Horst, B. A. NATURE PUBLISHING GROUP. 2008: 103A–104A

  View details for Web of Science ID 000252180200469

• Laminin/integrin expression profile in malignant melanoma Wong, K., Marinkovich, M. P., Horst, B. A. NATURE PUBLISHING GROUP. 2008: 103A–104A

  View details for Web of Science ID 000252181100469

• Discovery of basement membrane components. journal of investigative dermatology Marinkovich, M. P. 2008; 128 (E2): E3-4

  View details for DOI 10.1038/skinbio.6250012

  View details for PubMedID 21233818

• A processed laminin-332 domain selectively localizes to and potentiates carcinoma development 37th Annual Meeting of the European-Society-for-Dermatological-Research Tran, M. M., Rousselle, P., Nokelainen, N. P., Talapragada, S., Nguyen, N., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2007: S90–S90

  View details for Web of Science ID 000249933400536

• What's new in blistering disorders? CURRENT ALLERGY AND ASTHMA REPORTS Chaudbari, P., Marinkovich, M. P. 2007; 7 (4): 255-263

  Abstract

  From the characterization of new animal models for the study of disease pathogenesis, to the demonstration of new therapeutic modalities, many developments have revolutionized the field of autoimmune bullous diseases in the past several years. This review highlights many of the significant advances that have taken place in the diagnosis, pathogenesis, and treatment options for pemphigus, pemphigoid, epidermolysis bullosa acquisita, and immunoglobulin (Ig) A-mediated bullous disorders.

  View details for PubMedID 17547846

• A laminin-collagen complex drives human epidermal carcinogenesis through phosphoinositol-3-kinase activation CANCER RESEARCH Waterman, E. A., Sakai, N., Nguyen, N. T., Horst, B. A., Veitch, D. P., Dey, C. N., Ortiz-Urda, S., Khavari, P. A., Marinkovich, M. P. 2007; 67 (9): 4264-4270

  Abstract

  Laminin-332 (formerly laminin-5) and collagen VII are basement membrane proteins expressed at the invasive front of human squamous cell carcinoma (SCC) tumors. These proteins have protumorigenic properties, but whether laminin-332 and collagen VII promote SCC tumors by providing adhesion or other nonadhesive extracellular cues, or whether laminin-332 and collagen VII interact together in this process remains unknown. In this study, we examined the role of these molecules by a structural approach using an in vivo model of human SCC tumorigenesis. Here, we show that individual domains (VI and V-III) on the laminin-332 beta3 chain provide distinct and highly divergent cell adhesion and tumor-promoting functions. We found that laminin beta3 domain VI provided a critical role in the assembly of stable adhesion complexes, but this domain was not required in SCC tumors. Instead, we found that laminin beta3 domain V-III played an essential role in SCC carcinogenesis/invasion through binding to collagen VII, which in turn, led to phosphoinositol-3-kinase activation and protection from apoptosis. Overexpression of constitutively active p110 phosphoinositol-3-kinase subunit was sufficient to restore invasion and tumorigenesis in transformed cells lacking laminin-332/collagen VII interaction in a manner independent of cellular adhesion. These studies show distinctive adhesive and signaling functions in individual domains of laminin-332, one which is required for normal epithelial adhesion and one which is required for SCC tumorigenesis. This uncoupling of stable adhesion from tumor progression in our studies suggests that laminin-332/collagen VII interaction promotes epidermal carcinogenesis through signaling rather than adhesion.

  View details for DOI 10.1158/0008-5472.CAN-06-4141

  View details for Web of Science ID 000246330300034

  View details for PubMedID 17483338

• The role of collagen XVII in ras-driven human epidermal tumorigenesis Makino, T., Ngyun, N. T., Chen, C. H., Shimizu, T., Rizzo, A. C., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2007: S24

  View details for Web of Science ID 000245387800143

• Critical domains of a6 beta 4 integrin for squamous cell carcinoma progression 96th Annual Meeting of the United-States-and-Canadian-Academy-of-Pathology Horst, B. A., Russell, A., Nguyen, N., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2007: 94A–94A

  View details for Web of Science ID 000244935300414

• Critical domains of a6 beta 4 integrin for squamous cell carcinoma progression Horst, B. A., Russell, A., Nguyen, N., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2007: 94A

  View details for Web of Science ID 000244922400414

• Integrin beta 4 regulates migratory behavior of keratinocytes by determining laminin-332 organization JOURNAL OF BIOLOGICAL CHEMISTRY Sehgal, B. U., DeBiase, P. J., Matzno, S., Chew, T., Claiborne, J. N., Hopkinson, S. B., Russell, A., Marinkovich, M. P., Jones, J. C. 2006; 281 (46): 35487-35498

  Abstract

  Whether alpha6beta4 integrin regulates migration remains controversial. beta4 integrin-deficient (JEB) keratinocytes display aberrant migration in that they move in circles, a behavior that mirrors the circular arrays of laminin (LM)-332 in their matrix. In contrast, wild-type keratinocytes and JEB keratinocytes, induced to express beta4 integrin, assemble laminin-332 in linear tracks over which they migrate. Moreover, laminin-332-dependent migration of JEB keratinocytes along linear tracks is restored when cells are plated on wild-type keratinocyte matrix, whereas wild-type keratinocytes show rotation over circular arrays of laminn-332 in JEB keratinocyte matrix. The activities of Rac1 and the actin cytoskeleton-severing protein cofilin are low in JEB keratinocytes compared with wild-type cells but are rescued following expression of wild-type beta4 integrin in JEB cells. Additionally, in wild-type keratinocytes Rac1 is complexed with alpha6beta4 integrin. Moreover, Rac1 or cofilin inactivation induces wild-type keratinocytes to move in circles over rings of laminin-332 in their matrix. Together these data indicate that laminin-332 matrix organization is determined by the alpha6beta4 integrin/actin cytoskeleton via Rac1/cofilin signaling. Furthermore, our results imply that the organizational state of laminin-332 is a key determinant of the motility behavior of keratinocytes, an essential element of skin wound healing and the successful invasion of epidermal-derived tumor cells.

  View details for DOI 10.1074/jbc.M606317200

  View details for Web of Science ID 000241933700082

  View details for PubMedID 16973601

  View details for PubMedCentralID PMC2820731

• beta 4 integrin and epidermal growth factor coordinately regulate electric field-mediated directional migration via Rac1 MOLECULAR BIOLOGY OF THE CELL Pullar, C. E., Baier, B. S., Kariya, Y., Russell, A. J., Horst, B. A., Marinkovich, M. P., Isseroff, R. R. 2006; 17 (11): 4925-4935

  Abstract

  Endogenous DC electric fields (EF) are present during embryogenesis and are generated in vivo upon wounding, providing guidance cues for directional cell migration (galvanotaxis) required in these processes. To understand the role of beta (beta)4 integrin in directional migration, the migratory paths of either primary human keratinocytes (NHK), beta4 integrin-null human keratinocytes (beta4-), or those in which beta4 integrin was reexpressed (beta4+), were tracked during exposure to EFs of physiological magnitude (100 mV/mm). Although the expression of beta4 integrin had no effect on the rate of cell movement, it was essential for directional (cathodal) migration in the absence of epidermal growth factor (EGF). The addition of EGF potentiated the directional response, suggesting that at least two distinct but synergistic signaling pathways coordinate galvanotaxis. Expression of either a ligand binding-defective beta4 (beta4+AD) or beta4 with a truncated cytoplasmic tail (beta4+CT) resulted in loss of directionality in the absence of EGF, whereas inhibition of Rac1 blinded the cells to the EF even in the presence of EGF. In summary, both the beta4 integrin ligand-binding and cytoplasmic domains together with EGF were required for the synergistic activation of a Rac-dependent signaling pathway that was essential for keratinocyte directional migration in response to a galvanotactic stimulus.

  View details for DOI 10.1091/mbc.E06-05-0433

  View details for Web of Science ID 000241993500030

  View details for PubMedID 16914518

  View details for PubMedCentralID PMC1635387

• Keratinocyte-secreted laminin 5 can function as a transient receptor for human papillomaviruses by binding virions and transferring them to adjacent cells JOURNAL OF VIROLOGY Culp, T. D., Budgeon, L. R., Marinkovich, M. P., Meneguzzi, G., Christensen, N. D. 2006; 80 (18): 8940-8950

  Abstract

  Human papillomaviruses (HPVs) replicate only in the terminally differentiating epithelium of the skin and mucosa. While infection of basal keratinocytes is considered a requirement for permissive infection, it remains unclear whether virions can specifically target basal cells for adsorption and uptake following epithelial wounding. We present evidence that HPV binds specifically to laminin 5 (LN5), a component of the extracellular matrix (ECM) secreted by migrating and basal keratinocytes. HPV type 11 capsids colocalized with LN5 in the ECM secreted by vaginal keratinocytes. Binding of both virions and virus-like particles to purified LN5 and to the LN5-rich ECM secreted by cultured keratinocytes was effectively blocked by pretreatment with anti-LN5 antibodies. HPV capsid binding to human cervical mucosa sections included the basement membrane which contains LN5. Cultured keratinocytes expressing alpha6 integrin, a transmembrane protein known to bind LN5, were readily infected by virions preadsorbed to LN5-containing substrates, whereas mutant keratinocytes lacking alpha6 integrin were relatively resistant to infection via this route. These findings suggest a model of natural HPV infection in which proliferating keratinocytes expressing alpha6 integrin at the site of epithelial wounding might be targeted by virions adsorbed transiently to LN5 secreted by migrating keratinocytes.

  View details for DOI 10.1128/JVI.00724-06

  View details for Web of Science ID 000240384800010

  View details for PubMedID 16940506

  View details for PubMedCentralID PMC1563898

• Laminin-5 alpha 3 G4-5 inhibition ablates epidermal tumorigenesis through PI3K-Akt pathway inactivation but does not disrupt normal epithelial cohesion 67th Annual Meeting of the Society-for-Investigative-Dermatology Tran, M. M., Rousselle, P., Nokelainen, P., Nguyen, N., Keene, D. R., Fincher, E. F., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2006: 24–24

  View details for Web of Science ID 000242891500140

• Laminin-5 Beta 3 chain promotes epidermal carcinogenesis through type VII collagen binding and pi3k activation Sakai, N., Waterman, E. A., Nguyen, N. E., Horst, B. A., Veitch, D. P., Dey, C. N., Ortiz-Urda, S., Khavari, P. A., Marinkovich, M. NATURE PUBLISHING GROUP. 2006: 24

  View details for Web of Science ID 000242891500143

• Overexpression of laminin-8 in human dermal microvascular endothelial cells promotes angiogenesis-related functions JOURNAL OF INVESTIGATIVE DERMATOLOGY Li, J., Zhou, L., Tran, H. T., Chen, Y., Nguyen, N. E., Karasek, M. A., Marinkovich, M. P. 2006; 126 (2): 432-440

  Abstract

  This study examined the effects of endogenous overexpression of laminin-8 on angiogenesis and wound healing in primary human dermal microvascular endothelial cells (HDMECs). HDMECs expressed laminin-8 and laminin-10, but no other laminins, as determined by radioimmunoprecipitation assay using a panel of antibodies to individual laminin chains. To study laminin-8 function, full-length human laminin alpha4 cDNA was retrovirally transferred to HDMEC, and specific overexpression of laminin-8 was verified by Western blot. Laminin-8 overexpression promoted endothelial cell spreading and migration in scratch assays and accelerated angiogenic tubule formation in collagen gel overlay assays. Strong inhibitory effect of beta1 integrin and weak inhibition by alphavbeta3 integrin antibodies were observed in laminin-8-stimulated cell migration, but only beta1 integrin antibodies affected tubule formation. These studies suggest that laminin-8 overexpression may prove to be a useful method to engineer HDMECs to promote angiogenesis and wound repair.

  View details for DOI 10.1038/sj.jid.5700089

  View details for PubMedID 16374451

• A simplified laminin nomenclature MATRIX BIOLOGY Aumailley, M., Bruckner-Tuderman, L., Carter, W. G., Deutzmann, R., Edgar, D., Ekblom, P., Engel, J., ENGVALL, E., Hohenester, E., Jones, J. C., Kleinman, H. K., Marinkovich, M. P., Martin, G. R., Mayer, U., Meneguzzi, G., Miner, J. H., Miyazaki, K., PATARROYO, M., Paulsson, M., Quaranta, V., Sanes, J. R., Sasaki, T., Sekiguchi, K., Sorokin, L. M., Talts, J. F., Tryggvason, K., UITTO, J., VIRTANEN, I., von der Mark, K., Wewer, U. M., Yamada, Y., Yurchenco, P. D. 2005; 24 (5): 326-332

  Abstract

  A simplification of the laminin nomenclature is presented. Laminins are multidomain heterotrimers composed of alpha, beta and gamma chains. Previously, laminin trimers were numbered with Arabic numerals in the order discovered, that is laminins-1 to -5. We introduce a new identification system for a trimer using three Arabic numerals, based on the alpha, beta and gamma chain numbers. For example, the laminin with the chain composition alpha5beta1gamma1 is termed laminin-511, and not laminin-10. The current practice is also to mix two overlapping domain and module nomenclatures. Instead of the older Roman numeral nomenclature and mixed nomenclature, all modules are now called domains. Some domains are renamed or renumbered. Laminin epidermal growth factor-like (LE) domains are renumbered starting at the N-termini, to be consistent with general protein nomenclature. Domain IVb of alpha chains is named laminin 4a (L4a), domain IVa of alpha chains is named L4b, domain IV of gamma chains is named L4, and domain IV of beta chains is named laminin four (LF). The two coiled-coil domains I and II are now considered one laminin coiled-coil domain (LCC). The interruption in the coiled-coil of beta chains is named laminin beta-knob (Lbeta) domain. The chain origin of a domain is specified by the chain nomenclature, such as alpha1L4a. The abbreviation LM is suggested for laminin. Otherwise, the nomenclature remains unaltered.

  View details for DOI 10.1016/j.matbio.2005.05.006

  View details for Web of Science ID 000231205300002

  View details for PubMedID 15979864

• Type VII collagen is required for cellular invasiveness in epidermal carcinogenesis 66th Annual Meeting of the Society-for-Investigative-Dermatology Ortiz-Urda, S., Garcia, J., Marinkovich, M., Khavari, P. NATURE PUBLISHING GROUP. 2005: A25–A25

  View details for Web of Science ID 000228179900149

• Laminin 10 in the angiogenesis and invasion of squamous cell carcinoma Li, J., Zhang, Y., Romagosa, R., Saghari, S., Elgart, G., Miner, J. H., Marinkovich, M. P., Nouri, K. BLACKWELL PUBLISHING INC. 2005: A21

  View details for Web of Science ID 000228179900124

• Two distinct roles for the laminin-5 beta 3 chain in epidermal carcinogenesis and adhesion 66th Annual Meeting of the Society-for-Investigative-Dermatology Waterman, E. A., Ortiz-Urda, S., Nguyen, N. T., Veitch, D. P., Horst, B. A., Dey, C. N., Khavari, P. A., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2005: A22–A22

  View details for Web of Science ID 000228179900128

• Type VII collagen is required for Ras-driven human epidermal tumorigenesis SCIENCE Ortiz-Urda, S., Garcia, J., Green, C. L., Chen, L., Lin, Q., Veitch, D. P., Sakai, L. Y., LEE, H., Marinkovich, M. P., Khavari, P. A. 2005; 307 (5716): 1773-1776

  Abstract

  Type VII collagen defects cause recessive dystrophic epidermolysis bullosa (RDEB), a blistering skin disorder often accompanied by epidermal cancers. To study the role of collagen VII in these cancers, we examined Ras-driven tumorigenesis in RDEB keratinocytes. Cells devoid of collagen VII did not form tumors in mice, whereas those retaining a specific collagen VII fragment (the amino-terminal noncollagenous domain NC1) were tumorigenic. Forced NC1 expression restored tumorigenicity to collagen VII-null epidermis in a non-cell-autonomous fashion. Fibronectin-like sequences within NC1 (FNC1) promoted tumor cell invasion in a laminin 5-dependent manner and were required for tumorigenesis. Tumor-stroma interactions mediated by collagen VII thus promote neoplasia, and retention of NC1 sequences in a subset of RDEB patients may contribute to their increased susceptibility to squamous cell carcinoma.

  View details for DOI 10.1126/science.1106209

  View details for Web of Science ID 000227883900044

  View details for PubMedID 15774758

• Advances in inherited epidermolysis bullosa. Advances in dermatology McAllister, J. C., Peter Marinkovich, M. 2005; 21: 303-334

  View details for PubMedID 16350448

• Involvement of p53 and p16 tumor suppressor genes in recessive dystrophic epidermolysis bullosa-associated squamous cell carcinoma JOURNAL OF INVESTIGATIVE DERMATOLOGY Arbiser, J. L., Fan, C. Y., Su, X. B., van Emburgh, B. O., Cerimele, F., Miller, M. S., Harvell, J., Marinkovich, M. P. 2004; 123 (4): 788-790

  Abstract

  Recessive dystrophic epidermolysis bullosa (RDEB) is an autosomal recessive disorder characterized by the loss of collagen type VII, an intrinsic component of the anchoring fibrils, which attach the epidermis to the dermis. Of the genetic blistering disorders, RDEB has the highest rate of morbidity and mortality, with morbidity arising from fusion of digits in a mitten-glove deformity and growth retardation associated with anemia. The leading cause of death in RDEB is cutaneous squamous cell carcinoma, which causes death through invasion and metastasis. In order to better understand the pathogenesis of these rare but aggressive squamous cell carcinoma (SCC), we analyzed them for mutations in p53 and loss of p16ink4a. Three tumors demonstrated mutations in the p53 tumor suppressor gene. We also analyzed SCC from patients with RDEB for the presence of p16ink4a hypermethylation, and found two tumors that have loss of p16ink4a through hypermethylation. This is the first description of specific abnormalities in tumor suppressor genes in RDEB associated SCC, and demonstrates that alterations in both p53 and p16ink4a can contribute to RDEB associated SCC.

  View details for DOI 10.1111/j.0022-202X.2004.23418.x

  View details for Web of Science ID 000223982900031

  View details for PubMedID 15373786

• Identification of critical domains of beta 4 integrin and laminin-5 required for human SCC development 65th Annual Meeting of the Society-for-Investigative-Dermatology Horst, B. A., Nokelainen, P., Fincher, E. F., Nguyen, N. T., Russell, A. J., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2004: A19–A19

  View details for Web of Science ID 000220660500167

• Role of laminin-10 in hair development Gao, J., Chen, C., Nguyen, N. T., Leopold, P. L., Crystal, R. G., Miner, J. H., Oro, A. E., Li, J., Marinkovich, M. P. BLACKWELL PUBLISHING INC. 2004: A112

  View details for Web of Science ID 000220660500720

• Laminins 8 and 10 in skin basement membrane reconstitution and wound healing 65th Annual Meeting of the Society-for-Investigative-Dermatology Li, J., Zhu, L., Kirsner, R. S., Davis, S. C., Mertz, P. M., Eaglstein, W. H., Miner, J. H., Marinkovich, M. P., Zhang, Y. NATURE PUBLISHING GROUP. 2004: A31–A31

  View details for Web of Science ID 000220660500237

• Kinetics and specificity of Fas ligand induction in toxic epidermal necrolysis ARCHIVES OF DERMATOLOGY Chang, H. Y., Cooper, Z. A., Swetter, S. A., Marinkovich, M. P. 2004; 140 (2): 242-244

  View details for Web of Science ID 000188877100024

  View details for PubMedID 14967808

• Mature human thymocytes migrate on laminin-5 with activation of metalloproteinase-14 and cleavage of CD44 JOURNAL OF IMMUNOLOGY Vivinus-Nebot, M., Rousselle, P., Breittmayer, J. P., Cenciarini, C., Berrih-Aknin, S., Spong, S., Nokelainen, P., Cottrez, F., Marinkovich, M. P., Bernard, A. 2004; 172 (3): 1397-1406

  Abstract

  We have previously shown that laminin-5 is expressed in the human thymic medulla, in which mature thymocytes are located. We now report that laminin-5 promotes migration of mature medullary thymocytes, whereas it has no effect on cortical immature thymocytes. Migration was inhibited by blocking mAbs directed against laminin-5 integrin receptors and by inhibitors of metalloproteinases. Interactions of thymocytes with laminin-5 induced a strong up-regulation of active metalloproteinase-14. However, we found that thymocytes did not cleave the laminin-5 gamma(2) chain, suggesting that they do not use the same pathway as epithelial cells to migrate on laminin-5. Interactions of thymocytes with laminin-5 also induced the release of a soluble fragment of CD44 cell surface molecule. Moreover, CD44-rich supernatants induced thymocyte migration in contrast with supernatants depleted in CD44 by immunoadsorption. CD44 cleavage was recently reported to be due to metalloproteinase-14 activation and led to increased migration in cancer cells. Thus, in this study, we show that laminin-5 promotes human mature thymocyte migration in vitro via a multimolecular mechanism involving laminin-5 integrin receptors, metalloproteinase-14 and CD44. These data suggest that, in vivo, laminin-5 may function in the migration of mature thymocytes within the medulla and be part of the thymic emigration process.

  View details for Web of Science ID 000188378700009

  View details for PubMedID 14734715

• Autocrine laminin-5 ligates alpha 6 beta 4 integrin and activates RAC and NF kappa B to mediate anchorage-independent survival of mammary tumors JOURNAL OF CELL BIOLOGY Zahir, N., Lakins, J. N., Russell, A., Ming, W. Y., Chatterjee, C., Rozenberg, G. I., Marinkovich, M. P., Weaver, V. M. 2003; 163 (6): 1397-1407

  Abstract

  Invasive carcinomas survive and evade apoptosis despite the absence of an exogenous basement membrane. How epithelial tumors acquire anchorage independence for survival remains poorly defined. Epithelial tumors often secrete abundant amounts of the extracellular matrix protein laminin 5 (LM-5) and frequently express alpha6beta4 integrin. Here, we show that autocrine LM-5 mediates anchorage-independent survival in breast tumors through ligation of a wild-type, but not a cytoplasmic tail-truncated alpha6beta4 integrin. alpha6beta4 integrin does not mediate tumor survival through activation of ERK or AKT. Instead, the cytoplasmic tail of beta4 integrin is necessary for basal and epidermal growth factor-induced RAC activity, and RAC mediates tumor survival. Indeed, a constitutively active RAC sustains the viability of mammary tumors lacking functional beta1 and beta4 integrin through activation of NFkappaB, and overexpression of NFkappaB p65 mediates anchorage-independent survival of nonmalignant mammary epithelial cells. Therefore, epithelial tumors could survive in the absence of exogenous basement membrane through autocrine LM-5-alpha6beta4 integrin-RAC-NFkappaB signaling.

  View details for DOI 10.1083/jcb.200302023

  View details for Web of Science ID 000187583500021

  View details for PubMedID 14691145

  View details for PubMedCentralID PMC2173718

• alpha 6 beta 4 integrin regulates keratinocyte chemotaxis through differential GTPase activation and antagonism of alpha 3 beta 1 integrin JOURNAL OF CELL SCIENCE Russell, A. J., Fincher, E. F., Millman, L., Smith, R., Vela, V., Waterman, E. A., Dey, C. N., Guide, S., Weaver, V. M., Marinkovich, M. P. 2003; 116 (17): 3543-3556

  Abstract

  Growth factor-induced cell migration and proliferation are essential for epithelial wound repair. Cell migration during wound repair also depends upon expression of laminin-5, a ligand for alpha 6 beta 4 integrin. We investigated the role of alpha 6 beta 4 integrin in laminin-5-dependent keratinocyte migration by re-expressing normal or attachment-defective beta 4 integrin in beta 4 integrin null keratinocytes. We found that expression of beta 4 integrin in either a ligand bound or ligand unbound state was necessary and sufficient for EGF-induced cell migration. In a ligand bound state, beta 4 integrin supported EGF-induced cell migration though sustained activation of Rac1. In the absence of alpha 6 beta 4 integrin ligation, Rac1 activation became tempered and EGF chemotaxis proceeded through an alternate mechanism that depended upon alpha 3 beta 1 integrin and was characterized by cell scattering. alpha 3 beta 1 integrin also relocalated from cell-cell contacts to sites of basal clustering where it displayed increased conformational activation. The aberrant distribution and activation of alpha 3 beta 1 integrin in attachment-defective beta 4 cells could be reversed by the activation of Rac1. Conversely, in WT beta 4 cells the normal cell-cell localization of alpha 3 beta 1 integrin became aberrant after the inhibition of Rac1. These studies indicate that the extracellular domain of beta 4 integrin, through its ability to bind ligand, functions to integrate the divergent effects of growth factors on the cytoskeleton and adhesion receptors so that coordinated keratinocyte migration can be achieved.

  View details for DOI 10.1242/jcs.00663

  View details for PubMedID 12865436

• Topical tacrolimus is a useful adjunctive therapy for bullous pemphigoid ARCHIVES OF DERMATOLOGY Chu, J., Bradley, M., Marinkovich, M. P. 2003; 139 (6): 813-815

  View details for Web of Science ID 000183478000027

  View details for PubMedID 12810523

• Laminin-10 is crucial for hair morphogenesis EMBO JOURNAL Li, J., Tzu, J., Chen, Y., Zhang, Y. P., Nguyen, N. T., Gao, J., Bradley, M., Keene, D. R., Oro, A. E., Miner, J. H., Marinkovich, M. P. 2003; 22 (10): 2400-2410

  Abstract

  The role of the extracellular matrix in cutaneous morphogenesis is poorly understood. Here, we describe the essential role of laminin-10 (alpha5beta1gamma1) in hair follicle development. Laminin-10 was present in the basement membrane of elongating hair germs, when other laminins were downregulated, suggesting a role for laminin-10 in hair development. Treatment of human scalp xenografts with antibodies to laminin-10, or its receptor beta1 integrin, produced alopecia. E16.5 Lama5 -/- mouse skin, lacking laminin-10, contained fewer hair germs compared with controls, and after transplantation, Lama5 -/- skin showed a failure of hair germ elongation followed by complete hair follicle regression. Lama5 -/- skin showed defective basement membrane assembly, without measurable increases in anoikis. Instead, Lama5 -/- skin showed decreased expression of early hair markers including sonic hedgehog and Gli1, implicating laminin-10 in developmental signaling. Intriguingly, treatment of Lama5 -/- skin with purified laminin-10 corrected basement membrane defects and restored hair follicle development. We conclude that laminin-10 is required for hair follicle development and report the first use of exogenous protein to correct a cutaneous developmental defect.

  View details for PubMedID 12743034

• Mammalian tolloid metalloproteinase, and not matrix metalloprotease 2 or membrane type 1 metalloprotease, processes laminin-5 in keratinocytes and skin JOURNAL OF BIOLOGICAL CHEMISTRY Veitch, D. P., Nokelainen, P., McGowan, K. A., Nguyen, T. T., Nguyen, N. E., Stephenson, R., Pappano, W. N., Keene, D. R., Spong, S. M., Greenspan, D. S., Findell, P. R., Marinkovich, P. M. 2003; 278 (18): 15661-15668

  Abstract

  Laminin-5, a major adhesive ligand for epithelial cells, undergoes processing of its gamma2 and alpha3 chains. This study investigated the mechanism of laminin-5 processing by keratinocytes. BI-1 (BMP-1 isoenzyme inhibitor-1), a selective inhibitor of a small group of astacin-like metalloproteinases, which includes bone morphogenetic protein 1 (BMP-1), mammalian Tolloid (mTLD), mammalian Tolloid-like 1 (mTLL-1), and mammalian Tolloid-like 2 (mTLL-2), inhibited the processing of laminin-5 gamma2 and alpha3 chains in keratinocyte cultures in a dose-dependent manner. In a proteinase survey, all BMP-1 isoenzymes processed human laminin-5 gamma2 and alpha3 chains to 105- and 165-kDa fragments, respectively. In contrast, MT1-MMP and MMP-2 did not cleave the gamma2 chain of human laminin-5 but processed the rat laminin gamma2 chain to an 80-kDa fragment. An immunoblot and quantitative PCR survey of the BMP-1 isoenzymes revealed expression of mTLD in primary keratinocyte cultures but little or no expression of BMP-1, mTLL-1, or mTLL-2. mTLD was shown to cleave the gamma2 chain at the same site as the previously identified BMP-1 cleavage site. In addition, mTLD/BMP-1 null mice were shown to have deficient laminin-5 processing. Together, these data identify laminin-5 as a substrate for mTLD, suggesting a role for laminin-5 processing by mTLD in the skin.

  View details for DOI 10.1074/jbc.M210588200

  View details for PubMedID 12473650

• NF-kappa B blockade and oncogenic Ras trigger invasive human epidermal neoplasia NATURE Dajee, M., Lazarov, M., Zhang, J. Y., Cai, T., Green, C. L., Russell, A. J., Marinkovich, M. P., Tao, S. Y., Lin, Q., Kubo, Y., Khavari, P. A. 2003; 421 (6923): 639-643

  Abstract

  The nuclear factor NF-kappaB and oncogenic Ras can alter proliferation in epidermis, the most common site of human cancer. These proteins are implicated in epidermal squamous cell carcinoma in mice, however, the potential effects of altering their function are uncertain. Whereas inhibition of NF-kappaB enhances apoptosis in certain tumours, blockade of NF-kappaB predisposes murine skin to squamous cell carcinoma. Because therapeutics inhibiting Ras and NF-kappaB pathways are being developed to treat human cancer, it is essential to assess the effects of altering these regulators. The medical relevance of murine studies is limited, however, by differences between mouse and human skin, and by the greater ease of transforming murine cells. Here we show that in normal human epidermal cells both NF-kappaB and oncogenic Ras trigger cell-cycle arrest. Growth arrest triggered by oncogenic Ras can be bypassed by IkappaBalpha-mediated blockade of NF-kappaB, generating malignant human epidermal tissue resembling squamous cell carcinoma. Human cell tumorigenesis is dependent on laminin 5 and alpha6beta4 integrin. Thus, IkappaBalpha circumvents restraints on growth promotion induced by oncogenic Ras and can act with Ras to induce invasive human tissue neoplasia.

  View details for DOI 10.1038/nature01283

  View details for PubMedID 12571598

• Injection of genetically engineered fibroblasts corrects regenerated human epidermolysis bullosa skin tissue JOURNAL OF CLINICAL INVESTIGATION Ortiz-Urda, S., Lin, Q., Green, C. L., Keene, D. R., Marinkovich, M. P., Khavari, P. A. 2003; 111 (2): 251-255

  Abstract

  Current therapeutic strategies for genetic skin disorders rely on the complex process of grafting genetically engineered tissue to recipient wound beds. Because fibroblasts synthesize and secrete extracellular matrix, we explored their utility in recessive dystrophic epidermolysis bullosa (RDEB), a blistering disease due to defective extracellular type VII collagen. Intradermal injection of RDEB fibroblasts overexpressing type VII collagen into intact RDEB skin stably restored correctly localized type VII collagen expression in vivo and normalized hallmark RDEB disease features, including subepidermal blistering and anchoring fibril defects. This article was published online in advance of the print edition. The date of publication is available from the JCI website, http://www.jci.org.

  View details for DOI 10.1172/JCI.200317193

  View details for Web of Science ID 000180672900016

  View details for PubMedID 12531881

  View details for PubMedCentralID PMC151880

• Activation of the small GTPase Rac1 alters localization of integrin alpha 3 beta 1 leading to disruption of Hemidesmosome formation Chu, J., Russell, A. J., Marinkovich, M. P. NATURE PUBLISHING GROUP. 2002: 229–29

  View details for Web of Science ID 000177428100130

• Laminin 10 in wound re-epithelialization and angiogenesis Li, J., Kirsner, R. S., Marinkovich, M. P., Miner, J. H., Zhang, Y. BLACKWELL PUBLISHING INC. 2002: 233

  View details for Web of Science ID 000177428100156

• Laminin 10 is essential for hair development Tzu, J. E., Li, J., Lehman, D., Chen, Y., Nguyen, N. T., Keene, D. R., Miner, J. H., Oro, A. E., Marinkovich, M. NATURE PUBLISHING GROUP. 2002: 291–91

  View details for Web of Science ID 000177428100503

• The first international consensus on mucous membrane pemphigoid - Definition, diagnostic criteria, pathogenic factors, medical treatment, and prognostic indicators ARCHIVES OF DERMATOLOGY Chan, L. S., Ahmed, A. R., ANHALT, G. J., Bernauer, W., Cooper, K. D., Elder, M. J., Fine, J. D., Foster, S., Ghohestani, R., Hashimoto, T., Hoang-Xuan, T., Kirtschig, G., Korman, N. J., Lightman, S., Lozado-Nur, F., Marinkovich, M. P., Mondino, B. J., Prost-Squarcioni, C., Rogers, R. S., Setterfield, J. F., West, D. P., Wojnarowska, F., Woodley, D. T., Yancey, K. B., Zillikens, D., Zone, J. J. 2002; 138 (3): 370-379

  Abstract

  We aimed to develop consensus-based recommendations for streamlining medical communication among various health care professionals, to improve accuracy of diagnosis and treatment, and to facilitate future investigations for mucous membrane pemphigoid.Because of the highly specific nature of this group of diseases, the 26 invited participants included either international scholars in the field of mucous membrane pemphigoid or experts in cutaneous pharmacology representing the 3 medical disciplines ophthalmology, oral medicine, and dermatology.The first author (L.S.C.) conducted a literature search. Based on the information obtained, international experts who had contributed to the literature in the clinical care, diagnosis, and laboratory investigation for mucous membrane pemphigoid were invited to participate in a consensus meeting aimed at developing a consensus statement.A consensus meeting was convened and conducted on May 10, 1999, in Chicago, Ill, to discuss the relevant issues. The first author drafted the statement based on the consensus developed at the meeting and the participants' written comments. The draft was submitted to all participants for 3 separate rounds of review, and disagreements were reconciled based on literature evidence. The third and final statement incorporated all relevant evidence obtained in the literature search and the consensus developed by the participants. The final statement was approved and endorsed by all 26 participants.Specific consensus-based recommendations were made regarding the definition, diagnostic criteria, pathogenic factors, medical treatment, and prognostic indicators for mucous membrane pemphigoid. A system of standard reporting for these patients was proposed to facilitate a uniform data collection.

  View details for Web of Science ID 000174367300010

  View details for PubMedID 11902988

• Epidermolysis bullosa: new and emerging trends. American journal of clinical dermatology Pai, S., Marinkovich, M. P. 2002; 3 (6): 371-380

  Abstract

  Epidermolysis bullosa is a family of inherited blistering skin disorders characterized by blister formation in response to mechanical trauma. Major types of epidermolysis bullosa include epidermolysis bullosa simplex, hemidesmosomal epidermolysis bullosa, junctional epidermolysis bullosa, and dystrophic epidermolysis bullosa. Current treatment for epidermolysis bullosa consists of supportive care for skin and other organ systems and entails a combination of wound management, infection support for chronic wounds, surgical management as needed, nutritional support, and preventative screening for squamous cell carcinoma in recessive dystrophic epidermolysis bullosa. The regimen must be tailored specifically to the severity and extent of skin and systemic involvement in each case. Recent studies have identified specific protein and genetic abnormalities for most epidermolysis bullosa subtypes. These new advancements in the understanding of molecular pathophysiology have provided much of the basis for current efforts to develop effective gene and protein therapy for epidermolysis bullosa.

  View details for PubMedID 12113646

• Collagen XVII (BP180, BPAG2) is the most common epidermal basement membrane autoantigen in humans and other animals 4th World Conference of Veterinary Dermatology Olivry, T., Marinkovich, M. P. BLACKWELL SCIENCE PUBL. 2002: 20–29

  View details for Web of Science ID 000180110700002

• Linear IgA bullous dermatosis CLINICS IN DERMATOLOGY Guide, S. V., Marinkovich, M. P. 2001; 19 (6): 719-727

  View details for PubMedID 11705681

• Multiple functions for beta 4 integrin in human microvascular endothelial cells Marinkovich, M., Zhou, L., Russell, A., Li, J., Karasek, M. NATURE PUBLISHING GROUP. 2001: 391–91

  View details for Web of Science ID 000170668300046

• Integrin alpha 6 beta 4 ligation controls keratinocyte morphology and chemotaxis through opposing stimulation of the small GTPases, Rac and Rho Russell, A., Fincher, E., Vela, Smith, R., Marinkovich, M. BLACKWELL SCIENCE INC. 2001: 477

  View details for Web of Science ID 000170668300558

• Role of tyrosine kinase in endothelial cell migration and angiogenesis Zhou, L., Marinkovich, M., Karasek, M. BLACKWELL SCIENCE INC. 2001: 392

  View details for Web of Science ID 000170668300047

• Laminin 10 plays a critical role in the development of normal skin and hair follicles Li, J., Chen, Y., Miner, J., Marinkovich, M. BLACKWELL SCIENCE INC. 2001: 426

  View details for Web of Science ID 000170668300253

• Bone morphogenic protein-1 inhibitors block human squamous cell carcinoma invasion in vitro Pai, S., Veitch, D., Ho, W., Fincher, E., Russell, A., Findell, P., Marinkovich, M. BLACKWELL SCIENCE INC. 2001: 398

  View details for Web of Science ID 000170668300083

• A spontaneous canine model of mucous membrane (cicatricial) pemphigoid, an autoimmune blistering disease affecting mucosae and mucocutaneous junctions JOURNAL OF AUTOIMMUNITY Olivry, T., Dunston, S. M., Schachter, M., Xu, L. T., Nguyen, N., Marinkovich, M. P., Chan, L. S. 2001; 16 (4): 411-421

  Abstract

  Mucous membrane pemphigoid (MMP) is a rare autoimmune blistering dermatosis of humans that was previously known as cicatricial pemphigoid. It is characterized by vesicles, ulcers and scarring that affect predominantly mucosae and mucocutaneous junctions. Circulating autoantibodies recognize epitopes on basement membrane proteins such as collagen XVII or laminin-5/6. Herein, we describe the clinico-pathological and immunological characteristics of 17 dogs afflicted with a dermatosis homologous to MMP of humans. Patients exhibited vesicles and erosions predominantly on mucous membranes or mucocutaneous junctions of the mouth, nose, eyes, genitalia or anus. Histopathology revealed subepithelial vesicles with variable dermal inflammation. Direct immunofluorescence demonstrated IgG or complement at the dermoepithelial junction. Indirect immunofluorescence using salt-split epithelia permitted the detection of circulating basement membrane-specific IgG autoantibodies in 15 cases. In 11 patients, autoantibodies recognized the NC16A segment of collagen XVII, as determined by salt-split indirect immunofluorescence, immunoblotting using canine keratinocytes and ELISA with synthetic canine peptides. In one dog, autoantiodies bound to the dermal side of salt-split epithelia and recognized epitopes within the 30 kDa carboxy-terminal segment of human collagen XVII. Canine MMP, like its human counterpart, exhibits distinctive clinical signs and histopathological lesions, yet circulating autoantibodies target different antigenic epitopes. This spontaneous canine model of MMP could prove useful for studies on the pathogenesis or therapy of this human disease.

  View details for DOI 10.1006/jaut.2001.0510

  View details for Web of Science ID 000169515500005

  View details for PubMedID 11437489

• IgG anti-LABD97 antibodies in bullous pemphigoid patients' sera react with the mid-portion of the BPAg2 ectodomain JOURNAL OF INVESTIGATIVE DERMATOLOGY Egan, C. A., Reddy, E. D., Nie, Z. X., Taylor, T. B., Schmidt, L. A., Meyer, L. J., Petersen, M. J., Hashimoto, T., Marinkovich, M. P., Zone, J. J. 2001; 116 (2): 348-350

  View details for Web of Science ID 000167170700021

  View details for PubMedID 11180014

• Properties of the collagen type XVII ectodomain - Evidence for N- to C-terminal triple helix folding JOURNAL OF BIOLOGICAL CHEMISTRY Areida, S. K., Reinhardt, D. P., Muller, P. K., FIETZEK, P. P., Kowitz, J., Marinkovich, M. P., Notbohm, H. 2001; 276 (2): 1594-1601

  Abstract

  Collagen XVII is a transmembrane component of hemidesmosomal cells with important functions in epithelial-basement membrane interactions. Here we report on properties of the extracellular ectodomain of collagen XVII, which harbors multiple collagenous stretches. We have recombinantly produced subdomains of the collagen XVII ectodomain in a mammalian expression system. rColXVII-A spans the entire ectodomain from deltaNC16a to NC1, rColXVII-B is similar but lacks the NC1 domain, a small N-terminal polypeptide rColXVII-C encompasses domains deltaNC16a to C15, and a small C-terminal polypeptide rColXVII-D comprises domains NC6 to NC1. Amino acid analysis of rColXVII-A and -C demonstrated prolyl and lysyl hydroxylation with ratios for hydroxyproline/proline of 0.4 and for hydroxylysine/lysine of 0.5. A small proportion of the hydroxylysyl residues in rColXVII-C ( approximately 3.3%) was glycosylated. Limited pepsin and trypsin degradation assays and analyses of circular dichroism spectra clearly demonstrated a triple-helical conformation for rColXVII-A, -B, and -C, whereas the C-terminal rColXVII-D did not adopt a triple-helical fold. These results were further substantiated by electron microscope analyses, which revealed extended molecules for rColXVII-A and -C, whereas rColXVII-D appeared globular. Thermal denaturation experiments revealed melting temperatures of 41 degrees C (rColXVII-A), 39 degrees C (rColXVII-B), and 35 degrees C (rColXVII-C). In summary, our data suggest that triple helix formation in the ectodomain of ColXVII occurs with an N- to C-terminal directionality.

  View details for Web of Science ID 000166430900098

  View details for PubMedID 11042218

• Processing of laminin-5 in keratinocyte migration Veitch, D. P., McGowan, K., Findell, P., Sharma, P., Marinkovich, M. P. AMER SOC CELL BIOLOGY. 2000: 47A–47A

  View details for Web of Science ID 000165525900245

• Phenotypic reversion of alpha 3-deficient human keratinocytes and functional studies of laminin 5 Fincher, E. F., Russell, A. J., Marinkovich, M. P. AMER SOC CELL BIOLOGY. 2000: 391A–392A

  View details for Web of Science ID 000165525902037

• Laminins and human disease MICROSCOPY RESEARCH AND TECHNIQUE McGowan, K. A., Marinkovich, M. P. 2000; 51 (3): 262-279

  Abstract

  The laminin protein family has diverse tissue expression patterns and is involved in the pathology of a number of organs, including skin, muscle, and nerve. In the skin, laminins 5 and 6 contribute to dermal-epidermal cohesion, and mutations in the constituent chains result in the blistering phenotype observed in patients with junctional epidermolysis bullosa (JEB). Allelic heterogeneity is observed in patients with JEB: mutations that results in premature stop codons produce a more severe phenotype than do missense mutations. Gene therapy approaches are currently being studied in the treatment of this disease. A blistering phenotype is also observed in patients with acquired cicatricial pemphigoid (CP). Autoantibodies targeted against laminins 5 and 6 destabilize epithelial adhesion and are pathogenic. In muscle cells, laminin alpha 2 is a component of the bridge that links the actin cytoskeleton to the extracellular matrix. In patients with laminin alpha 2 mutations, the bridge is disrupted and mature muscle cells apoptose. Congenital muscular dystrophy (CMD) results. The role of laminin in diseases of the nervous system is less well defined, but the extracellular protein has been shown to serve an important role in peripheral nerve regeneration. The adhesive molecule influences neurite outgrowth, neural differentiation, and synapse formation. The broad spatial distribution of laminin gene products suggests that laminin may be involved in a number of diseases for which pathogenic mechanisms are still being unraveled.

  View details for PubMedID 11054876

• Autoantibodies to BP180 associated with bullous pemphigoid release interleukin-6 and interleukin-8 from cultured human keratinocytes JOURNAL OF INVESTIGATIVE DERMATOLOGY Schmidt, E., Reimer, S., Kruse, N., Jainta, S., Brocker, E. B., Marinkovich, M. P., Giudice, G. J., Zillikens, D. 2000; 115 (5): 842-848

  Abstract

  Bullous pemphigoid is an inflammatory subepidermal blistering disease that is associated with auto- antibodies to the keratinocyte surface protein, BP180. In addition to the binding of autoantibodies, the infiltration of inflammatory cells is necessary for blister formation. Cytokines, including interleukin-6 and interleukin-8, have been implicated in the disease process of both human and experimental murine bullous pemphigoid. This study was aimed at testing the hypothesis that the binding of anti-BP180 antibodies to their target antigen triggers a signal transduction event that results in the secretion of these pro-inflammatory cytokines. Consistent with this hypothesis, treatment of cultured normal human epidermal keratinocytes with bullous pemphigoid IgG, but not control IgG, led to increased levels of interleukin-6 and interleukin-8, but not interleukin-1alpha, interleukin-1beta, tumor necrosis factor-alpha, interleukin-10, or monocyte chemoattractant protein-1, in the culture medium. This effect was concentration- and time-dependent and was abolished by depleting the bullous pemphigoid IgG of reactivity to two distinct epitopes on the BP180 NC16A domain. Upregulation of interleukin-6 and interleukin-8 was found at both protein and mRNA levels. In addition, bullous pemphigoid IgG did not induce the release of interleukin-6 and interleukin-8 from BP180-deficient keratinocytes obtained from a patient with generalized atrophic benign epidermolysis bullosa. These data indicate that bullous pemphigoid-associated autoantibodies to the human BP180 ectodomain trigger a signal transducing event that leads to expression and secretion of interleukin-6 and interleukin-8 from human keratinocytes.

  View details for Web of Science ID 000165219000010

  View details for PubMedID 11069622

• Autoantibodies against the processed ectodomain of collagen XVII (BPAG2, BP180) define a canine homologue of linear IgA disease of humans VETERINARY PATHOLOGY Olivry, T., Dunston, S. M., Fahey, M., Nguyen, N., Marinkovich, M. P. 2000; 37 (4): 302-309

  Abstract

  Linear IgA disease (LAD) is an acquired autoimmune subepidermal blistering dermatosis that affects human children and adults. In contrast to bullous pemphigoid, in which autoantibodies recognize transmembrane type XVII collagen (BP180, BPAG2), LAD is associated with skin-fixed and circulating IgA autoantibodies that target LAD-1, the processed extracellular form of type XVII collagen. An immunologic homologue of LAD in humans was identified in two dogs according to the following criteria: 1) erosive, ulcerative, and crusted lesions seen on the face, in the oral cavity, and on the extremities, 2) dermoepidermal clefting present in the basement membrane lamina lucida without inflammation or with mild neutrophilic infiltration, 3) basement membrane-fixed IgG and/or IgA antibodies, and 4) circulating IgA and IgG autoantibodies that target the 120-kd soluble protein LAD-1. The present study establishes unequivocally the existence of a naturally occurring canine model of LAD of humans.

  View details for Web of Science ID 000087996900003

  View details for PubMedID 10896391

• Subepidermal blistering disease with autoantibodies against a novel dermal 200-kDa antigen JOURNAL OF DERMATOLOGICAL SCIENCE Kawahara, Y., Zillikens, D., Yancey, K. B., Marinkovich, M. P., Nie, Z., Hashimoto, T., Nishikawa, T. 2000; 23 (2): 93-102

  Abstract

  A number of autoimmune subepidermal blistering diseases are characterized by the distinct autoantigens of the cutaneous basement membrane zone. Recently, a few cases with autoantibodies against a novel 200-kDa dermal protein have been reported. We collected nine cases of subepidermal blistering disease with IgG antibodies against this 200-kDa antigen. In this report, we describe the clinical and immunological appearances in these cases. Five cases showed bullous pemphigoid-like features, one case resembled dermatitis herpetiformis, and another case showed mixed features of bullous pemphigoid and linear IgA bullous dermatosis. It was interesting to note that psoriasis coexisted in four cases. By indirect immunofluorescence on 1 M NaCl split skin, IgG antibodies from all sera reacted with the dermal side of the split. By immunoblot analysis, IgG antibodies recognized a 200-kDa protein of dermal extract. IgG affinity-purified antibodies on the 200-kDa immunoblot membrane stained the dermal side of 1 M NaCl split skin. Various examinations suggested that the 200-kDa antigen is not identical to any chains of laminins-1, -5 or -6. This autoimmune subepidermal blistering disease against the dermal 200-kDa protein may form a new distinct entity, which often associates with psoriasis.

  View details for Web of Science ID 000087425100003

  View details for PubMedID 10808126

• Compound heterozygosity for novel splice site mutations in the BPAG2/COL17A1 gene underlies generalized atrophic benign epidermolysis bullosa JOURNAL OF INVESTIGATIVE DERMATOLOGY Pulkkinen, L., Marinkovich, M. P., Tran, H. T., Lin, L., Herron, G. S., Uitto, J. 1999; 113 (6): 1114-1118

  Abstract

  Generalized atrophic benign epidermolysis bullosa, GABEB (OMIM# 226650), is a nonlethal variant of epidermolysis bullosa with autosomal recessive inheritance pattern. The pathogenesis of this disorder can be caused by mutations affecting two different gene/protein systems. Most of the mutations have been identified in the BPAG2/COL17A1 gene encoding a hemidesmosomal transmembrane protein, the 180 kDa bullous pemphigoid antigen (BP180), also known as type XVII collagen. The minority of the mutations are localized in the LAMB3 gene encoding the beta3 polypeptide of laminin 5. In In this study we describe a GABEB patient who showed absent expression of BP180 in the cultured keratinocytes as well as in the skin. The patient was a compound heterozygote for two different splice site mutations, 3053-1G-->C and 3871+1G-->C, affecting the extra-cellular domain of the protein. These mutations resulted in multiple aberrant splice variants, three of them causing premature termination codons for translation. This case, dealing with out-of-frame splice site mutations in BPAG2/COL17A1, attests to the molecular heterogeneity of GABEB.

  View details for Web of Science ID 000084436600038

  View details for PubMedID 10636730

• Reduced anchoring fibril formation and collagen VII immunoreactivity in feline dystrophic epidermolysis bullosa VETERINARY PATHOLOGY Olivry, T., Dunston, S. M., Marinkovich, M. P. 1999; 36 (6): 616-618

  Abstract

  Dystrophic epidermolysis bullosa was diagnosed in a cat with juvenile-onset epithelial sloughing of the oral mucosa, footpads, and haired skin. Dermoepidermal separation occurred in the absence of inflammation or cytolysis of basal epidermal cells. Collagen IV-specific immunostaining corroborated the fact that clefting took place below the epidermal basement membrane. Ultrastructural examination revealed that the proband's anchoring fibrils exhibited a filamentous morphology and were decreased in number compared with those in a normal cat. Finally, the attenuated immunoreactivity for collagen VII in our patient led us to suspect that its encoding gene, COL7A1, could be mutated in this case of feline dystrophic epidermolysis bullosa.

  View details for Web of Science ID 000085281300014

  View details for PubMedID 10568446

• Antibodies to BP180 induce the release of IL-6 and IL-8 from cultured normal human keratinocytes Schmidt, E., Reimer, S., Kruse, N., Brocker, E. B., Marinkovich, M. P., Giudice, G., Zillikens, D. BLACKWELL SCIENCE INC. 1999: 438

  View details for Web of Science ID 000082406000073

• Update on inherited bullous dermatoses DERMATOLOGIC CLINICS Marinkovich, M. P. 1999; 17 (3): 473-?

  Abstract

  Bullous diseases are becoming increasingly better understood owing to the active research which has taken place in this field over the past decade. Advances in understanding of bullous disease pathophysiology is translating into clinical applications for diagnosis and therapy that will greatly enhance the quality of care bullous disease patients may receive now and in the future. This review focuses on the progress which has been achieved in inherited bullous dermatoses.

  View details for PubMedID 10410853

• Novel feline autoimmune blistering disease resembling bullous pemphigoid in humans: IgG autoantibodies target the NC16A ectodomain of type XVII collagen (BP180/BPAG2) VETERINARY PATHOLOGY Olivry, T., Chan, L. S., Xu, L., Chace, P., Dunston, S. M., Fahey, M., Marinkovich, M. P. 1999; 36 (4): 328-335

  Abstract

  In humans and dogs, bullous pemphigoid (BP) is an autoimmune blistering disease associated with the production of basement membrane autoantibodies that target the 180-kd type XVII collagen (BP180, BPAG2) and/or the 230-kd plakin epidermal isoform BPAG1e (BP230). In two adult cats, an acquired dermatosis and stomatitis was diagnosed as BP subsequent to the fulfillment of the following criteria: 1) presence of cutaneous vesicles, erosions, and ulcers; 2) histologic demonstration of subepidermal vesiculation with inflammatory cells, including eosinophils; 3) in vivo deposition of IgG autoantibodies at the epidermal basement membrane zone; and 4) serum IgG autoantibodies targeting a 180-kd epidermal protein identified as type XVII collagen. In both cats, the antigenic epitopes targeted by IgG autoantibodies were shown to be situated in the NC16A ectodomain of type XVII collagen, a situation similar to that of humans and dogs with BP. Feline BP therefore can be considered a clinical, histopathologic, and immunologic homologue of BP in humans and dogs.

  View details for Web of Science ID 000083807900008

  View details for PubMedID 10421100

• Melanocytes adhere to and synthesize laminin-5 in vitro EXPERIMENTAL DERMATOLOGY Scott, G. A., Cassidy, L., Tran, H., Rao, S. K., Marinkovich, M. P. 1999; 8 (3): 212-221

  Abstract

  Melanocytes arise from the neural crest, migrate to the skin, and can be detected in the basal layer of the epidermis in skin biopsies of human fetuses as early as 11 weeks gestational age. During post-natal life, melanocytes reside at the basal layer of the epidermis, but the ligands to which they attach are unknown. Laminin-5 is a component of anchoring filaments of the lamina lucida of the epidermal basement membrane. In this report we show that human melanocytes adhere to purified laminin-5 to a level comparable with normal human keratinocytes. Blocking antibodies to the 165 kDa subunit of laminin-5 significantly inhibited fetal and neonatal melanocyte attachment to the surface of salt-split skin, which exposes laminin-5 on its surface, suggesting that laminin-5 is a ligand for melanocyte attachment to the basement membrane in vivo. Western blotting of concentrated culture supernatant of fetal and neonatal melanocytes with anti-laminin-5 antibodies demonstrated a single immunoreactive band of the expected size of laminin-5. In contrast, 3 human metastatic melanoma cell lines did not produce laminin-5. Immunofluorescence microscopy with antibodies to each of the three chains of laminin-5 confirmed the presence of laminin-5 in a peri-cellular distribution around melanocytes, but not melanoma cells. Our results suggest that laminin-5 may be a ligand for normal human melanocytes in the basement membrane, and that loss of laminin-5 production by melanoma cells may be a marker for malignant transformation.

  View details for Web of Science ID 000080694800007

  View details for PubMedID 10389639

• Bullous systemic lupus erythematosus with autoantibodies recognizing multiple skin basement membrane components, bullous pemphigoid antigen 1, laminin-5, laminin-6, and type VII collagen International Investigative Dermatology Meeting Chan, L. S., Lapiere, J. C., Chen, M., Traczyk, T., Mancini, A. J., Paller, A. S., Woodley, D. T., Marinkovich, M. P. AMER MEDICAL ASSOC. 1999: 569–73

  Abstract

  Bullous systemic lupus erythematosus is a generalized subepidermal blistering skin eruption in patients suffering from systemic lupus erythematosus. Type VII collagen was initially identified as the target antigen.We studied an unusual patient who had bullous systemic lupus crythematosus. The patient fulfilled the criteria of systemic lupus with an antinuclear antibody titer of 1:5120. Immunopathological testing revealed in vivo deposition of all IgG subclasses, secretory IgA1, and both light chains at the patient's skin basement membrane. The in vivo-bound IgG and IgA were localized at the hemidesmosomes and lamina densa. The patient's IgG and IgA circulating autoantibodies labeled both the epidermal roof and the dermal floor of salt-split skin and recognized the hemidesmosomal protein BP230 as well as the full-length native form and the recombinant noncollagenous domain 1 of type VII collagen (anchoring fibril). In addition, the patient's IgG autoantibodies recognized the anchoring filament proteins laminin-5 and laminin-6 (alpha3 chain and gamma2 chain).We conclude that patients with bullous systemic lupus erythematosus may have autoantibodies to multiple basement membrane components critical for epidermal-dermal junctional adhesion. Possible pathogenic mechanisms in this patient's clinical diseases include provocation of organ-specific disease (bullous disease) by systemic autoimmunity (lupus) and the "epitope spreading" immune phenomenon.

  View details for Web of Science ID 000080167000012

  View details for PubMedID 10328198

• Disruption of integrin a6b4 ligand binding through point mutation in human keratinocytes: Effects upon hemidesmosome formation and cell migration Russell, A. J., Ghaed, S. V., Marinkovich, M. P. BLACKWELL SCIENCE INC. 1999: 537

  View details for Web of Science ID 000079495800111

• Durable and efficient corrective keratin 14 gene therapy in recessive epidermolysis bullosa simplex (EBS) Pereira, P., Bruckner-Tuderman, L., Zabel, B., Marinkovich, M. P. BLACKWELL SCIENCE INC. 1999: 640

  View details for Web of Science ID 000079495800729

• NC1 domain of type VII collagen binds to the beta 3 chain of laminin 5 via a unique subdomain within the fibronectin-like repeats JOURNAL OF INVESTIGATIVE DERMATOLOGY Chen, M., Marinkovich, M. P., Jones, J. C., O'Toole, E. A., Li, Y. Y., Woodley, D. T. 1999; 112 (2): 177-183

  Abstract

  Type VII collagen, the major component of anchoring fibrils, consists of a central collagenous triple-helical domain flanked by two noncollagenous, globular domains, NC1 and NC2. Approximately 50% of the molecular mass of the molecule is consumed by the NC1 domain. We previously demonstrated that NC1 binds to various extracellular matrix components including a complex of laminin 5 and laminin 6 (Chen et al, 1997a). In this study, we examined the interaction of NC1 with laminin 5 (a component of anchoring filaments). Both authentic and purified recombinant NC1 bound to human and rat laminin 5 as measured by enzyme-linked immunosorbant assay and by binding of 125I-radiolabeled NC1 to laminin 5-coated wells, but not to laminin 1 or albumin. NC1 bound predominantly to the beta3 chain of laminin 5, but also to the gamma2 chain when examined by a protein overlay assay. The binding of 125I-NC1 to laminin 5 was inhibited by a 50-fold excess of unlabeled NC1 or de-glycosylated NC1, as well as a polyclonal antibody to laminin 5 or a monoclonal antibody to the beta3 chain. In contrast, the NC1-laminin 5 interaction was not affected by a monoclonal antibody to the alpha3 chain. Using NC1 deletion mutant recombinant proteins, a 285 AA (residues 760-1045) subdomain of NC1 was identified as the binding site for laminin 5. IgG from an epidermolysis bullosa acquisita serum containing autoantibodies to epitopes within NC1 that colocalized with the laminin 5 binding site inhibited the binding of NC1 to laminin 5. Thus, perturbation of the NC1-laminin 5 interaction may contribute to the pathogenesis of epidermolysis bullosa acquisita.

  View details for Web of Science ID 000078287000007

  View details for PubMedID 9989793

• Gene therapy for a lethal genetic blistering disease: a status report. Transactions of the American Clinical and Climatological Association BAUER, E. A., Herron, G. S., Marinkovich, M. P., Khavari, P. A., Lane, A. T. 1999; 110: 86-92

  View details for PubMedID 10344009

• BP180 gene delivery in junctional epidermolysis bullosa GENE THERAPY Seitz, C. S., Giudice, G. J., Balding, S. D., Marinkovich, M. P., Khavari, P. A. 1999; 6 (1): 42-47

  Abstract

  Epidermolysis bullosa (EB) comprises a family of inherited blistering skin diseases for which current therapy is only palliative. Junctional EB (JEB) involves dissociation of the dermal-epidermal junction and results from mutations in a number of genes that encode vital structural proteins, including BP180 (type XVII collagen/BPAG2). In order to develop a model of corrective gene delivery for JEB, we produced a retroviral expression vector for wild-type human BP180 and used it to restore BP180 protein expression to primary keratinocytes from BP180-negative patients with generalized atrophic JEB. Restoration of full-length BP180 protein expression was associated with adhesion parameter normalization of primary JEB keratinocytes in vitro. These cells were then used to regenerate human skin on immune-deficient mice. BP180 gene-transduced tissue demonstrated restoration of BP180 gene expression at the dermal-epidermal junction in vivo while untransduced regenerated JEB skin entirely lacked BP180 expression. These findings provide a basis for future efforts to achieve gene delivery in human EB skin tissue.

  View details for Web of Science ID 000078169300007

  View details for PubMedID 10341874

• NC1 domain of type VII (anchoring fibril) collagen binds to the beta 3 chain of laminin 5 via a unique subdomain within the fibronectin-like repeats. Chen, M., Marinkovich, M. P., Jones, J. C., O'Toole, E. A., Li, Y. Y., Woodley, D. T. BLACKWELL SCIENCE INC. 1998: 496

  View details for Web of Science ID 000072738200174

• Self-assembly of laminin isoforms JOURNAL OF BIOLOGICAL CHEMISTRY Cheng, Y. S., Champliaud, M. F., Burgeson, R. E., Marinkovich, M. P., Yurchenco, P. D. 1997; 272 (50): 31525-31532

  Abstract

  The alpha, beta, and gamma subunits of basement membrane laminins can combine into different heterotrimeric molecules with either three full short arms (e.g. laminins-1-4), or molecules containing one (laminins-6-9) or more (laminin-5) short arm truncations. Laminin-1 (alpha1beta1gamma1), self-assembles through a calcium-dependent thermal gelation requiring binding interactions between N-terminal short arm domains, forming a meshwork polymer thought to contribute to basement membrane architecture (Yurchenco, P. D., and Cheng, Y. S. (1993) J. Biol. Chem. 268, 17286-17299). However, it has been unclear whether other isoforms share this property, and if so, which ones. To begin to address this, we evaluated laminin-2 (alpha2beta1gamma1), laminin-4 (alpha2beta2gamma1), laminin-5 (alpha3Abeta3gamma2), and laminin-6 (alpha3Abeta1gamma1). The first two isoforms were found to self-aggregate in a concentration- and temperature-dependent manner and a close self-assembly relationship among laminins-1, -2, and -4 were demonstrated by: (a) polymerization of all three proteins was inhibited by EDTA and laminin-1 short arm fragments, (b) polymerization of laminin-1 was inhibited by fragments of laminins-2 and -4, (c) laminin-2 and, to a lesser degree, laminin-4, even well below their own critical concentration, co-aggregated with laminin-1, evidence for co-polymerization. Laminin-5, on the other hand, neither polymerized nor co-polymerized with laminin-1. Laminin-6 failed to co-aggregate with laminin-1 at all concentrations evaluated, evidence for a lack of a related self-assembly activity. The data support the hypothesis that all three short arms are required for self-assembly and suggest that the short arm domain structure of laminin isoforms affect their architecture-forming properties in basement membranes.

  View details for Web of Science ID A1997YL41900048

  View details for PubMedID 9395489

• Hypoxia increases human keratinocyte motility on connective tissue 56th Annual Meeting of the Society-for-Investigative-Dermatology O'Toole, E. A., Marinkovich, M. P., Peavey, C. L., Amieva, M. R., FURTHMAYR, H., Mustoe, T. A., Woodley, D. T. AMER SOC CLINICAL INVESTIGATION INC. 1997: 2881–91

  Abstract

  Re-epithelialization of skin wounds depends upon the migration of keratinocytes from the cut margins of the wound and is enhanced when human keratinocytes are covered with occlusive dressings that induce hypoxia. In this study, two independent migration assays were used to compare cellular motility on connective tissue components under normoxic or hypoxic conditions. Human keratinocytes apposed to collagens or fibronectin exhibited increased motility when subjected to hypoxic (0.2 or 2% oxygen) conditions compared with normoxic (9 or 20% oxygen) conditions. When compared with normoxic cells, hypoxic keratinocytes exhibited increased expression and redistribution of the lamellipodia-associated proteins (ezrin, radixin, and moesin). Furthermore, hypoxic keratinocytes demonstrated decreased secretion of laminin-5, a laminin isoform known to inhibit keratinocyte motility. Hypoxia did not alter the number of integrin receptors on the cell surface, but did induce enhanced secretion of the 92-kD type IV collagenase. These data demonstrate that hypoxia promotes human keratinocyte motility on connective tissue. Hypoxia-driven motility is associated with increased expression of lamellipodia proteins, increased expression of collagenase and decreased expression of laminin-5, the locomotion brake for keratinocytes.

  View details for Web of Science ID 000071007300031

  View details for PubMedID 9389755

  View details for PubMedCentralID PMC508495

• LAD-1 is absent in a subset of junctional epidermolysis bullosa patients 57th Annual Meeting of the Society-for-Investigative-Dermatology Marinkovich, M. P., Tran, H. H., Rao, S. K., Giudice, G. J., Balding, S., Jonkman, M. F., Pas, H. H., McGuire, J. S., Herron, G. S., BRUCKNERTUDERMAN, L. NATURE PUBLISHING GROUP. 1997: 356–59

  Abstract

  The anchoring filament protein LAD-1 has been recently identified as the target of autoantibodies in the acquired blistering disorder linear IgA bullous dermatosis. Because this protein appears to be involved in the process of dermal-epidermal cohesion, this study sought to determine the involvement of LAD-1 in the pathology of junctional epidermolysis bullosa (JEB). To this end, 44 patients with a variety of subtypes of JEB were analyzed by indirect immunofluorescence microscopy with antibodies to LAD-1, BP180, and laminin-5. We found that only patients with generalized atrophic benign epidermolysis bullosa (GABEB) contained LAD-1 defects. Of the 16 GABEB patients studied, 13 showed absent or greatly reduced expression of LAD-1 (including 2 patients with a peculiar interrupted staining pattern) and 3 patients showed defects of laminin-5 expression with normal LAD-1 expression. Patients who showed LAD-1 defects also showed abnormal expression of BP180. Keratinocytes were cultured from the skin of two GABEB patients and analyzed by indirect immunofluorescent microscopy. One culture demonstrated defects of BP180 and LAD-1 expression (which was also verified by radioimmunoprecipitation assay), and one culture showed decreased laminin-5 expression but normal BP180 and LAD-1 expression. Thus, these studies demonstrate that: (i) LAD-1 and BP180 are normally expressed in all subtypes of JEB except GABEB, (ii) the majority of GABEB patients show absent or near absent expression of both LAD-1 and BP180 but normal expression of laminin-5, and (iii) a smaller subset of GABEB patients show normal LAD-1 and BP180 expression but express persistent but reduced levels of laminin-5.

  View details for PubMedID 9284104

• Immunodissection of the connective tissue matrix in human skin MICROSCOPY RESEARCH AND TECHNIQUE Keene, D. R., Marinkovich, M. P., Sakai, L. Y. 1997; 38 (4): 394-406

  Abstract

  Much of what has been learned of the components and structure of human skin over the past few years has been accomplished with the aid of antibody technology. Antibodies are used in techniques such as affinity chromatography to isolate individual molecules and by immunofluorescence and immunoelectron microscopy to identify each of those molecules as components of specific macromolecular assemblies present within the dermis. This manuscript is meant not as a review of technique but instead as a summary of recent progress made in the understanding of dermal matrix architecture.

  View details for Web of Science ID A1997XU93000007

  View details for PubMedID 9297689

• Laminin-5 inhibits human keratinocyte migration 57th Annual Meeting of the Society-for-Investigative-Dermatology OToole, E. A., Marinkovich, M. P., Hoeffler, W. K., FURTHMAYR, H., Woodley, D. T. ELSEVIER INC. 1997: 330–39

  Abstract

  Laminin-5 (previously known as kalinin, epiligrin, and nicein) is an adhesive protein localized to the anchoring filaments within the lamina lucida space of the basement membrane zone lying between the epidermis and dermis of human skin. Anchoring filaments are structures within the lamina lucida and lie immediately beneath the hemidesmosomes of the overlying basal keratinocytes apposed to the basement membrane zone. Human keratinocytes synthesize and deposit laminin-5. Laminin-5 is present at the wound edge during reepithelialization. In this study, we demonstrate that laminin-5, a powerful matrix attachment factor for keratinocytes, inhibits human keratinocyte migration. We found that the inhibitory effect of laminin-5 on keratinocyte motility can be reversed by blocking the alpha3 integrin receptor. Laminin-5 inhibits keratinocyte motility driven by a collagen matrix in a concentration-dependent fashion. Using antisense oligonucleotides to the alpha3 chain of laminin-5 and an antibody that inhibits the cell binding function of secreted laminin-5, we demonstrated that the endogenous laminin-5 secreted by the keratinocyte also inhibits the keratinocyte's own migration on matrix. These findings explain the hypermotility that characterizes keratinocytes from patients who have forms of junctional epidermolysis bullosa associated with defects in one of the genes encoding for laminin-5 chains, resulting in low expression and/or functional inadequacy of laminin-5 in these patients. These studies also suggest that during reepithelialization of human skin wounds, the secreted laminin-5 stabilizes the migrating keratinocyte to establish the new basement membrane zone.

  View details for Web of Science ID A1997XE31700010

  View details for PubMedID 9194495

• Interactions of the amino-terminal noncollagenous (NC1) domain of type VII collagen with extracellular matrix components - A potential role in epidermal-dermal adherence in human skin JOURNAL OF BIOLOGICAL CHEMISTRY Chen, M., Marinkovich, M. P., Veis, A., Cai, X. Y., Rao, C. N., OToole, E. A., Woodley, D. T. 1997; 272 (23): 14516-14522

  Abstract

  Type VII collagen, the major component of anchoring fibrils, consists of a central collagenous triple-helical domain flanked by two noncollagenous domains, NC1 and NC2. The NC1 domain contains multiple submodules with homology to known adhesive molecules including fibronectin type III-like repeats and the A domain of von Willebrand factor. In this study, we produced the entire NC1 domain of human type VII collagen in the stably transfected human kidney 293 cell clones and purified large quantities of the recombinant NC1 protein from serum-free culture media. The recombinant NC1 formed interchain disulfide-bonded dimers and trimers and was N-linked glycosylated. Tunicamycin inhibited the cellular secretion of NC1, suggesting that N-linked glycosylation may play a role in NC1 secretion. The recombinant NC1 was indistinguishable from the authentic NC1 obtained from human amnions or WISH cells with respect to N-linked sugar content, electrophoretic mobility, rotary shadow imaging, and binding affinity to type IV collagen. Purified recombinant NC1, like authentic NC1, also bound specifically to fibronectin, collagen type I, and a laminin 5/6 complex. Both monomeric and trimeric forms of NC1 exhibited equal affinity for these extracellular matrix components, suggesting that the individual arms of NC1 can function independently. The multiple interactions of NC1 with other extracellular matrix components may support epidermal-dermal adhesion.

  View details for Web of Science ID A1997XC32700005

  View details for PubMedID 9169408

• Laminin-6 and laminin-5 are recognized by autoantibodies in a subset of cicatricial pemphigoid Annual Meeting of the Society-for-Investigative-Dermatology Chan, L. S., MAJMUDAR, A. A., Tran, H. H., Meier, F., SCHAUMBURGLEVER, G., Chen, M., Anhalt, G., Woodley, D. T., Marinkovich, M. P. NATURE PUBLISHING GROUP. 1997: 848–53

  Abstract

  We characterized basement membrane zone (BMZ) autoantigens targeted by autoantibodies (AAb) from patients with cicatricial pemphigoid. Serum from a patient with severe oral cicatricial pemphigoid contained IgG anti-BMZ AAb. The AAb labeled a lower BMZ component on salt-split skin and localized to the lower lamina lucida/lamina densa by direct and indirect immunoelectron microscopy (IEM) but did not label blood vessels. The AAb did not react with EHS laminin-1 and type IV collagen, pepsinized human type IV collagen, recombinant entactin, or NC1 domain of type VII collagen by dot blotting and western blotting. We focused our studies on the laminin family, as laminin-5 was identified as an autoantigen in cicatricial pemphigoid. Culture-conditioned media from normal keratinocytes (containing laminin-6 and laminin-5) and JEB keratinocytes (containing laminin-6 but not laminin-5) were studied by western blotting. Under nonreducing conditions, the patient's AAb recognized a 600-kDa protein (laminin-6) intensely and a 400-kDa protein (laminin-5) weakly in normal keratinocyte medium even though abundant laminin-5 was present. InJEB keratinocyte medium, however, the 600-kDa protein (laminin-6) alone was recognized by the patient's AAb. The AAb also immunolabeled BMZ of JEB skin that lacked laminin-5. The AAb from this patient and two other patients with anti-laminin-5 cicatricial pemphigoid immunoprecipitated both laminin-6 and laminin-5. Taken together, the results of IEM, non-reducing western blotting, immunoprecipitation, and JEB skin BMZ immunolabeling indicate that laminin-6, as well as laminin-5, is identified by the AAb from a subset of cicatricial pemphigoid patients. We propose the name "anti-laminin cicatricial pemphigoid" for this subset.

  View details for Web of Science ID A1997XB83600003

  View details for PubMedID 9182809

• LAD-1 is a collagenous component of keratinocyte adhesion complexes which assembles into a high molecular weight complex and which has homology to BP180. Tran, H. H., Schumann, H., Balding, S., Giudice, G. J., BRUCKNERTUDERMAN, L., Marinkovich, M. P. NATURE PUBLISHING GROUP. 1997: 289–89

  View details for Web of Science ID A1997WP04000312

• Identification of binding sites of noncollagenous (NCl) domain of type VII collagen with laminin 5/6 and other extracellular matrix components. Chen, M., Marinkovich, M. P., Cai, X. Y., Woodley, D. T. NATURE PUBLISHING GROUP. 1997: 461–61

  View details for Web of Science ID A1997WP04000482

• BP180 gene transfer and expression in benign junctional epidermolysis bullosa. Seitz, C. S., Marinkovich, M. P., Tran, H., Janoff, M., Diaz, L. A., OToole, E. A., Woodley, D. T., Giudice, G. J., Khavari, P. A. NATURE PUBLISHING GROUP. 1997: 305–

  View details for Web of Science ID A1997WP04000326

• A novel subepidermal blistering disease with autoantibodies to a 200-kDa antigen of the basement membrane zone. journal of investigative dermatology Zillikens, D., Kawahara, Y., Ishiko, A., Shimizu, H., Mayer, J., Rank, C. V., Liu, Z., Giudice, G. J., Tran, H. H., Marinkovich, M. P., Brocker, E. B., Hashimoto, T. 1996; 106 (6): 1333-1338

  Abstract

  Several components of the basement membrane zone (BMZ) have been identified as antigenic targets in autoimmune bullous diseases. We report a novel disease with autoantibodies to a BMZ antigen that is different from the targets described so far. The patient suffering from this disorder showed tense bullae and severe mucous membrane involvement rapidly responding to oral tetracyclines and colchicine. Histopathologic findings resembled those of dermatitis herpetiformis. Direct immunofluorescence microscopy showed linear deposits of IgG and C3 at the BMZ. By indirect immunofluorescence studies on split human skin, using both 1 M NaCl and suction blistering for dermal-epidermal separation, IgG antibodies localized exclusively to the dermal side of the split. The antibodies were mainly of the IgG4 subclass. By Western blot analysis of epidermal and dermal extracts, the patient's serum unequivocally reacted with a dermal antigen of 200 kDa. It did not recognize bullous pemphigoid antigens, the autoantigen of epidermolysis bullosa acquisita, purified preparations of laminin-1 and laminin-5, or the recently described 105-kDa BMZ antigen. By immunoblotting of concentrated conditioned SCC-25 medium, the patient's antibodies reacted with a band of 200 kDa and several hands of lower molecular weight. No reactivity was seen with extracts of cultured human fibroblasts. By indirect immunogold electron microscopy, immunoreactants localized to the lower lamina lucida. After clearance of skin lesions, both indirect immunofluorescence and Western blot analysis became negative. This patient suffers from a novel autoimmune bullous disease with autoantibodies to a 200-kDa antigen of the BMZ.

  View details for PubMedID 8752680

• A novel subepidermal blistering disease with autoantibodies to a 200-kDa antigen of the basement membrane zone (vol 106, pg 465, 1996) JOURNAL OF INVESTIGATIVE DERMATOLOGY Zillikens, D., Kawahara, Y., Ishiko, A., Shimizu, H., Mayer, J., Rank, C. V., Liu, Z., Giudice, G. J., Tran, H. H., Marinkvch, M. P., Brocker, E. B., Hashimoto, T. 1996; 106 (6): 1332-1338

  Abstract

  Several components of the basement membrane zone (BMZ) have been identified as antigenic targets in autoimmune bullous diseases. We report a novel disease with autoantibodies to a BMZ antigen that is different from the targets described so far. The patient suffering from this disorder showed tense bullae and severe mucous membrane involvement rapidly responding to oral tetracyclines and colchicine. Histopathologic findings resembled those of dermatitis herpetiformis. Direct immunofluorescence microscopy showed linear deposits of IgG and C3 at the BMZ. By indirect immunofluorescence studies on split human skin, using both 1 M NaCl and suction blistering for dermal-epidermal separation, IgG antibodies localized exclusively to the dermal side of the split. The antibodies were mainly of the IgG4 subclass. By Western blot analysis of epidermal and dermal extracts, the patient's serum unequivocally reacted with a dermal antigen of 200 kDa. It did not recognize bullous pemphigoid antigens, the autoantigen of epidermolysis bullosa acquisita, purified preparations of laminin-1 and laminin-5, or the recently described 105-kDa BMZ antigen. By immunoblotting of concentrated conditioned SCC-25 medium, the patient's antibodies reacted with a band of 200 kDa and several hands of lower molecular weight. No reactivity was seen with extracts of cultured human fibroblasts. By indirect immunogold electron microscopy, immunoreactants localized to the lower lamina lucida. After clearance of skin lesions, both indirect immunofluorescence and Western blot analysis became negative. This patient suffers from a novel autoimmune bullous disease with autoantibodies to a 200-kDa antigen of the BMZ.

  View details for Web of Science ID A1996UR40400031

• LAD-1, the linear IgA bullous dermatosis autoantigen, is a novel 120-kDa anchoring filament protein synthesized by epidermal cells JOURNAL OF INVESTIGATIVE DERMATOLOGY Marinkovich, M. P., Taylor, T. B., Keene, D. R., Burgeson, R. E., Zone, J. J. 1996; 106 (4): 734-738

  Abstract

  This study characterizes a novel basement membrane component that is the target of autoantibodies in patients with linear IgA bullous dermatosis. Tissue surveys showed that this protein localized to the epidermal side of 1 M NaCl split skin and to basement membranes in cornea, oral mucosa, esophagus, intestine, kidney collecting ducts, ureter, bladder, urethra, and thymus, but was absent in lung, blood vessels, skeletal muscle, and nerve. Monoclonal antibody 123, which recognizes this protein, induced dermal-epidermal separation of human skin in situ, and this protein was found, by immunoelectron microscopy, to localize exclusively to anchoring filaments. This protein was secreted as as a 120-kDa peptide from primary cultures of keratinocytes as determined by radioimmunoprecipitation. Monoclonal antibody 123 recognized this protein as a 120-kDa band from conditioned cell culture medium and a 97-kDa band from human skin extracts as shown by immunoblot. Serum from five patients with the autoimmune blistering disorder linear IgA bullous dermatosis specifically recognized bands of 120 and 97 kDa from culture medium and skin extracts, respectively, that were of identical electrophoretic migration to the bands recognized by monoclonal antibody 123. In summary, this study characterizes a novel anchoring filament protein that is the target of linear IgA bullous dermatosis autoantibodies. Because monoclonal antibody 123 induces blistering of human skin, we hypothesize that this protein functions to maintain dermal-epidermal cohesion and that autoantibodies in this disease are themselves pathogenic. We propose LAD-1 as the name for this protein.

  View details for PubMedID 8618013

• Molecular mechanisms of hypoxia-driven keratinocyte motility OToole, E. A., Peavey, C., Marinkovich, M. P., Furthmayr, H., Mustoe, T., Woodley, D. T. BLACKWELL SCIENCE INC. 1996: 141

  View details for Web of Science ID A1996UC78700182

• Laminin-5 inhibits keratinocyte motility OToole, E. A., Marinkovich, M. P., Shervin, N., Woodley, D. T. BLACKWELL SCIENCE INC. 1996: 98

  View details for Web of Science ID A1996UC78700142

• Therapy for junctional epidermolysis bullosa: Functional assays and a grafted scid mouse animal model Hoffler, W. K., Nelson, C. F., Matsui, C., Marinkovich, M., Mak, L. L., Wang, C. K. BLACKWELL SCIENCE INC. 1996: 10

  View details for Web of Science ID A1996UC78700053

• Identification of multiple, distinct cicatritial pemphigoid autoantigens. Rao, S. K., Tran, H. H., Allen, J., Wojnarowska, F., Marinkovich, M. P. NATURE PUBLISHING GROUP. 1996: 39–39

  View details for Web of Science ID A1996UC78700080

• Laminin-5 is synthesized by normal human melanocytes, but not melanoma cells, and is a ligand for melanocytes in vivo. Scott, G. A., Cassidy, L., Tran, H. H., Rao, S. K., Marinkovich, M. P. NATURE PUBLISHING GROUP. 1996: 130–30

  View details for Web of Science ID A1996UC78700173

• Human microvascular endothelial cells produce a novel laminin variant. Tran, H. H., Rao, S. K., Zhang, D. N., Romero, L., Khush, K., Herron, G. S., Marinkovich, M. P. NATURE PUBLISHING GROUP. 1996: 143–43

  View details for Web of Science ID A1996UC78700183

• Laminin-B is a lamina, densa autoantigen ih oral pemphigoid. Chan, L. S., MAJMUDAR, A. A., Tran, H. H., Meier, F., SCHAUMBURGLEVER, G., Chen, M., Woodley, D. T., Marinkovich, M. P. NATURE PUBLISHING GROUP. 1996: 41–41

  View details for Web of Science ID A1996UC78700081

• LAD-1 is absent in a subset of generalized atrophic benign junctional epidermolysis bullosa patients Marinkovich, M. P., Tran, H. H., Rao, S. K., Giudice, G. J., Balding, S., Jonkman, M. F., Pas, H. H., BRUCKNERTUDERMAN, L. NATURE PUBLISHING GROUP. 1996: 281–81

  View details for Web of Science ID A1996UC78700321

• A novel subepidermal blistering disease with autoantibodies to a 200-kDa antigen of the basement membrane zone JOURNAL OF INVESTIGATIVE DERMATOLOGY Zillikens, D., Kawahara, Y., Ishiko, A., Shimizu, H., Mayer, J., Rank, C. V., Liu, Z., Giudice, G. J., Tran, H. H., Marinkovich, M. P., Brocker, E. B., Hashimoto, T. 1996; 106 (3): 465-470

  Abstract

  Several components of the basement membrane zone (BMZ) have been identified as antigenic targets in autoimmune bullous diseases. We report a novel disease with autoantibodies to a BMZ antigen that is different from the targets described so far. The patient suffering from this disorder showed tense bullae and severe mucous membrane involvement rapidly responding to oral tetracyclines and colchicine. Histopathologic findings resembled those of dermatitis herpetiformis. Direct immunofluorescence microscopy showed linear deposits of IgG and C3 at the BMZ. By indirect immunofluorescence studies on split human skin, using both 1 M NaCl and suction blistering for dermal-epidermal separation, IgG antibodies localized exclusively to the dermal side of the split. The antibodies were mainly of the IgG4 sub-class. By Western blot analysis of epidermal and dermal extracts, the patient's serum unequivocally reacted with a dermal antigen of 200 kDa. It did not recognize bullous pemphigoid antigens (the autoantigen of epidermolysis bullosa acquisita), purified preparations of laminin-1 and laminin-5, or the recently described 105-kDa BMZ antigen. By immunoblotting of concentrated conditioned SCC-25 medium, the patient's antibodies reacted with a band of 200 kDa and several bands of lower molecular weight. No reactivity was seen with extracts of cultured human fibroblasts. By indirect immunogold electron microscopy, immunoreactants localized to the lower lamina lucida. After clearance of skin lesions, both indirect immunofluorescence and Western blot analysis became negative. This patient suffers from a novel autoimmune bullous disease with autoantibodies to a 200-kDa antigen of the BMZ.

  View details for Web of Science ID A1996UB96800014

  View details for PubMedID 8648178

• LAMININ-5 INHIBITS KERATINOCYTE MOTILITY OTOOLE, E. A., MARINKOVICH, M. P., YURKO, M. A., WOODLEY, D. T. BLACKWELL SCIENCE PUBL INC CAMBRIDGE. 1995: 871

  View details for Web of Science ID A1995TH09000094

• PRENATAL-DIAGNOSIS OF HERLITZ JUNCTIONAL EPIDERMOLYSIS-BULLOSA BY AMNIOCENTESIS PRENATAL DIAGNOSIS Marinkovich, M. P., Meneguzzi, G., Burgeson, R. E., BLANCHETBARDON, C., Holbrook, K. A., Smith, L. T., Christiano, A. M., Ortonne, J. P. 1995; 15 (11): 1027-1034

  Abstract

  Herlitz junctional epidermolysis bullosa (HJEB) is a severe blistering disorder which usually results in death during infancy. We have previously shown that the anchoring filament protein laminin-5 (kalinin/nicein), which mediates keratinocyte attachment and dermal-epidermal cohesion, is abnormally expressed in individuals with HJEB. Laminin-5 was detected by Western blot analysis in amniotic fluid from 44 consecutive normal second-trimester control pregnancies, but was undetectable in second-trimester amniotic fluid from four pregnancies with fetuses affected by HJEB. In one case of severe non-Herlitz JEB, laminin-5 was detected in both amniotic fluid and skin. In human amniotic fluid, the laminin-5 a3 subunit was processed to a major 165 kD species and a minor 145 kD species and the beta 2 subunit was partially processed to 105 kD. Although laminin-5 was covalently associated with laminin-6 (K-laminin) in amniotic membrane, no covalent interaction was detected in amniotic fluid. Laminin-5 from amniotic fluid strongly supported keratinocyte attachment. These results suggest that Western blot analysis of second-trimester amniotic fluid is useful in determining the prenatal diagnosis of HJEB and that laminin-5 may serve a physiologically important function in amniotic fluid.

  View details for PubMedID 8606881

• ANTIBASEMENT MEMBRANE AUTOANTIBODIES IN PATIENTS WITH ANTI-EPILIGRIN CICATRICIAL PEMPHIGOID BIND THE ALPHA-SUBUNIT OF LAMININ-5 JOURNAL OF INVESTIGATIVE DERMATOLOGY Kirtschig, G., Marinkovich, M. P., Burgeson, R. E., Yancey, K. B. 1995; 105 (4): 543-548

  Abstract

  Recent studies have identified a group of cicatricial pemphigoid patients who have IgG anti-basement membrane autoantibodies that recognize epiligrin, a set of disulfide-linked polypeptides closely related if not identical to laminin 5 (formerly called kalinin, nicein, or BM600). To further understand the pathophysiology of blister formation in these patients, we have sought to identify the specific polypeptide(s) targeted by their autoantibodies. Comparative studies show that sera from these patients (nine of nine), P1E1 monoclonal anti-epiligrin antibody, and polyclonal as well as monoclonal anti-laminin 5 antibodies immunoprecipitate the same set of disulfide-linked polypeptides from media of biosynthetically radiolabeled human keratinocytes. Moreover, sera from eight of nine patients with anti-epiligrin cicatricial pemphigoid immunoblot the alpha subunit of laminin 5 but show no reactivity to its beta or gamma subunits. In addition, circulating IgG from a representative patient was affinity-purified against the alpha subunit of laminin 5 and shown to bind the dermal side of 1 M NaC1 split skin in the same manner as autoantibodies from all patients with anti-epiligrin cicatricial pemphigoid. Sera from patients with bullous pemphigoid (n = 5), other forms of cicatricial pemphigoid (n = 5), epidermolysis bullosa acquisita (n = 4), or bullous systemic lupus erythematosus (n = 1) show no reactivity against any subunit of this laminin isoform in immunoprecipitation or immunoblot experiments. These findings correlate with prior reports showing that a monoclonal antibody directed against the alpha subunit of laminin 5 (i.e., laminin subunit alpha 3) induces detachment of human keratinocytes from extracellular matrix in vitro as well as epidermis from human skin in situ. Together, these studies suggest that laminin subunit alpha 3 mediates attachment of basal keratinocytes to epidermal basement membrane and that autoantibodies directed against it may be pathogenic.

  View details for Web of Science ID A1995RY59800004

  View details for PubMedID 7561156

• A NEWLY IDENTIFIED 105-KD LOWER LAMINA-LUCIDA AUTOANTIGEN IS AN ACIDIC PROTEIN DISTINCT FROM THE 105-KD GAMMA-2 CHAIN OF LAMININ-5 JOURNAL OF INVESTIGATIVE DERMATOLOGY Chan, L. S., Wang, X. S., Lapiere, J. C., Marinkovich, M. P., Jones, J. C., Woodley, D. T. 1995; 105 (1): 75-79

  Abstract

  A 105-kD lower lamina lucida antigen (p105) has been detected by autoantibodies (anti-p105) from patients with a novel immunobullous disease. To distinguish p105 from other known lamina lucida components, we performed comparative immunoblotting on purified human amniotic laminin-5 (kalinin), 804G matrix (enriched in laminin-5), and keratinocyte and fibroblast proteins using anti-804G matrix antibody (J-18) and anti-p105. J-18 labeled the truncated laminin-5 gamma 2 chain in amniotic laminin-5, 804G matrix, and keratinocyte conditioned medium, but did not label fibroblast cytosol. Conversely, anti-p105 did not label amniotic laminin-5 or 804G matrix, but did label p105 in both keratinocyte conditioned medium and fibroblast cytosol. J-18 labeled the 105-kD laminin-5 gamma 2 chain in reduced keratinocyte proteins and a 400-kD laminin-5 complex under non-reducing conditions. In contrast, anti-p105 labeled p105 under both reducing and non-reducing conditions but did not label a 400-kD protein complex. Similarly, comparative immunoblotting on keratinocyte proteins using anti-p105 and anti-laminin-1 revealed no commonly labeled protein bands. Electrophoretic fractionations by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting of these fractions revealed that the peak fractions of keratinocyte proteins reactive with anti-p105 are different from those reactive with J-18. Furthermore, keratinocyte proteins fractionated by Mono Q anion-exchange chromatography revealed fractions immunoreactive with anti-p105, whereas J-18 showed no reactivity with these fractions. Two-dimensional gel electrophoresis and immunoblotting with anti-p105 revealed p105 to be an acidic protein with isoelectric points between 5.7 and 6.3, distinct from the isoelectric points of laminin-5 gamma 2 chain. We conclude that p105 is an acidic protein located in the lamina lucida and distinct from the truncated laminin-5 gamma 2 chain and the laminin-1 family.

  View details for Web of Science ID A1995RJ63200014

  View details for PubMedID 7615980

• NECROLYTIC MIGRATORY ERYTHEMA WITHOUT GLUCAGONOMA IN PATIENTS WITH LIVER-DISEASE JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Marinkovich, M. P., Botella, R., DATLOFF, J., Sangueza, O. P. 1995; 32 (4): 604-609

  Abstract

  Necrolytic migratory erythema (NME) is an uncommon inflammatory dermatosis with a distinct clinical and histologic appearance. NME is usually associated with glucagonoma. Only a few cases of NME in the absence of glucagonoma have been previously reported.We sought to understand further the pathogenesis of NME by analyzing data from three patients.Three patients were examined both clinically and histopathologically.Each patient had an extensive erythematous scaling eruption in intertriginous, perioral, and acral areas, and a markedly red, smooth tongue. Skin biopsy specimens showed confluent parakeratosis, epidermal pallor, papillary edema, and a lymphohistiocytic infiltrate. Two patients had alcoholic liver disease and one had liver dysfunction as a result of hemochromatosis. Serum albumin level was depressed, and liver enzyme values were increased in all three patients. Glucagonoma was undetectable in these patients.In the absence of glucagonoma, hepatocellular dysfunction and hypoalbuminemia appear to be the most common factors associated with NME.

  View details for PubMedID 7896950

• JUNCTIONAL EPIDERMOLYSIS-BULLOSA SKIN GRAFTED TO THE SCID MOUSE SHOWS PRESERVATION OF THE DISEASE PHENOTYPE MARINKOVICH, M. P., WOODLEY, D. T., WATANABE, G. L., JENSEN, R. A., CANTRELL, C. F., BANER, E. A., HOEFFLER, W. K., KIM, Y. H. BLACKWELL SCIENCE PUBL INC CAMBRIDGE. 1995: 603

  View details for Web of Science ID A1995QR53200311

• THE MOLECULAR-GENETICS OF BASEMENT-MEMBRANE DISEASES ARCHIVES OF DERMATOLOGY Marinkovich, M. P. 1993; 129 (12): 1557-1565

  Abstract

  Keratin filaments, hemidesmosomes, anchoring filaments, the lamina densa, and anchoring fibrils each function to maintain different levels of basement membrane cohesion.Keratin 5 or 14 mutations are present in epidemiolysis bullosa simplex. Herlitz junctional epidermolysis bullosa is characterized by defects of the anchoring filament protein kalinin (alternatively known as nicein). Mutations of the type VII collagen gene appear to be the primary cause of dominant and recessive dystrophic epidermolysis bullosa. Two hemidesmosomal components are the bullous pemphigoid (BP) antigens: BP230 shows homology to desmoplakin, a desmosomal component; BP180 contains extracellular collagen domains. The autoantigens in cicatricial pemphigiod and IgA-mediated autoimmune diseases are less well understood. Type IV collagen chains are affected in Alport's and Goodpasture's syndromes.New diagnostic and therapeutic techniques based on these genetic/biochemical advances are currently being developed.

  View details for PubMedID 8250577

• BASEMENT-MEMBRANE PROTEINS KALININ AND NICEIN ARE STRUCTURALLY AND IMMUNOLOGICALLY IDENTICAL LABORATORY INVESTIGATION Marinkovich, M. P., Verrando, P., Keene, D. R., Meneguzzi, G., Lunstrum, G. P., Ortonne, J. P., Burgeson, R. E. 1993; 69 (3): 295-299

  Abstract

  The purpose of this investigation was 2-fold: (a) to compare two recently described proteins, the anchoring filament protein, kalinin and the hemidesmosome-associated protein, nicein (formerly called BM-600) which are both absent in junctional epidermolysis bullosa (JEB) Herlitz's disease; (b) to further define the structural defect in JEB Herlitz's disease.Cultured keratinocytes were analyzed with monoclonal antibodies (mAbs) against kalinin and nicein by indirect immunofluorescence. These mAbs were also used to immunoprecipitate radiolabeled proteins from keratinocyte cultures and to immunoaffinity purify proteins from keratinocyte conditioned culture medium. The precipitated or purified products were compared by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Western blotting, partial V8 protease digestion, and rotary shadowing.Kalinin and nicein mAbs show identical immunofluorescent staining patterns on cultured keratinocytes. Kalinin and nicein mAbs immunoprecipitate peptides from radiolabeled normal human keratinocyte cell and medium fractions that are electrophoretically identical. Partial V8 protease digestion patterns of the reduced 140 kilodalton peptides precipitated by nicein and kalinin mAbs are identical. Kalinin (like nicein) is absent from JEB Herlitz keratinocyte conditioned medium although K-laminin, another anchoring filament component, is present in these cultures. Kalinin, purified from conditioned keratinocyte medium by antibody affinity chromatography with K140-Sepharose (mAb against kalinin) and nicein purified from conditioned keratinocyte medium with GB3-Sepharose (mAb against nicein) are electrophoretically identical by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunologically identical by immunoblotting using kalinin and nicein mAbs. Rotary shadowed images of kalinin and nicein molecules are identical.We demonstrate that kalinin and nicein are identical by biochemical and immunologic analysis. We also verify that kalinin, like nicein, is absent in the conditioned medium of cultured JEB Herlitz keratinocytes, although another anchoring filament protein, K-laminin, is secreted by these cultures. These results correlate with previous immunofluorescent findings that show that while kalinin or nicein is absent in basement membranes of individuals with JEB Herlitz's disease, K-laminin appears to be present.

  View details for PubMedID 8377472

• CELLULAR-ORIGIN OF THE DERMAL-EPIDERMAL BASEMENT-MEMBRANE DEVELOPMENTAL DYNAMICS Marinkovich, M. P., Keene, D. R., RIMBERG, C. S., Burgeson, R. E. 1993; 197 (4): 255-267

  Abstract

  The basement membrane underlying epithelium of skin is generally believed to be of epithelial origin, but a mesenchymal contribution to the basement membrane has not been directly examined. The purpose of this study was to directly evaluate both epithelial and mesenchymal contributions to the basement membrane. Fetal bovine keratinocytes cultured on the surface of collagen gels in the absence of fibroblasts did not produce an ultrastructurally recognizable basement membrane; however, when these cells were cultured in the presence of dermal fibroblasts a basement membrane at the keratinocyte-fibroblast interface was produced after 1 week which was very similar in biochemical composition and ultrastructural appearance to dermal-epidermal basement membrane in human skin. When dual species cultures of bovine keratinocytes and human fibroblasts were analyzed by indirect immunofluorescent microscopy (IF)1 with human specific antibodies against basement membrane components, dermal fibroblasts were shown to synthesize and deposit type IV collagen, type VII collagen, and laminin in a linear manner into the basement membrane zone. Fetal bovine keratinocytes cultured in the presence or absence of fibroblasts synthesized and deposited type IV collagen, type VII collagen, laminin, K-laminin, kalinin, and basement membrane associated heparan sulfate proteoglycan (HSPG) into the underlying basement membrane zone. In organ culture, a subpopulation of fibroblasts initially migrating from human foreskin explants was found to stain strongly for types VII and IV collagen and laminin by IF whereas after subculture all cells showed a uniform low staining. Based on these observations we propose that differentiated fibroblasts exist adjacent to epithelial tissues in vivo which produce basement membrane components and assist in basement membrane assembly.

  View details for PubMedID 8292823

• Kalinin is abnormally expressed in epithelial basement membranes of Herlitz's junctional epidermolysis bullosa patients. Experimental dermatology Meneguzzi, G., Marinkovich, M. P., Aberdam, D., Pisani, A., Burgeson, R., Ortonne, J. P. 1992; 1 (5): 221-229

  Abstract

  Kalinin is an extracellular adhesion molecule specific to epithelial basement membranes (BM) identified as a component of anchoring filaments of hemidesmosomes. This heterotrimeric protein is synthesized by cultured normal human keratinocytes and is involved in cell attachment. In indirect immunofluorescence studies, the epidermal BM of patients with junctional epidermolysis bullosa (JEB) of Herlitz's type were found not to be reactive with the anti-kalinin monoclonal antibodies ka146 and K140 and displayed a decreased immunoreactivity to two anti-kalinin antibodies cross-reacting with K-laminin, an anchoring filament component recently described. The intrinsic defect of JEB keratinocytes in the synthesis of kalinin was further documented by indirect immunofluorescence on in vitro cultures of these cells. In non-Herlitz JEB patients, staining of BM was constantly detected. Impairment of expression of kalinin correlated with the lack of reactivity to the monoclonal antibody GB3, which detects the BM component nicein/BM600. These results clearly demonstrate a defect of kalinin expression in epithelial basement membranes of Herlitz JEB patients and suggest that kalinin may play a role in the pathogenesis of the disease. Further studies are in progress to define possible relationships between kalinin and nicein.

  View details for PubMedID 1365323

• IDENTIFICATION AND PARTIAL-PURIFICATION OF A LARGE, VARIANT FORM OF TYPE-XII COLLAGEN JOURNAL OF BIOLOGICAL CHEMISTRY Lunstrum, G. P., McDonough, A. M., Marinkovich, M. P., Keene, D. R., Morris, N. P., Burgeson, R. E. 1992; 267 (28): 20087-20092

  Abstract

  A large, alternate form of type XII collagen has been identified in cultures of the human epidermoid cell line WISH. This form, designated XIIA, is comprised of alpha chains that are approximately 90 kDa larger than the 220-kDa alpha chain previously characterized in extracts of fetal chicken and bovine tissues. Results from both collagenase digestion and rotary shadow analysis of partially purified material show that the increase is due to a larger NC3 domain. While both the large (XIIA) and the small (XIIB) forms of type XII collagen are identified in pulse-chase radiolabeling of fetal bovine skin explant culture, they are not related in a precursor-product fashion. Inhibition studies with alpha, alpha'-dipyridyl indicate that proper folding of the collagen helix is required for complete assembly and secretion of type XIIA in WISH cell culture. The 310-kDa alpha 1A chain is likely to represent the bovine equivalent of a second translation product, estimated to be 340 kDa, predicted from analysis of one complete chick cDNA sequence. Additionally, the amino-terminal amino acid sequence of the 220-kDa bovine alpha 1B chain was determined. This sequence is very near a potential alternate splice site predicted from analysis of chicken type XII cDNA.

  View details for Web of Science ID A1992JR85800055

  View details for PubMedID 1400326

• THE ANCHORING FILAMENT PROTEIN KALININ IS SYNTHESIZED AND SECRETED AS A HIGH-MOLECULAR-WEIGHT PRECURSOR JOURNAL OF BIOLOGICAL CHEMISTRY Marinkovich, M. P., Lunstrum, G. P., Burgeson, R. E. 1992; 267 (25): 17900-17906

  Abstract

  Kalinin, a recently characterized novel protein component of anchoring filaments, has been shown to be involved in keratinocyte attachment to culture substrates and to dermis in vivo, and to exist in keratinocyte-conditioned culture medium in two heterotrimeric forms of 440 and 400 kDa (Rousselle, P., Lunstrum, G.P., Keene, D.R., and Burgeson, R.E. (1991) J. Cell Biol. 114, 567-576). This study demonstrates that kalinin is initially synthesized in a cell-associated form estimated to be 460 kDa. By second dimension reduced electrophoresis, V8 protease digestion, and immunoblot analysis, we demonstrate that the cell form contains nonidentical subunits of 200, 155, and 140 kDa. The 440-kDa medium form is derived from the cell form by extracellular processing of the 200-kDa subunit to 165 kDa, a step which also occurs in skin organ culture. The 400-kDa form is derived from the 440-kDa form by extracellular processing of the 155 kDa-subunit to 105 kDa. The cell form is secreted by keratinocytes, deposited onto culture substratum, and is the form which facilitates attachment and adhesion of growing and spreading keratinocytes. It is also the form initially synthesized in skin organ culture. Kalinin purified from tissue, which appears to facilitate epithelial-mesenchymal cohesion in vivo, is closely related to the 400-kDa medium form purified from culture.

  View details for Web of Science ID A1992JM22300064

  View details for PubMedID 1517226

• SYNTHESIS OF CALELECTRINS AND CALPACTIN-I DURING CYTOCHALASIN MEDIATED CELL SPREADING INHIBITION CELL CALCIUM Hom, Y. K., Marinkovich, M. P., Lozano, J. J., Rocha, V. 1989; 10 (3): 135-144

  Abstract

  Mammary epithelial cell spreading on collagen gels has previously been shown to be correlated with the synthesis of a group of calcium-binding proteins (CBPs) which we have identified as the calcium-binding proteins termed calelectrins and calpactin I monomer/p36. To determine whether cell spreading per se is required for CBP synthesis, we examined the effect of cytochalasin D on these two events. Concentrations of cytochalasin D that did not reduce total protein synthesis, caused inhibition of cell spreading in a dose-dependent manner, but did not cause inhibition of CBP synthesis. Synthesis of collagen also continued during cytochalasin inhibition of cell spreading. Removal of the inhibitor from the cultures initiated cell spreading and CBP synthesis continued. Membrane-cytoskeleton complexes from control and CD treated cells were identical in regard to binding CBPs in a calcium-dependent manner. Colchicine, which inhibited cell spreading, was shown to be toxic to general protein synthesis at 75 nM. The data clearly indicate that mere inhibition of epithelial cell spreading does not automatically suppress CBP synthesis.

  View details for Web of Science ID A1989T948700002

  View details for PubMedID 2524259

• BASAL LAMINA INHIBITION SUPPRESSES SYNTHESIS OF CALCIUM-DEPENDENT PROTEINS ASSOCIATED WITH MAMMARY EPITHELIAL-CELL SPREADING EXPERIMENTAL CELL RESEARCH Rocha, V., Hom, Y. K., Marinkovich, M. P. 1986; 165 (2): 450-460

  Abstract

  Spreading of mouse mammary epithelial cells on collagen gels is closely correlated with the synthesis of a group of putative calcium-binding proteins (CBP) (Braslau et al., Exp cell res 155 (1984) 213). Collagen synthesis was shown to occur during cell spreading, while omission of serum prevented cell spreading and the synthesis of collagen. The proline analogues cis-hydroxyproline and L-azetidine-2-carboxylic acid were shown to inhibit epithelial cell spreading and to suppress the collagen synthesis that occurs during serum-supported cell spreading. Inhibition of collagen synthesis resulted in the inhibition of CBP synthesis associated with cell spreading. In contrast, the collagen cross-linking inhibitor B-aminopropionitrile did not inhibit cell spreading nor did it suppress collagen synthesis; CBP synthesis was also normal during treatment with this inhibitor. Thus, mammary epithelial cell spreading on collagen gels and CBP synthesis can both be suppressed by inhibition of collagen synthesis indicating that they may be integrated in some manner. It is suggested that inhibition of cell spreading during inhibition of collagen synthesis results from failure to assemble a normal basal lamina; this may in turn signal suppression of CBP synthesis.

  View details for Web of Science ID A1986D238000016

  View details for PubMedID 3720859

• COLLAGEN-SYNTHESIS AND DEPOSITION DURING MAMMARY EPITHELIAL-CELL SPREADING ON COLLAGEN GELS JOURNAL OF CELLULAR PHYSIOLOGY Marinkovich, M. P., Rocha, V. 1986; 128 (1): 61-70

  Abstract

  Mouse mammary epithelial cultivated on collagen gels demonstrate active spreading as the cells form monolayers. In this novel system, initiation of cell spreading is preceded by de novo synthesis of type IV collagen. The newly synthesized collagen is partitioned such that after 48 hr, approximately 24% is found in the culture medium, 35% is intracellular, and 41% is deposited in the extracellular matrix of the developing epithelium. Cultures deprived of serum failed to spread and to synthesize collagen. Proline analogues were shown to inhibit cell spreading and to suppress collagen synthesis in a dose-dependent manner. Cytochalasin D inhibition of F-actin elongation was shown to prevent cell spreading but not to suppress total collagen synthesis. During cytochalasin D treatment, inhibition of cell spreading was shown to result from failure to deposit or to maintain deposited collagen in the epithelium extracellular matrix. The data indicate that synthesis and extracellular deposition of a major basal lamina component (viz. type IV collagen) must precede and then accompany epithelial cell spreading in collagen gel culture. It is suggested that the microfilament apparatus, through some hypothetical integral membrane protein, can anchor extracellular type IV collagen, which then provides a necessary condition for cell spreading.

  View details for Web of Science ID A1986D155700010

  View details for PubMedID 3722273