Glenn Rosen

All Publications

• Defining Improvement in Nonalcoholic Steatohepatitis for Treatment Trial Endpoints: Recommendations From the Liver Forum HEPATOLOGY Cheung, A., Neuschwander-Tetri, B. A., Kleiner, D. E., Schabel, E., Rinella, M., Harrison, S., Ratziu, V., Sanyal, A. J., Loomba, R., Megnien, S., Torstenson, R., Miller, V., Abdelmalek, M., Anstee, Q., Banerjee, R., Bashir, M., Bedossa, P., Berner-Hansen, M., Chakravarthy, M., Charles, E., Dimick-Santos, L., Ertle, J., Francque, S., Friedman, S., Gannedahl, G., Greene, K., Hambleton, M., Hum, D., Imperial, J., Leeming, D., Mehta, R., Naoumov, N., Omokaro, S., Palmer, M., Peres, D., Powell, M., Regev, A., Rosen, G., Sanyal, A., Schoelch, C., Schwimmer, J., Shringarpure, R., Schwander-Tetri, B., Ukomadu, C., Wong, V., Wright, T., Chan, J., Tai, D., Liver Forum Case Definitions Worki 2019; 70 (5): 1841–55

  View details for DOI 10.1002/hep.30672

  View details for Web of Science ID 000493023400026

• Defining Improvement in Nonalcoholic Steatohepatitis for Treatment Trial Endpoints: Recommendations from the Liver Forum. Hepatology (Baltimore, Md.) Cheung, A., Neuschwander-Tetri, B. A., Kleiner, D. E., Schabel, E., Rinella, M., Harrison, S., Ratziu, V., Sanyal, A. J., Loomba, R., Jeannin Megnien, S., Torstenson, R., Miller, V., Liver Forum Case Definitions Working Group, Abdelmalek, M., Anstee, Q., Banerjee, R., Bashir, M., Bedossa, P., Berner-Hansen, M., Chakravarthy, M., Chan, J., Charles, E., Cheung, A., Dimick-Santos, L., Ertle, J., Francque, S., Friedman, S., Gannedahl, G., Greene, K., Hambleton, M., Harrison, S., Hum, D., Imperial, J., Kleiner, D., Leeming, D. J., Loomba, R., Megnien, S. J., Mehta, R., Miller, V., Naoumov, N., Omokaro, S., Palmer, M., Peres, D., Powell, M., Ratziu, V., Regev, A., Rinella, M., Rosen, G., Sanyal, A., Schabel, E., Scholch, C., Schwimmer, J., Shapiro, D., Shringarpure, R., Tai, D., Neuschwander-Tetri, B., Torstenson, R., Ukomadu, C., Wong, V., Wright, T. 2019

  Abstract

  Identifying effective therapies for nonalcoholic steatohepatitis (NASH) with fibrosis is a pressing challenge, with 1-2% of the population in developed nations at risk of developing NASH cirrhosis and its complications. The design of NASH clinical therapeutic trials is hampered by the long period of minimally symptomatic disease that typically precedes the development of decompensated cirrhosis, and the accompanying uncertainties regarding the best pre-cirrhotic trial endpoints that reliably reflect a subsequent reduction in liver-related morbidity and mortality. The Liver Forum is a multi-stakeholder organization comprised of academic, industry, and regulatory experts working from a regulatory science perspective to identify barriers, prioritize research, and identify solutions to accelerate therapeutic development for NASH. Prior work of The Liver Forum has focused on recommendations for disease definitions and baseline parameters to be implemented in clinical trials that are designed to assess disease status and prevent progression to cirrhosis, liver transplantation, hepatocellular carcinoma, and death. The purpose of this summary is to review currently available clinical data to identify parameters that change in parallel with liver histology and are likely to reflect clinically meaningful reductions in the risk of developing cirrhosis and its complications. We review available data on exploratory histologic, blood-based and imaging pharmacodynamic biomarkers that may reflect meaningful treatment responses and provide recommendations regarding measurements to be considered in phase 2 and 3 trials as well as during post-marketing monitoring trials. This article is protected by copyright. All rights reserved.

  View details for PubMedID 31034092

• Non-invasive Imaging of Idiopathic Pulmonary Fibrosis Using Cathepsin Protease Probes SCIENTIFIC REPORTS Withana, N. P., Ma, X., McGuire, H. M., Verdoes, M., van der Linden, W. A., Ofori, L. O., Zhang, R., Li, H., Sanman, L. E., Wei, K., Yao, S., Wu, P., Li, F., Huang, H., Xu, Z., Wolters, P. J., Rosen, G. D., Collard, H. R., Zhu, Z., Cheng, Z., Bogyo, M. 2016; 6

  Abstract

  Idiopathic pulmonary fibrosis (IPF) is a lethal, chronic, progressive disease characterized by formation of scar tissue within the lungs. Because it is a disease of unknown etiology, it is difficult to diagnose, to predict disease course and to devise treatment strategies. Recent evidence suggests that activated macrophages play key roles in the pathology of IPF. Therefore, imaging probes that specifically recognize these pools of activated immune cells could provide valuable information about how these cells contribute to the pathobiology of the disease. Here we demonstrate that cysteine cathepsin-targeted imaging probes can be used to monitor the contribution of macrophages to fibrotic disease progression in the bleomycin-induced murine model of pulmonary fibrosis. Furthermore, we show that the probes highlight regions of macrophage involvement in fibrosis in human biopsy tissues from IPF patients. Finally, we present first-in-human results demonstrating non-invasive imaging of active cathepsins in fibrotic lesions of patients with IPF. Together, our findings validate small molecule cysteine cathepsin probes for clinical PET imaging and suggest that they have the potential to be used to generate mechanistically-informative molecular information regarding cellular drivers of IPF disease severity and progression.

  View details for DOI 10.1038/srep19755

  View details for Web of Science ID 000368667700001

  View details for PubMedCentralID PMC4726431

• Non-invasive Imaging of Idiopathic Pulmonary Fibrosis Using Cathepsin Protease Probes. Scientific reports Withana, N. P., Ma, X., McGuire, H. M., Verdoes, M., van der Linden, W. A., Ofori, L. O., Zhang, R., Li, H., Sanman, L. E., Wei, K., Yao, S., Wu, P., Li, F., Huang, H., Xu, Z., Wolters, P. J., Rosen, G. D., Collard, H. R., Zhu, Z., Cheng, Z., Bogyo, M. 2016; 6: 19755-?

  Abstract

  Idiopathic pulmonary fibrosis (IPF) is a lethal, chronic, progressive disease characterized by formation of scar tissue within the lungs. Because it is a disease of unknown etiology, it is difficult to diagnose, to predict disease course and to devise treatment strategies. Recent evidence suggests that activated macrophages play key roles in the pathology of IPF. Therefore, imaging probes that specifically recognize these pools of activated immune cells could provide valuable information about how these cells contribute to the pathobiology of the disease. Here we demonstrate that cysteine cathepsin-targeted imaging probes can be used to monitor the contribution of macrophages to fibrotic disease progression in the bleomycin-induced murine model of pulmonary fibrosis. Furthermore, we show that the probes highlight regions of macrophage involvement in fibrosis in human biopsy tissues from IPF patients. Finally, we present first-in-human results demonstrating non-invasive imaging of active cathepsins in fibrotic lesions of patients with IPF. Together, our findings validate small molecule cysteine cathepsin probes for clinical PET imaging and suggest that they have the potential to be used to generate mechanistically-informative molecular information regarding cellular drivers of IPF disease severity and progression.

  View details for DOI 10.1038/srep19755

  View details for PubMedID 26797565

• Pleiotropic effect of the proton pump inhibitor esomeprazole leading to suppression of lung inflammation and fibrosis JOURNAL OF TRANSLATIONAL MEDICINE Ghebremariam, Y. T., Cooke, J. P., Gerhart, W., Griego, C., Brower, J. B., Doyle-Eisele, M., Moeller, B. C., Zhou, Q., Ho, L., De Andrade, J., Raghu, G., Peterson, L., Rivera, A., Rosen, G. D. 2015; 13

  Abstract

  The beneficial outcome associated with the use of proton pump inhibitors (PPIs) in idiopathic pulmonary fibrosis (IPF) has been reported in retrospective studies. To date, no prospective study has been conducted to confirm these outcomes. In addition, the potential mechanism by which PPIs improve measures of lung function and/or transplant-free survival in IPF has not been elucidated.Here, we used biochemical, cell biological and preclinical studies to evaluate regulation of markers associated with inflammation and fibrosis. In our in vitro studies, we exposed primary lung fibroblasts, epithelial and endothelial cells to ionizing radiation or bleomycin; stimuli typically used to induce inflammation and fibrosis. In addition, we cultured lung fibroblasts from IPF patients and studied the effect of esomeprazole on collagen release. Our preclinical study tested efficacy of esomeprazole in a rat model of bleomycin-induced lung injury. Furthermore, we performed retrospective analysis of interstitial lung disease (ILD) databases to examine the effect of PPIs on transplant-free survival.The cell culture studies revealed that esomeprazole controls inflammation by suppressing the expression of pro-inflammatory molecules including vascular cell adhesion molecule-1, inducible nitric oxide synthase, tumor necrosis factor-alpha (TNF-α) and interleukins (IL-1β and IL-6). The antioxidant effect is associated with strong induction of the stress-inducible cytoprotective protein heme oxygenase-1 (HO1) and the antifibrotic effect is associated with potent inhibition of fibroblast proliferation as well as downregulation of profibrotic proteins including receptors for transforming growth factor β (TGFβ), fibronectin and matrix metalloproteinases (MMPs). Furthermore, esomeprazole showed robust effect in mitigating the inflammatory and fibrotic responses in a murine model of acute lung injury. Finally, retrospective analysis of two ILD databases was performed to assess the effect of PPIs on transplant-free survival in IPF patients. Intriguingly, this data demonstrated that IPF patients on PPIs had prolonged survival over controls (median survival of 3.4 vs 2 years).Overall, these data indicate the possibility that PPIs may have protective function in IPF by directly modulating the disease process and suggest that they may have other clinical utility in the treatment of extra-intestinal diseases characterized by inflammatory and/or fibrotic phases.

  View details for DOI 10.1186/s12967-015-0614-x

  View details for Web of Science ID 000358774500001

  View details for PubMedID 26231702

  View details for PubMedCentralID PMC4522053

• Automated classification of usual interstitial pneumonia using regional volumetric texture analysis in high-resolution computed tomography. Investigative radiology Depeursinge, A., Chin, A. S., Leung, A. N., Terrone, D., Bristow, M., Rosen, G., Rubin, D. L. 2015; 50 (4): 261-267

  Abstract

  We propose a novel computational approach for the automated classification of classic versus atypical usual interstitial pneumonia (UIP).Thirty-three patients with UIP were enrolled in this study. They were classified as classic versus atypical UIP by a consensus of 2 thoracic radiologists with more than 15 years of experience using the American Thoracic Society evidence-based guidelines for computed tomography diagnosis of UIP. Two cardiothoracic fellows with 1 year of subspecialty training provided independent readings. The system is based on regional characterization of the morphological tissue properties of lung using volumetric texture analysis of multiple-detector computed tomography images. A simple digital atlas with 36 lung subregions is used to locate texture properties, from which the responses of multidirectional Riesz wavelets are obtained. Machine learning is used to aggregate and to map the regional texture attributes to a simple score that can be used to stratify patients with UIP into classic and atypical subtypes.We compared the predictions on the basis of regional volumetric texture analysis with the ground truth established by expert consensus. The area under the receiver operating characteristic curve of the proposed score was estimated to be 0.81 using a leave-one-patient-out cross-validation, with high specificity for classic UIP. The performance of our automated method was found to be similar to that of the 2 fellows and to the agreement between experienced chest radiologists reported in the literature. However, the errors of our method and the fellows occurred on different cases, which suggests that combining human and computerized evaluations may be synergistic.Our results are encouraging and suggest that an automated system may be useful in routine clinical practice as a diagnostic aid for identifying patients with complex lung disease such as classic UIP, obviating the need for invasive surgical lung biopsy and its associated risks.

  View details for DOI 10.1097/RLI.0000000000000127

  View details for PubMedID 25551822

  View details for PubMedCentralID PMC4355184

• Proton pump inhibitors inhibit the cardiovascular enzyme DDAH and regulate processes underlying idiopathic pulmonary fibrosis Ghebremariam, Y. T., Zhou, Q., de Andrade, J., Ho, L., Rosen, G. D., Cooke, J. P. SAGE PUBLICATIONS LTD. 2014: 223

  View details for Web of Science ID 000337579800012

• Connective tissue disease related interstitial lung diseases and idiopathic pulmonary fibrosis: provisional core sets of domains and instruments for use in clinical trials THORAX Saketkoo, L. A., Mittoo, S., Huscher, D., Khanna, D., Dellaripa, P. F., Distler, O., Flaherty, K. R., Frankel, S., Oddis, C. V., Denton, C. P., Fischer, A., Kowal-Bielecka, O. M., LeSage, D., Merkel, P. A., Phillips, K., Pittrow, D., Swigris, J., Antoniou, K., Baughman, R. P., Castelino, F. V., Christmann, R. B., Christopher-Stine, L., Collard, H. R., Cottin, V., Danoff, S., Highland, K. B., Hummers, L., Shah, A. A., Kim, D. S., Lynch, D. A., Miller, F. W., Proudman, S. M., Richeldi, L., Ryu, J. H., Sandorfi, N., Sarver, C., Wells, A. U., Strand, V., Matteson, E. L., Brown, K. K., Seibold, J. R. 2014; 69 (5): 428-436

  Abstract

  Clinical trial design in interstitial lung diseases (ILDs) has been hampered by lack of consensus on appropriate outcome measures for reliably assessing treatment response. In the setting of connective tissue diseases (CTDs), some measures of ILD disease activity and severity may be confounded by non-pulmonary comorbidities.The Connective Tissue Disease associated Interstitial Lung Disease (CTD-ILD) working group of Outcome Measures in Rheumatology-a non-profit international organisation dedicated to consensus methodology in identification of outcome measures-conducted a series of investigations which included a Delphi process including >248 ILD medical experts as well as patient focus groups culminating in a nominal group panel of ILD experts and patients. The goal was to define and develop a consensus on the status of outcome measure candidates for use in randomised controlled trials in CTD-ILD and idiopathic pulmonary fibrosis (IPF).A core set comprising specific measures in the domains of lung physiology, lung imaging, survival, dyspnoea, cough and health-related quality of life is proposed as appropriate for consideration for use in a hypothetical 1-year multicentre clinical trial for either CTD-ILD or IPF. As many widely used instruments were found to lack full validation, an agenda for future research is proposed.Identification of consensus preliminary domains and instruments to measure them was attained and is a major advance anticipated to facilitate multicentre RCTs in the field.

  View details for DOI 10.1136/thoraxjnl-2013-204202

  View details for Web of Science ID 000347648600009

  View details for PubMedID 24368713

  View details for PubMedCentralID PMC3995282

• Reconciling Healthcare Professional and Patient Perspectives in the Development of Disease Activity and Response Criteria in Connective Tissue Disease-related Interstitial Lung Diseases JOURNAL OF RHEUMATOLOGY Saketkoo, L. A., Mittoo, S., Frankel, S., LeSage, D., Sarver, C., Phillips, K., Strand, V., Matteson, E. L. 2014; 41 (4): 792-798

  Abstract

  Interstitial lung diseases (ILD), including those related to connective tissue disease (CTD), and idiopathic pulmonary fibrosis (IPF) carry high morbidity and mortality. Great efforts are under way to develop and investigate meaningful treatments in the context of clinical trials. However, efforts have been challenged by a lack of validated outcome measures and by inconsistent use of measures in clinical trials. Lack of consensus has fragmented effective use of strategies in CTD-ILD and IPF, with a history of resultant difficulties in obtaining agency approval of treatment interventions. Until recently, the patient perspective to determine domains and outcome measures in CTD-ILD and IPF had never been applied. Efforts described here demonstrate unequivocally the value and influence of patient involvement on core set development. Regarding CTD-ILD, this is the first OMERACT working group to directly address a manifestation/comorbidity of a rheumatic disease (ILD) as well as a disease not considered rheumatic (IPF). The OMERACT 11 proceedings of the CTD-ILD Working Group describe the forward and lateral process to include both the medical and patient perspectives in the urgently needed identification of a core set of preliminary domains and outcome measures in CTD-ILD and IPF.

  View details for DOI 10.3899/jrheum.131251

  View details for Web of Science ID 000333906400024

  View details for PubMedID 24488412

  View details for PubMedCentralID PMC4369780

• Transcriptome analysis reveals differential splicing events in IPF lung tissue. PloS one Nance, T., Smith, K. S., Anaya, V., Richardson, R., Ho, L., Pala, M., Mostafavi, S., Battle, A., Feghali-Bostwick, C., Rosen, G., Montgomery, S. B. 2014; 9 (3)

  Abstract

  Idiopathic pulmonary fibrosis (IPF) is a complex disease in which a multitude of proteins and networks are disrupted. Interrogation of the transcriptome through RNA sequencing (RNA-Seq) enables the determination of genes whose differential expression is most significant in IPF, as well as the detection of alternative splicing events which are not easily observed with traditional microarray experiments. We sequenced messenger RNA from 8 IPF lung samples and 7 healthy controls on an Illumina HiSeq 2000, and found evidence for substantial differential gene expression and differential splicing. 873 genes were differentially expressed in IPF (FDR<5%), and 440 unique genes had significant differential splicing events in at least one exonic region (FDR<5%). We used qPCR to validate the differential exon usage in the second and third most significant exonic regions, in the genes COL6A3 (RNA-Seq adjusted pval = 7.18e-10) and POSTN (RNA-Seq adjusted pval = 2.06e-09), which encode the extracellular matrix proteins collagen alpha-3(VI) and periostin. The increased gene-level expression of periostin has been associated with IPF and its clinical progression, but its differential splicing has not been studied in the context of this disease. Our results suggest that alternative splicing of these and other genes may be involved in the pathogenesis of IPF. We have developed an interactive web application which allows users to explore the results of our RNA-Seq experiment, as well as those of two previously published microarray experiments, and we hope that this will serve as a resource for future investigations of gene regulation in IPF.

  View details for DOI 10.1371/journal.pone.0092111

  View details for PubMedID 24647608

• A Critical Role for the mTORC2 Pathway in Lung Fibrosis. PloS one Chang, W., Wei, K., Ho, L., Berry, G. J., Jacobs, S. S., Chang, C. H., Rosen, G. D. 2014; 9 (8): e106155

  Abstract

  A characteristic of dysregulated wound healing in IPF is fibroblastic-mediated damage to lung epithelial cells within fibroblastic foci. In these foci, TGF-β and other growth factors activate fibroblasts that secrete growth factors and matrix regulatory proteins, which activate a fibrotic cascade. Our studies and those of others have revealed that Akt is activated in IPF fibroblasts and it mediates the activation by TGF-β of pro-fibrotic pathways. Recent studies show that mTORC2, a component of the mTOR pathway, mediates the activation of Akt. In this study we set out to determine if blocking mTORC2 with MLN0128, an active site dual mTOR inhibitor, which blocks both mTORC1 and mTORC2, inhibits lung fibrosis. We examined the effect of MLN0128 on TGF-β-mediated induction of stromal proteins in IPF lung fibroblasts; also, we looked at its effect on TGF-β-mediated epithelial injury using a Transwell co-culture system. Additionally, we assessed MLN0128 in the murine bleomycin lung model. We found that TGF-β induces the Rictor component of mTORC2 in IPF lung fibroblasts, which led to Akt activation, and that MLN0128 exhibited potent anti-fibrotic activity in vitro and in vivo. Also, we observed that Rictor induction is Akt-mediated. MLN0128 displays multiple anti-fibrotic and lung epithelial-protective activities; it (1) inhibited the expression of pro-fibrotic matrix-regulatory proteins in TGF-β-stimulated IPF fibroblasts; (2) inhibited fibrosis in a murine bleomycin lung model; and (3) protected lung epithelial cells from injury caused by TGF-β-stimulated IPF fibroblasts. Our findings support a role for mTORC2 in the pathogenesis of lung fibrosis and for the potential of active site mTOR inhibitors in the treatment of IPF and other fibrotic lung diseases.

  View details for DOI 10.1371/journal.pone.0106155

  View details for PubMedID 25162417

  View details for PubMedCentralID PMC4146613

• A critical role for the mTORC2 pathway in lung fibrosis. PloS one Chang, W., Wei, K., Ho, L., Berry, G. J., Jacobs, S. S., Chang, C. H., Rosen, G. D. 2014; 9 (8)

  Abstract

  A characteristic of dysregulated wound healing in IPF is fibroblastic-mediated damage to lung epithelial cells within fibroblastic foci. In these foci, TGF-β and other growth factors activate fibroblasts that secrete growth factors and matrix regulatory proteins, which activate a fibrotic cascade. Our studies and those of others have revealed that Akt is activated in IPF fibroblasts and it mediates the activation by TGF-β of pro-fibrotic pathways. Recent studies show that mTORC2, a component of the mTOR pathway, mediates the activation of Akt. In this study we set out to determine if blocking mTORC2 with MLN0128, an active site dual mTOR inhibitor, which blocks both mTORC1 and mTORC2, inhibits lung fibrosis. We examined the effect of MLN0128 on TGF-β-mediated induction of stromal proteins in IPF lung fibroblasts; also, we looked at its effect on TGF-β-mediated epithelial injury using a Transwell co-culture system. Additionally, we assessed MLN0128 in the murine bleomycin lung model. We found that TGF-β induces the Rictor component of mTORC2 in IPF lung fibroblasts, which led to Akt activation, and that MLN0128 exhibited potent anti-fibrotic activity in vitro and in vivo. Also, we observed that Rictor induction is Akt-mediated. MLN0128 displays multiple anti-fibrotic and lung epithelial-protective activities; it (1) inhibited the expression of pro-fibrotic matrix-regulatory proteins in TGF-β-stimulated IPF fibroblasts; (2) inhibited fibrosis in a murine bleomycin lung model; and (3) protected lung epithelial cells from injury caused by TGF-β-stimulated IPF fibroblasts. Our findings support a role for mTORC2 in the pathogenesis of lung fibrosis and for the potential of active site mTOR inhibitors in the treatment of IPF and other fibrotic lung diseases.

  View details for DOI 10.1371/journal.pone.0106155

  View details for PubMedID 25162417

  View details for PubMedCentralID PMC4146613

• Transcriptome Analysis Reveals Differential Splicing Events in IPF Lung Tissue. PloS one Nance, T., Smith, K. S., Anaya, V., Richardson, R., Ho, L., Pala, M., Mostafavi, S., Battle, A., Feghali-Bostwick, C., Rosen, G., Montgomery, S. B. 2014; 9 (3): e92111

  Abstract

  Idiopathic pulmonary fibrosis (IPF) is a complex disease in which a multitude of proteins and networks are disrupted. Interrogation of the transcriptome through RNA sequencing (RNA-Seq) enables the determination of genes whose differential expression is most significant in IPF, as well as the detection of alternative splicing events which are not easily observed with traditional microarray experiments. We sequenced messenger RNA from 8 IPF lung samples and 7 healthy controls on an Illumina HiSeq 2000, and found evidence for substantial differential gene expression and differential splicing. 873 genes were differentially expressed in IPF (FDR<5%), and 440 unique genes had significant differential splicing events in at least one exonic region (FDR<5%). We used qPCR to validate the differential exon usage in the second and third most significant exonic regions, in the genes COL6A3 (RNA-Seq adjusted pval = 7.18e-10) and POSTN (RNA-Seq adjusted pval = 2.06e-09), which encode the extracellular matrix proteins collagen alpha-3(VI) and periostin. The increased gene-level expression of periostin has been associated with IPF and its clinical progression, but its differential splicing has not been studied in the context of this disease. Our results suggest that alternative splicing of these and other genes may be involved in the pathogenesis of IPF. We have developed an interactive web application which allows users to explore the results of our RNA-Seq experiment, as well as those of two previously published microarray experiments, and we hope that this will serve as a resource for future investigations of gene regulation in IPF.

  View details for DOI 10.1371/journal.pone.0092111

  View details for PubMedID 24647608

  View details for PubMedCentralID PMC3960165

• Proton Pump Inhibitors Inhibit DDAH and Processes Underlying Idiopathic Pulmonary Fibrosis Ghebrernariam, Y. T., Ho, L., Rosen, G. D., Cooke, J. P. LIPPINCOTT WILLIAMS & WILKINS. 2013

  View details for Web of Science ID 000332162900435

• Connective Tissue Disease-associated Interstitial Lung Disease: A review. Current respiratory care reports Gutsche, M. n., Rosen, G. D., Swigris, J. J. 2012; 1: 224–32

  Abstract

  Interstitial lung disease (ILD) is commonly encountered in patients with connective tissue diseases (CTD). Besides the lung parenchyma, the airways, pulmonary vasculature and structures of the chest wall may all be involved, depending on the type of CTD. As a result of this so-called multi-compartment involvement, airflow limitation, pulmonary hypertension, vasculitis and extrapulmonary restriction can occur alongside fibro-inflammatory parenchymal abnormalities in CTD. Rheumatoid arthritis (RA), systemic sclerosis (SSc), poly-/dermatomyositis (PM/DM), Sjögren's syndrome (SjS), systemic lupus erythematosus (SLE), and undifferentiated (UCTD) as well as mixed connective tissue disease (MCTD) can all be associated with the development of ILD. Non-specific interstitial pneumonia (NSIP) is the most commonly observed histopathological pattern in CTD-ILD, but other patterns including usual interstitial pneumonia (UIP), organizing pneumonia (OP), diffuse alveolar damage (DAD) and lymphocytic interstitial pneumonia (LIP) may occur. Although the majority of patients with CTD-ILD experience stable or slowly advancing ILD, a small yet significant group exhibits a more severe and progressive course. Randomized placebo-controlled trials evaluating the efficacy of immunomodulatory treatments have been conducted only in SSc-associated ILD. However, clinical experience suggests that a handful of immunosuppressive medications are potentially effective in a sizeable portion of patients with ILD caused by other CTDs. In this manuscript, we review the clinical characteristics and management of the most common CTD-ILDs.

  View details for PubMedID 23125954

  View details for PubMedCentralID PMC3486427

• An Analysis of Connective Tissue Disease-associated Interstitial Lung Disease at a US Tertiary Care Center: Better Survival in Patients with Systemic Sclerosis JOURNAL OF RHEUMATOLOGY Su, R., Bennett, M., Jacobs, S., Hunter, T., Bailey, C., Krishnan, E., Rosen, G., Chung, L. 2011; 38 (4): 693-701

  Abstract

  To compare survival of patients with connective tissue disease-associated interstitial lung disease (CTD-ILD) versus idiopathic pulmonary fibrosis (IPF) and patients with systemic sclerosis-associated ILD (SSc-ILD) versus other CTD-ILD followed at our center.We used the Stanford ILD database, which contains prospectively collected information on patients with ILD evaluated at our tertiary care center from 2002 to 2009. Survival at last followup from time of ILD diagnosis was calculated using the Kaplan-Meier estimator. Prognostic factors for survival in the overall cohort (IPF and CTD-ILD) and in the CTD-ILD group were identified with univariate and multivariate Cox regression models.Of 427 patients with ILD, 148 (35%) had IPF and 76 (18%) had CTD-ILD at the baseline visit. The cumulative incidence of CTD was 4%. After a median followup of 4 years, 67 patients (36.4%) had died and 4 (2.2%) were lost to followup. Patients with IPF (n = 122) and CTD-ILD (n = 62) experienced similar survival rates (5-year survival about 50%). Patients with SSc-ILD (n = 24) experienced better survival than those with other CTD-ILD (n = 38), with 1-year, 3-year, and 5-year survival rates of 100%, 90%, and 77%, respectively, versus 78%, 42%, and 38% (p = 0.01). The presence of SSc in patients with CTD-ILD decreased the risk of death by > 80% even after correcting for age at ILD diagnosis, sex, and ethnicity (HR = 0.17, 95% CI 0.04-0.83).Survival in patients with SSc-ILD was better than in patients with other CTD-ILD, potentially related to routine screening for and early detection of ILD in patients with SSc at our center.

  View details for DOI 10.3899/jrheum.100675

  View details for Web of Science ID 000289333800018

  View details for PubMedID 21285162

• The HLA Class II Allele DRB1*1501 Is Over-Represented in Patients with Idiopathic Pulmonary Fibrosis PLOS ONE Xue, J., Gochuico, B. R., Alawad, A. S., Feghali-Bostwick, C. A., Noth, I., Nathan, S. D., Rosen, G. D., Rosas, I. O., Dacic, S., Ocak, I., Fuhrman, C. R., Cuenco, K. T., Smith, M. A., Jacobs, S. S., Zeevi, A., Morel, P. A., Pilewski, J. M., Valentine, V. G., Gibson, K. F., Kaminski, N., Sciurba, F. C., Zhang, Y., Duncan, S. R. 2011; 6 (2)

  Abstract

  Idiopathic pulmonary fibrosis (IPF) is a progressive and medically refractory lung disease with a grim prognosis. Although the etiology of IPF remains perplexing, abnormal adaptive immune responses are evident in many afflicted patients. We hypothesized that perturbations of human leukocyte antigen (HLA) allele frequencies, which are often seen among patients with immunologic diseases, may also be present in IPF patients.HLA alleles were determined in subpopulations of IPF and normal subjects using molecular typing methods. HLA-DRB1*15 was over-represented in a discovery cohort of 79 Caucasian IPF subjects who had lung transplantations at the University of Pittsburgh (36.7%) compared to normal reference populations. These findings were prospectively replicated in a validation cohort of 196 additional IPF subjects from four other U.S. medical centers that included both ambulatory patients and lung transplantation recipients. High-resolution typing was used to further define specific HLA-DRB1*15 alleles. DRB1*1501 prevalence in IPF subjects was similar among the 143 ambulatory patients and 132 transplant recipients (31.5% and 34.8%, respectively, p = 0.55). The aggregate prevalence of DRB1*1501 in IPF patients was significantly greater than among 285 healthy controls (33.1% vs. 20.0%, respectively, OR 2.0; 95%CI 1.3-2.9, p = 0.0004). IPF patients with DRB1*1501 (n = 91) tended to have decreased diffusing capacities for carbon monoxide (DL(CO)) compared to the 184 disease subjects who lacked this allele (37.8±1.7% vs. 42.8±1.4%, p = 0.036).DRB1*1501 is more prevalent among IPF patients than normal subjects, and may be associated with greater impairment of gas exchange. These data are novel evidence that immunogenetic processes can play a role in the susceptibility to and/or manifestations of IPF. Findings here of a disease association at the HLA-DR locus have broad pathogenic implications, illustrate a specific chromosomal area for incremental, targeted genomic study, and may identify a distinct clinical phenotype among patients with this enigmatic, morbid lung disease.

  View details for DOI 10.1371/journal.pone.0014715

  View details for Web of Science ID 000287657500003

  View details for PubMedID 21373184

  View details for PubMedCentralID PMC3044131

• SPARC Suppresses Apoptosis of Idiopathic Pulmonary Fibrosis Fibroblasts through Constitutive Activation of beta-Catenin JOURNAL OF BIOLOGICAL CHEMISTRY Chang, W., Wei, K., Jacobs, S. S., Upadhyay, D., Weill, D., Rosen, G. D. 2010; 285 (11): 8196-8206

  Abstract

  Idiopathic pulmonary fibrosis (IPF) is a poorly understood progressive disease characterized by the accumulation of scar tissue in the lung interstitium. A hallmark of the disease is areas of injury to type II alveolar epithelial cells with attendant accumulation of fibroblasts in areas called fibroblastic foci. In an effort to better characterize the lung fibroblast phenotype in IPF patients, we isolated fibroblasts from patients with IPF and looked for activation of signaling proteins, which could help explain the exaggerated fibrogenic response in IPF. We found that IPF fibroblasts constitutively expressed increased basal levels of SPARC, plasminogen activator inhibitor-1 (PAI-1), and active beta-catenin compared with control cells. Control of basal PAI-1 expression in IPF fibroblasts was regulated by SPARC-mediated activation of Akt, leading to inhibition of glycogen synthase kinase-3beta and activation of beta-catenin. Additionally, IPF fibroblasts (but not control fibroblasts) were resistant to plasminogen-induced apoptosis and were sensitized to plasminogen-mediated apoptosis by inhibition of SPARC or beta-catenin. These findings uncover a newly discovered regulatory pathway in IPF fibroblasts that is characterized by elevated SPARC, giving rise to activated beta-catenin, which regulates expression of downstream genes, such as PAI-1, and confers an apoptosis-resistant phenotype. Disruption of this pathway may represent a novel therapeutic target in IPF.

  View details for DOI 10.1074/jbc.M109.025684

  View details for Web of Science ID 000275413800038

  View details for PubMedID 20061390

  View details for PubMedCentralID PMC2832971

• Nicotine Induces Resistance to Chemotherapy by Modulating Mitochondrial Signaling in Lung Cancer AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Zhang, J., Kamdar, O., Le, W., Rosen, G. D., Upadhyay, D. 2009; 40 (2): 135-146

  Abstract

  Continued smoking causes tumor progression and resistance to therapy in lung cancer. Carcinogens possess the ability to block apoptosis, and thus may induce development of cancers and resistance to therapy. Tobacco carcinogens have been studied widely; however, little is known about the agents that inhibit apoptosis, such as nicotine. We determine whether mitochondrial signaling mediates antiapoptotic effects of nicotine in lung cancer. A549 cells were exposed to nicotine (1 muM) followed by cisplatin (35 muM) plus etoposide (20 muM) for 24 hours. We found that nicotine prevented chemotherapy-induced apoptosis, improved cell survival, and caused modest increases in DNA synthesis. Inhibition of mitogen-activated protein kinase (MAPK) and Akt prevented the antiapoptotic effects of nicotine and decreased chemotherapy-induced apoptosis. Small interfering RNA MAPK kinase-1 blocked antiapoptotic effects of nicotine, whereas small interfering RNA MAPK kinase-2 blocked chemotherapy-induced apoptosis. Nicotine prevented chemotherapy-induced reduction in mitochondrial membrane potential and caspase-9 activation. Antiapoptotic effects of nicotine were blocked by mitochondrial anion channel inhibitor, 4,4'diisothiocyanatostilbene-2,2'disulfonic acid. Chemotherapy enhanced translocation of proapoptotic Bax to the mitochondria, whereas nicotine blocked these effects. Nicotine up-regulated Akt-mediated antiapoptotic X-linked inhibitor of apoptosis protein and phosphorylated proapoptotic Bcl2-antagonist of cell death. The A549-rho0 cells, which lack mitochondrial DNA, demonstrated partial resistance to chemotherapy-induced apoptosis, but blocked the antiapoptotic effects of nicotine. Accordingly, we provide evidence that nicotine modulates mitochondrial signaling and inhibits chemotherapy-induced apoptosis in lung cancer. The mitochondrial regulation of nicotine imposes an important mechanism that can critically impair the treatment of lung cancer, because many cancer-therapeutic agents induce apoptosis via the mitochondrial death pathway. Strategies aimed at understanding nicotine-mediated signaling may facilitate the development of improved therapies in lung cancer.

  View details for DOI 10.1165/rcmb.2007-0277OC

  View details for Web of Science ID 000262872200002

  View details for PubMedID 18676776

  View details for PubMedCentralID PMC2633138

• Air pollution induces enhanced mitochondrial oxidative stress in cystic fibrosis airway epithelium FEBS LETTERS Kamdar, O., Le, W., Zhang, J., Ghio, A. J., Rosen, G. D., Upadhyay, D. 2008; 582 (25-26): 3601-3606

  Abstract

  We studied the effects of airborne particulate matters (PM) on cystic fibrosis (CF) epithelium. We noted that PM enhanced human CF bronchial epithelial apoptosis, activated caspase-9 and PARP-1; and reduced mitochondrial membrane potential. Mitochondrial inhibitors (4,4-diisothiocyanatostilbene-2,2'disulfonic acid, rotenone and thenoyltrifluoroacetone) blocked PM-induced generation of reactive oxygen species and apoptosis. PM upregulated pro-apoptotic Bad, Bax, p53 and p21; and enhanced mitochondrial localization of Bax. The anti-apoptotic Bcl-2, Bcl-xl, Mcl-1 and Xiap remained unchanged; however, overexpression of Bcl-xl blocked PM-induced apoptosis. Accordingly, we provide the evidence that PM enhances oxidative stress and mitochondrial signaling mediated apoptosis via the modulation of Bcl family proteins in CF.

  View details for DOI 10.1016/j.febslet.2008.09.030

  View details for Web of Science ID 000260807500006

  View details for PubMedID 18817777

• Ambient particulate matter induces alveolar epithelial cell cycle arrest: Role of G1 cyclins FEBS LETTERS Zhang, J., Ghio, A. J., Gao, M., Wei, K., Rosen, G. D., Upadhyay, D. 2007; 581 (27): 5315-5320

  Abstract

  We hypothesized that the ambient air pollution particles (particulate matter; PM) induce cell cycle arrest in alveolar epithelial cells (AEC). Exposure of PM (25microg/cm(2)) to AEC induced cells cycle arrest in G1 phase, inhibited DNA synthesis, blocked cell proliferation and caused decrease in cyclin E, A, D1 and Cyclin E- cyclin-dependent kinase (CDK)-2 kinase activity after 4h. PM induced upregulation of CDK inhibitor, p21 protein and p21 activity in AEC. SiRNAp21 blocked PM-induced downregulation of cyclins and AEC G1 arrest. Accordingly, we provide the evidence that PM induces AEC G1 arrest by altered regulation of G1 cyclins and CDKs.

  View details for DOI 10.1016/j.febslet.2007.10.020

  View details for Web of Science ID 000253487700023

  View details for PubMedID 17977533

  View details for PubMedCentralID PMC2140149

• Bim mediates mitochondria-regulated particulate matter-induced apoptosis in alveolar epithelial cells FEBS LETTERS Zhang, J., Ghio, A. J., Chang, W., Kamdar, O., Rosen, G. D., Upadhyay, D. 2007; 581 (22): 4148-4152

  Abstract

  We studied the role of Bim, a pro-apoptotic BCL-2 family member in Airborne particulate matter (PM 2.5 microm)-induced apoptosis in alveolar epithelial cells (AEC). PM induced AEC apoptosis by causing significant reduction of mitochondrial membrane potential and increase in caspase-9, caspase-3 and PARP-1 activation. PM upregulated pro-apoptotic protein Bim and enhanced translocation of Bim to the mitochondria. ShRNABim blocked PM-induced apoptosis by preventing activation of the mitochondrial death pathway suggesting a role of Bim in the regulation of mitochondrial pathway in AEC. Accordingly, we provide the evidence that Bim mediates PM-induced apoptosis via mitochondrial pathway.

  View details for DOI 10.1016/j.febslet.2007.07.080

  View details for Web of Science ID 000249476900004

  View details for PubMedID 17716672

• Fibroblast growth factor-10 prevents H2O2-induced cell cycle arrest by regulation of GI cyclins and cyclin dependent kinases FEBS LETTERS Upadhyay, D., Chang, W., Wei, K., Gao, M., Rosen, G. D. 2007; 581 (2): 248-252

  Abstract

  We studied the effects of fibroblast growth factor (FGF-10) on H2O2-induced alveolar epithelial cell (AEC) G1 arrest and the role of G1 cyclins. FGF-10 prevented H2O2-induced AEC G1 arrest. FGF-10 induced 2-4-fold increase in cyclin E, cyclin A and CDKs (2,4) alone and in AEC treated with H2O2. H2O2 downregulated cyclin D1; FGF-10 blocked these effects. FGF-10 prevented H2O2-induced upregulation of CDK inhibitor, p21. SiRNAp21 blocked H2O2-induced downregulation of cyclins, CDKs and AEC G1 arrest. Accordingly, we provide first evidence that FGF-10 regulates G1 cyclins and CDKs, and prevents H2O2-induced AEC G1 arrest.

  View details for DOI 10.1016/j.febslet.2006.12.020

  View details for Web of Science ID 000244188700012

  View details for PubMedID 17188682

• Recurrence of pulmonary intravascular bronchoalveolar tumor with mediastinal metastasis 20 years later RESPIRATORY MEDICINE Chen, T. M., Donington, J., Mak, G., Berry, G. J., Ruoss, S. J., Rosen, G. D., Upadhyay, D. 2006; 100 (2): 367-370

  Abstract

  Pulmonary intravascular bronchoalveolar tumor (IVBAT) also recognized as pulmonary epithelioid hemangioendothelioma, is a rare malignant vascular tumor of unknown etiology. IVBAT is a tumor of multicentric origin and the lungs are rarely involved, with only about 60 cases of pulmonary IVBAT described in the literature. The prognosis is unpredictable, with life expectancy ranging from 1 to 15 years. We report an unusual case of pulmonary IVBAT that recurred in the lung with metastasis to the mediastinum.

  View details for DOI 10.1016/j.rmed.2005.05.010

  View details for Web of Science ID 000235243000024

  View details for PubMedID 15990286

• A 35-year-old man, with fever, dyspnea, and diffuse reticular opacities CHEST Katikireddy, C. K., Krishna, G., Keifer, T., Kuschner, W., Rosen, G. 2006; 129 (2): 482-487

  View details for Web of Science ID 000235646100041

  View details for PubMedID 16478870

• A placebo-controlled trial of interferon gamma-1b in patients with idiopathic pulmonary fibrosis NEW ENGLAND JOURNAL OF MEDICINE Raghu, G., Brown, K. K., Bradford, W. Z., Starko, K., Noble, P. W., Schwartz, D. A., King, T. E., Adlakha, A., Tarczynski, S., Ainslie, G., Kalam, R., Bai, T., Truchan, H., BAUGHMAN, R., Wingst, D., Bhorade, S., Norwick, L., Brown, K. K., Kervitsky, D., Calhoun, W., DiNella, L., Chan, C., Jamieson, L., Chan, K., Turpen, T., Chapman, J., Slattery, S., Chen, L., Turner, J., Clark, M., Sanders, R., Crain, M., Pate, D., Davis, G., Lynn, M., Dhar, A., Hrytsytk, M., Drent, M., Horr, E. T., du Bois, R., Goh, N., EGAN, J., Anthony, N., Enelow, R., Haram, T., Ettinger, N., MERLI, S., Frost, A., Holy, R., Glassberg, M., Brown, A., Golden, J., DesMarais, M., Haim, Y. D., Rzeszutko, B., Hassoun, P., Finlay, G., Lawler, L., Hollingsworth, H., Goncalves, P., Jackson, R., Meadows, T., Kallay, M., Celebie, A., King-Biggs, M., Beyle, A., Lancaster, L., Sanderson, R., Lasky, J., Ditta, S., Lorch, D., Schoh, T., Lynch, J., Alousha, A., Mageto, Y., Meachom, M., Martinez, F., Dahlgren, D., McKee, C., Hamilton, T., Meyer, K., Wilson, S., Millar, A., Hann, A., Mohsenifar, Z., Balfe, D., Morell, F., Reyes, L., Nathan, S., Theodoslod, S., Noble, P. W., Shrauger, K., Noth, I., Strek, M., Au, J., Olivier, K., Wells, R., Padilla, M., Behnegar, A., Polito, A., Bloom, C., Raghu, G., Snydsman, A., Rosen, G., Jacobs, S., Ryu, J. H., Carlson, K., Sahn, S. A., Oser, R., Schaumberg, T., Chesnutt, M., Leonard, S., Schluger, N., Jellen, P., Steele, M., Willis, C., Strieter, R., Christedoulou, V., Valentine, V., Lanning, T., Wencel, M., Sturgeon, D., Xaubert, A., Zisman, D., Douglas, L., Lynch, D. A., Godwin, J. D., Webb, W. R., Fleming, T., Reis, A., Riley, D. 2004; 350 (2): 125-133

  Abstract

  Idiopathic pulmonary fibrosis is a progressive, fatal disease with no known efficacious therapy.In a double-blind, multinational trial, we randomly assigned 330 patients with idiopathic pulmonary fibrosis that was unresponsive to corticosteroid therapy to receive subcutaneous interferon gamma-1b or placebo.Over a median of 58 weeks, interferon gamma-1b therapy did not significantly affect the primary end point of progression-free survival, defined as the time to disease progression or death, and no significant treatment effect was observed on measures of lung function, gas exchange, or the quality of life. Ten percent of patients in the interferon gamma-1b group died, as compared with 17 percent of patients in the placebo group (P=0.08). Treatment with interferon gamma-1b was associated with more frequent constitutional symptoms. However, the rates of treatment adherence and premature discontinuation of treatment were similar in the two groups. More pneumonias were reported among patients in the interferon gamma-1b group, but the incidence of severe or life-threatening respiratory tract infections was similar in the two groups.In a well-defined population of patients with idiopathic pulmonary fibrosis, interferon gamma-1b did not affect progression-free survival, pulmonary function, or the quality of life. Owing to the size and duration of the trial, a clinically significant survival benefit could not be ruled out.

  View details for Web of Science ID 000187858500006

  View details for PubMedID 14711911

• PG490-88, a derivative of triptolide, causes tumor regression and sensitizes tumors to chemotherapy MOLECULAR CANCER THERAPEUTICS Fidler, J. M., Li, K., Chung, C., Wei, K., Ross, J. A., Gao, M. X., Rosen, G. D. 2003; 2 (9): 855-862

  Abstract

  Treatment of solid tumors with combinations of chemotherapeutic agents has not led to significant increases in long-term survival. Recent studies support a role for inhibitors of checkpoint arrest as a means to enhance the cytotoxicity of chemotherapy. We have shown previously that triptolide (PG490), an oxygenated diterpene derived from a Chinese medicinal plant, induces apoptosis in cultured tumor cells and sensitizes tumor cells to topoisomerase inhibitors by blocking p53-mediated induction of p21. Here we extend our studies to a tumor xenograft model and evaluate the efficacy and safety of PG490-88 (14-succinyl triptolide sodium salt), a water-soluble prodrug of PG490. We also look at the combination of PG490 or PG490-88 with CPT-11, a topoisomerase I inhibitor, in cultured cells and in the tumor xenograft model. We show that PG490-88 is a safe and potent antitumor agent when used alone, causing tumor regression of lung and colon tumor xenografts. We also show that PG490-88 acts in synergy with CPT-11 to cause tumor regression. A phase I trial of PG490-88 for solid tumors began recently and safety and optimal dosing data should accrue within the next 12 months. Our findings that PG490-88 causes tumor regression and that it acts in synergy with DNA-damaging chemotherapeutic agents suggest a role as an antineoplastic agent and chemosensitizer for the treatment of patients with solid tumors.

  View details for Web of Science ID 000185440400004

  View details for PubMedID 14555704

• Triptolide sensitizes lung cancer cells to TNF-related apoptosis-inducing ligand (TRAIL)-induced apoptosis by inhibition of NF-kappa B activation EXPERIMENTAL AND MOLECULAR MEDICINE Lee, K. Y., Park, J. S., Jee, Y. K., Rosen, G. D. 2002; 34 (6): 462-468

  Abstract

  TNF-related apoptosis-inducing ligand (TRAIL/Apo- 2L), a newly identified member of the TNF family promotes apoptosis by binding to the transmembrane receptors (TRAIL-R1/DR4 and TRAIL-R2/DR5). TRAIL known to activate NF-kappaB in number of tumor cells including A549 (wt p53) and NCI-H1299 (null p53) lung cancer cells exerts relatively selective cytotoxic affects to the human tumor cell lines without much effect on the normal cells. We set out to identify an agent that would sensitize lung cancer cells to TRAIL-induced apoptosis through inhibition of NF-kappaB activation. We found that triptolide, an oxygenated diterpene extracted and purified from the Chinese herb Tripterygium wilfordii sensitized A549 and NCI-H1299 cells to TRAIL-induced apoptosis through inhibition of NF-kappaB activation. Pretreatment with MG132 which is a well-known NF-kappaB inhibitor by blocking degradation of IkappaBalpha also greatly sensitized lung cancer cells to TRAIL-induced apoptosis. Triptolide did not block DNA binding of NF-kappaB activated by TRAIL as in the case of TNF-alpha. It has been already proven that triptolide blocks transactivation of p65 which plays a key role in NF-kappaB activation. These observations suggest that triptolide may be a potentially useful drug to enhance TRAIL-induced tumor killing in lung cancer.

  View details for Web of Science ID 000180504600009

  View details for PubMedID 12526088

• PG490-88, a derivative of triptolide, attenuates obliterative airway disease in a mouse heterotopic tracheal allograft model JOURNAL OF HEART AND LUNG TRANSPLANTATION Leonard, C. T., Soccal, P. M., Berry, G. J., Doyle, R. L., Theodore, J., Duncan, S. R., Rosen, G. D. 2002; 21 (12): 1314-1318

  Abstract

  The current treatment of obliterative bronchiolitis in lung transplant recipients is sub-optimal. Triptolide is a novel immunosuppressant that has a mechanism of action distinct from currently available immunosuppressants, including induction of T-cell apoptosis, blockade of fibroblast proliferation/maturation and inhibition of transforming growth factor-beta (TGF-beta) mRNA production. We hypothesized that triptolide may be helpful in blocking obliterative airway disease in lung transplant recipients. We investigated the effect of PG490-88, a water-soluble derivative of triptolide, in a mouse heterotopic tracheal allograft model of obliterative airway disease. We show that PG490-88 attenuates airway obliteration in this model and inhibits accumulation of inflammatory cells, and therefore may have preventive or therapeutic benefits for patients with obliterative airway disease (OAD) following lung transplantation.

  View details for Web of Science ID 000179959800009

  View details for PubMedID 12490278

• The molecular biology of lung cancer CURRENT OPINION IN PULMONARY MEDICINE Ross, J. A., Rosen, G. D. 2002; 8 (4): 265-269

  Abstract

  Lung cancer is the result of molecular changes that occur in the cell, resulting in the deregulation of pathways which control normal cellular growth, differentiation, and apoptosis. Several of these pathways contain well-characterized proto-oncogenes and tumor suppressor genes which are found to be mutated or have abnormal expression patterns in lung cancer. The molecular changes that characterize lung cancer are complex, but it is known that cigarette smoking causes most squamous cell and small-cell carcinomas. However, the association between cigarette smoke and adenocarcinoma is less clear. Environmental factors, such as air pollutants, radon, and asbestos, likely contribute to the development of lung cancer. In this review, we discuss the major molecular abnormalities in lung cancer with a review of recent studies that begin to decipher the role that different tumor suppressor genes and oncogenes play in the pathogenesis of lung cancer. Also, we highlight the research that has identified new genes which may play a role in lung cancer pathogenesis or progression.

  View details for PubMedID 12055387

• Oligoclonal CD4(+) T cell expansions in lung transplant recipients with obliterative bronchiolitis AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE Duncan, S. R., Leonard, C., Theodore, J., Lega, M., Girgis, R. E., Rosen, G. D., Theofilopoulos, A. N. 2002; 165 (10): 1439-1444

  Abstract

  Obliterative bronchiolitis (OB) is a dreaded and frequent complication of lung transplantation with a poorly understood immunopathogenesis. To further evaluate disease mechanisms, we used T cell antigen receptor (TCR) beta-chain variable region RNase protection assays, after polymerase chain reaction amplification of TCR cDNA, to quantitate circulating CD4(+) and CD8(+) repertoires of transplant recipients with OB or no evidence of rejection (NER). All six recipients with OB had markedly abnormal CD4 expansions (2.5 +/- 0.5 expansions/recipient) attributable to oligoclonal proliferations. Only two of six recipients with NER had a single, much lesser, CD4(+) abnormality each (p < 0.01). Moreover, one of these patients developed OB shortly thereafter, and the other NER abnormality may have predated transplantation. In contrast, CD8(+) expansions were common in both recipient populations. Findings of CD4(+) expansions had 100% sensitivity and 80% specificity for the presence or imminent development of OB. These data suggest proliferations of CD4(+) T cells are important in OB pathogenesis, and these are most likely part of a major histocompatibility complex Class II-dependent process of indirect alloantigen presentation. These CD4(+) clones are likely to have facultative helper functions for the multiple and diverse immune processes that have been implicated in OB. Furthermore, the close association of CD4(+) expansions with OB raises possibilities of development of novel diagnostic and therapeutic approaches.

  View details for DOI 10.1164/rccm.2107009

  View details for Web of Science ID 000175661600018

  View details for PubMedID 12016109

• Pharmacogenomics and the challenge to privacy. pharmacogenomics journal Vaszar, L. T., Rosen, G. D., Raffin, T. A. 2002; 2 (3): 144-147

  View details for PubMedID 12082584

• Diversity of gene expression in adenocarcinoma of the lung PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Garber, M. E., Troyanskaya, O. G., Schluens, K., Petersen, S., Thaesler, Z., Pacyna-Gengelbach, M., van de Rijn, M., Rosen, G. D., Perou, C. M., Whyte, R. I., Altman, R. B., Brown, P. O., Botstein, D., Petersen, I. 2001; 98 (24): 13784-13789

  Abstract

  The global gene expression profiles for 67 human lung tumors representing 56 patients were examined by using 24,000-element cDNA microarrays. Subdivision of the tumors based on gene expression patterns faithfully recapitulated morphological classification of the tumors into squamous, large cell, small cell, and adenocarcinoma. The gene expression patterns made possible the subclassification of adenocarcinoma into subgroups that correlated with the degree of tumor differentiation as well as patient survival. Gene expression analysis thus promises to extend and refine standard pathologic analysis.

  View details for PubMedID 11707590

• Tumor necrosis factor-related apoptosis-inducing ligand and chemotherapy cooperate to induce apoptosis in mesothelioma cell lines AMERICAN JOURNAL OF RESPIRATORY CELL AND MOLECULAR BIOLOGY Liu, W. H., Bodle, E., Chen, J. Y., Gao, M. X., Rosen, G. D., Broaddus, V. C. 2001; 25 (1): 111-118

  Abstract

  Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) can induce apoptosis in certain tumor cells. In addition, TRAIL and chemotherapy can act cooperatively, possibly as a result of chemotherapy-induced increases in expression of a TRAIL receptor, DR5. We used cell lines derived from a highly chemoresistant tumor, malignant mesothelioma, to learn whether TRAIL was effective alone or together with chemotherapy and whether cooperativity depended on increases in DR5 expression. TRAIL (codons 95-285) was expressed in a bacterial expression vector and purified by nickel affinity chromatography. TRAIL alone (25 to 500 ng/ml) had little effect on mesothelioma cells. TRAIL plus chemotherapy (doxorubicin, cis-platinum, etoposide, or gemcitabine) acted cooperatively to induce apoptosis in mesothelioma cells (M28, REN, VAMT, and MS-1). For example, in M28 cells treated for 18 h, apoptosis from TRAIL (100 ng/ml) plus doxorubicin (0.6 microg/ml; 71 +/- 11%) greatly exceeded that from TRAIL alone (21 +/- 8%) or from doxorubicin alone (6 +/- 2%) (means +/- standard deviation; P < 0.03). Mesothelioma cells treated with chemotherapy showed no change in DR5 protein by Western analysis or by immunocytochemistry. TRAIL plus chemotherapy was associated with an increase in mitochondrial cytochrome c release and mitochondrial depolarization. We conclude that TRAIL and chemotherapy act cooperatively to kill mesothelioma cell lines, not by increases in DR5 receptor but in association with mitochondrial amplification of apoptotic signals.

  View details for Web of Science ID 000170221800018

  View details for PubMedID 11472983

• PG490-88, a derivative of triptolide, blocks bleomycin-induced lung fibrosis AMERICAN JOURNAL OF PATHOLOGY Krishna, G., Liu, K. L., Shigemitsu, H., Gao, M. X., Raffin, T. A., Rosen, G. D. 2001; 158 (3): 997-1004

  Abstract

  In this study we evaluate the antifibrotic properties of PG-490-88, a water-soluble derivative of triptolide. Triptolide is an oxygenated diterpene that is derived from a traditional Chinese herb that has potent immunosuppressive and antitumor activity. We used the intratracheal bleomycin mouse model and found that PG490-88 inhibits fibrosis in the bleomycin group when given the same day or 5 days after bleomycin. PG490-88 also markedly reduced the number of myofibroblasts in the bleomycin treatment group. An enzyme-linked immunosorbent assay of transforming growth factor (TGF)-beta in the bronchoalveolar lavage fluid showed a significant decrease in TGF-beta in the PG490-88-treated groups compared to the bleomycin-treated group. Additionally, triptolide blocked bleomycin-induced increase in TGF-beta mRNA in cultured normal human lung fibroblasts. The efficacy of PG490-88 when administered late after bleomycin installation suggests a potential role in the treatment of idiopathic pulmonary fibrosis.

  View details for Web of Science ID 000167411100023

  View details for PubMedID 11238047

  View details for PubMedCentralID PMC1850337

• Triptolide and chemotherapy cooperate in tumor cell apoptosis - A role for the p53 pathway JOURNAL OF BIOLOGICAL CHEMISTRY Chang, W. T., Kang, J. J., Lee, K. Y., Wei, K., Anderson, E., Gotmare, S., Ross, J. A., Rosen, G. D. 2001; 276 (3): 2221-2227

  Abstract

  Triptolide (PG490), a diterpene triepoxide, is a potent immunosuppressive agent extracted from the Chinese herb Tripterygium wilfordii. We have previously shown that triptolide blocks NF-kappaB activation and sensitizes tumor necrosis factor (TNF-alpha)-resistant tumor cell lines to TNF-alpha-induced apoptosis. We show here that triptolide enhances chemotherapy-induced apoptosis. In triptolide-treated cells, the expression of p53 increased but the transcriptional function of p53 was inhibited, and we observed a down-regulation of p21(waf1/cip1), a p53-responsive gene. The increase in levels of the p53 protein was mediated by enhanced translation of the p53 protein. Additionally, triptolide induced accumulation of cells in S phase and blocked doxorubicin-mediated accumulation of cells in G(2)/M and doxorubicin-mediated induction of p21. Our data suggest that triptolide, by blocking p21-mediated growth arrest, enhances apoptosis in tumor cells.

  View details for PubMedID 11053449

• Dendritic cells and macrophages in lung allografts a role in chronic rejection? AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE Leonard, C. T., Soccal, P. M., Singer, L., Berry, G. J., Theodore, J., Holt, P. G., Doyle, R. L., Rosen, G. D. 2000; 161 (4): 1349-1354

  Abstract

  Antigen presentation by lung macrophages/dendritic cells (DC) is thought to be important in obliterative bronchiolitis/bronchiolitis obliterans syndrome (OB/BOS), which severely limits survival post-lung transplantation. However, a recent study found minimal numbers of DC in lung allografts. We looked at numbers and phenotype of macrophages/DC in lung allografts using endobronchial biopsy (EBB) and transbronchial biopsy (TBB) from 22 lung transplant patients. Biopsies were stained with monoclonal markers of DC (CD1a, RFD1, and major histocompatibility complex [MHC] Class II), and "suppressor macrophages" (RFD1 and RFD7). Dendritic cells were also stained for the costimulatory molecules CD80 and CD86. Significantly greater numbers of DC/high-power field (HPF) were seen in biopsies when we defined DC using dendritic morphology and Class II MHC expression instead of CD1a expression. Dendritic cell numbers were significantly higher in eight patients with OB/BOS compared with 14 stable patients. Fifty percent of DC expressed CD86 and 20% expressed CD80. There was no difference in CD80 or CD86 expression between OB/BOS patients and stable patients. There was no correlation between DC numbers and presence or absence of acute rejection (AR), and/or cytomegalovirus (CMV) pneumonitis on current or prior biopsies. There were significantly more MHC Class II DC in EBB compared with TBB. We found minimal staining for lung macrophages capable of suppressing T-cell inflammation. We conclude that studies of lung allografts may underestimate DC numbers if relying on CD1a as the sole marker of DC. DC are increased in patients with OB/BOS compared with stable patients. EBB may be more important than TBB in looking for inflammatory changes of OB. DC expressing costimulatory molecules are present in lung allografts, and costimulatory pathway blockade may be useful in human lung allografts. Also, the absence of "suppressor" macrophages may increase susceptibility of human lung allografts to the rejection process.

  View details for Web of Science ID 000086573400044

  View details for PubMedID 10764333

• Thoracic lymphangiomas, lymphangiectasis, lymphangiomatosis, and lymphatic dysplasia syndrome AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE Faul, J. L., Berry, G. J., Colby, T. V., Ruoss, S. J., Walter, M. B., Rosen, G. D., Raffin, T. A. 2000; 161 (3): 1037-1046

  View details for Web of Science ID 000085996400058

  View details for PubMedID 10712360

• Loss of STAT1 expression confers resistance to IFN-gamma-induced apoptosis in ME180 cells FEBS LETTERS Lee, K. Y., Anderson, E., Madani, K., Rosen, G. D. 1999; 459 (3): 323-326

  Abstract

  Interferon gamma (IFN-gamma) induces apoptosis in many tumor cell lines and sensitizes tumor cells to apoptosis by tumor necrosis factor family members. IFN-gamma induces the expression of many early response genes such as interferon regulatory factor-1 (IRF-1) by activation of signal transducer and activator of transcription (STAT) factor proteins. We found that ME180 cells became resistant to IFN-gamma-induced cell death after 4-5 passages in culture. These resistant cells were characterized by a loss of STAT1 expression and a loss of inducible IRF-1 expression. We describe for the first time the emergence of a STAT1-deficient ME180 cell line.

  View details for Web of Science ID 000083204500008

  View details for PubMedID 10526158

• PG490 (triptolide) cooperates with tumor necrosis factor-alpha to induce apoptosis in tumor cells JOURNAL OF BIOLOGICAL CHEMISTRY Lee, K. Y., Chang, W. T., Qiu, D. M., Kao, P. N., Rosen, G. D. 1999; 274 (19): 13451-13455

  Abstract

  Progress in the treatment of solid tumors has been slow and sporadic. The efficacy of conventional chemotherapy in solid tumors is limited because tumors frequently have mutations in the p53 gene. Also, chemotherapy only kills rapidly dividing cells. Members of the tumor necrosis factor (TNF) family, however, induce apoptosis regardless of the p53 phenotype. Unfortunately, the cytotoxicity of TNF-alpha is limited by its activation of NF-kappaB and activation of NF-kappaB is proinflammatory. We have identified a compound called PG490, that is composed of purified triptolide, which induces apoptosis in tumor cells and sensitizes tumor cells to TNF-alpha-induced apoptosis. PG490 potently inhibited TNF-alpha-induced activation of NF-kappaB. PG490 also blocked TNF-alpha-mediated induction of c-IAP2 (hiap-1) and c-IAP1 (hiap-2), members of the inhibitor of apoptosis (IAP) family. Interestingly, PG490 did not block DNA binding of NF-kappaB, but it blocked transactivation of NF-kappaB. Our identification of a compound that blocks TNF-alpha-induced activation of NF-kappaB may enhance the cytotoxicity of TNF-alpha on tumors in vivo and limit its proinflammatory effects.

  View details for PubMedID 10224110

• Proteolytic cleavage of Ras GTPase-activating protein during apoptosis CELL DEATH AND DIFFERENTIATION Wen, L. P., Madani, K., Martin, G. A., Rosen, G. D. 1998; 5 (9): 729-734

  Abstract

  p120-ras GTPase-activating protein (rasGAP) associates with Ras and negatively regulates Ras signaling by stimulating the intrinsic rate of Ras GTPase activity. rasGAP also associates with other cellular signaling proteins which suggest that rasGAP may play a role in coordinating other signal transduction pathways. Disruption of rasGAP in vivo results in extensive apoptosis. Fas-mediated apoptosis results in the activation of caspases that cleave cellular substrates which are important for maintaining cytoplasmic and nuclear integrity. We show here that rasGAP is proteolytically cleaved by caspases early in Fas-induced apoptosis of Jurkat cells. rasGAP was also cleaved by DNA-damaging chemotherapeutic agents and TNF-related apoptosis inducing ligand (TRAIL), also known as Apo2L. Based on the size of the products generated by cleavage of deletion mutants of rasGAP we predict that cleavage of rasGAP occurs in the hydrophobic region and between the SH2(2) and ras-p21 interacting domain which would leave an intact ras-p21 interacting domain. Interestingly, cleavage of rasGAP in vitro enhanced rasGAP hydrolysis activity. Our results demonstrate that diverse apoptotic stimuli cause caspase-mediated cleavage of rasGAP early in apoptosis.

  View details for Web of Science ID 000075787200003

  View details for PubMedID 10200531

• Dexamethasone inhibits lung epithelial cell apoptosis induced by IFN-gamma and Fas. American journal of physiology. Lung cellular and molecular physiology Wen, L., Madani, K., Fahrni, J. A., Duncan, S. R., Rosen, G. D. 1997; 273 (5): L921–L929

  Abstract

  Lung epithelium plays a central role in modulation of the inflammatory response and in lung repair. Airway epithelial cells are targets in asthma, viral infection, acute lung injury, and fibrotic lung disease. Activated T lymphocytes release cytokines such as interferon-gamma (IFN-gamma) that can cooperate with apoptotic signaling pathways such as the Fas-APO-1 pathway to induce apoptosis of damaged epithelial cells. We report that IFN-gamma alone and in combination with activation of the Fas pathway induced apoptosis in A549 lung epithelial cells. Interestingly, the corticosteroid dexamethasone was the most potent inhibitor of IFN-gamma- and IFN-gamma plus anti-Fas-induced apoptosis. IFN-gamma induced expression of an effector of apoptosis, the cysteine protease interleukin-1beta-converting enzyme, in A549 cells. Dexamethasone, in contrast, induced expression of an inhibitor of apoptosis, human inhibitor of apoptosis (hIAP-1), also known as cIAP2. We suggest that the inhibition of epithelial cell apoptosis by corticosteroids may be one mechanism by which they suppress the inflammatory response.

  View details for DOI 10.1152/ajplung.1997.273.5.L921

  View details for PubMedID 29586270

• Dexamethasone inhibits lung epithelial cell apoptosis induced by IFN-gamma and Fas AMERICAN JOURNAL OF PHYSIOLOGY-LUNG CELLULAR AND MOLECULAR PHYSIOLOGY Wen, L. P., Madani, K., Fahrni, J. A., Duncan, S. R., Rosen, G. D. 1997; 273 (5): L921-L929

  Abstract

  Lung epithelium plays a central role in modulation of the inflammatory response and in lung repair. Airway epithelial cells are targets in asthma, viral infection, acute lung injury, and fibrotic lung disease. Activated T lymphocytes release cytokines such as interferon-gamma (IFN-gamma) that can cooperate with apoptotic signaling pathways such as the Fas-APO-1 pathway to induce apoptosis of damaged epithelial cells. We report that IFN-gamma alone and in combination with activation of the Fas pathway induced apoptosis in A549 lung epithelial cells. Interestingly, the corticosteroid dexamethasone was the most potent inhibitor of IFN-gamma- and IFN-gamma plus anti-Fas-induced apoptosis. IFN-gamma induced expression of an effector of apoptosis, the cysteine protease interleukin-1 beta-converting enzyme, in A549 cells. Dexamethasone, in contrast, induced expression of an inhibitor of apoptosis, human inhibitor of apoptosis (hIAP-1), also known as cIAP2. We suggest that the inhibition of epithelial cell apoptosis by corticosteroids may be one mechanism by which they suppress the inflammatory response.

  View details for Web of Science ID A1997YF56800003

  View details for PubMedID 9374718

• Cleavage of focal adhesion kinase by caspases during apoptosis JOURNAL OF BIOLOGICAL CHEMISTRY Wen, L. P., Fahrni, J. A., Troie, S., Guan, J. L., Orth, K., Rosen, G. D. 1997; 272 (41): 26056-26061

  Abstract

  Apoptotic cells undergo characteristic morphological changes that include detachment of cell attachment from the substratum and loss of cell-cell interactions. Attachment of cells to the extracellular matrix and to other cells is mediated by integrins. The interactions of integrins with the extracellular matrix activates focal adhesion kinase (FAK) and suppresses apoptosis in diverse cell types. Members of the tumor necrosis family such as Fas and Apo-2L, also known as tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), induce apoptosis in both suspension and adherent cells through the activation of caspases. These caspases, when activated, cleave substrates that are important for the maintenance of nuclear and membrane integrity. In this study, we show that FAK is sequentially cleaved into two different fragments early in Apo-2L-induced apoptosis. We also demonstrate that FAK cleavage is mediated by caspases and that FAK shows unique sensitivity to different caspases. Our results suggest that disruption of FAK may contribute to the morphological changes observed in apoptotic suspension and adherent cells.

  View details for Web of Science ID A1997YA35800098

  View details for PubMedID 9325343

• Rapamycin inhibits development of obliterative airway disease in a murine heterotopic airway transplant model TRANSPLANTATION Fahrni, J. A., Berry, G. J., Morris, R. E., Rosen, G. D. 1997; 63 (4): 533-537

  Abstract

  Obliterative bronchiolitis is the major cause of long-term morbidity and mortality in heart-lung and lung transplant recipients. There is presently no completely effective therapy for the treatment of obliterative bronchiolitis. We have examined the effects of rapamycin (RPM) on the development of obliterative airway disease in murine recipients of heterotopically transplanted allograft tracheas. In this model, an untreated allograft develops almost complete occlusion of the airway lumen with fibroblastic tissue and collagen scar by day 28 after transplantation. RPM administered intraperitoneally at the time of transplantation or even as late as day 14 after transplantation markedly inhibited obliteration of the airway lumen by fibroblastic tissue. Also, RPM significantly inhibited infiltration of the graft by macrophages. In the RPM-treated animals, the airway was reconstituted with an attenuated squamous epithelium rather than a normal pseudostratified epithelium. No adverse side effects were observed with RPM doses up to 12 mg/kg/ day. These findings suggest a potential role for RPM, perhaps in combination with cyclosporine, in preventing and treating obliterative bronchiolitis in heart-lung and lung allograft recipients.

  View details for Web of Science ID A1997WL85500008

  View details for PubMedID 9047146

• Airway epithelial cells produce stem cell factor BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH Wen, L. P., Fahrni, J. A., Matsui, S., Rosen, G. D. 1996; 1314 (3): 183-186

  Abstract

  Airway epithelial cells modulate the inflammatory response in asthmatic, allergic and fibrotic lung diseases through the secretion of cytokines that regulate the movement and activation of inflammatory cells. Mast cells play an important role in the pathogenesis of these lung diseases. In this study we report that normal airway epithelial cells express stem cell factor which is a critical mediator of mast cell growth and differentiation and that transforming growth factor-beta inhibits secretion of stem cell factor by airway epithelial cells.

  View details for Web of Science ID A1996VX94800001

  View details for PubMedID 8982273

• Acid aspiration induced lung injury - New insights and therapeutic options AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE Matthay, M. A., Rosen, G. D. 1996; 154 (2): 277-278

  View details for Web of Science ID A1996VB85800001

  View details for PubMedID 8756794

• MULTIPLE REGULATORY ELEMENTS CONTROL THE BASAL PROMOTER ACTIVITY OF THE HUMAN ALPHA-4 INTEGRIN GENE DNA AND CELL BIOLOGY Audet, J. F., Masson, J. Y., Rosen, G. D., Salesse, C., Guerin, S. L. 1994; 13 (11): 1071-1085

  Abstract

  It has been suggested that expression of the genes encoding the alpha 4/beta 1 integrin increases during wound healing of the cornea. As a first step in understanding the mechanisms required to stimulate alpha 4 gene expression during this process, we defined the minimal upstream sequence required to direct basal promoter activity for this gene. Using deletion analyses of the alpha 4 gene upstream sequence, we identified two functionally important negative regulatory elements. Dimethylsulfate (DMS) methylation interference assays provided evidence for the binding of a single nuclear protein to tandemly repeated homologous cis-acting elements (designated alpha 4.1 and alpha 4.2) from the alpha 4 basal promoter that share the core sequence 5'-GTGGGT-3'. The formation of a protection only at alpha 4.1 in DNase I footprinting suggested that it is the primary target element for the binding of nuclear proteins. Three distinct nuclear proteins bound a double-stranded oligonucleotide bearing the DNA sequence of alpha 4.1 to produce specific DNA-protein complexes (R1 to R3) in gel-shift assays, from which that producing R3 was identified as the protein yielding DNase I protection at alpha 4.1. Detailed mutational analysis of alpha 4.1 and alpha 4.2 indicated that both elements positively regulate gene expression in primary cultures of corneal epithelial cells and Jurkat tissue culture cells, which is consistent with the deletion analysis. However, when transiently transfected into pituitary GH4C1, the alpha 4.2 mutants yielded increased chloramphenicol acetyl transferase activity therefore demonstrating that these elements have the ability to function either as positive or negative regulators of gene transcription in a manner that is dependent on the type of cell transfected.

  View details for Web of Science ID A1994PW25100002

  View details for PubMedID 7702751

• AN INTRICATE ARRANGEMENT OF BINDING-SITES FOR THE ETS FAMILY OF TRANSCRIPTION FACTORS REGULATES ACTIVITY OF THE ALPHA-4 INTEGRIN GENE PROMOTER JOURNAL OF BIOLOGICAL CHEMISTRY Rosen, G. D., Barks, J. L., Iademarco, M. F., Fisher, R. J., Dean, D. C. 1994; 269 (22): 15652-15660

  Abstract

  alpha 4 integrins mediate cell-cell and cell-extracellular matrix interactions that are critical for maturation and function of the immune system as well as differentiation of skeletal muscle. Here we examine molecular mechanisms controlling the pattern of alpha 4 expression. The activity of constructs containing 5' deletion mutants of the alpha 4 gene promoter was compared in transfection assays into cell lines that express alpha 4 and cell lines that do not. The sequence between position -42 and -76 base pairs (bp) was required for efficient transcription in cells that express alpha 4, but it showed no activity in HeLa cells, which do not express alpha 4. Three binding sites for the Ets family of transcription factors are found in this region: two adjacent sites at positions -50 and -54 bp and a more 5' site at position -67 bp. Using a series of constructs containing deletions and mutations in this region, we found that the 3'-most site alone was sufficient for binding GA-binding protein alpha (GABP alpha)/GABP beta and for a low level of transcriptional activation. When all three sites were present, a second complex "a" was detected, which contains an unknown member of the Ets family. Formation of complex a was cell-type specific and correlated with a high level of transcription. Deletion of the 5'-most Ets site had no effect on binding to GABP alpha/GABP beta, but it eliminated a. Concomitant with this loss of a, a new Ets-1-containing complex "c" appeared. Complex c substituted efficiently for complex a in transcriptional activation. We conclude that although neither of the two 5'-most Ets sites alone binds nuclear protein, they appear to act as modulators which control the pattern of Ets proteins that bind the alpha 4 gene promoter. This arrangement of Ets sites, coupled with the tissue- and developmental-specific expression of Ets members, likely play a key role in defining the pattern of alpha 4 integrin.

  View details for Web of Science ID A1994NP51300043

  View details for PubMedID 8195215

• EXPANDING ROLES FOR ALPHA-4 INTEGRIN AND ITS LIGANDS IN DEVELOPMENT CELL ADHESION AND COMMUNICATION Sheppard, A. M., Onken, M. D., Rosen, G. D., Noakes, P. G., Dean, D. C. 1994; 2 (1): 27-43

  Abstract

  Interaction of alpha 4 integrins with vascular cell adhesion molecule-1 (VCAM-1) is classically important for immune function. However, we found recently that these receptors have a second role, in embryogenesis, where they mediate cell-cell interactions that are important for skeletal muscle differentiation. Here, we present evidence of an expanding role for these receptors in murine development. alpha 4 and VCAM-1 were found at embryonic sites of hematopoiesis, suggesting a role for these receptors during embryogenesis that parallels their hematopoietic function in adult bone marrow. During angiogenesis in the lung, alpha 4 and VCAM-1 were found on mesenchyme that gives rise to vascular endothelium and smooth muscle. alpha 4 persisted on the smooth muscle and the endothelium of newly forming vessels where it colocalized with its extracellular matrix ligand, fibronectin (FN). These patterns suggest several roles for alpha 4 integrins and their ligands in angiogenesis. alpha 4 was also found on neural crest derivatives where it colocalized with FN. alpha 4 was expressed selectively on cells in the dorsal root ganglia: it was apparent along ventral projections, but absent from dorsal projections, suggesting that alpha 4 integrins could be involved in defining neuronal fates. Although VCAM-1 was not expressed on most neural crest derivatives, it was found in the neural crest-derived outflow tract of the embryonic heart, where it colocalized with alpha 4. These results imply that alpha 4 integrins and their ligands could be important for migration or differentiation of neural crest. alpha 4 was also expressed on embryonic retina and FN was found on inductive mesenchyme surrounding the eye, suggesting a role for these proteins in eye development. Finally, based on their patterns of expression, we conclude that VCAM-1 only participates in a subset of interactions involving alpha 4 integrins, whereas FN appears to be the more general ligand.

  View details for Web of Science ID A1994NL40700004

  View details for PubMedID 7526952

• THE INTEGRIN ALPHA-4-BETA-1 AND ITS COUNTER-RECEPTOR VCAM-1 IN DEVELOPMENT AND IMMUNE FUNCTION 8th Transatlantic Airway Conference on Integrins and Other Adhesion Molecules Dean, D. C., Iademarco, M. F., Rosen, G. D., Sheppard, A. M. AMER THORACIC SOC. 1993: S43–S46

  Abstract

  The integrin alpha 4 beta 1 and its counter receptor vascular cell adhesion molecule-1 (VCAM-1) mediate well-described cell-cell interactions that are critical for immune function. However, these receptors also mediate cell-cell interactions that are important for skeletal muscle differentiation. We have found that contrasting transcriptional mechanisms control their patterns of expression in the immune system and in muscle. Recent studies indicate that alpha 4 beta 1 and VCAM-1 are also expressed in a number of developing tissues, implying that these receptors have a general role in facilitating cell-cell interactions during development.

  View details for Web of Science ID A1993NH92300005

  View details for PubMedID 7504895

• CHARACTERIZATION OF THE PROMOTER FOR VASCULAR CELL-ADHESION MOLECULE-1 (VCAM-1) JOURNAL OF BIOLOGICAL CHEMISTRY Iademarco, M. F., McQuillan, J. J., Rosen, G. D., Dean, D. C. 1992; 267 (23): 16323-16329

  Abstract

  Vascular cell adhesion molecule-1 (VCAM-1) was first identified as a protein that appears on the surface of endothelial cells after exposure to inflammatory cytokines. Through interaction with its integrin counter receptor VLA-4, VCAM-1 mediates cell-cell interactions important for immune function. We have cloned and begun characterization of the promoter for the VCAM-1 gene. In a series of transfection assays into human umbilical vein endothelial cells (HUVECs), we find that silencers between positions -1.641 kilobases and -288 base pairs restrict promoter activity, and that treatment with tumor necrosis factor-alpha overcomes this inhibition and activates the promoter through two NF kappa B sites located at positions -77 and -63 base pairs of the VCAM-1 gene. This responsiveness appears cell-specific since constructs containing the VCAM-1 NF kappa B sites are not responsive to tumor necrosis factor alpha in the T-cell line Jurkat. The two VCAM-1 NF kappa B sites, which differ slightly in their sequence, form distinct complexes in gel retardation assays, suggesting that they interact with different NF kappa B-site binding proteins. The distribution of these proteins could then control activity of the NF kappa B sites. We conclude that the pattern of VCAM-1 expression in HUVECs is controlled by a combination of these silencers and NF kappa B sites.

  View details for Web of Science ID A1992JJ45800053

  View details for PubMedID 1379595

• ROLES FOR THE INTEGRIN VLA-4 AND ITS COUNTER RECEPTOR VCAM-1 IN MYOGENESIS CELL Rosen, G. D., Sanes, J. R., Lachance, R., Cunningham, J. M., Roman, J., Dean, D. C. 1992; 69 (7): 1107-1119

  Abstract

  Mammalian myogenesis is biphasic: primary myoblasts fuse to form primary myotubes, then secondary myoblasts align along the primary myotubes and form secondary myotubes, which comprise most of adult muscle. We provide evidence that an integrin (VLA-4) and its counter receptor (VCAM-1) have a role in secondary myogenesis. Both receptors are synthesized by cultured muscle cells: VLA-4 is induced as myotubes form, whereas VCAM-1 is present on myoblasts and myotubes. In vivo, both molecules are expressed at sites of secondary myogenesis, VLA-4 on primary and secondary myotubes, and VCAM-1 on secondary myoblasts and on regions of secondary myotubes apposed to primary myotubes. These patterns suggest that VLA-4-VCAM-1 interactions influence alignment of secondary myoblasts along primary myotubes and/or the fusion of secondary myoblasts. In support of the latter possibility, antibodies to VLA-4 or VCAM-1 inhibit myotube formation in culture.

  View details for Web of Science ID A1992JA43100006

  View details for PubMedID 1377605

• IDENTIFICATION OF A PROTEIN THAT INTERACTS WITH THE NUCLEAR FACTOR-I (NF-1) BINDING-SITE IN CELLS THAT DO NOT EXPRESS NF-1 - COMPARISON TO NF-1, CELLULAR-DISTRIBUTION, AND EFFECT ON TRANSCRIPTION NUCLEIC ACIDS RESEARCH McQuillan, J. J., Rosen, G. D., BIRKENMEIER, T. M., Dean, D. C. 1991; 19 (23): 6627-6631

  Abstract

  We examined expression of nuclear factor-1 (NF-1) in different cell lines. Expression was low or undetectable in T and B lymphocyte cell lines, whereas fibroblasts and other adherent cell lines generally had a relatively high level of NF-1 mRNA. In cell lines that did not express NF-1, gel retardation assays, nevertheless, indicated complexes between a protein or proteins and the NF-1 site. These complexes were less abundant than those formed with NF-1, they migrated more slowly, and they appeared as single species instead of the multiple species observed with NF-1. NF-1 site-binding proteins were compared in the fibrosarcoma cell line HT-1080 (expressed the highest level of NF-1 in our study) and the B cell line Raji (does not express NF-1). UV-crosslinking studies indicated that the NF-1 site-binding proteins in both cell lines were similar in size. Proteolytic clipping band shift assays suggested that the Raji protein and NF-1 share structural similarity in their DNA binding domains, but are distinct proteins. The NF-1 site mediated transcriptional stimulation in cell lines where NF-1 is expressed; however, this element did not affect transcription in cell lines that do not express NF-1, suggesting that the NF-1 site-binding protein in these cells is functionally distinct from NF-1.

  View details for Web of Science ID A1991GV81600039

  View details for PubMedID 1754398

• EXPRESSION OF FIBRONECTIN AND ITS CELL-SURFACE RECEPTORS 6TH TRANSATLANTIC AIRWAY CONF : AIRWAY MUCIN Dean, D. C., BIRKENMEIER, T. M., Rosen, G. D., Weintraub, S. J. AMER LUNG ASSOC. 1991: S25–S28

  View details for Web of Science ID A1991GG42400007

• Glycoprotein synthesis and secretion. Expression of fibronectin and its cell surface receptors. American review of respiratory disease Dean, D. C., BIRKENMEIER, T. M., Rosen, G. D., Weintraub, S. J. 1991; 144 (3): S25-8

  Abstract

  Fibronectin (FN) is an extracellular matrix protein that acts as a substrate for cell migration and adhesion during development. Cells adhere to FN through integral membrane proteins that are members of the integrin family of adhesion molecules. The interaction between cells and FN is important in a number of biologic processes, including gastrulation, hematopoietic differentiation, neural crest cell migration, cardiac development, branching morphogenesis in lung, wound healing, tumorigenesis, and metastasis. Expression of FN and its receptors is controlled by a number of hormones and growth factors as well as by tissue-specific factors. Here, the molecular aspects of how expression of these genes is controlled are reviewed, with particular emphasis on promoter regulator elements that modulate expression.

  View details for PubMedID 1832529

• CHARACTERIZATION OF THE ALPHA-4 INTEGRIN GENE PROMOTER PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Rosen, G. D., BIRKENMEIER, T. M., Dean, D. C. 1991; 88 (10): 4094-4098

  Abstract

  A cDNA for the alpha 4 chain of the alpha 4 beta 1 integrin was described previously [Takada, Y., Elices, M. J., Crouse, C. & Hemler, M. E. (1989) EMBO J. 8, 1361-1368]. Primer extension analysis indicated that alpha 4 mRNA extended well beyond the 5' end of this cDNA. To clone this 5' sequence, a primer extension cDNA library was constructed, and a cDNA extending an additional 660 base pairs was isolated. This cDNA hybridized to multiple mRNAs in both T and B lymphocytes, but no alpha 4 mRNA was found in different tissues or in adherent cell lines. A single alpha 4 gene was detected in a genomic Southern blot when hybridization was done at high stringency; however, additional bands were observed at lower stringency, indicating the presence of alpha 4-related genes. Some of the different mRNAs that hybridize to the alpha 4 cDNA may then be the products of these related genes. Analysis of the alpha 4 genomic sequence revealed a large first exon of 958 base pairs. Interestingly, translation of alpha 4 initiates from the second ATG in this exon (nucleotide + 744). The first ATG (nucleotide +21) is followed by a termination codon 21 amino acids downstream. Such upstream ATG codons have been implicated in translational control of protooncogenes. One major transcriptional start site was identified by using S1 nuclease and primer extension mapping. Consensus sequences for DNA regulatory elements were found upstream of the gene and in exon 1 and intron 1. The alpha 4 gene 5' flanking region acted as a promoter in transfection assays. Detailed characterization of the promoter should provide insight into molecular events regulating expression and tissue specificity of alpha 4.

  View details for Web of Science ID A1991FM04200008

  View details for PubMedID 2034655

• IDENTIFICATION OF A CYCLOOXYGENASE-RELATED GENE AND ITS POTENTIAL ROLE IN PROSTAGLANDIN FORMATION BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Rosen, G. D., BIRKENMEIER, T. M., Raz, A., Holtzman, M. J. 1989; 164 (3): 1358-1365

  Abstract

  Regulation of cyclooxygenase expression was studied in homogenous preparations of epithelial cells isolated from sheep tracheal mucosa. Cellular capacity to generate cyclooxygenase-derived arachidonate metabolites (predominantly prostaglandin E2) increased markedly in cultured compared to freshly isolated cells. A 70 kDa cyclooxygenase protein and corresponding 2.8 kb mRNA were coordinately expressed but their levels did not increase proportionately to the increase in cyclooxygenase activity. Rehybridization of Northern blots at lower stringency revealed the presence of a new tissue-specific 4.0 kb mRNA species exhibiting increased expression during cell culture. Hybridization of the 4.0 kb mRNA with two nonoverlapping cDNA probes at only low stringency conditions suggests that it is derived from a distinct gene. Its relatedness to cyclooxygenase and its increase in parallel with enzymatic activity further suggest that the larger mRNA may encode for a cyclooxygenase.

  View details for Web of Science ID A1989AY83200058

  View details for PubMedID 2480117

• UPTAKE, RELEASE AND NOVEL SPECIES-DEPENDENT OXYGENATION OF ARACHIDONIC-ACID IN HUMAN AND ANIMAL AIRWAY EPITHELIAL-CELLS BIOCHIMICA ET BIOPHYSICA ACTA Holtzman, M. J., Hansbrough, J. R., Rosen, G. D., Turk, J. 1988; 963 (3): 401-413

  Abstract

  To determine identities of mediators and mechanisms for their release from pulmonary airway epithelial cells, we examined the capacities of epithelial cells from human, dog and sheep airways to incorporate, release and oxygenate arachidonic acid. Purified cell suspensions were incubated with radiolabeled arachidonic acid and/or ionophore A23187; fatty acid esterification and hydrolysis were traced chromatographically, and oxygenated metabolites were identified using high-pressure liquid chromatography and mass-spectrometry. In each species, cellular uptake of 10 nM arachidonic acid was concentrated in the phosphatidylcholine, phosphatidylinositol and phosphatidylethanolamine fractions, and subsequent incubation with 5 microM A23187 caused release of 10-12% of the radiolabeled pool selectively from phosphatidylcholine and phosphatidylinositol. By contrast, the products of arachidonic acid oxygenation were species-dependent and in the case of human cells were also novel: A23187-stimulated human epithelial cells converted arachidonic acid predominantly to 15-hydroxyeicosatetraenoic acid (15-HETE) and two distinct 8,15-diols in addition to prostaglandin (PG) E2 and PGF2 alpha. Cell incubation with exogenous arachidonic acid (2.0-300 microM) led to progressively larger amounts of 15-HETE and the dihydroxy, epoxyhydroxy and keto acids characteristic of arachidonate 15-lipoxygenase. Both dog and sheep cells converted exogenous or endogenous arachidonic acid to low levels of 5-lipoxygenase products, including leukotriene B4 without significant 15-lipoxygenase activity. In the cyclooxygenase series, sheep cells selectively released PGE2, while dog cells generated predominantly PGD2. The findings demonstrate that stereotyped esterification and phospholipase activities are expressed at uniform levels among airway epithelial cells from these species, but pathways for oxygenating arachidonic acid allow mediator diversity depending greatly on species and little on arachidonic acid presentation.

  View details for Web of Science ID A1988R586300002

  View details for PubMedID 2848586