- Pediatric Infectious Disease
- Primary Immunodeficiency
Chief, Stanford Department of Pediatrics, Division of Immunology and Allergy (2008 - Present)
Professor, Stanford University, Department of Pediatrics (2005 - Present)
Director of the Jeffrey Modell Primary Immunodeficiency Center at Stanford, Supported by the Jeffrey Modell Foundation and the Lucile Packard Fund of Children's Health (2002 - Present)
Associate Professor, Stanford University, Department of Pediatrics (1997 - 2005)
Acting Associate Professor, Stanford University, Department of Pediatrics (1997 - 1997)
Honors & Awards
Henry J. Kaiser Award for Excellence in Preclinical Teaching (Immunology), Kaiser Foundation (2005)
Henry J. Kaiser Award for Excellence in Preclinical Teaching (Immunology), Kaiser Foundation (2001)
Board Certification: Pediatric Infectious Disease, American Board of Pediatrics (1994)
Fellowship:Children's Hospital and Regional Medical Center (1988) WA
Board Certification: Pediatrics, American Board of Pediatrics (1986)
Residency:UCSF Medical Center (1984) CA
Internship:Children's Hospital and Regional Medical Center (1982) WA
Medical Education:UCSF Medical Center (1981) CA
M.D., Univ.Calif at San Francisco, Medicine (1981)
B.S., Yale University, Biology (1976)
Current Research and Scholarly Interests
A long-standing interest is to understand the cellular and molecular basis for this vulnerability of the human neonate to infection with intracellular pathogens that require T helper 1 (Th1) cells [CD4 T cell producing interferon-gamma (IFN-gamma)] for effective immune control. We have previously shown that CD4 T cells of the newborn have a unique limitation in the ability to produce certain effector molecules, such as CD40-ligand (CD154) and IFN-gamma compared to these cells in adults due to both reduced gene transcriptional and impaired signals that lead to gene transcription. Recently, we have shown that these limitations apply to physiological T-cell activation, e.g., using allogeneic dendritic cells. Defining the molecular mechanisms for decreased IFN-gamma production by neonatal CD4 T cells is a current focus.
We have also found that recent thymic emigrants, which predominate in the newborn and young infant, are less able to differentiate into T helper 1 cells, which produce IFN-gamma. These studies required the development of a novel marker for recent thymic emigrants (RTEs) of the CD4 T-cell lineage in humans. Using a combination of approaches, we have identified protein tyrosine kinase 7 (PTK7) as such a marker. In progress are to studies to define the role of PTK7, an orphan member (no known ligand) of the receptor tyrosine kinase family, in T-cell development and immunity, and to determine how this marker can be used to follow the output of recent thymic emigrants in health and disease. We are also interesed in determining the molecular mechanisms for the reduced RTE function and to what extent these mechanisms are shared by neonatal CD4 T cells and CD4+CD8-CD3+ thymocytes, the immediate precursors of antigenically naive CD4 T cells.
We have also found that limitations in T-cell immunogenicity to viruses and viral vaccines extend beyond the neonatal period to childhood. These studies highlight a need to develop more potent vaccines to overcome developmental and other factors, such as genetic inheritance, in mounting adaptive immunity. With this as an ultimate goal, we are examining the ability of a novel adjuvant, cationic liposome DNA complexes (CLDC)(Juvaris Biotherapeutics), to induce durable CD4 and CD8 T-cell immunity and humoral immunity to influenza A. The molecular and cellular components of the innate immune system that are required for immunogenicity are of particular interest. Our preliminary results in mice suggest that the CLDC adjuvant will be substantially more robust than any adjuvants currently approved for clinical use or in clinical trials.
In collaboration with Dr. Neal Boerkoel, University of British Columbia, we are defining the mechanism of T-cell lymphopenia in genetic deficiency of SMARCAL1, a protein that may play a novel role in PolII gene transcription using T cells from patients with SMARCAL1 deficiency (Schimke immuno-osseous dysplasia) as well as SMARCAL1 knockout mice. We hypothesize that the peripheral T-cell lymphopenia is due to attenuation of transcriptional efficiency for multiple genes required for T-cell development and peripheral homeostasis, e.g., cytokines and cytokine receptors.
Independent Studies (11)
- Directed Reading in Immunology
IMMUNOL 299 (Aut, Win, Spr, Sum)
- Directed Reading in Pediatrics
PEDS 299 (Aut, Win, Spr, Sum)
- Early Clinical Experience
PEDS 280 (Aut, Win, Spr, Sum)
- Early Clinical Experience in Immunology
IMMUNOL 280 (Aut, Win, Spr, Sum)
- Graduate Research
IMMUNOL 399 (Aut, Win, Spr, Sum)
- Graduate Research
PEDS 399 (Aut, Win, Spr, Sum)
- Medical Scholars Research
PEDS 370 (Aut, Win, Spr, Sum)
- Out-of-Department Advanced Research Laboratory in Experimental Biology
BIO 199X (Sum)
- Teaching in Immunology
IMMUNOL 290 (Aut, Win, Spr, Sum)
- Undergraduate Directed Reading/Research
PEDS 199 (Aut, Win, Spr, Sum)
- Undergraduate Research
IMMUNOL 199 (Aut, Win, Spr, Sum)
- Directed Reading in Immunology
- Prior Year Courses
Bayesian immunological model development from the literature: Example investigation of recent thymic emigrants.
Journal of immunological methods
2014; 414: 32-50
Bayesian estimation techniques offer a systematic and quantitative approach for synthesizing data drawn from the literature to model immunological systems. As detailed here, the practitioner begins with a theoretical model and then sequentially draws information from source data sets and/or published findings to inform estimation of model parameters. Options are available to weigh these various sources of information differentially per objective measures of their corresponding scientific strengths. This approach is illustrated in depth through a carefully worked example for a model of decline in T-cell receptor excision circle content of peripheral T cells during development and aging. Estimates from this model indicate that 21years of age is plausible for the developmental timing of mean age of onset of decline in T-cell receptor excision circle content of peripheral T cells.
View details for DOI 10.1016/j.jim.2014.08.001
View details for PubMedID 25179832
Somatic reversion in dedicator of cytokinesis 8 immunodeficiency modulates disease phenotype
JOURNAL OF ALLERGY AND CLINICAL IMMUNOLOGY
2014; 133 (6): 1667-1675
Autosomal recessive loss-of-function mutations in dedicator of cytokinesis 8 (DOCK8) cause a combined immunodeficiency characterized by atopy, recurrent infections, and cancer susceptibility. A genotype-phenotype explanation for the variable disease expression is lacking.We investigated whether reversions contributed to the variable disease expression.Patients followed at the National Institutes of Health's Clinical Center were studied. We performed detailed genetic analyses and intracellular flow cytometry to detect DOCK8 protein expression within lymphocyte subsets.We identified 17 of 34 DOCK8-deficient patients who had germline mutations with variable degrees of reversion caused by somatic repair. Somatic repair of the DOCK8 mutations resulted from second-site mutation, original-site mutation, gene conversion, and intragenic crossover. Higher degrees of reversion were associated with recombination-mediated repair. DOCK8 expression was restored primarily within antigen-experienced T cells or natural killer cells but less so in naive T or B cells. Several patients exhibited multiple different repair events. Patients who had reversions were older and had less severe allergic disease, although infection susceptibility persisted. No patients were cured without hematopoietic cell transplantation.In patients with DOCK8 deficiency, only certain combinations of germline mutations supported secondary somatic repair. Those patients had an ameliorated disease course with longer survival but still had fatal complications or required hematopoietic cell transplantation. These observations support the concept that some DOCK8-immunodeficient patients have mutable mosaic genomes that can modulate disease phenotype over time.
View details for DOI 10.1016/j.jaci.2014.03.025
View details for Web of Science ID 000336672500020
View details for PubMedID 24797421
ICOS Regulates the Generation and Function of Human CD4(+) Treg in a CTLA-4 Dependent Manner
2013; 8 (12)
Inducible co-stimulator (ICOS) is a member of CD28/Cytotoxic T-lymphocyte Antigen-4 (CTLA-4) family and broadly expressed in activated CD4(+) T cells and induced regulatory CD4(+) T cells (CD4(+) iTreg). ICOS-related signal pathway could be activated by the interaction between ICOS and its ligand (ICOSL). In our previous work, we established a cost-effective system to generate a novel human allo-antigen specific CD4(hi) Treg by co-culturing their naïve precursors with allogeneic CD40-activated B cells in vitro. Here we investigate the role of ICOS in the generation and function of CD4(hi) Treg by interrupting ICOS-ICOSL interaction with ICOS-Ig. It is found that blockade of ICOS-ICOSL interaction impairs the induction and expansion of CD4(hi) Treg induced by allogeneic CD40-activated B cells. More importantly, CD4(hi) Treg induced with the addition of ICOS-Ig exhibits decreased suppressive capacity on alloantigen-specific responses. Dysfunction of CD4(hi) Treg induced with ICOS-Ig is accompanied with its decreased exocytosis and surface CTLA-4 expression. Through inhibiting endocytosis with E64 and pepstatin A, surface CTLA-4 expression and suppressive functions of induced CD4(hi) Treg could be partly reversed. Conclusively, our results demonstrate the beneficial role of ICOS-ICOSL signal pathway in the generation and function of CD4(hi) Treg and uncover a novel relationship between ICOS and CTLA-4.
View details for DOI 10.1371/journal.pone.0082203
View details for Web of Science ID 000327944500124
View details for PubMedID 24312642
- Defective sphingosine 1-phosphate receptor 1 (S1P(1)) phosphorylation exacerbates T(H)17-mediated autoimmune neuroinflammation NATURE IMMUNOLOGY 2013; 14 (11): 1166-U128
Newborn screening for severe combined immunodeficiency and T-cell lymphopenia in California: Results of the first 2 years
JOURNAL OF ALLERGY AND CLINICAL IMMUNOLOGY
2013; 132 (1): 140-U245
Assay of T-cell receptor excision circles (TRECs) in dried blood spots obtained at birth permits population-based newborn screening (NBS) for severe combined immunodeficiency (SCID).We sought to report the first 2 years of TREC NBS in California.Since August 2010, California has conducted SCID NBS. A high-throughput TREC quantitative PCR assay with DNA isolated from routine dried blood spots was developed. Samples with initial low TREC numbers had repeat DNA isolation with quantitative PCR for TRECs and a genomic control, and immunophenotyping was performed within the screening program for infants with incomplete or abnormal results. Outcomes were tracked.Of 993,724 infants screened, 50 (1/19,900 [0.005%]) had significant T-cell lymphopenia. Fifteen (1/66,250) required hematopoietic cell or thymus transplantation or gene therapy; these infants had typical SCID (n = 11), leaky SCID or Omenn syndrome (n = 3), or complete DiGeorge syndrome (n = 1). Survival to date in this group is 93%. Other T-cell lymphopenic infants had variant SCID or combined immunodeficiency (n = 6), genetic syndromes associated with T-cell impairment (n = 12), secondary T-cell lymphopenia (n = 9), or preterm birth (n = 8). All T-cell lymphopenic infants avoided live vaccines and received appropriate interventions to prevent infections. TREC test specificity was excellent: only 0.08% of infants required a second test, and 0.016% required lymphocyte phenotyping by using flow cytometry.TREC NBS in California has achieved early diagnosis of SCID and other conditions with T-cell lymphopenia, facilitating management and optimizing outcomes. Furthermore, NBS has revealed the incidence, causes, and follow-up of T-cell lymphopenia in a large diverse population.
View details for DOI 10.1016/j.jaci.2013.04.024
View details for Web of Science ID 000321052300019
View details for PubMedID 23810098
Retinoic Acid Hypersensitivity Promotes Peripheral Tolerance in Recent Thymic Emigrants
JOURNAL OF IMMUNOLOGY
2013; 190 (6): 2603-2613
Whereas thymic education eliminates most self-reactive T cells, additional mechanisms to promote tolerance in the periphery are critical to prevent excessive immune responses against benign environmental Ags and some self-Ags. In this study we show that murine CD4(+) recent thymic emigrants (RTEs) are programmed to facilitate tolerance in the periphery. Both in vitro and in vivo, naive RTEs more readily upregulate Foxp3 than do mature naive cells after stimulation under tolerogenic conditions. In RTEs, a relatively high sensitivity to retinoic acid contributes to decreased IFN-? production, permitting the expression of Foxp3. Conversely, mature naive CD4 cells have a lower sensitivity to retinoic acid, resulting in increased IFN-? production and subsequent IFN-?-mediated silencing of Foxp3 expression. Enhanced retinoic acid signaling and Foxp3 induction in RTEs upon Ag encounter in the periphery may serve as form of secondary education that complements thymic education and helps avoid inappropriate immune responses. This mechanism for tolerance may be particularly important in settings where RTEs comprise a large fraction of the peripheral T cell pool, such as in newborns or after umbilical cord blood transplant.
View details for DOI 10.4049/jimmunol.1200852
View details for Web of Science ID 000315657200018
View details for PubMedID 23401586
Human Neonatal Naive CD4(+) T Cells Have Enhanced Activation-Dependent Signaling Regulated by the MicroRNA miR-181a
JOURNAL OF IMMUNOLOGY
2013; 190 (6): 2682-2691
Compared with older children and adults, human neonates have reduced and delayed CD4(+) T cell immunity to certain pathogens, but the mechanisms for these developmental differences in immune function remain poorly understood. We investigated the hypothesis that impaired human neonatal CD4(+) T cell immunity was due to reduced signaling by naive CD4(+) T cells following engagement of the αβ-TCR/CD3 complex and CD28. Surprisingly, calcium flux following engagement of CD3 was significantly higher in neonatal naive CD4(+) T cells from umbilical cord blood (CB) compared with naive CD4(+) T cells from adult peripheral blood. Enhanced calcium flux was also observed in adult CD4(+) recent thymic emigrants. Neonatal naive CD4(+) T cells also had higher activation-induced Erk phosphorylation. The microRNA miR-181a, which enhances activation-induced calcium flux in murine thymocytes, was expressed at significantly higher levels in CB naive CD4(+) T cells compared with adult cells. Overexpression of miR-181a in adult naive CD4(+) T cells increased activation-induced calcium flux, implying that the increased miR-181a levels of CB naive CD4(+) T cells contributed to their enhanced signaling. In contrast, AP-1-dependent transcription, which is downstream of Erk and required for full T cell activation, was decreased in CB naive CD4(+) T cells compared with adult cells. Thus, CB naive CD4(+) T cells have enhanced activation-dependent calcium flux, indicative of the retention of a thymocyte-like phenotype. Enhanced calcium signaling and Erk phosphorylation are decoupled from downstream AP-1-dependent transcription, which is reduced and likely contributes to limitations of human fetal and neonatal CD4(+) T cell immunity.
View details for DOI 10.4049/jimmunol.1202534
View details for Web of Science ID 000315657200026
Genotype, phenotype, and outcomes of nine patients with T-B plus NK plus SCID
2011; 15 (7): 733-741
There are few reports of clinical presentation, genotype, and HCT outcomes for patients with T-B+NK+ SCID. Between 1981 and 2007, eight of 84 patients with SCID who received and/or were followed after HCT at UCSF had the T-B+NK+ phenotype. One additional patient with T-B+NK+ SCID was identified as the sibling of a patient treated at UCSF. Chart reviews were performed. Molecular analyses of IL7R, IL2RG, JAK3, and the genes encoding the CD3 T-cell receptor components ? (CD3D), ? (CD3E), and ? (CD3Z) were carried out. IL7R mutations were documented in four patients and CD3D mutations in two others. Three patients had no defects found. Only two of nine patients had an HLA-matched related HCT donor. Both survived, and neither developed GVHD. Five of seven recipients of haploidentical grafts survived. Although the majority of reported cases of T-B+NK+ SCID are caused by defects in IL7R, CD3 complex defects were also found in this series and should be considered when evaluating patients with T-B+NK+ SCID. Additional genes, mutations in which account for T-B+NK+ SCID, remain to be found. Better approaches to early diagnosis and HCT treatment are needed for patients lacking an HLA-matched related donor.
View details for DOI 10.1111/j.1399-3046.2011.01563.x
View details for Web of Science ID 000296049000020
View details for PubMedID 21883749
Protein tyrosine kinase 7: a novel surface marker for human recent thymic emigrants with potential clinical utility
JOURNAL OF PERINATOLOGY
2011; 31: S72-S81
Recent thymic emigrants (RTEs) are antigenically naive T cells that have recently completed intrathymic maturation and have emigrated from the thymus to the periphery. RTEs are clinically and immunologically important as they are essential for maintaining peripheral T cells in sufficient numbers in order to recognize, by their ??T-cell receptors (TCRs), a diverse array of foreign peptide antigens. However, RTE frequency and function has been poorly understood because of a lack of surface markers to distinguish them from older non-RTE naive T cells. This review summarizes the biology of the intrathymic generation and function of RTEs, including the recent identification of protein tyrosine kinase 7 (PTK7) as a novel marker for human RTEs of the CD4 (helper) T-cell lineage. PTK7+ RTEs in adults have a reduced capacity for activation-induced proliferation and cytokine production (interleukin-2 and interferon-?) than older PTK7- naive CD4 T cells. Importantly, this immaturity in CD4 RTE effector function may contribute to the reduced adaptive immune responses observed in situations in which CD4 RTEs predominate, including the fetus, neonate and young infant, and following immune reconstitution, such as post-hematopoietic stem cell transplant. The ability to identify viable CD4+ RTEs based on PTK7 surface staining may be particularly useful in the infant for better defining the impact of nutritional and environmental factors on thymic output, peripheral T-cell function and adaptive immune responses to vaccination and infection.
View details for DOI 10.1038/jp.2010.187
View details for Web of Science ID 000289236900012
View details for PubMedID 21448210
CROSS-REACTIVE NEUTRALIZING ANTIBODY AGAINST PANDEMIC 2009 H1N1 INFLUENZA A VIRUS IN INTRAVENOUS IMMUNOGLOBULIN PREPARATIONS
PEDIATRIC INFECTIOUS DISEASE JOURNAL
2011; 30 (1): 67-69
Prepandemic intravenous immunoglobulin (IVIG) and sera from Kawasaki disease patients treated with this IVIG were analyzed for 2009 H1N1-specific microneutralization and hemagglutination inhibition antibodies. All 6 different IVIG preparations tested had significant levels of cross-reactive-specific antibody at a concentration of 2.0 g/dL of immunoglobulin. Sera from 18 of 19 Kawasaki disease patients had significant increases of cross-reactive-specific antibody after 2.0 g/kg of prepandemic IVIG. These results suggest a role for adjunctive IVIG therapy for severe and/or drug-resistant 2009 H1N1 virus and other highly antigenically drifted influenza strains, particularly in the immunocompromised.
View details for DOI 10.1097/INF.0b013e3181f127be
View details for Web of Science ID 000285498800017
View details for PubMedID 20724956
Cationic Lipid/DNA Complex-Adjuvanted Influenza A Virus Vaccination Induces Robust Cross-Protective Immunity
JOURNAL OF VIROLOGY
2010; 84 (24): 12691-12702
Influenza A virus is a negative-strand segmented RNA virus in which antigenically distinct viral subtypes are defined by the hemagglutinin (HA) and neuraminidase (NA) major viral surface proteins. An ideal inactivated vaccine for influenza A virus would induce not only highly robust strain-specific humoral and T-cell immune responses but also cross-protective immunity in which an immune response to antigens from a particular viral subtype (e.g., H3N2) would protect against other viral subtypes (e.g., H1N1). Cross-protective immunity would help limit outbreaks from newly emerging antigenically novel strains. Here, we show in mice that the addition of cationic lipid/noncoding DNA complexes (CLDC) as adjuvant to whole inactivated influenza A virus vaccine induces significantly more robust adaptive immune responses both in quantity and quality than aluminum hydroxide (alum), which is currently the most widely used adjuvant in clinical human vaccination. CLDC-adjuvanted vaccine induced higher total influenza virus-specific IgG, particularly for the IgG2a/c subclass. Higher levels of multicytokine-producing influenza virus-specific CD4 and CD8 T cells were induced by CLDC-adjuvanted vaccine than with alum-adjuvanted vaccine. Importantly, CLDC-adjuvanted vaccine provided significant cross-protection from either a sublethal or lethal influenza A viral challenge with a different subtype than that used for vaccination. This superior cross-protection afforded by the CLDC adjuvant required CD8 T-cell recognition of viral peptides presented by classical major histocompatibility complex class I proteins. Together, these results suggest that CLDC has particular promise for vaccine strategies in which T cells play an important role and may offer new opportunities for more effective control of human influenza epidemics and pandemics by inactivated influenza virus vaccine.
View details for DOI 10.1128/JVI.00769-10
View details for Web of Science ID 000284469600023
View details for PubMedID 20943978
Revisiting Human IL-12R beta 1 Deficiency A Survey of 141 Patients From 30 Countries
2010; 89 (6): 381-402
Interleukin-12 receptor ?1 (IL-12R?1) deficiency is the most common form of Mendelian susceptibility to mycobacterial disease (MSMD). We undertook an international survey of 141 patients from 102 kindreds in 30 countries. Among 102 probands, the first infection occurred at a mean age of 2.4 years. In 78 patients, this infection was caused by Bacille Calmette-Guérin (BCG; n = 65), environmental mycobacteria (EM; also known as atypical or nontuberculous mycobacteria) (n = 9) or Mycobacterium tuberculosis (n = 4). Twenty-two of the remaining 24 probands initially presented with nontyphoidal, extraintestinal salmonellosis. Twenty of the 29 genetically affected sibs displayed clinical signs (69%); however 8 remained asymptomatic (27%). Nine nongenotyped sibs with symptoms died. Recurrent BCG infection was diagnosed in 15 cases, recurrent EM in 3 cases, recurrent salmonellosis in 22 patients. Ninety of the 132 symptomatic patients had infections with a single microorganism. Multiple infections were diagnosed in 40 cases, with combined mycobacteriosis and salmonellosis in 36 individuals. BCG disease strongly protected against subsequent EM disease (p = 0.00008). Various other infectious diseases occurred, albeit each rarely, yet candidiasis was reported in 33 of the patients (23%). Ninety-nine patients (70%) survived, with a mean age at last follow-up visit of 12.7 years ± 9.8 years (range, 0.5-46.4 yr). IL-12R?1 deficiency is characterized by childhood-onset mycobacteriosis and salmonellosis, rare recurrences of mycobacterial disease, and more frequent recurrence of salmonellosis. The condition has higher clinical penetrance, broader susceptibility to infections, and less favorable outcome than previously thought.
View details for DOI 10.1097/MD.0b013e3181fdd832
View details for Web of Science ID 000283840300002
View details for PubMedID 21057261
Alternate Mechanisms of Initial Pattern Recognition Drive Differential Immune Responses to Related Poxviruses
CELL HOST & MICROBE
2010; 8 (2): 174-185
Vaccinia immunization was pivotal to successful smallpox eradication. However, the early immune responses that distinguish poxvirus immunization from pathogenic infection remain unknown. To address this, we developed a strategy to map the activation of key signaling networks in vivo and applied this approach to define and compare the earliest signaling events elicited by immunizing (vaccinia) and lethal (ectromelia) poxvirus infections in mice. Vaccinia induced rapid TLR2-dependent responses, leading to IL-6 production, which then initiated STAT3 signaling in dendritic and T cells. In contrast, ectromelia did not induce TLR2 activation, and profound mouse strain-dependent responses were observed. In resistant C57BL/6 mice, the STAT1 and STAT3 pathways were rapidly activated, whereas in susceptible BALB/c mice, IL-6-dependent STAT3 activation did not occur. These data link early immune signaling events to infection outcome and suggest that activation of different pattern-recognition receptors early after infection may be important in determining vaccine efficacy.
View details for DOI 10.1016/j.chom.2010.07.008
View details for Web of Science ID 000281169600007
View details for PubMedID 20709294
Adult-onset presentations of genetic immunodeficiencies: genes can throw slow curves
CURRENT OPINION IN INFECTIOUS DISEASES
2010; 23 (4): 359-364
The molecular and genetic mechanisms behind adult presentations of primary immunodeficiency diseases are examined, with particular emphasis on cases where this was heralded by severe, recurrent, or opportunistic infection.A detailed analysis over the last two decades of the relationship between genotype and clinical phenotype for a number of genetic immunodeficiencies has revealed multiple mechanisms that can account for the delayed presentation of genetic disorders that typically present in childhood, including hypomorphic gene mutations and X-linked gene mutations with age-related skewing in random X-chromosome inactivation. Adult-onset presentations of chronic granulomatous disease, X-linked agammaglobulinemia, IL-12/Th1/IFN-gamma and IL-23/Th17/IL-17 pathway defects, and X-linked lymphoproliferative disorder are used to illustrate these mechanisms. Finally, certain genetic types of common variable immunodeficiency are used to illustrate that inherited null mutations can take decades to manifest immunologically.Both genetic mechanisms and environmental factors can account for adult-onset infectious and noninfectious complications as manifestations of disorders that are typically present in childhood. This emphasizes the potential complexity in the relationship between genotype and phenotype with natural human mutations.
View details for DOI 10.1097/QCO.0b013e32833bc1b0
View details for Web of Science ID 000279397600010
View details for PubMedID 20581672
A new genetic subgroup of chronic granulomatous disease with autosomal recessive mutations in p40(phox) and selective defects in neutrophil NADPH oxidase activity
2009; 114 (15): 3309-3315
Chronic granulomatous disease (CGD), an immunodeficiency with recurrent pyogenic infections and granulomatous inflammation, results from loss of phagocyte superoxide production by recessive mutations in any 1 of 4 genes encoding subunits of the phagocyte NADPH oxidase. These include gp91(phox) and p22(phox), which form the membrane-integrated flavocytochrome b, and cytosolic subunits p47(phox) and p67(phox). A fifth subunit, p40(phox), plays an important role in phagocytosis-induced superoxide production via a phox homology (PX) domain that binds to phosphatidylinositol 3-phosphate (PtdIns(3)P). We report the first case of autosomal recessive mutations in NCF4, the gene encoding p40(phox), in a boy who presented with granulomatous colitis. His neutrophils showed a substantial defect in intracellular superoxide production during phagocytosis, whereas extracellular release of superoxide elicited by phorbol ester or formyl-methionyl-leucyl-phenylalanine (fMLF) was unaffected. Genetic analysis of NCF4 showed compound heterozygosity for a frameshift mutation with premature stop codon and a missense mutation predicting a R105Q substitution in the PX domain. Parents and a sibling were healthy heterozygous carriers. p40(phox)R105Q lacked binding to PtdIns(3)P and failed to reconstitute phagocytosis-induced oxidase activity in p40(phox)-deficient granulocytes, with premature loss of p40(phox)R105Q from phagosomes. Thus, p40(phox) binding to PtdIns(3)P is essential for phagocytosis-induced oxidant production in human neutrophils and its absence can be associated with disease.
View details for DOI 10.1182/blood-2009-07-231498
View details for Web of Science ID 000270595700024
View details for PubMedID 19692703
Efficient Induction and Expansion of Human Alloantigen-Specific CD8 Regulatory T Cells from Naive Precursors by CD40-Activated B Cells
JOURNAL OF IMMUNOLOGY
2009; 183 (6): 3742-3750
Although recent studies have focused on CD4(+) regulatory T cells (Treg), CD8(+) Treg have also been reported to play important roles in the induction and maintenance of immune tolerance. Adoptive transfer of CD8(+) Treg in rodents or induction of CD8(+) Treg in humans can prevent or treat allograft rejection and autoimmune diseases. However, no approaches have been reported for the generation of human Ag-specific CD8(+) Treg at a practical scale for clinical use. Here, we found that two novel CD8(+) T cell subsets with different levels of CD8 surface expression, CD8(high) and CD8(low), could be induced from naive CD8(+) precursors in vitro by allogeneic CD40-activated B cells, whereas only CD8(high) T cells were alloantigen-specific Treg with relatively poor alloantigen-specific cytotoxicity. Importantly, alloantigen-specific CD8(high) Treg could be induced and expanded from naive CD8(+)CD25(-) T cells at a large scale after 3 wk of culture without exogenous cytokines. These induced alloantigen-specific Treg were CD45RO(+) and CCR7(-) memory cells, and they expressed Foxp3, CD25, CD27, CD28, and CD62L. The induction and expansion of CD8(high) Treg by CD40-activated B cells were dependent on endogenously expressed IFN-gamma, IL-2, IL-4, and CTLA-4. This approach may facilitate the clinical application of CD8(+) Treg-based immunotherapy in transplantation and autoimmune diseases.
View details for DOI 10.4049/jimmunol.0901329
View details for Web of Science ID 000270179700025
View details for PubMedID 19684082
Cationic lipid/DNA complexes (JVRS-100) combined with influenza vaccine (Fluzone (R)) increases antibody response, cellular immunity, and antigenically drifted protection
2009; 27 (29): 3811-3820
Safe and effective adjuvants for influenza vaccines that could increase both the levels of neutralizing antibody, including against drifted viral subtypes, and T-cell immunity would be a major advance in vaccine design. The JVRS-100 adjuvant, consisting of DOTIM/cholesterol cationic liposome-DNA complexes, is particularly promising for vaccines that require induction of high levels of antibody and T-cell immunity, including CD8(+) cytotoxic T lymphocytes (CTL). Inclusion of protein antigens with JVRS-100 results in the induction of enhanced humoral and cell-mediated (i.e., CD4(+) and CD8(+) T cells) immune responses. The JVRS-100 adjuvant combined with a split trivalent influenza vaccine (Fluzone-sanofi pasteur) elicited increased antibody and T-cell responses in mice and non-human primates compared to vaccination with Fluzone alone. Mice vaccinated with JVRS-100-Fluzone and challenged with antigenically drifted strains of H1N1 (PR/8/34) and influenza B (B/Lee/40) viruses had higher grade protection, as measured by attenuation of weight loss and increased survival, compared to recipients of unadjuvanted vaccine. The results indicate that the JVRS-100 adjuvant substantially increases immunogenicity and protection from drifted-strain challenge using an existing influenza vaccine.
View details for DOI 10.1016/j.vaccine.2009.04.054
View details for Web of Science ID 000267581300003
View details for PubMedID 19406188
Human CD4(+) T cell recent thymic emigrants are identified by protein tyrosine kinase 7 and have reduced immune function
JOURNAL OF EXPERIMENTAL MEDICINE
2009; 206 (2): 275-285
CD4(+) recent thymic emigrants (RTEs) comprise a clinically and immunologically important T cell population that indicates thymic output and that is essential for maintaining a diverse alphabeta-T cell receptor (TCR) repertoire of the naive CD4(+) T cell compartment. However, their frequency and function are poorly understood because no known surface markers distinguish them from older non-RTE naive CD4(+) T cells. We demonstrate that protein tyrosine kinase 7 (PTK7) is a novel marker for human CD4(+) RTEs. Consistent with their recent thymic origin, human PTK7(+) RTEs contained higher levels of signal joint TCR gene excision circles and were more responsive to interleukin (IL)-7 compared with PTK7(-) naive CD4(+) T cells, and rapidly decreased after complete thymectomy. Importantly, CD4(+) RTEs proliferated less and produced less IL-2 and interferon-gamma than PTK7(-) naive CD4(+) T cells after alphabeta-TCR/CD3 and CD28 engagement. This immaturity in CD4(+) RTE effector function may contribute to the reduced CD4(+) T cell immunity observed in contexts in which CD4(+) RTEs predominate, such as in the fetus and neonate or after immune reconstitution. The ability to identify viable CD4(+) RTEs by PTK7 staining should be useful for monitoring thymic output in both healthy individuals and in patients with genetic or acquired CD4(+) T cell immunodeficiencies.
View details for DOI 10.1084/jem.20080996
View details for Web of Science ID 000266008800004
View details for PubMedID 19171767
Efficient generation of human alloantigen-specific CD4(+) regulatory T cells from naive precursors by CD40-activated B cells
2008; 112 (6): 2554-2562
CD4(+)CD25(+)Foxp3(+) regulatory T cells (Treg) play an important role in the induction and maintenance of immune tolerance. Although adoptive transfer of bulk populations of Treg can prevent or treat T cell-mediated inflammatory diseases and transplant allograft rejection in animal models, optimal Treg immunotherapy in humans would ideally use antigen-specific rather than polyclonal Treg for greater specificity of regulation and avoidance of general suppression. However, no robust approaches have been reported for the generation of human antigen-specific Treg at a practical scale for clinical use. Here, we report a simple and cost-effective novel method to rapidly induce and expand large numbers of functional human alloantigen-specific Treg from antigenically naive precursors in vitro using allogeneic nontransformed B cells as stimulators. By this approach naive CD4(+)CD25(-) T cells could be expanded 8-fold into alloantigen-specific Treg after 3 weeks of culture without any exogenous cytokines. The induced alloantigen-specific Treg were CD45RO(+)CCR7(-) memory cells, and had a CD4(high), CD25(+), Foxp3(+), and CD62L (L-selectin)(+) phenotype. Although these CD4(high)CD25(+)Foxp3(+) alloantigen-specific Treg had no cytotoxic capacity, their suppressive function was cell-cell contact dependent and partially relied on cytotoxic T lymphocyte antigen-4 expression. This approach may accelerate the clinical application of Treg-based immunotherapy in transplantation and autoimmune diseases.
View details for DOI 10.1182/blood-2008-04-152041
View details for Web of Science ID 000259088000052
View details for PubMedID 18599794
Focus on FOCIS: The continuing diagnostic challenge of autosomal recessive chronic granulomatous disease
2008; 128 (2): 117-126
Chronic granulomatous disease (CGD) is a primary immunodeficiency of defective neutrophil oxidative burst activity due to mutations in the genes CYBA, NCF-1, NCF-2, and CYBB, which respectively encode the p22-phox, p47-phox, p67-phox, and gp91-phox subunits. CGD usually presents in early childhood with recurrent or severe infection with catalase-positive bacteria and fungi. We present an unusual case of CGD in which Burkholderia cepacia lymphadenitis developed in a previously healthy 10-year-old girl. Flow cytometric analysis of dihydrorhodamine (DHR)-labeled neutrophils performed by a CLIA-approved outside reference laboratory was reported as normal. However, we found that this patient's neutrophil oxidative burst activity in DHR assays was substantially reduced but not absent. A selective decrease in intracellular staining for p67-phox suggested the diagnosis of autosomal recessive CGD due to NCF-2 gene mutations, and a novel homozygous and hypomorphic NCF-2 gene mutation was found. The potential mechanisms for this delayed and mild presentation of CGD are discussed.
View details for DOI 10.1016/j.clim.2008.05.008
View details for Web of Science ID 000257941400001
View details for PubMedID 18625437
Maternal T-cell engraftment associated with severe hemophagocytosis of the bone marrow in untreated X-linked severe combined immunodeficiency
JOURNAL OF PEDIATRIC HEMATOLOGY ONCOLOGY
2008; 30 (5): 396-400
Maternal engraftment of T cells in severe combined immunodeficiency can lead to graft-versus-host disease of the skin and liver. We report the case of an infant with X-linked severe combined immunodeficiency, confirmed by DNA sequencing of the common gamma chain gene locus, in which this disorder's characteristic peripheral lymphocyte phenotype [T(-)B(+)NK(-)] was obscured by the postnatal onset of hemophagocytic syndrome that included severe B-cell lymphopenia, neutropenia, and anemia. Hemophagocytosis was most likely owing to maternal graft-versus-host disease, as perforin-expressing CD8 T cells, presumably of maternal origin, were prominent in the bone marrow and there was no concurrent severe infection.
View details for Web of Science ID 000255614700014
View details for PubMedID 18458578
Asymmetric dimethylarginine and cardiac allograft vasculopathy progression: Modulation by sirolimus
2008; 85 (6): 827-833
Cardiac allograft vasculopathy (CAV) is a major cause of death after heart transplantation (HT). The reduced bioavailability of endothelium-derived nitric oxide may play a role in endothelial vasodilator dysfunction and thus in the structural changes characterizing CAV. A potential contributor to endothelial pathobiology is asymmetric dimethylarginine (ADMA), an endogenous nitric oxide synthase inhibitor. It was hypothesized that ADMA concentrations may influence CAV progression during the first postoperative year.Thirty-two consecutive HT recipients underwent intravascular ultrasound evaluation at month 1 and year 1 after HT. Immunosuppression included mycophenolate mofetil (MMF, n=16) and sirolimus (n=16). Change in intimal volume greater than the median and vascular remodeling were major outcome measures.Plasma ADMA levels were associated with subsequent development of intimal hyperplasia (risk ratio [95% confidence interval] =2.72 [1.06-6.94]; P=0.038), and plasma ADMA levels greater than 0.70 micromol/L most accurately identified patients who would have developed intimal hyperplasia. However, ADMA levels did not correlate with negative coronary remodeling. Treatment with sirolimus, as compared with MMF, was associated with significantly lower ADMA levels (0.65+/-0.12 vs. 0.77+/-0.10 micromol/L; P<0.01) and less intimal hyperplasia (risk ratio [95% confidence interval] = 0.08 [0.01-0.56]; P=0.01).Elevated plasma ADMA is associated with coronary intimal hyperplasia, supporting the importance of nitric oxide synthase inhibition in CAV pathogenesis. Treatment with sirolimus (rather than MMF) is associated with lower ADMA levels and reduced risk of accelerated CAV.
View details for DOI 10.1097/TP.0b013e318166a3a4
View details for Web of Science ID 000254592000008
View details for PubMedID 18360263
- A novel mutation associated with autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy ANNALS OF ALLERGY ASTHMA & IMMUNOLOGY 2008; 100 (2): 169-169
Influenza A virus elevates active cathepsin B in primary murine DC
2007; 19 (5): 645-655
Dendritic cells (DCs) act as a first-line recognition system for invading pathogens, such as influenza A. The interaction of DC with influenza A virus results in DC activation via endosomal Toll-like receptors and also leads to presentation of viral peptides on MHC class II molecules. Prior work demonstrated that influenza A virus (A/HKx31; H3N2) infection of BALB/c mice activates lung DCs for antigen presentation, and that the enhanced function of these cells persists long after viral clearance and resolution of the virus-induced inflammatory response. Whether influenza A virus has acute or longer-lasting effects on the endo/lysosomal antigen-processing machinery of DCs has not been studied. Here, we show that antigen presentation from intact protein antigen, but not peptide presentation, results in increased T cell stimulation by influenza-exposed lung DCs, suggesting increased antigen processing/loading in these DCs. We find that cathepsin (Cat) B levels and activity are substantially up-regulated in murine lung DCs, harvested 30 days after A/HKx31 infection. CatB levels and activity are also increased in murine splenic and bone marrow-derived DCs, following short-term in vitro exposure to UV-inactivated influenza A virus. Modest effects on CatX are also seen during in vivo and in vitro exposure to influenza A virus. Using a cell permeable Cat inhibitor, we show Cats in influenza-exposed DCs to be functional and required for generation of a T cell epitope from intact ovalbumin. Our findings indicate that influenza A virus affects the MHC class II antigen-processing pathway, an essential pathway for CD4(+) T cell activation.
View details for DOI 10.1093/intimm/dxm030
View details for Web of Science ID 000246964500007
View details for PubMedID 17446210
Humoral and cellular immune responses in children given annual immunization with trivalent inactivated influenza vaccine
PEDIATRIC INFECTIOUS DISEASE JOURNAL
2007; 26 (2): 107-115
There have been no prior reports of the frequency of circulating influenza-specific, interferon gamma-producing memory CD4+ and CD8+ T-cells in healthy children who have received multiple influenza immunizations.We evaluated 21 previously immunized children, ages 3 to 9 years, before and 1 month after administration of trivalent inactivated influenza vaccine. Frequencies of influenza-specific CD4+ and CD8+ T-cells stimulated with trivalent inactivated influenza vaccine or A/Panama (H3N2) virus were determined by flow cytometry, and antibody responses to vaccine strains and a drifted H3N2 strain were measured by hemagglutination inhibition assay and neutralizing antibody assays.Mean change in CD4+ and in CD8+ T-cell frequencies after immunization was 0.01% (P > 0.39) with postimmunization CD4+ frequencies higher than CD8+ frequencies. Children with more previous vaccinations had a higher baseline frequency of CD4+ T-cells (P = 0.0002) but a smaller increase or even a decline from baseline after immunization (P = 0.003). An association between age and change in frequency was not detected. Baseline geometric mean titers (GMTs) and seroprotection rates were significantly higher in older children against A/Panama (neutralizing baseline GMT, P = 0.0488) and A/New Caledonia (hemagglutination inhibition baseline GMT and seroprotection, P < 0.0297). Baseline GMTs against B/Hong Kong were not associated with age or quantity of prior vaccinations.These findings suggest that children may plateau in CD4+ T-cell responses to influenza antigens with repeated exposures and that the number of exposures may play a large role in building a memory CD4+ T-cell response to influenza A, perhaps independently from age.
View details for DOI 10.1097/01.inf.0000253251.03785.9b
View details for Web of Science ID 000243985700002
View details for PubMedID 17259871
[Coronary allograft vasculopathy: pathophysiological interaction between the immune system, infections and metabolic syndrome].
Giornale italiano di cardiologia (2006)
2007; 8 (2): 73-82
Cardiac allograft vasculopathy is still the main cause of long-term graft loss after heart transplantation. Indeed, recent advances in immunosuppression management led to a significant improvement in short-term survival, while long-term death rate did not change significantly in the last 20 years. In this paper, we will review the latest advances in the understanding of this peculiar form of atherosclerosis, focusing on the mechanisms that can be potentially targeted by specific therapeutic interventions.
View details for PubMedID 17402351
Cellular immune responses in children and adults receiving inactivated or live attenuated influenza vaccines
JOURNAL OF VIROLOGY
2006; 80 (23): 11756-11766
The patterns of cellular immune responses induced by live attenuated influenza vaccine (LAIV) versus those of the trivalent inactivated influenza vaccine (TIV) have not been studied extensively, especially in children. The goals of this study were to evaluate the effects of TIV and LAIV immunization on cellular immunity to live influenza A virus in children and adults and to explore factors associated with variations in responses to influenza vaccines among individuals. A gamma interferon (IFN-gamma) flow cytometry assay was used to measure IFN-gamma-producing (IFN-gamma+) NK and T cells in peripheral blood mononuclear cell cultures stimulated with a live influenza A virus strain before and after LAIV or TIV immunization of children and adults. The mean percentages of influenza A virus-specific IFN-gamma+ CD4 and CD8 T cells increased significantly after LAIV, but not TIV, immunization in children aged 5 to 9 years. No increases in the mean levels of influenza A virus-reactive IFN-gamma+ T cells and NK cells were observed in adults given LAIV or TIV. TIV induced a significant increase in influenza A virus-reactive T cells in 6-month- to 4-year-old children; LAIV was not evaluated in this age group. The postvaccination changes (n-fold) in the percentages of influenza A virus-reactive IFN-gamma+ T and NK cells in adults were highly variable and correlated inversely with the prevaccination percentages, in particular with that of the CD56(dim) NK cell subset. In conclusion, our findings identify age, type of vaccine, and prevaccination levels of immune reactivity to influenza A virus as factors significantly associated with the magnitude of cellular immune responses to influenza vaccines.
View details for DOI 10.1128/JVI.01460-06
View details for Web of Science ID 000242222200032
View details for PubMedID 16971435
T-cell immunity to subclinical cytomegalovirus infection reduces cardiac allograft disease
2006; 114 (15): 1608-1615
Asymptomatic cytomegalovirus (CMV) replication is frequent after cardiac transplantation in recipients with pretransplantation CMV infection. How subclinical viral replication influences cardiac allograft disease remains poorly understood, as does the importance of T-cell immunity in controlling such replication.Thirty-nine cardiac recipients who were pretransplantation CMV antibody positive were longitudinally studied for circulating CMV-specific CD4 and CD8 T-cell responses, CMV viral load in blood neutrophils, and allograft rejection during the first posttransplantation year. Nineteen of these recipients were also analyzed for changes of coronary artery intimal, lumen, and whole-vessel area. All recipients received early prophylactic therapy with ganciclovir. No recipients developed overt CMV disease. Those with detectable levels of CMV-specific CD4 T cells in the first month after transplantation were significantly protected from high mean and peak posttransplantation viral load (P<0.05), acute rejection (P<0.005), and loss of allograft coronary artery lumen (P<0.05) and of whole-vessel area (P<0.05) compared with those who lacked this immune response. The losses of lumen and vessel area were both significantly correlated with the time after transplantation at which a CD4 T-cell response was first detected (P<0.05) and with the cumulative graft rejection score (P<0.05).The early control of subclinical CMV replication after transplantation by T-cell immunity may limit cardiac allograft rejection and vascular disease. Interventions to increase T-cell immunity might be clinically useful in limiting these adverse viral effects.
View details for DOI 10.1161/CIRCULATIONAHA.105.607549
View details for Web of Science ID 000241077600011
View details for PubMedID 17015794
Cutting edge: Decreased accumulation and regulatory function of CD4(+)CD25(high) T cells in human STAT 5b deficiency
JOURNAL OF IMMUNOLOGY
2006; 177 (5): 2770-2774
We show that STAT5b is important for the in vivo accumulation of CD4+ CD25(high) T cells with regulatory cell function. A patient homozygous for a missense A630P STAT5b mutation displayed immune dysregulation and decreased numbers of CD4+ CD25(high) T cells. STAT5b(A630P/A630P) CD4+ CD25(high) T cells had low expression of forkhead box P3 and an impaired ability to suppress the proliferation of or to kill CD4+ CD25- T cells. Expression of CD25, a component of the high-affinity IL-2R, was also reduced in response to IL-2 or after in vitro propagation. The impact of the STAT5b mutation was selective in that IL-2-mediated up-regulation of the common gamma-chain cytokine receptor and perforin, and activation-induced expressions of CD154 and IFN-gamma were normal. These results indicate that STAT5b propagates an important IL-2-mediated signal for the in vivo accumulation of functional regulatory T cells.
View details for Web of Science ID 000240002800007
View details for PubMedID 16920911
Acute rejection and cardiac allograft vascular disease is reduced by suppression of subclinical cytomegalovirus infection
2006; 82 (3): 398-405
Anticytomegalovirus (CMV) prophylaxis prevents the acute disease but its impact on subclinical infection and allograft outcome is unknown. We sought to determine whether CMV prophylaxis administered for three months after heart transplant would improve patient outcomes.This prospective cohort study of 66 heart transplant recipients compared aggressive CMV prophylaxis (n = 21, CMV hyperimmune globulin [CMVIG] plus four weeks of intravenous ganciclovir followed by two months of valganciclovir); with standard prophylaxis (n = 45, intravenous ganciclovir for four weeks). Prophylaxis was based on pretransplant donor (D) and recipient (R) CMV serology: R-/D+ received aggressive prophylaxis; R+ received standard prophylaxis. Outcome measures were: CMV infection assessed by DNA-polymerase chain reaction on peripheral blood polymorphonuclear leukocytes, acute rejection, and cardiac allograft vascular disease (CAV) assessed by intravascular ultrasound. All patients completed one year of follow-up. RESULTS.: CMV infection was subclinical in all but four patients (two in each group). Aggressively treated patients had a lower incidence of CMV infection (73 +/- 10% vs. 94 +/- 4%; P = 0.038), and an independent reduced relative risk for acute rejection graded > or =3A (relative risk [95% CI] = 0.55 [0.26-0.96]; P = 0.03), as compared with the standard prophylaxis group. Aggressively prophylaxed patients also showed a slower progression of CAV, in terms of coronary artery lumen loss (lumen volume change=-21 +/- 13% vs. -10+/-14%; P = 0.05); and vessel shrinkage (vessel volume change = -15 +/- 11% vs. -3 +/- 18%; P = 0.03).Prolonged (val)ganciclovir plus CMVIG reduces viral levels, acute rejection, and allograft vascular disease, suggesting a role for chronic subclinical infection in the pathophysiology of the most common diseases affecting heart transplant recipients.
View details for DOI 10.1097/01.tp.0000229039.87735.76
View details for Web of Science ID 000239884800018
View details for PubMedID 16906040
Newborn immunology: relevance to the clinician.
Current problems in pediatric and adolescent health care
2006; 36 (5): 189-204
View details for PubMedID 16631097
Prevention of subclinical CMV infection reduces cardiac allograft disease progression by positively affecting coronary remodeling
ELSEVIER SCIENCE INC. 2006: S138-S139
View details for Web of Science ID 000203407400273
Impaired allogeneic activation and T-helper differentiation of human cord blood naive CD4 T cells
BIOLOGY OF BLOOD AND MARROW TRANSPLANTATION
2006; 12 (2): 160-171
CD4 T cells, particularly those of the T-helper 1 (Th1) subset, are important effectors in alloimmune diseases, such as graft-versus-host disease, and in controlling infections with intracellular pathogens. Thus, it is plausible that impaired neonatal CD4 T-cell immunity might contribute to the low incidence of acute graft-versus-host disease after allogeneic transplantation of hematopoietic stem cells using cord blood (CB) compared with adult sources of hematopoietic stem cells. In support of this hypothesis, we found that CB naive CD4 T cells had reduced activation and impaired early Th1 differentiation compared with adult peripheral blood naive CD4 T cells after stimulation by allogeneic dendritic cells derived from adult monocytes. Early Th1 polarization was dependent on interleukin-12 and CD154, and CB CD4 T cell/dendritic cell co-cultures had impaired expression of both proteins. CB naive CD4 T cells had low basal levels of signal transduction and activation of transcription 4 messenger RNA and protein, and, after alloantigen stimulation, reduced interleukin-12-induced signal transduction and activation of transcription 4 tyrosine phosphorylation, compared with adult peripheral blood naive T cells. Lastly, FoxP3 protein expression, a marker for regulatory CD25(high) CD4 T cells, was lower for naive CD4 T cells of CB compared with those of adult peripheral blood, which argued against increased T-regulatory activity as a mechanism for the decreased Th1 differentiation of CB CD4 T cells. Together, these intrinsic limitations in T-cell activation and Th1 differentiation may impair the ability of T cells in CB and the neonate to respond to allogeneic or infectious challenges.
View details for DOI 10.1016/j.bbmt.2005.10.027
View details for Web of Science ID 000235284900005
View details for PubMedID 16443514
Chemotactic responses of IL-4-, IL-10-, and IFN-gamma-producing CD4(+) T cells depend on tissue origin and microbial stimulus
JOURNAL OF IMMUNOLOGY
2006; 176 (1): 557-566
Th1- and Th2-polarized immune responses are crucial in the defense against pathogens but can also promote autoimmunity and allergy. The chemokine receptors CXCR3 and CCR4 have been implicated in differential trafficking of IFN-gamma- and IL-4-producing T cells, respectively, but also in tissue and inflammation-specific homing independent of cytokine responses. Here, we tested whether CD4+ T cells isolated from murine tissues under homeostatic or inflammatory conditions exhibit restricted patterns of chemotactic responses that correlate with their production of IFN-gamma, IL-4, or IL-10. In uninfected mice, IL-4-producing T cells preferentially migrated to the CCR4 ligand, CCL17, whereas IFN-gamma-expressing T cells as well as populations of IL-4+ or IL-10+ T cells migrated to the CXCR3 ligand, CXCL9. All cytokine-producing T cell subsets strongly migrated to the CXCR4 ligand, CXCL12. We assessed chemotaxis of T cells isolated from mice infected with influenza A virus or the nematode Nippostrongylus brasiliensis, which induce a strong Th1 or Th2 response in the lung, respectively. Unexpectedly, the chemotactic responses of IL-4+ T cells and T cells expressing the immunosuppressive cytokine IL-10 were influenced not only by the strongly Th1- or Th2-polarized environments but also by their anatomical localization, i.e., lung or spleen. In contrast, IFN-gamma+ T cells exhibited robust chemotaxis toward CXCL9 and had the most consistent migration pattern in both infection models. The results support a model in which the trafficking responses of many effector and regulatory T cells are regulated as a function of the infectious and tissue environments.
View details for Web of Science ID 000234262600064
View details for PubMedID 16365450
Avian flu to human influenza
ANNUAL REVIEW OF MEDICINE
2006; 57: 139-154
Influenza A viral infection causes substantial annual morbidity and mortality worldwide, particularly for infants, the elderly, and the immunocompromised. The virus mainly replicates in the respiratory tract and is spread by respiratory secretions. A growing concern is the recent identification of H5N1 strains of avian influenza A in Asia that were previously thought to infect only wild birds and poultry, but have now infected humans, cats, pigs, and other mammals, often with fatal results, in an ongoing outbreak. A human pandemic with H5N1 virus could potentially be catastrophic because most human populations have negligible antibody-mediated immunity to the H5 surface protein and this viral subtype is highly virulent. Whether an H5N1 influenza pandemic will occur is likely to hinge on whether the viral strains involved in the current outbreak acquire additional mutations that facilitate efficient human-to-human transfer of infection. Although there is no historical precedent for an H5N1 avian strain causing widespread human-to-human transmission, some type of influenza A pandemic is very likely in the near future. The possibility of an H5N1 influenza pandemic has highlighted the many current limitations of treatment with antiviral agents and of vaccine production and immunogenicity. Future vaccine strategies that may include more robust induction of T-cell responses, such as cytotoxic T lymphocytes, may provide better protection than is offered by current vaccines, which rely solely or mainly on antibody neutralization of infection.
View details for DOI 10.1146/annurev.med.57.121304.131333
View details for Web of Science ID 000235981900009
View details for PubMedID 16409141
- Transient deficiencies of T-cell-mediated immunity in the neonate HOT TOPICS IN INFECTION AND IMMUNITY IN CHILDREN III 2006; 582: 55-69
Thymic stromal lymphopoietin as a key initiator of allergic airway inflammation in mice
2005; 6 (10): 1047-1053
The cytokine thymic stromal lymphopoietin (TSLP) has been linked to human allergic inflammatory diseases. We show here that TSLP expression was increased in the lungs of mice with antigen-induced asthma, whereas TSLP receptor-deficient mice had considerably attenuated disease. Lung-specific expression of a Tslp transgene induced airway inflammation and hyperreactivity characterized by T helper type 2 cytokines and increased immunoglobulin E. The lungs of Tslp-transgenic mice showed massive infiltration of leukocytes, goblet cell hyperplasia and subepithelial fibrosis. TSLP was capable of activating bone marrow-derived dendritic cells to upregulate costimulatory molecules and produce the T helper type 2 cell-attracting chemokine CCL17. These findings suggest that TSLP is an important factor necessary and sufficient for the initiation of allergic airway inflammation.
View details for DOI 10.1038/ni1247
View details for Web of Science ID 000232027200024
View details for PubMedID 16142237
Systemic inflammation links impaired glucose metabolism to cardiac allograft vasculopathy development
LIPPINCOTT WILLIAMS & WILKINS. 2004: 753-753
View details for Web of Science ID 000224783504063
Antiviral CD8 T cells in the control of primary human cytomegalovirus infection in early childhood
JOURNAL OF INFECTIOUS DISEASES
2004; 189 (9): 1619-1627
Human cytomegalovirus (CMV) establishes persistent infection, with control of replication thought to be mediated by CMV-specific CD8 T cells. Primary CMV infection commonly affects young children and causes prolonged viral shedding in saliva and urine. We investigated whether this virus-host interaction pattern reflects a developmental deficiency of antiviral CD8 T cell-mediated immunity during childhood. CMV-specific CD8 T cell responses in asymptomatic children with active infection were not different from adults with recent or long-term infection in frequency and functional analyses. High urine CMV concentrations were detected, despite these CMV-specific CD8 T cell responses. We conclude that delayed resolution of primary CMV infection in young children is not caused by a deficient CMV-specific CD8 T cell response. Because these healthy children continue to have local CMV replication, we suggest that CD8 T cells may function primarily to prevent symptomatic, disseminated disease.
View details for Web of Science ID 000220951300010
View details for PubMedID 15116298
Cytomegalovirus infectious burden is proportional to cardiac allograft vasculopathy in heart transplant recipients
ELSEVIER SCIENCE INC. 2004: 185A-185A
View details for Web of Science ID 000189388500790
Viral-induced T helper type 1 responses enhance allergic disease by effects on lung dendritic cells
2004; 5 (3): 337-343
It is widely accepted that T helper type 1 (T(H)1) cytokines such as interferon-gamma (IFN-gamma) antagonize allergic diseases mediated by T(H)2 cytokines. The 'hygiene hypothesis' has also proposed that decreased childhood exposure to pathogen-derived T(H)1 cytokines may underlie the recent increased prevalence of asthma, a T(H)2-mediated disease. We show here that influenza A viral infection, which induces large amounts of intrapulmonary IFN-gamma production, unexpectedly enhanced later allergen-specific asthma and promoted dual allergen-specific T(H)1 and T(H)2 responses. Pulmonary dendritic cells obtained from the lung after viral clearance and resolution of acute inflammation conferred enhanced allergic disease and concurrent T(H)1 and T(H)2 immune responses, and these effects were dependent on IFN-gamma secreted during the acute viral infection. Thus, respiratory viral infection and the acute T(H)1 response can positively regulate T(H)2-dependent allergic pulmonary disease in vivo, at least in part, by altering pulmonary dendritic cell function.
View details for DOI 10.1038/ni1041
View details for Web of Science ID 000189344600017
View details for PubMedID 14973436
Persistent and selective deficiency of CD4(+) T cell immunity to cytomegalovirus in immunocompetent young children
JOURNAL OF IMMUNOLOGY
2004; 172 (5): 3260-3267
Healthy young children who acquire CMV have prolonged viral shedding into the urine and saliva, but whether this is attributable to limitations in viral-specific immune responses has not been explored. In this study, we found that otherwise immunocompetent young children after recent primary CMV infection accumulated markedly fewer CMV-specific CD4(+) T cells that produced IFN-gamma than did adults. These differences in CD4(+) T cell function persisted for more than 1 year after viral acquisition, and did not apply to CMV-specific IFN-gamma production by CD8(+) T cells. The IFN-gamma-producing CD4(+) T cells of children or adults that were reactive with CMV Ags were mainly the CCR7(low) cell subset of memory (CD45R0(high)CD45RA(low)) cells. The decreased IFN-gamma response to CMV in children was selective, because their CCR7(low) memory CD4(+) T cells and those of adults produced similar levels of this cytokine after stimulation with staphylococcal enterotoxin B superantigen. CD4(+) T cells from children also had reduced CMV-specific IL-2 and CD154 (CD40 ligand) expression, suggesting an early blockade in the differentiation of viral-specific CD4(+) T cells. Following CMV acquisition, children, but not adults, persistently shed virus in urine, and this was observable for at least 29 mo postinfection. Thus, CD4(+) T cell-mediated immunity to CMV in humans is generated in an age-dependent manner, and may have a substantial role in controlling renal viral replication and urinary shedding.
View details for Web of Science ID 000189186000069
View details for PubMedID 14978134
Asymptomatic cytomegalovirus activation leads to acute rejection in heart transplant recipients despite anti-viral prophylaxis.
WILEY-BLACKWELL. 2004: 453-453
View details for Web of Science ID 000221322501076
Decreased CD154 expression by neonatal CD4(+) T cells is due to limitations in both proximal and distal events of T cell activation
2003; 15 (12): 1461-1472
Neonatal CD4(+) T cells express less CD154 protein and mRNA than adult CD4(+) T cells after activation by calcium ionophore and phorbol ester, but the mechanism for this reduced expression and its relevance to the primary immune response remain unclear. We compared expression of CD154 protein and mRNA and CD154 gene promoter activity by purified naive (CD45RA(high)CD45RO(low)) neonatal and adult CD4(+) T cells after activation by calcium ionophore (ionomycin) and phorbol myristate acetate (PMA) treatment or by engagement of alphabeta TCR-CD3 complex. Substantial and consistent reductions in expression by neonatal cells were found in all cases and were paralleled by decreased CD154-dependent activation of a B cell line. CD69 expression by neonatal CD4(+) T cells after alphabeta TCR-CD3 engagement was also reduced compared to adult cells, which suggested that limitations in activation-induced signaling by neonatal CD4(+) T cells occurred at a point upstream of where the signaling pathways leading to CD154 and CD69 expression diverge. Decreased CD154 expression by neonatal cells after alphabeta TCR-CD3 engagement was paralleled by a lower free intracellular calcium concentration, a key event for CD154 gene transcription. Reduced CD154 promoter activity by neonatal cells persisted when proximal signaling events were bypassed using ionomycin and PMA, suggesting an additional and more distal mechanism for decreased transcription. In contrast, CD154 mRNA stability was similar in neonatal and adult cells after either ionomycin and PMA stimulation or engagement of the alphabeta TCR-CD3 complex. We conclude that decreased CD154 production by neonatal CD4(+) T cells is due to limitations in both proximal and distal activation events, which together ultimately limit CD154 gene transcription.
View details for DOI 10.1093/intimm/dxg145
View details for Web of Science ID 000187225000007
View details for PubMedID 14645155
Toll-like receptors and T-helper-1/T-helper-2 responses
CURRENT OPINION IN INFECTIOUS DISEASES
2003; 16 (3): 199-204
Toll-like receptors (TLRs) are a family of pattern recognition receptors that are activated by specific components of microbes and certain host molecules. They constitute the first line of defense against many pathogens and play a crucial role in the function of the innate immune system. Recently, TLRs were observed to influence the development of adaptive immune responses, presumably by activating antigen-presenting cells. This has important implications for our understanding of how the host tailors its immune response as a function of specific pathogen recognition. The present review discusses the recent studies that demonstrate the role of TLRs in the regulation of adaptive T-helper-1 (Th1) and Th2 responses, and the mechanisms by which the effects are carried out.Most studies have focused on the role of TLRs and components of their signaling pathways in the control of Th1-type immune responses, and on the implications for their use as antimicrobial agents, such as adjuvants in vaccines, or to treat or prevent the Th2-type dominated immune responses seen in allergies. TLR-deficient mice have been described and used to come to these conclusions. Although controversial, there is also evidence that TLRs may be important for Th2-type responses, possibly by augmenting the overall maturity of dendritic cells.A greater understanding of the processes by which TLRs regulate adaptive immunity may yield not only improved ways to treat infectious diseases but also new approaches to the treatment and prevention of allergic and certain autoimmune disorders.
View details for DOI 10.1097/01.qco.0000073767.11390.47
View details for Web of Science ID 000183434100003
View details for PubMedID 12821808
Deficient cytomegalovirus-specific Th1 responses in young children with urine virus shedding
FEDERATION AMER SOC EXP BIOL. 2003: C301-C301
View details for Web of Science ID 000182367001403
Impaired accumulation and function of memory CD4 T cells in human IL-12 receptor beta 1 deficiency
JOURNAL OF IMMUNOLOGY
2003; 170 (1): 597-603
Defects in IL-12 production or IL-12 responsiveness result in a vulnerability to infection with non-viral intracellular organisms, but the immunological mechanisms responsible for this susceptibility remain poorly understood. We present an immunological analysis of a patient with disseminated Salmonella enteritidis and a homozygous splice acceptor mutation in the IL-12Rbeta1-chain gene. This mutation resulted in the absence of IL-12Rbeta1 protein on PBMC and an inability of T cells to specifically bind IL-12 or produce IFN-gamma in response to either IL-12 or IL-23. The accumulation of memory (CD45R0(high)) CD4 T cells that were CCR7(high) (putative central memory cells) was normal or increased for age. Central memory CD4 T cells of the patient and age-matched controls were similar in having a low to undetectable capacity to produce IFN-gamma after polyclonal stimulation. In contrast, the patient had a substantial decrease in the number of CCR7(neg/dull) CD45R0(high) memory CD4 T cells (putative effector memory cells), and these differed from control cells in having a minimal ability to produce IFN-gamma after polyclonal stimulation. Importantly, tetanus toxoid-specific IFN-gamma production by PBMC from the patient was also significantly reduced compared with that in age-matched controls, indicating that signaling via the IL-12Rbeta1-chain is generally necessary for the in vivo accumulation of human memory CD4 T cells with Th1 function. These results are also consistent with a model in which the IL-12Rbeta1 subunit is necessary for the conversion of central memory CD4 T cells into effector memory cells.
View details for Web of Science ID 000180106600075
View details for PubMedID 12496448
Allergy immunotherapy and inhibition of Th2 immune responses: a sufficient strategy?
CURRENT OPINION IN IMMUNOLOGY
2002; 14 (5): 644-651
Th2 immune responses mediated by the secretion of IL-4, IL-5 and IL-13 are key in the pathogenesis of atopic disorders, including allergen-induced asthma, rhinoconjunctivitis and anaphylaxis. Although such responses are downregulated to some degree by conventional specific immunotherapy, this approach is only partially effective and has a substantial risk of adverse effects. Many strategies for immunotherapeutic prophylaxis and for treatment of atopic diseases have been devised on the basis of mouse allergy and autoimmune models, including the downregulation of Th2 responses by the induction of regulatory T cell activity, Th2 to Th1 immune deviation, Th1 crossregulation of Th2 immune responses, anergy and immunosuppressive cytokines. The blockade of events that are not allergen-specific, such as T cell costimulation and downstream events dependent on IgE, cytokines and chemokines, has also been pursued. With the exception of monoclonal antibody therapy for the blockade of IgE effector function, the application of most of these strategies to humans is at an early stage. Whether the inhibition of Th2 responses without concurrent downregulation of Th1 responses will be sufficient for allergic immunotherapy, particularly for atopic dermatitis and asthma, is an important but unresolved issue.
View details for Web of Science ID 000177716100017
View details for PubMedID 12183167
Dynamics of CD4 and CD8 T cell responses to cytomegalovirus in healthy human donors
JOURNAL OF INFECTIOUS DISEASES
2002; 186 (1): 15-22
To study the dynamics of cytomegalovirus (CMV) immunity in healthy immunocompetent hosts, interferon-gamma-producing CD4 and CD8 T cell responses in the presence or absence of CMV antigens were measured from 15 CMV-seropositive donors and 13 CMV-seronegative donors. Cytokine responses in the absence of antigen were significantly higher in CMV-seropositive donors. Also, a disproportionate number of CD69(+) cells isolated ex vivo from CMV-seropositive donors were specific for CMV, suggesting recent reactivation in vivo. To examine changes in cellular responses over time, 10 seropositive donors were tested over a 6-month period. About half of the donors showed significant variability over time, but staphylococcal enterotoxin B responses remained relatively constant. These findings suggest that CMV can present a considerable and recurrent burden to the human immune system. By understanding the normal dynamics of CMV responses over time, it may be possible to better identify aberrant responses to CMV in immunocompromised hosts.
View details for Web of Science ID 000176307800003
View details for PubMedID 12089657
- Simple febrile seizures - Do polymorphisms of the interleukin 1 gene cluster simplify our understanding? ARCHIVES OF PEDIATRICS & ADOLESCENT MEDICINE 2002; 156 (6): 529-530
Toll-like receptor 4 is required for optimal development of Th2 immune responses: Role of dendritic cells
JOURNAL OF IMMUNOLOGY
2002; 168 (9): 4524-4530
LPS potently induces dendritic cell maturation and the production of proinflammatory cytokines, such as IL-12, by activation of Toll-like receptor 4 (TLR4). Since IL-12 is important for the generation and maintenance of Th1 responses and may also inhibit Th2 cell generation from naive CD4 T cell precursors, it has been inferred that TLR4 signaling would have similar effects via the induction of IL-12 secretion. Surprisingly, we found that TLR4-defective mice subjected to sensitization and pulmonary challenge with a protein allergen had reductions in airway inflammation with eosinophils, allergen-specific IgE levels, and Th2 cytokine production, compared with wild-type mice. These reduced responses were attributable, at least in part, to decreased dendritic cell function: Dendritic cells from TLR4-defective mice expressed lower levels of CD86, a costimulatory molecule important for Th2 responses. They also induced less Th2 cytokine production by antigenically naive CD4 T cells in vitro and mediated diminished CD4 T cell Ag-specific pulmonary inflammation in vivo. These results indicate that TLR4 is required for optimal Th2 responses to Ags from nonpathogenic sources and suggest a role for TLR4 ligands, such as LPS derived from commensal bacteria or endogenously derived ligands, in maturation of the innate immune system before pathogen exposure.
View details for Web of Science ID 000175263000036
View details for PubMedID 11970998
A T cell-specific enhancer of the human CD40 ligand gene
JOURNAL OF BIOLOGICAL CHEMISTRY
2002; 277 (9): 7386-7395
We observed that the human CD40 ligand (CD40L) gene 5'-flanking region conferred weak promoter activity in activated CD4 T cells, suggesting that additional regions are required for optimal CD40L gene transcription. We therefore examined a 3'-flanking segment of the CD40L gene, which contained a putative NF-kappaB/Rel cis-element, for its ability to enhance CD40L promoter function. This segment augmented CD40L promoter activity in an orientation-independent manner in CD4 T-lineage cells but not in human B cell or monocyte cell lines. Mapping of CD4 T-lineage cell nuclei identified a DNase I-hypersensitive site in the flanking region near the NF-kappaB/Rel sequence, suggesting a transcriptional regulatory role. This was further supported by truncation analysis and site-directed mutagenesis, which indicated that the CD40L 3'-flanking NF-kappaB/Rel cis-element was critical for enhancer function. Electrophoretic mobility shift assays showed that the cis-element preferentially bound the p50 form of the NF-kappaB1 gene contained in human T cell nuclear protein extracts. This binding also appeared to occur in vivo in CD4 T cells based on chromatin immunoprecipitation assays using NF-kappaB p50-specific antiserum. Together, these results suggest that the CD40L gene 3'-flanking region acts as a T cell-specific classical transcriptional enhancer by a NF-kappaB p50-dependent mechanism.
View details for DOI 10.1074/jbc.M110350200
View details for Web of Science ID 000174104300083
View details for PubMedID 11751888
Group A streptococcal meningitis: Report of a case and review of literature since 1976
PEDIATRIC EMERGENCY CARE
2001; 17 (6): 430-434
Group A streptococcal (GAS) invasive disease has become increasingly common in recent years. However, acute bacterial meningitis caused by this pathogen is unusual. We report a case of GAS meningitis in a previously healthy 21/2-year-old child associated with a dramatically rapid course and fatal outcome. A literature review of previously reported cases is presented. This case serves as a reminder that GAS can cause severe meningitis in otherwise healthy hosts.
View details for Web of Science ID 000173001500007
View details for PubMedID 11753187
Octamer proteins inhibit IL-4 gene transcription in normal human CD4 T cells
GENES AND IMMUNITY
2001; 2 (8): 464-468
The balance of Th1 (eg, interleukin-2 (IL-2)) and Th2 (eg, IL-4) cytokines produced by CD4 T cells markedly influences the outcome of the adaptive immune response. Although octamer transcription factor proteins increase IL-2 transcription in T cells, their role in IL-4 gene transcription remains controversial. We have previously shown and now confirm that the proximal octamer binding site of the human IL-4 promoter, which separates the two most proximal NFAT binding sites, is bound prior to, but not after, activation in vivo. Since these two NFAT sites are essential for optimal IL-4 promoter activity, this suggested that prior engagement by octamer proteins might prevent adjacent NFAT binding and inhibit IL-4 gene transcription. In support of this hypothesis, here we show that NFAT proteins are unable to bind to a combined octamer/NFAT site unless the octamer proteins are competed away. Moreover, activity of an IL-4 reporter gene mutated in the proximal octamer binding site is increased compared to the wild-type promoter in human peripheral blood CD4 T cells. In addition, over-expression of either Oct-1 or Oct-2 decreased wild-type IL-4 promoter activity, while increasing IL-2 promoter activity. No decrease in promoter activity was seen when Oct-1 or Oct-2 was over-expressed with the octamer-mutant IL-4 reporter gene. Thus, octamer proteins are candidates to promote a Th1 rather than Th2 pattern of cytokine gene expression by activated CD4 T cells.
View details for Web of Science ID 000172443000008
View details for PubMedID 11781715
Respiratory infection with influenza A virus interferes with the induction of tolerance to aeroallergens
JOURNAL OF IMMUNOLOGY
2000; 165 (6): 3484-3491
Viral respiratory infections have been implicated in influencing allergen sensitization and the development of asthma, but their exact role remains controversial. Because respiratory exposure to Ag normally engenders T cell tolerance and prevents the development of airway hyperreactivity (AHR) and inflammation, we examined the effects of influenza A virus infection on tolerance induced by exposure to intranasal (i.n.) OVA and the subsequent development of AHR. We found that concurrent infection with influenza A abrogated tolerance induced by exposure to i.n. OVA, and instead led to the development of AHR accompanied by the production of OVA-specific IgE, IL-4, IL-5, IL-13, and IFN-gamma. When both IL-4 and IL-5 were neutralized in this system, AHR was still induced, suggesting that influenza-induced cytokines such as IL-13, or mechanisms unrelated to cytokines, might be responsible for the development of AHR. The length of time between influenza A infection and i.n. exposure to OVA was crucial, because mice exposed to i.n. OVA 15-30 days after viral inoculation developed neither AHR nor OVA-specific tolerance. These mice instead acquired Th1-biased OVA-specific immune responses associated with vigorous OVA-induced T cell proliferation, and reduced production of OVA-specific IgE. The protective effect of influenza A on AHR was dependent on IFN-gamma, because protection was abrogated with a neutralizing anti-IFN-gamma mAb. These results suggest that viral respiratory infection interferes with the development of respiratory allergen-induced tolerance, and that the time interval between viral infection and allergen exposure is critical in determining whether viral infection will enhance, or protect against, the development of respiratory allergen sensitization and AHR.
View details for Web of Science ID 000165938100073
View details for PubMedID 10975869
Local blockade of allergic airway hyperreactivity and inflammation by the poxvirus-derived pan-CC-chemokine inhibitor vCCI
JOURNAL OF IMMUNOLOGY
2000; 165 (6): 3418-3422
Allergen-induced asthma is characterized by chronic pulmonary inflammation, reversible bronchoconstriction, and airway hyperreactivity to provocative stimuli. Multiple CC-chemokines, which are produced by pulmonary tissue in response to local allergen challenge of asthmatic patients or experimentally sensitized rodents, chemoattract leukocytes from the circulation into the lung parenchyma and airway, and may also modify nonchemotactic function. To determine the therapeutic potential of local intrapulmonary CC-chemokine blockade to modify asthma, a recombinant poxvirus-derived viral CC-chemokine inhibitor protein (vCCI), which binds with high affinity to rodent and human CC-chemokines in vitro and neutralizes their biological activity, was administered by the intranasal route. Administration of vCCI to the respiratory tract resulted in dramatically improved pulmonary physiological function and decreased inflammation of the airway and the lung parenchyma. In contrast, vCCI had no significant effect on the circulating levels of total or allergen-specific IgE, allergen-specific cytokine production by peripheral lymph node T cells, or peritoneal inflammation after local allergen challenge, indicating that vCCI did not alter systemic Ag-specific immunity or chemoattraction at extrapulmonary sites. Together, these findings emphasize the importance of intrapulmonary CC-chemokines in the pathogenesis of asthma, and the therapeutic potential of generic and local CC-chemokine blockade for this and other chronic diseases in which CC-chemokines are locally produced.
View details for Web of Science ID 000165938100065
View details for PubMedID 10975861
An immunosuppressive and anti-inflammatory HLA class I-derived peptide binds vascular cell adhesion molecule-1
2000; 70 (4): 662-667
A synthetic peptide corresponding to residues 75-84 of HLA-B2702 modulates immune responses in rodents and humans both in vitro and in vivo.We used a yeast two-hybrid screening, an in vitro biochemical method, and an in vivo animal model.Two cellular receptors for this novel immunomodulatory peptide were identified using a yeast two-hybrid screen: immunoglobulin binding protein (BiP), a member of the heat shock protein 70 family, and vascular cell adhesion molecule (VCAM)-1. Identification of BiP as a ligand for this peptide confirms earlier biochemical findings, while the interaction with VCAM-1 suggests an alternative mechanism of action. Binding to the B2702 peptide but not to closely related variants was confirmed by ligand Western blot analysis and correlated with immunomodulatory activity of each peptide. In mice, an ovalbumin-induced allergic pulmonary response was blocked by in vivo administration of either the B2702 peptide or anti-VLA-4 antibody.We propose that the immunomodulatory effect of the B2702 peptide is caused, in part, by binding to VCAM-1, which then prevents the normal interaction of VCAM-1 with VLA-4.
View details for Web of Science ID 000088986100021
View details for PubMedID 10972226
NFAT1 enhances HIV-1 gene expression in primary human CD4 T cells
2000; 94 (3): 179-191
Cyclosporin A (CsA) is a potent inhibitor of the NFAT family of transcription factors that enhance T cell activation. The observation that human immunodeficiency virus type 1 (HIV-1)-positive transplant recipients have a reduced HIV-1 viral burden during treatment with CsA suggested that NFAT may play a direct role in enhancing transcription of the HIV-1 viral genome. Two sets of NFAT binding sites were identified in the HIV-1 long terminal repeat (LTR) promoter by in vitro footprinting with full-length recombinant NFAT protein, and gel shift analysis of nuclear protein from polyclonally activated primary CD4 T cells revealed specific binding of NFAT1 to the NFkappaB binding sites of the HIV-1 LTR. Activation of primary CD4 T cells transiently transfected with a HIV-1 LTR luciferase reporter plasmid, lacking the NFAT binding sites in the upstream putative negative regulatory element but maintaining the NFkappaB/NFAT sites, demonstrated increased HIV-1 gene expression when cotransfected with a NFAT1 expression vector. Moreover, CsA, FK506, and a dominant-negative NFAT1 protein independently inhibited HIV-1 LTR promoter activity in CD4 T cells stimulated with phorbol ester and calcium ionophore. In primary human CD4 T cells, CsA also inhibited promoter activity directed by multimers of binding sites for NFAT, while having no effect on NFkappaB multimer-driven promoter activity. Increasing NFAT1 levels in CD4 T cells transiently transfected with a HIV-1 provirus also increased p24 protein expression. Thus, NFAT may be a target for prevention of HIV-1 LTR-directed gene expression in human CD4 T cells.
View details for Web of Science ID 000085927300004
View details for PubMedID 10692237
Distinct types of diffuse large B-cell lymphoma identified by gene expression profiling
2000; 403 (6769): 503-511
Diffuse large B-cell lymphoma (DLBCL), the most common subtype of non-Hodgkin's lymphoma, is clinically heterogeneous: 40% of patients respond well to current therapy and have prolonged survival, whereas the remainder succumb to the disease. We proposed that this variability in natural history reflects unrecognized molecular heterogeneity in the tumours. Using DNA microarrays, we have conducted a systematic characterization of gene expression in B-cell malignancies. Here we show that there is diversity in gene expression among the tumours of DLBCL patients, apparently reflecting the variation in tumour proliferation rate, host response and differentiation state of the tumour. We identified two molecularly distinct forms of DLBCL which had gene expression patterns indicative of different stages of B-cell differentiation. One type expressed genes characteristic of germinal centre B cells ('germinal centre B-like DLBCL'); the second type expressed genes normally induced during in vitro activation of peripheral blood B cells ('activated B-like DLBCL'). Patients with germinal centre B-like DLBCL had a significantly better overall survival than those with activated B-like DLBCL. The molecular classification of tumours on the basis of gene expression can thus identify previously undetected and clinically significant subtypes of cancer.
View details for Web of Science ID 000085227300039
View details for PubMedID 10676951
Bone marrow NK1.1(-) and NK1.1(+) T cells reciprocally regulate acute graft versus host disease
JOURNAL OF EXPERIMENTAL MEDICINE
1999; 189 (7): 1073-1081
Sorted CD4(+) and CD8(+) T cells from the peripheral blood or bone marrow of donor C57BL/6 (H-2(b)) mice were tested for their capacity to induce graft-versus-host disease (GVHD) by injecting the cells, along with stringently T cell-depleted donor marrow cells, into lethally irradiated BALB/c (H-2(d)) host mice. The peripheral blood T cells were at least 30 times more potent than the marrow T cells in inducing lethal GVHD. As NK1.1(+) T cells represented <1% of all T cells in the blood and approximately 30% of T cells in the marrow, the capacity of sorted marrow NK1.1(-) CD4(+) and CD8(+) T cells to induce GVHD was tested. The latter cells had markedly increased potency, and adding back marrow NK1.1(+) T cells suppressed GVHD. The marrow NK1.1(+) T cells secreted high levels of both interferon gamma (IFN-gamma) and interleukin 4 (IL-4), and the NK1.1(-) T cells secreted high levels of IFN-gamma with little IL-4. Marrow NK1.1(+) T cells obtained from IL-4(-/-) rather than wild-type C57BL/6 donors not only failed to prevent GVHD but actually increased its severity. Together, these results demonstrate that GVHD is reciprocally regulated by the NK1.1(-) and NK1.1(+) T cell subsets via their differential production of cytokines.
View details for Web of Science ID 000079648700008
View details for PubMedID 10190898
T cell priming enhances IL-4 gene expression by increasing nuclear factor of activated T cells
JOURNAL OF IMMUNOLOGY
1999; 162 (2): 860-870
The repetitive activation of T cells (priming) enhances the expression of many cytokines, such as IL-4, but not others, such as IL-2. Molecular mechanisms underlying selective expression of cytokines by T cells remain poorly understood. Here we show that priming of CD4 T cells selectively enhances IL-4 expression relative to IL-2 expression by a transcriptional mechanism involving nuclear factor of activated T cells (NFAT) proteins. As detected by in vivo footprinting, priming markedly increases the activation-dependent engagement of the P0 and P1 NFAT-binding elements of the IL-4 promoter. Moreover, each proximal P element is essential for optimal IL-4 promoter activity. Activated primed CD4 T cells contain more NFAT1 and support greater NFAT-directed transcription than unprimed CD4 T cells, while activator protein 1 binding and activator protein 1-mediated transcription by both cell types is similar. Increased expression of wild-type NFAT1 substantially increases IL-4 promoter activity in unprimed CD4 T cells, suggesting NFAT1 may be limiting for IL-4 gene expression in this cell type. Furthermore, a truncated form of NFAT1 acts as a dominant-negative, reducing IL-4 promoter activity in primed CD4 T cells and confirming the importance of endogenous NFAT to increased IL-4 gene expression by effector T cells. NFAT1 appears to be the major NFAT family member responsible for the initial increased expression of IL-4 by primed CD4 T cells.
View details for Web of Science ID 000077951400031
View details for PubMedID 9916709
Diminished expression of CD40 ligand may contribute to the defective humoral immunity in patients with MHC class II deficiency
EUROPEAN JOURNAL OF IMMUNOLOGY
1998; 28 (2): 589-598
Major histocompatibility complex (MHC) class II deficiency (bare lymphocyte syndrome, BLS) is a rare primary immunodeficiency classified as a subgroup of severe combined immunodeficiency. We studied T and B lymphocyte function by examining the CD40 ligand/CD40 system in three BLS patients from two unrelated families. CD40 ligand expression by maximally activated BLS T cells was diminished. This abnormality may represent immunological naïveté rather than a general T cell defect, since expression of activation marker CD69 and proliferative responses to PHA or anti-CD3 were normal, and BLS T cells primed and restimulated in vitro expressed normal amounts of CD40 ligand. BLS B cells proliferated and produced IgE if stimulated with anti-CD40 or soluble CD40 ligand and IL-4. Activation of BLS B cells with soluble CD40 ligand and IL-4 induced normal expression of activation markers, although MHC class II expression remained absent. Depressed antibody titers, lack of amplification and failure to undergo isotype switching in response to immunization with bacteriophage phi x 174 demonstrated defective T cell help. We conclude that BLS B cells are functionally normal if appropriately stimulated, and that the defective humoral immunity observed may be related to diminished expression of CD40 ligand on BLS T cells.
View details for Web of Science ID 000072307700023
View details for PubMedID 9521069
Consistent transient transfection of DNA into non-transformed human and murine T-lymphocytes
JOURNAL OF IMMUNOLOGICAL METHODS
1997; 205 (2): 145-150
The ability to analyze transcriptional regulation in non-transformed T-cells has been hampered by the inability to reproducibly transiently transfect these cells with DNA constructs. We have previously demonstrated that normal human whole mononuclear and CD4 T-cells can be consistently transiently transfected with plasmid DNA. Human cells were most receptive to plasmid DNA uptake between 19.5 and 20 h after prestimulation with a submitogenic dose of the polyclonal T-cell activator, PHA. Here we report an alteration and optimization of this protocol for non-transformed murine splenic T-cells, using concanavalin A instead of PHA as the preactivation stimulus. When coupled with the high sensitivity of luciferase reporter gene constructs, this protocol facilitates the analysis of a variety of T-cell-specific promoters in non-transformed T-cells. In addition, we directly demonstrate that murine T-cells are specifically transiently transfected among a population of whole mononuclear cells by using an expression vector for green fluorescent protein.
View details for Web of Science ID A1997XR21700005
View details for PubMedID 9294595
Influence of the route of allergen administration and genetic background on the murine allergic pulmonary response
AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE
1997; 155 (2): 661-669
We used various ovalbumin sensitization and challenge protocols to determine the importance of the route of allergen administration and the genetic background in modulating the physiologic, inflammatory, and immunologic features characteristic of allergen-induced asthma. In BALB/c mice, induction of maximal airway hyperresponsiveness and airspace eosinophilia required administration of ovalbumin by both the intraperitoneal and the intranasal routes (combination protocol), whereas intraperitoneal immunization alone resulted in maximal ovalbumin-specific IgE plasma levels. Thus, a systemic immune response to allergen, in addition to, or independent of IgE production, as well as local allergen challenge were necessary for maximal induction of pulmonary disease. BALB/c mice treated with ovalbumin by the combination protocol had increased Th2-type cytokine mRNA levels in bronchial lymph node tissue compared with control mice. In contrast, C57BL/6 mice treated with ovalbumin by the combination protocol had significantly decreased responses compared with BALB/c mice for all parameters of allergic pulmonary disease examined, with the exception of airspace eosinophilia. Genetic background has a striking and selective effect on the phenotype of murine allergic pulmonary disease. Further analysis of this murine model should be useful in helping define the critical pathogenetic events in allergen-induced asthma.
View details for Web of Science ID A1997WH87100041
View details for PubMedID 9032210
The importance of leukotrienes in airway inflammation in a mouse model of asthma
JOURNAL OF EXPERIMENTAL MEDICINE
1996; 184 (4): 1483-1494
Inhalation of antigen in immunized mice induces an infiltration of eosinophils into the airways and increased bronchial hyperreactivity as are observed in human asthma. We employed a model of late-phase allergic pulmonary inflammation in mice to address the role of leukotrienes (LT) in mediating airway eosinophilia and hyperreactivity to methacholine. Allergen intranasal challenge in OVA-sensitized mice induced LTB4 and LTC4 release into the airspace, widespread mucus occlusion of the airways, leukocytic infiltration of the airway tissue and broncho-alveolar lavage fluid that was predominantly eosinophils, and bronchial hyperreactivity to methacholine. Specific inhibitors of 5-lipoxygenase and 5-lipoxygenase-activating protein (FLAP) blocked airway mucus release and infiltration by eosinophils indicating a key role for leukotrienes in these features of allergic pulmonary inflammation. The role of leukotrienes or eosinophils in mediating airway hyperresponsiveness to aeroallergen could not be established, however, in this murine model.
View details for Web of Science ID A1996VP23000030
View details for PubMedID 8879219
beta 2-microglobulin-dependent T cells are dispensable for allergen-induced T helper 2 responses
JOURNAL OF EXPERIMENTAL MEDICINE
1996; 184 (4): 1507-1512
CD4+ and CD8+ alpha/beta+ T cells of the T helper cell (Th)2 phenotype produce the cytokines IL-4, IL-5, and IL-13 that promote IgE production and eosinophilic inflammation. IL-4 may play an important role in mediating the differentiation of antigenically naive alpha/beta+ T cells into Th2 cells. Murine NK1.1+ (CD4+ or CD4-CD8-) alpha/beta+ T cells comprise a beta 2-microglobulin (beta 2m)-dependent cell population that rapidly produces IL-4 after cell activation in vitro and in vivo and has been proposed as a source of IL-4 for Th2 cell differentiation. alpha/beta+ CD8+ T cells, most of which require beta 2m for their development, have also been proposed as positive regulators of allergen-induced Th2 responses. We tested whether beta 2m-dependent T cells were essential for Th2 cell-mediated allergic reactions by treating wild-type, beta 2m-deficient (beta 2m -/-), and IL-4-deficient (IL-4 -/-) mice of the C57BL/6 genetic background with ovalbumin (OVA), using a protocol that induces robust allergic pulmonary disease in wild-type mice. OVA-treated beta 2m -/- mice had circulating levels of total and OVA-specific IgE, pulmonary eosinophilia, and expression of IL-4, IL-5, and IL-13 mRNA in bronchial lymph node tissue similar to that of OVA-treated wild-type mice. In contrast, these responses in OVA-treated IL-4 -/- mice were all either undetectable or markedly reduced compared with wild-type mice, confirming that IL-4 was required in this allergic model. These results indicate that the NK1.1+ alpha/beta+ T cell population, as well as other beta 2m-dependent populations, such as most peripheral alpha/beta+ CD8+ T cells, are dispensable for the Th2 pulmonary response to protein allergens.
View details for Web of Science ID A1996VP23000032
View details for PubMedID 8879221
Coexpression of the interleukin-13 and interleukin-4 genes correlates with their physical linkage in the cytokine gene cluster on human chromosome 5q23-31
1996; 87 (8): 3316-3326
Interleukin-13 (IL-13) and IL-4 are cytokines produced by T cells that are encoded by the q23-31 region of human chromosome 5. To investigate the regulation of IL-13 gene expression by T cells, we isolated and sequenced the human IL-13 gene, analyzed its 5'-flanking region for potential transcriptional activation elements, and examined its expression in nontransformed T-lineage cell populations. The human IL-13 gene was located 12.5-kb upstream of the IL-4 gene and 2-kb downstream of a CpG island. The IL-13 gene 5' flank region included a segment with sequence homology to P elements of the IL-4 promoter involved in transcriptional activation in T cells. Mutation of the IL-13 P element site significantly reduced IL-13 promoter activity in response to T-cell activation. Oligonucleotides containing the IL-13 or IL-4 P element sites specifically bound the transcriptional activator protein, nuclear factor-activated T cells, preformed (NF-ATp), when incubated with nuclear protein extracts from activated T cells. Similar to IL-4, IL-13 mRNA expression was highest in T-cell populations enriched for cells that had previously been primed in vivo or in vitro, indicating that priming increases the expression of the IL-13 and IL-4 genes in a coordinate manner. Because the primed T cells contain higher levels of nuclear NF-ATp, capable of binding to P elements of the IL-4 and IL-13 promoters, than do freshly-isolated T cells, the NF-AT-binding P elements are attractive candidates to mediate the coordinate expression of these two cytokine genes.
View details for Web of Science ID A1996UF38100030
View details for PubMedID 8605348
THE HUMAN GP39 PROMOTER - 2 DISTINCT NUCLEAR FACTORS OF ACTIVATED T-CELL PROTEIN-BINDING ELEMENTS CONTRIBUTE INDEPENDENTLY TO TRANSCRIPTIONAL ACTIVATION
JOURNAL OF BIOLOGICAL CHEMISTRY
1995; 270 (50): 29624-29627
gp39, a cytokine expressed on the surface of activated T cells, is essential for T cell-dependent antibody responses in vivo. We cloned and sequenced 1.2 kilobases of the 5' flank region of the human gp39 gene promoter and determined its transcription start site. When used in reporter gene assays, this DNA segment conferred promoter activity in response to T cell activation. gp39 promoter function in transfectants was inhibited by cyclosporin A, as is expression of the endogenous gp39 gene in T-lineage cells. At least 0.5 kilobase of the 5' flank region was required for promoter activity. Two putative binding sites for the NF-AT family of transcriptional activator proteins were identified at -259 to -265 and -62 to -69 with respect to the transcription start site. Both sites contributed significantly and independently to promoter activity in response to T cell activation. Additionally, when incubated in vitro with nuclear protein purified from activated human CD4 T cells, both of these sites preferentially bound the NF-AT family member, NF-ATp. These results suggest that NF-ATp, via binding to at least two cis-elements, is essential for the induction of gp39 gene expression in response to T cell activation.
View details for Web of Science ID A1995TK38000002
View details for PubMedID 8530342
HYPOMETHYLATION OF THE INTERFERON-GAMMA GENE CORRELATES WITH ITS EXPRESSION BY PRIMARY T-LINEAGE CELLS
EUROPEAN JOURNAL OF IMMUNOLOGY
1995; 25 (2): 426-430
To determine the potential role of methylation in the regulation of interferon-gamma (IFN-gamma) gene transcription by T cells, primary T-lineage cell populations were analyzed for the extent of methylation of three CpG sites within or near transcriptional activator elements in the 5' flank and first intron of the human IFN-gamma gene. A striking correlation was observed between the capacity of the IFN-gamma gene to be expressed and the degree of hypomethylation. The IFN-gamma gene was virtually completely methylated at all sites in thymocytes, neonatal T cells, and adult CD45RAhiCD45R0lo (antigenically naive) CD4 T cells, cell types that all have a low or undetectable capacity to express the IFN-gamma gene. In contrast, there was substantial hypomethylation in T-lineage cell types with relatively high capacities to express the IFN-gamma gene, including adult CD8 T cells and adult CD45RAloCD45R0hi (memory/effector) CD4 T cells. These results suggest that hypomethylation of the IFN-gamma genetic locus may be an important determinant of IFN-gamma gene expression in vivo by T-lineage cells.
View details for Web of Science ID A1995QY50600017
View details for PubMedID 7875204
DIMINISHED EXPRESSION OF CD40 LIGAND BY ACTIVATED NEONATAL T-CELLS
JOURNAL OF CLINICAL INVESTIGATION
1995; 95 (1): 66-75
CD40 and CD40 ligand (gp39) mediate contact-dependent T-B cell interaction. We determined the expression of CD40 ligand by activated neonatal T cells and the response of neonatal B cells when activated through CD40. Although expression of CD40 ligand peaked simultaneously in both activated adult and neonatal cells, neonatal T cells expressed significantly less CD40 ligand surface protein and mRNA than adult T cells. Activated thymocytes also expressed far less CD40 ligand than adult T cells. Consistent with these results, activated neonatal T cells exhibited less helper function than activated adult T cells. Neonatal T cells primed and restimulated in vitro expressed CD40 ligand in amounts comparable with adult T cells and provided B cell help more effectively. This suggests that the poor expression of CD40 ligand reflects antigenic naiveté rather than an intrinsic defect of neonatal T cells. Neonatal B cells cultured with soluble CD40 ligand (sgp39) and IL-10 produced IgM in amounts comparable with adult cells, but much less IgG and IgA. Nevertheless, neonatal B cells were capable of proliferation and class switching, since sgp39 and IL-4 induced proliferation and IgE production comparable to adult B cells and production of modest amounts of IgG. Together, these results indicate that diminished CD40 ligand expression, along with decreased production of lymphokines, may be responsible, at least in part, for the transient immunodeficiency observed in human neonates.
View details for Web of Science ID A1995QA88200012
View details for PubMedID 7814647
DIFFERENTIATION OF THE T-HELPER PHENOTYPES BY ANALYSIS OF THE METHYLATION STATE OF THE IFN-GAMMA GENE
JOURNAL OF IMMUNOLOGY
1994; 153 (8): 3603-3610
Th1 and Th2 CD4+ T cell clones have been defined by their ability to produce different lymphokines. However, the processes by which CD4+ T cells differentially regulate lymphokine gene expression have not been well defined. In this report, we demonstrate that the methylation status of a CpG dinucleotide contained within a TATA proximal regulatory element of the IFN-gamma promoter correlates with the transcription of the gene. In murine Th1 clones and two human CD4+ Th0 clones, this site is either completely or partially hypomethylated, whereas in murine Th2 clones this site is > 98% methylated. Treatment of murine Th2 clones with 5-azacytidine, an agent that inhibits methylation of the DNA, converts these cells to IFN-gamma producers. Additional targets for methylation outside the transcriptional control regions of the IFN-gamma genetic locus were found to be hypomethylated in Th2 cells but not in Th1 cells. Electrophoretic mobility shift assays (EMSA) revealed at least five distinct protein-DNA complexes that are formed with an oligonucleotide containing the IFN-gamma promoter TATA proximal regulatory element, and in vitro methylation of this site results in a loss of these three complexes. Furthermore, a comparison of nuclear extracts prepared from Th1 and Th2 clones revealed that the EMSA patterns were qualitatively similar but differed quantitatively. In addition, transient transfection of a murine IFN-gamma promoter-chloramphenicol acetyl transferase (CAT) gene construct into both Th1 and Th2 clones produced CAT activity that was not inducible by anti-CD3, indicating that hypomethylation per se of the promoter alone is not sufficient for inducible gene expression.
View details for Web of Science ID A1994PM58100026
View details for PubMedID 7523497
OSTEOPOROSIS INDUCED IN MICE BY OVERPRODUCTION OF INTERLEUKIN-4
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
1993; 90 (24): 11618-11622
Osteoporosis is a common disease in which loss of bone mass results in skeletal fragility. The development of therapies for this disorder has been hampered by the lack of a convenient animal model. Here we describe a disorder in bone homeostasis in transgenic mice that inappropriately express the cytokine interleukin 4 (IL-4) under the direction of the lymphocyte-specific proximal promoter for the lck gene. Bone disease in lck-IL-4 mice appeared to result from markedly decreased bone formation by osteoblasts, features strikingly similar to those observed in cases of severe low-turnover human involutional osteoporosis. By 2 months of age, female and male lck-IL-4 mice invariably developed severe osteoporosis of both cortical and trabecular bone. Osteoporosis was observed in two independently derived founder animals, indicating that this phenotype was directly mediated by the IL-4 transgene.
View details for Web of Science ID A1993MM51500038
View details for PubMedID 8265598
LYMPHOKINE REGULATION AND THE ROLE OF ABNORMAL REGULATION IN IMMUNODEFICIENCY
ACADEMIC PRESS INC ELSEVIER SCIENCE. 1993: S25-S32
T cell-derived lymphokines mediate or modulate various aspects of the immune response and immunodeficiency states related to abnormalities in lymphokine production or regulation are now being recognized. One example of this is seen in the fetus and neonate, in whom a physiologic immunodeficiency appears to reflect in part deficient production of certain lymphokines, including interferon-gamma, IL-4, and IL-5. The deficiency in production of these lymphokines appears to reflect to a large extent the paucity of memory T cells during these periods of life. Diminished lymphokine production has also recently been implicated as the cause for three cases of primary severe combined immunodeficiency. In disorders associated with excess IgE production, including allergy, hyper IgE syndrome, and Omenn's syndrome, excess IL-4 production relative to the production of interferon-gamma may play a contributory role. Regulation of the production of these and other T cell-derived lymphokines appears to be affected predominantly by control of lymphokine gene transcription, the basis for which is just now becoming understood at a molecular level. The elucidation of these regulatory mechanisms offers the promise for understanding the basis for disordered lymphokine production in immunodeficiency states.
View details for Web of Science ID A1993LF99900004
View details for PubMedID 8500278
ONTOGENY OF LYMPHOCYTE-T FUNCTION IN THE NEONATE
MUNKSGAARD INT PUBL LTD. 1992: 132-135
T cell precursors are first detected in the thymus at eight weeks of gestation. By 15 to 20 weeks of gestation, T-cell precursors expressing alpha beta and gamma delta T-cell receptors are present in the thymus in numbers relatively similar to those found in postnatal life. However, recent data suggest that T-cell receptor diversity is more limited during fetal and neonatal life than in adults. Additionally, the functional capacity of T cells in the fetus and neonate is immature, in that neonatal T cells express a limited repertoire of lymphokines in response to activation. Specifically, the production of the lymphokines, interferon-gamma and interleukin-4, which participate in the maturation of cytotoxic cells, activation of macrophages, and the maturation and modulation of B cell function and isotype expression, is reduced more than tenfold compared to cells from adults. This appears to result primarily from the lack of memory T cells in the fetus and neonate, reflecting their antigenic naivete. The difference in lymphokine expression is due to diminished transcription of these genes in neonatal T cells in response to activation. Preliminary data indicate that differences in essential promoter elements regulating transcription of these lymphokine genes plays a role in their differential expression in T cells.
View details for Web of Science ID A1992KH72800005
View details for PubMedID 1285862
HAWKINSINURIA IN 2 FAMILIES
AMERICAN JOURNAL OF MEDICAL GENETICS
1992; 44 (1): 52-56
Hawkinsinuria, a disorder of tyrosine metabolism has been documented in two families in the United States, in one of which there was clear evidence of autosomal dominant inheritance. Metabolic acidosis and failure to thrive appear to be confined to infancy. Tyrosyl metabolites and 5-oxoproline are also found only in infancy, while 4-hydroxycyclohexylacetic acid was present only with time. The disease may be detected by organic acid analysis or by staining an electropherogram for sulfur containing compounds.
View details for Web of Science ID A1992JJ63400012
View details for PubMedID 1519651
CLINICAL AND MICROBIOLOGIC CHARACTERISTICS OF CUTANEOUS INFECTION WITH YERSINIA-ENTEROCOLITICA
JOURNAL OF INFECTIOUS DISEASES
1992; 165 (4): 740-743
The clinical and microbiologic features of primary cutaneous infections by Yersinia enterocolitica are described in three children. Vesiculobullous lesions developed in two patients, and an intense granulation response followed incision and drainage. In the third child, cellulitis and abscess formation developed at the site of minor skin trauma. Y. enterocolitica isolates from the patients were extensively serologically, biochemically, and molecularly analyzed and compared with virulent Y. enterocolitica strains. The ability of the isolates to adhere to and invade eukaryotic cells was determined using in vitro assays; virulence was assessed by inoculation of suckling mice. The resulting data suggest that primary cutaneous infections by Y. enterocolitica involve strains that are as virulent as pathogenic gastrointestinal isolates.
View details for Web of Science ID A1992HJ64200023
View details for PubMedID 1552204
DECREASED GRANULOCYTE-MACROPHAGE COLONY-STIMULATING FACTOR PRODUCTION BY HUMAN NEONATAL BLOOD MONONUCLEAR-CELLS AND T-CELLS
1992; 31 (3): 211-216
Impaired production and delivery of neutrophils to the site of infection have been implicated in the increased susceptibility of the neonate to infection. Because granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte colony-stimulating factor (G-CSF) play critical roles in the production of neutrophils from marrow precursors, we assessed the ability of leukocytes from neonates and adults to produce GM-CSF, G-CSF, and, for comparison, macrophage colony-stimulating factor (M-CSF) after stimulation with concanavalin A +/- phorbol myristate acetate [blood mononuclear cells (MC) and T lymphocytes] or lipopolysaccharide (monocytes). MC and monocytes from adult and neonatal subjects produced mRNA for GM-CSF, G-CSF, and M-CSF, whereas T cells produced only GM-CSF mRNA. Neonatal MC and T cells accumulated only approximately 30% as much GM-CSF mRNA as did adult MC and T cells. In contrast, the accumulation of GM-CSF mRNA by neonatal and adult monocytes was similar. Neonatal MC also accumulated similar amounts of G-CSF mRNA and somewhat more M-CSF mRNA than did adult MC; results with monocytes were similar to those with MC. Results of colony-stimulating activity bioassays on supernatants from neonatal and adult MC stimulated with concanavalin A paralleled the mRNA results.(ABSTRACT TRUNCATED AT 250 WORDS)
View details for Web of Science ID A1992HF34600003
View details for PubMedID 1373232
DEVELOPMENTAL REGULATION OF LCK GENE-EXPRESSION IN LYMPHOCYTES-T
JOURNAL OF EXPERIMENTAL MEDICINE
1991; 173 (2): 383-393
In the mouse and human, mRNA transcripts encoding the lymphocyte-specific protein tyrosine kinase p56lck are derived from two separate promoters resulting in heterogeneity in the 5' untranslated region sequence. The proximal promoter lies just 5' to the coding region for the gene and is active only in thymocytes. In contrast, the distal promoter lies 34 kilobases (kb) 5' in the human, and is active both in thymocytes and mature peripheral T cells. As previously reported, transgenic mice bearing functional proximal promoter sequence juxtaposed with the SV40 large T antigen gene invariably develop lymphoid tumors confined to the thymus. In the current work, transgenic mice bearing a 2.6-kb fragment of the human distal promoter fused to the SV40 large T antigen gene express large T antigen in thymocytes and in peripheral lymphoid cells, and develop tumors of both the thymus and the peripheral lymphoid organs. The ability of the human distal promoter to function appropriately in transgenic mice is consistent with the strong similarity observed between the mouse and human distal promoter sequences. With the exception of a single short interval that serves as a target for binding of nuclear factors, significant sequence similarity is not seen when the distal and proximal promoter sequences are compared. Hence, developmentally regulated, lineage-specific transcription of the lck gene is mediated by distinct promoter sequences that appear to be capable of functioning independently.
View details for Web of Science ID A1991EV17500013
View details for PubMedID 1988541
INTERLEUKIN-4 EXPRESSED INSITU SELECTIVELY ALTERS THYMOCYTE DEVELOPMENT
JOURNAL OF EXPERIMENTAL MEDICINE
1991; 173 (1): 89-100
Using a transgenic mouse model we show that increased intrathymic expression of interleukin 4 (IL-4) significantly perturbs the development of thymocytes. Transgenic double-positive (CD4+CD8+) thymocytes, which are present in dramatically reduced numbers, exhibit increased T cell receptor (TCR) expression and increased mobilization of calcium mediated by these receptors. In contrast, transgenic single-positive (CD4+CD8- and CD4-CD8+) thymocytes and peripheral T cells exhibit decreased TCR-mediated calcium mobilization. The development of CD4-CD8+ thymocytes is significantly perturbed by IL-4 expressed in vivo; only peripheral CD4+ T cells are found in significant numbers in transgenic mice, while CD4-CD8+ thymocytes are present in increased numbers, apparently because of their failure to emigrate to the periphery. In contrast to these selective effects on T cell development, no significant differences in the numbers of B cells or mast cells, or in the plasma levels of IgE and IgG1 are observed between transgenic and control mice. These observations suggest that IL-4 in vivo exerts its major effects locally rather than systemically, even when its expression is constitutively increased.
View details for Web of Science ID A1991EP65900010
View details for PubMedID 1824637
T cell development in the fetus and neonate.
Advances in experimental medicine and biology
1991; 310: 17-27
View details for PubMedID 1808993
CELLULAR AND MOLECULAR MECHANISMS FOR REDUCED INTERLEUKIN-4 AND INTERFERON-GAMMA PRODUCTION BY NEONATAL T-CELLS
JOURNAL OF CLINICAL INVESTIGATION
1991; 87 (1): 194-202
The mechanisms by which T lymphocytes acquire the capacity to produce interleukin 4 (IL-4) and other lymphokines during intrathymic and extrathymic development are poorly understood. To gain insight into this process, we determined the capacity of human neonatal and adult T lineage cell populations to produce IL-4 after polyclonal activation. IL-2 and interferon-gamma (IFN-gamma) production were studied in parallel, since their production by neonatal T cells is known to be similar or diminished, respectively, compared to adult T cells. Production of IL-4 by neonatal CD4+ T cells and IFN-gamma by neonatal CD4+ and CD8+ T cells was markedly lower compared with analogous adult cell populations, whereas IL-2 production was similar. Transcription of IL-4, as determined by nuclear run-on assays, and IL-4 mRNA-containing cells, as determined by in situ hybridization, were undetectable in neonatal T cells, whereas both were detectable in adult T cells. IFN-gamma transcription and IFN-gamma mRNA-containing cells were reduced in neonatal T cells compared with adult T cells. Reduced lymphokine production by neonatal T cells correlated with their lack of a CD45R- (putative memory T cell) population; cells with this surface phenotype comprised 30-40% of the adult CD4+ T cells and were highly enriched for IL-4 and IFN-gamma, but not IL-2 production. IL-4, IFN-gamma, and IL-2 mRNA expression by neonatal CD4+CD8- thymocytes was similar to that found in circulating neonatal CD4+ T cells. Taken together, these findings suggest that the extrathymic generation of memory T cells during postnatal life may result in an increased capacity for IL-4 and IFN-gamma gene expression. In addition, IFN-gamma and IL-2 mRNA were significantly more abundant than IL-4 mRNA in activated neonatal CD4+CD8- thymocytes and CD4+ T cells, as well as adult CD4+ CD45R- T cells. Therefore, the capacity of T lineage cells to express the IL-4 gene may be more restricted compared to other lymphokine genes beginning in intrathymic development. This restricted capacity appears to persist during postnatal extrathymic maturation of T cells.
View details for Web of Science ID A1991EQ97600026
View details for PubMedID 1824631
- GAMMA-INTERFERON - AN IMMUNOREGULATORY LYMPHOKINE WITH IMMUNOTHERAPEUTIC POTENTIAL PEDIATRIC INFECTIOUS DISEASE JOURNAL 1990; 9 (9): 642-651
REGULATION OF IMMUNOGLOBULIN (IG)E SYNTHESIS IN THE HYPER-IGE SYNDROME
JOURNAL OF CLINICAL INVESTIGATION
1990; 85 (5): 1666-1671
The hyper-IgE (HIE) syndrome is characterized by high IgE serum levels, chronic dermatitis, and recurrent infections. The mechanisms responsible for hyperproduction of IgE in HIE patients are presently unknown. We investigated whether spontaneous in vitro IgE synthesis by PBMC from seven HIE patients was sensitive to signals (cell adhesion, T/B cell cognate interaction and lymphokines: IL-4, IL-6, and IFN-gamma) known to regulate IgE induction in normals. Our results show that, unlike IL-4 dependent IgE synthesis induced in normals, spontaneous IgE production by PBMC from HIE patients was not blocked by monoclonal antibodies to CD2, CD4, CD3, and MHC class II antigens. Furthermore, antibodies to IL-4 and IL-6 did not significantly suppress IgE production. IFN-gamma had no significant effects on spontaneous in vitro IgE synthesis. To test whether an imbalance in lymphokine production might underlie hyperproduction of IgE in HIE patients, mitogen-induced secretion of IL-4 and IFN-gamma by PBMC was assessed. No significant difference was detected between HIE patients and normal controls. Thus, ongoing IgE synthesis in the HIE syndrome is largely independent of cell-cell interactions and endogenous lymphokines, and is due to a terminally differentiated B cell population, no longer sensitive to regulatory signals.
View details for Web of Science ID A1990DB95800041
View details for PubMedID 2110192
BASIS AND IMPLICATIONS OF SELECTIVELY DIMINISHED CYTOKINE PRODUCTION IN NEONATAL SUSCEPTIBILITY TO INFECTION
UNIV CHICAGO PRESS. 1990: S410-S420
The human neonate is unduly susceptible to infection with viruses and other pathogens, such as Toxoplasma and Listeria, that survive and replicate within cells. Cellular immunity is the major mechanism of host defense against these intracellular pathogens. Selective immaturity in certain functions of T lymphocytes appears to be a major factor in the neonate's susceptibility to these infections. Particularly striking is the deficiency in production of interferon-gamma. We review the data regarding the deficiency in the production of interferon-gamma by cells of healthy and infected neonates, discuss what is known regarding the cellular and molecular mechanisms for this deficiency, and review the unique role played by interferon-gamma in host defense against intracellular pathogens. The response of the neonate's effector cells to the immunoenhancing effects of interferon-gamma appears to be variable; diminished enhancement by interferon-gamma of cytotoxic cell function and the production of tumor necrosis factor by macrophages may further compound the effects of diminished production of interferon-gamma.
View details for Web of Science ID A1990DG33700005
View details for PubMedID 2114034
LYMPHOCYTE-ACTIVATION PROVOKES MODIFICATION OF A LYMPHOCYTE-SPECIFIC PROTEIN TYROSINE KINASE (P56LCK)
JOURNAL OF IMMUNOLOGY
1989; 142 (7): 2430-2437
The protein tyrosine kinase p56lck is implicated in the control of lymphocyte growth by virtue of its overexpression in some lymphoid malignancies and its transforming activity in heterologous systems. Previous studies have demonstrated that levels of lck mRNA and of p56lck decline rapidly after T cell activation. The disappearance of p56lck results primarily from post-translational conversion of p56lck to more slowly migrating forms with apparent sizes of approximately 60 kDa. This modification can be provoked by treatment of lymphocytes with PMA, and has been associated with increased serine phosphorylation of the p56lck molecule. Here we demonstrate that conversion of p56lck to p60lck is a feature of the physiologic activation of T lymphocytes by antigen-presenting cells. In addition, we show that the PMA-induced modification of p56lck proceeds via a mechanism distinct from conventional protein kinase C activation. The rapid conversion of p56lck to p60lck after antigenic stimulation is consistent with the view that this membrane-associated protein tyrosine kinase regulates some aspects of the lymphocyte activation sequence.
View details for Web of Science ID A1989T802000037
View details for PubMedID 2784463
RESTRICTED PRODUCTION OF INTERLEUKIN-4 BY ACTIVATED HUMAN T-CELLS
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
1988; 85 (24): 9743-9747
Interleukin 4 (IL-4) is secreted by activated T cells and pleiotropically modulates both B- and T-lymphocyte function. In murine helper (CD4+) T-cell clones IL-4 production appears to be regulated independently of interferon gamma and interleukin 2. To determine whether production of these lymphokines is also differentially regulated in uncloned human T cells, we studied lymphokine production by normal human peripheral T cells and T-cell subsets after in vitro polyclonal activation. After maximal induction of lymphokine expression, IL-4 mRNA was detectable in less than 5% of CD4+ and 1-2% of unfractionated T cells, whereas approximately 33% and 60% of CD4+ cells expressed detectable mRNA for interferon gamma and interleukin 2, respectively. This finding correlated with dramatically lower production of IL-4 mRNA and protein than of interferon gamma and interleukin 2 by peripheral blood and tonsillar T cells. The helper-inducer (CD4+ CD45R-) T-cell subset, which significantly enhances in vitro immunoglobulin production, accounted for the preponderance of IL-4 mRNA accumulation and protein production by CD4+ T cells; nevertheless, cells with detectable IL-4 mRNA constituted less than 10% of the CD4+ CD45R- subset. Limitation of IL-4 production to a comparatively small population of normal human T cells could selectively regulate the effects of this lymphokine in T-cell-mediated immune responses; such selective regulation may be a fundamental mechanism for restricting the potentially pleiotropic effects of certain lymphokines to appropriate responder cells.
View details for Web of Science ID A1988R518700080
View details for PubMedID 3144002
STRUCTURE AND EXPRESSION OF ICK - TRANSCRIPTS IN HUMAN LYMPHOID-CELLS
JOURNAL OF CELLULAR BIOCHEMISTRY
1988; 38 (2): 117-126
The murine lck gene encodes a membrane-associated protein tyrosine kinase that has been implicated in lymphocyte oncogenesis. Here we report the structure of normal human lck transcripts and the pattern of expression of these transcripts in developing thymus and in peripheral T cell subsets. The human lck gene encodes a 509 amino acid polypeptide that is closely related to the murine lck-encoded protein throughout its length. Analysis of the deduced amino acid sequence of human p56lck demonstrates that an amino-terminal domain, widely divergent among the seven known src family members, has been conserved between murine and human p56lck, and thus probably includes sequences crucial to the lymphocyte-specific function of this molecule. Human lck transcripts were detected in CD4+ and CD8+ T cells, in partially purified B cells, and in Epstein-Barr virus-immortalized B cell lines, but not in monocytes, granulocytes, or in nonhematopoetic cell types. Human lck transcripts are readily detectable in fetal thymocytes at 70 days of gestation, but not at 57 days of gestation, indicating that lck expression appears coordinately with the appearance of lymphoid cells in the developing thymus. Thus lck gene expression is a marker for cells of the lymphocyte lineage in man. We conclude that the lck gene probably participates in a signal transduction pathway uniquely present in lymphoid cells.
View details for Web of Science ID A1988Q654800005
View details for PubMedID 3265417
REGULATION OF PP56LCK DURING T-CELL ACTIVATION - FUNCTIONAL IMPLICATIONS FOR THE SRC-LIKE PROTEIN TYROSINE KINASES
1987; 6 (9): 2727-2734
The lymphocyte-specific protein tyrosine kinase pp56lck, encoded by a member of the src gene family, is implicated in the control of T-cell growth and differentiation. Purified resting human T lymphocytes contain appreciable levels of lck mRNA and of pp56lck. Upon activation of these T cells, levels of lck mRNA and of pp56lck promptly decline. These reductions in lck mRNA and protein expression are closely correlated with the induction of lymphokine production. Both require identical stimuli and follow a similar time course of response. Down-regulation of lck expression, however, is not correlated with proliferation. Our results provide an example of regulation of a src-like protein tyrosine kinase in a normal fully differentiated cell population and suggest that modulation of lck RNA and protein expression is an important feature of the lymphocyte activation sequence leading to lymphokine production.
View details for Web of Science ID A1987J692700030
View details for PubMedID 3119327
NOVEL PROTEIN-TYROSINE KINASE GENE (HCK) PREFERENTIALLY EXPRESSED IN CELLS OF HEMATOPOIETIC ORIGIN
MOLECULAR AND CELLULAR BIOLOGY
1987; 7 (6): 2276-2285
Protein-tyrosine kinases are implicated in the control of cell growth by virtue of their frequent appearance as products of retroviral oncogenes and as components of growth factor receptors. Here we report the characterization of a novel human protein-tyrosine kinase gene (hck) that is primarily expressed in hematopoietic cells, particularly granulocytes. The hck gene encodes a 505-residue polypeptide that is closely related to pp56lck, a lymphocyte-specific protein-tyrosine kinase. The exon breakpoints of the hck gene, partially defined by using murine genomic clones, demonstrate that hck is a member of the src gene family and has been subjected to strong selection pressure during mammalian evolution. High-level expression of hck transcripts in granulocytes is especially provocative since these cells are terminally differentiated and typically survive in vivo for only a few hours. Thus the hck gene, like other members of the src gene family, appears to function primarily in cells with little growth potential.
View details for Web of Science ID A1987H488700030
View details for PubMedID 3453117
REDUCED INTERFERON-GAMMA MESSENGER-RNA LEVELS IN HUMAN NEONATES - EVIDENCE FOR AN INTRINSIC T-CELL DEFICIENCY INDEPENDENT OF OTHER GENES INVOLVED IN T-CELL ACTIVATION
JOURNAL OF EXPERIMENTAL MEDICINE
1986; 163 (4): 1018-1023
IFN-gamma mRNA levels in human neonatal blood mononuclear cells or highly purified T cells were markedly lower than those of adult cells after incubation with Con A and PMA. In contrast, IL-2, IL-2-R, and T3 delta chain mRNA levels were kinetically and quantitatively similar in neonatal and adult T cells. The peak amount of IFN-gamma and IL-2 mRNA correlated well with IFN-gamma and IL-2 detected in supernatants of both neonatal and adult T cells. These results suggest that reduced IFN-gamma mRNA levels in neonatal T cells is due to an intrinsic deficiency at the pretranslational level and indicate that the magnitude of IL-2 and IFN-gamma gene expression can be independently modulated pretranslationally.
View details for Web of Science ID A1986A688300019
View details for PubMedID 3081678
DECREASED PRODUCTION OF INTERFERON-GAMMA BY HUMAN NEONATAL CELLS - INTRINSIC AND REGULATORY DEFICIENCIES
JOURNAL OF CLINICAL INVESTIGATION
1986; 77 (3): 860-867
Human neonatal lymphocytes produced little macrophage activation factor in response to mitogens. This correlated with decreased production of interferon-gamma (IFN gamma): adult lymphokines contained 894.2 +/- 177.1 U/ml, whereas neonatal cord and peripheral lymphokines contained 66.9 +/- 17.0 and 116.7 +/- 29.6 U/ml by bioassay. Results by radioimmunoassay (RIA) for IFN gamma were similar. In contrast, the interleukin 2 content of cord lymphokines was greater (P less than 0.01) and that of neonatal peripheral blood lymphokines similar to that of adults. Interleukin 1 production and interleukin 2 receptor expression and affinity were similar for adult and neonatal cells. Interleukins 1 and 2 in amounts comparable to those in adult lymphokines did not increase production of macrophage activation factor or IFN gamma by neonatal cells. Neonatal cells did not contain intracellular IFN or degrade exogenous IFN. Excess suppressor activity was not found in neonatal cultures. Addition of IFN alpha, 10,000-50,000 U/ml of interleukin 2 or phorbol myristate acetate (PMA) to cord mononuclear cells or of adult monocytes or PMA to cord T cells increased IFN gamma production compared to cells stimulated with concanavalin A (ConA) alone. Nevertheless, under optimal conditions (T cells + PMA + Con A), adult cells produced much more IFN gamma (1,360 +/- 261 U/ml by RIA) than cord cells (122 +/- 37 U/ml). Staphylococcal enterotoxin A (SEA) stimulated cord cell IFN gamma production at low cell densities; nevertheless, adult cells produced more IFN in response to SEA 1,341 +/- 350 U/ml) than cord cells (350 +/- 33 U/ml). Decreased production of IFN gamma by neonatal cells appears to be due both to differences in their intrinsic capacity to produce IFN gamma and to differences in regulatory mechanisms.
View details for Web of Science ID A1986A542900025
View details for PubMedID 3081575