Biomedical subjects

L H Glimcher

Publications and source records attributed to L H Glimcher.

At least 73 records · Page 4Linked to original sources

PubMed1995

The proximal promoter of the IL-4 gene is composed of multiple essential regulatory sites that bind at least two distinct factors.

Immune responses to pathogens often lead to the generation of polarized T helper subsets designated Th1 and Th2. Th1 cells, characterized by the production of IL-2 and IFN-gamma, stimulate cellular immune responses important for protection against intracellular pathogens. In contrast, Th2 cells, which produce IL-4, are potent stimulators of B cells and stimulate protection against extracellular pathogens. IL-4 has also emerged as a key cytokine in T cell differentiation since it has been shown to direct the development of naive T cells toward a Th2 phenotype. Recent studies have provided insights into the transcriptional regulation of IL-4, including the identification of multiple binding sites for a subunit of the IL-2 transcription factor NF-AT. In this study we describe the characterization of an essential region of the IL-4 promoter located immediately upstream of the TATA element. High-resolution mutagenesis of this 33-bp region revealed multiple sites indispensable for inducible IL-4 transcription. Included in this region are overlapping binding sites for the cyclosporin A-sensitive factor NF-ATp and a novel constitutively expressed factor designated PCC. An additional sequence adjacent to the TATA element is also shown to be critical for IL-4 transcription in Th2 cells.

Animals↗

PubMed1995

B7-1 and B7-2 costimulatory molecules activate differentially the Th1/Th2 developmental pathways: application to autoimmune disease therapy.

CD4 T helper precursor cells mature along two alternative pathways, Th1 and Th2. Here we show that these pathways are differentially activated by two costimulatory molecules, B7-1 and B7-2. Using anti-B7 antibodies, this developmental step was manipulated both in vitro and in vivo in experimental allergic encephalomyelitis (EAE). Anti-B7-1 reduced the incidence of disease while anti-B7-2 increased disease severity. Neither antibody affected overall T cell induction but rather altered cytokine profile. Administration of anti-B7-1 at immunization resulted in predominant generation of Th2 clones whose transfer both prevented induction of EAE and abrogated established disease. Since co-treatment with anti-IL-4 antibody prevented disease amelioration, costimulatory molecules may directly affect initial cytokine secretion. Thus, interaction of B7-1 and B7-2 with shared counterreceptors CD28 and CTLA-4 results in very different outcomes in clinical disease by influencing commitment of precursors to a Th1 or Th2 lineage.

Amino Acid Sequence↗

PubMed1995

Coordinate and cooperative roles for NF-AT and AP-1 in the regulation of the murine IL-4 gene.

The transcription factor NF-AT plays an essential role in the inducible transcription of several cytokine genes during T cell activation. The distal NF-AT site of the murine IL-2 promoter binds both NF-AT and AP-1 proteins, and thus represents a composite regulatory site that integrates Ca(2+)- and PKC-dependent signaling pathways in T cell activation. However, the individual contributions of the NF-AT and AP-1 components to promoter activity via this composite site have not been resolved, owing to the absence of a clearly defined AP-1 binding site, which, when mutated abolishes AP-1 binding. We describe here an apparently analogous NF-AT/AP-1 composite site in the murine IL-4 promoter, which can be mutated to selectively block the recruitment of each component. We show that the cooperative and coordinate involvement of both NF-AT and AP-1 is necessary for full activity of the NF-AT/AP-1 composite site, and, ultimately, the entire IL-4 promoter.

Animals↗

PubMed1995

Human MHC class II gene transcription directed by the carboxyl terminus of CIITA, one of the defective genes in type II MHC combined immune deficiency.

Type II major histocompatibility complex combined immune deficiency (type II MHC CID or bare lymphocyte syndrome) is a congenital immunodeficiency disease characterized by absent MHC class II expression. Four distinct complementation groups have been identified. Recently, the defective gene in group II type II MHC CID has been isolated and termed CIITA. Here, we demonstrate that CIITA is an MHC class II gene-specific transcription activator. The transcription activation function is provided by the N-terminal acidic domain (amino acids 26-137), which is experimentally exchangeable with a heterologous viral transcription-activating domain. The specificity of CIITA for three major MHC class II genes, DR, DQ and DP, is mediated by its remaining C-terminal residues (amino acids 317-1130). The transactivation of multiple cis elements, especially S and X2, of the DR alpha proximal promoter in group II CID cells is CIITA dependent. Since CIITA overexpression in normal cells did not increase class II expression, we propose that initiation of CIITA expression serves as the on-off switch, while availability of downstream interactor(s) limits transcription.

B-Lymphocytes↗

PubMed1995

Early murine Lyme carditis has a macrophage predominance and is independent of major histocompatibility complex class II-CD4+ T cell interactions.

To compare the role of macrophages and CD4+ T lymphocytes in early Lyme carditis, immunohistochemical techniques were used to analyze cardiac infiltrates in immunocompetent mice infected with Borrelia burgdorferi spirochetes. Macrophages predominated in the infiltrate during the first 4 weeks after infection. CD4+ and CD8+ lymphocytes each constituted < 5% of the infiltrate; B lymphocytes were rare. Infected mice deficient in class II major histocompatibility complex (MHC) antigen and depleted of CD4+ lymphocytes developed similar infiltrates, suggesting that class II MHC-CD4+ lymphocyte interactions do not play a critical role in disease initiation. Expression of mRNA encoding JE within areas of cardiac inflammation implicates this chemokine in the recruitment and activation of macrophages in this disease. These data demonstrate that early murine Lyme carditis requires neither class II antigen expression nor presentation of antigen to CD4+ T lymphocytes and suggest a direct response of macrophages to cardiac tissue invasion by B. burgdorferi.

Animals↗

PubMed1995

Class II major histocompatibility complex-deficient mice initially control an infection with Leishmania major but succumb to the disease.

Class II major histocompatibility complex (MHC)-deficient (H-2b) mice do not express I-A or I-E molecules and, as a result, do not develop CD4 cells. Thus, they represent the ideal model for examining the importance of CD4 cells and MHC class II molecules in resistance to infection with Leishmania major and the capacity of MHC class I-restricted T cells to mediate resistance to L. major. Class II MHC-deficient mice and control (C57BL/6, normal and nude) mice were infected with L. major. Although MHC class II-deficient mice were able to control infection more effectively than nude mice, cutaneous lesions on the mice eventually progressed, and parasite replication became uncontrolled. These results suggest that CD4 cells and MHC class II molecules are essential for resistance to L. major and that in the absence of these cells and molecules, such mice can transiently control infection with L. major but are unable to resolve such infections.

Animals↗

PubMed1995

Transgenic mice expressing MHC class II molecules with truncated A beta cytoplasmic domains reveal signaling-independent defects in antigen presentation.

The thymic development and peripheral activation of CD4+ T cells are critically dependent upon interactions with MHC class II molecules on the surface of antigen presenting cells (APC). In vitro studies involving transfection of cell lines with mutant MHC molecules have demonstrated that the cytoplasmic domains of class II molecules can be required for efficient antigen presentation. To address the role of class II cytoplasmic domains in physiological, non-transformed APC and in vivo immune responses, we have generated transgenic mice which express only truncated class II A beta molecules lacking the 13 membrane distal residues. In vivo, CD4+ T cell development and immune responses to conventional protein antigens, parasitic infections and skin grafts were indistinguishable between control and transgenic mice. Nevertheless, in vitro, APC from transgenic mice poorly stimulate T cell hybridomas and wild-type in vivo-primed T cells. Neither class II-mediated induction of B7-1 expression nor homotypic aggregation were diminished in transgenic B cells, suggesting that both cAMP and tyrosine kinase signaling pathways remain intact despite truncation of the A beta cytoplasmic domain. Furthermore, chemically-fixed cells from transgenic animals are impaired in their antigen presenting capacity. Thus, in contrast to previous studies with cell lines transfected with truncated class II molecules, these results suggest that signaling-independent mechanisms account for the defective in vitro antigen presenting capacity of physiological APC expressing truncated A beta proteins.

Amino Acid Sequence↗

PubMed1995

Novel NFAT sites that mediate activation of the interleukin-2 promoter in response to T-cell receptor stimulation.

The transcription factors NFAT and AP-1 have been shown to be essential for inducible interleukin-2 (IL-2) expression in activated T cells. NFAT has been previously reported to bind to two sites in the IL-2 promoter: in association with AP-1 at the distal antigen response element at -280 and at -135. On the basis of DNase I footprinting with recombinant NFAT and AP-1 proteins, gel shift assays, and transfection experiments, we have identified three additional NFAT sites in the IL-2 promoter. Strikingly, all five NFAT sites are essential for the full induction of promoter activity in response to T-cell receptor stimulation. Four of the five NFAT sites are part of composite elements able to bind AP-1 in association with NFAT. These sites display a diverse range of cooperativity and interdependency on NFAT and AP-1 proteins for binding. One of the NFAT sites directly overlaps the CD28-responsive element. We present evidence that CD28 inducibility is conferred by the AP-1 component in NFAT-AP-1 composite elements. These findings provide further insight into the mechanisms involved in the regulation of the IL-2 promoter.

Animals↗

PubMed1995

Immune responses in MHC class II-deficient mice.

Major histocompatibility complex (MHC) class II molecules are heterodimeric cell surface proteins that are critically important for the development and function of cells in the immune system. In particular, the maturation of CD4+ T cells is dependent on the expression of MHC class II molecules on thymic epithelium, while the activation of these cells requires the expression of class II molecules on specialized antigen-presenting cells in the periphery. The importance of class II molecules is especially evident in humans who are afflicted with MHC class II-deficient combined immunodeficiency, as these individuals die at an early age unless provided with a bone marrow transplant. Here we discuss the functional consequences of MHC class II deficiency in a mouse model generated by gene targeting in embryonic stem (ES) cells. These mice have proved to be valuable reagents for dissecting the mechanisms by which MHC class II molecules control the maturation and activation of lymphocytes as well as for elucidating the role of these cells in various immune responses.

Animals↗

PubMed1994

Activation of T cells by superantigen in class II-negative mice.

The ability of the staphylococcal enterotoxins to stimulate T cells has been thought to depend on their association with class II MHC products. Here, we demonstrate that a subgroup of staphylococcal enterotoxins, which includes staphylococcal enterotoxin C and staphylococcal enterotoxin E, stimulates strong MHC-independent responses, thereby resulting in T cell expansion and generation of CTL. The immunologic consequences of MHC-independent activation of T cells by superantigens differ from those of class II-dependent activation, inasmuch as this pathway does not result in detectable T cell deletion. These findings delineate a novel MHC-independent T cell activation pathway that leads to both clonal expansion and expression of CTL effector function in response to a subgroup of bacterial superantigens.

Animals↗

PubMed1994

Protective role of major histocompatibility complex class II Ebd transgene on collagen-induced arthritis.

Collagen-induced arthritis (CIA) is an animal model of autoimmune inflammatory polyarthritis that has features similar to rheumatoid arthritis (RA). Much like RA, susceptibility to mouse CIA is influenced by the major histocompatibility complex (MHC), H-2, and restricted to the H-2q and H-2r haplotypes. Whereas the role of the H-2A molecule in susceptibility to CIA is well established, little is known about the role of H-2E molecule in the disease. In this study, we analyzed the effect of a transgenic E beta d molecule on CIA susceptibility in a recombinant mouse B10.RQB3, which expresses the CIA susceptible Aq genes and an Eak gene, but does not produce an E molecule since Ebq is nonfunctional. In the presence of an Ebd transgene, a viable E molecule is generated. Whereas B10.RQB3 were susceptible to CIA, B10.RQB3-E beta d+ showed a dramatic reduction in the incidence of arthritis as well as a decrease in the level of anti-mouse and anti-bovine CII antibodies in their serum. No clear cut differences in the expression of T cell receptor (TCR) V beta was observed between E beta d+ and E beta d- transgenic mice. Mechanisms underlying the protective effect of E beta d transgenic molecule on CIA may shed light on how HLA-DR molecules influence human RA.

Animals↗

PubMed1994

Absence of MHC class II molecules reduces CNS demyelination, microglial/macrophage infiltration, and twitching in murine globoid cell leukodystrophy.

Globoid cell leukodystrophy (GLD) is a severe genetic demyelinating disorder with an increased number of Ia (immune response antigen) positive brain microglia/macrophages. To assess the role of aberrant Ia expression in the central nervous system (CNS), twitcher mice, which represent the murine model for GLD, were mated with Ia- transgenic mice. Compared with the Ia+ controls, Ia- twitcher mice showed a profound reduction in the severity of demyelinating lesions correlated with significantly fewer microglia/macrophages. Most importantly, Ia- twitcher mice showed significantly reduced twitching compared with ia+ twitcher mice. In contrast with experimental allergic encephalomyelitis (EAE), there was no significant amount of inflammatory T cell infiltrates, implying that T cells may not play a predominant role in this disease. These findings may have broad therapeutic implications for Alzheimer's disease, Parkinson's disease, and Huntington's disease, which display enhanced Ia expression in the CNS without obvious T cell infiltrates.

Animals↗

PubMed1994

Prevention of nephritis in major histocompatibility complex class II-deficient MRL-lpr mice.

MRL-lpr mice develop aggressive autoimmune kidney disease associated with increased or de novo renal expression of major histocompatibility complex (MHC) class II molecules and a massive systemic expansion of CD4-CD- double negative (DN) T cells. Whereas non-MHC linked genes can have a profound effect on the development of nephritis, lymphadenopathy, and anti-DNA antibody production in MRL-lpr mice, the role of MHC molecules has not been unequivocally established. To study the role of MHC class II in this murine model of systemic lupus erythematosis, class II-deficient MRL-lpr mice (MRL-lpr -/-) were created. MRL-lpr -/- mice developed lymphadenopathy but not autoimmune renal disease or autoantibodies. This study demonstrates that class II expression is critical for the development of autoaggressive CD4+ T cells involved in autoimmune nephritis and clearly dissociates DN T cell expansion from autoimmune disease initiation.

Animals↗

PubMed1994

B7 expression on thymic medullary epithelium correlates with epithelium-mediated deletion of V beta 5+ thymocytes.

Recent evidence suggests that I-E+ thymic epithelium, especially medullary epithelium, can induce partial deletion of superantigen-reactive T cells expressing TcR V beta 5, V beta 11, and V beta 17. To seek further information on this issue, we constructed bone marrow chimeras in which MHC class II I-E is expressed on thymic epithelial cells at various levels and locations; the chimeras were reconstituted with stem cells from TcR V beta 5 transgenic mice. Intrathymic deletion of V beta 5 T cells was restricted to relatively mature T cells (expressing high TcR levels), and the degree of deletion correlated with the density of I-E expression in the thymic medulla rather than in the thymic cortex; selective I-E expression in medullary epithelium caused prominent deletion. Interestingly, immunostaining of normal and chimeric mice revealed that expression of B7 (the ligand for CD28) is largely restricted to a subset of medullary epithelial cells; these cells are I-E+ and co-express a specific carbohydrate bound by the lectin UEA-1. B7 expression was lower in thymuses of class II-deficient mice (A beta b-/-) and T-cell-deficient mice (SCID), suggesting that B7 expression is up-regulated during CD4+ thymocyte selection. In support of this idea, B7 expression in the thymus was restored to a normal level in bone marrow reconstituted SCID mice. Because B7 expression correlates with a costimulatory signal for T cells, selective expression of B7 and related antigens on I-E+ medullary epithelium may explain why these cells play a more prominent role in V beta deletion than cortical epithelium.

Animals↗

PubMed1994

Indirect recognition by helper cells can induce donor-specific cytotoxic T lymphocytes in vivo.

In vitro studies have revealed that help for cytotoxic T lymphocyte (CTL) induction can be mediated through several pathways, including direct recognition of allogenic class II antigens by CD4+ cells, direct recognition of allogeneic class I antigens by "CD4-independent" CD8+ cells, and "indirect" recognition of peptides of alloantigens presented in association with self class II molecules. Whereas good evidence for the two direct pathways is available in vivo, there is relatively little evidence to show that indirect recognition can initiate graft rejection. This study examined the role of indirect allorecognition during the generation of CTLs in mice as they rejected major histocompatibility complex (MHC) class II-deficient skin after depletion of CD8+ T cells in vivo. Recipients were depleted of CD8+ T cells by in vivo treatment with anti-CD8 monoclonal antibody and then grafted with allogeneic skin lacking MHC class II antigens. The mice rejected the skin grafts rapidly. Although flow cytometry showed marked depletion of CD8+ T cells in these mice, we found that (a) CD8+ CTLs were generated and sensitized to MHC class I antigens of the donor; (b) the generation of the CD8+ CTLs required the help in vivo of CD4+ cells, as well as priming with the allogeneic skin graft; and (c) the CD4+ T helper cells were sensitized indirectly to donor peptides presented in association with class II antigens on recipient antigen-presenting cells. These results provide evidence that indirect recognition can provide effective help for CTL induction during graft rejection, even when the cytotoxic T cells are sensitized by determinants expressed only on the donor graft.

Animals↗

PubMed1994

A common factor regulates both Th1- and Th2-specific cytokine gene expression.

Murine T helper cell clones are classified into two distinct subsets, T helper 1 (Th1) and T helper 2 (Th2), on the basis of cytokine secretion patterns. Th1 clones produce interleukin-2 (IL-2), tumor necrosis factor-beta (TNF-beta) and interferon-gamma (IFN-gamma), while Th2 clones produce IL-4, IL-5, IL-6 and IL-10. These subsets differentially promote delayed-type hypersensitivity or antibody responses, respectively. The nuclear factor NF-AT is induced in Th1 clones stimulated through the T cell receptor-CD3 complex, and is required for IL-2 gene induction. The NF-AT complex consists of two components: NF-ATp, which pre-exists in the cytosol and whose appearance in the nucleus is induced by an increase of intracellular calcium, and a nuclear AP-1 component whose induction is dependent upon activation of protein kinase C (PKC). Here we report that the induction of the Th2-specific IL-4 gene in an activated Th2 clone involves an NF-AT complex that consists only of NF-ATp, and not the AP-1 component. On the basis of binding experiments we show that this 'AP-1-less' NF-AT complex is specific for the IL-4 promoter and does not reflect the inability of activated Th2 cells to induce the AP-1 component. We propose that NF-ATp is a common regulatory factor for both Th1 and Th2 cytokine genes, and that the involvement of PKC-dependent factors, such as AP-1, may help determine Th1-/Th2-specific patterns of gene expression.

Animals↗

PubMed1994

Regulation of cytokine gene expression in T helper cell subsets.

We have examined the transcriptional regulation of the Th1-specific IL-2 gene and the Th2-specific IL-4 gene by transient transfection of promoter-chloramphenicol acetyl transferase (CAT) constructs into T cell clones and by electrophoretic mobility shift assays. Transfection of the Th2 clone D10.G4 with IL-4 promoter-CAT constructs demonstrated anti-CD3 inducible IL-4 promoter activity. In contrast, CAT constructs containing murine IL-2 promoter sequences were not inducible in D10.G4 cells. Transfection analyses in the Th1 clone D1.1 demonstrated inducible IL-2 promoter activity but no IL-4 promoter activity. Electrophoretic mobility shift assays using well-defined regulatory elements in the murine IL-2 gene promoter showed that anti-CD3 stimulation of Th2 clones failed to induce a characteristic increase in the ratio of p65-p50:p50-p50 NF-kappa B binding complexes in the nucleus that did occur in IL-2-producing clones. In addition, other protein complexes with NF-kappa B binding sequences were seen using lysates from Th1 and Th0 cells but not Th2 cells. Thus, expression of the promoter-CAT constructs directly correlates with endogenous IL-2 and IL-4 gene expression in Th1 and Th2 clones, confirming that the differential expression of IL-2 and IL-4 genes in these T cells is transcriptionally controlled. Furthermore, the lack of IL-2 transcription in activated Th2 cells is associated with the failure to generate required IL-2 gene promoter binding proteins, particularly NF-kappa B in the nucleus.

Animals↗

PubMed1994

Expression of metalloproteinases in pigmented villonodular synovitis.

Pigmented villonodular synovitis (PVNS) is an idiopathic proliferative synovial process composed of two predominant cell types: mononuclear histiocytic cells and giant cells. This lesion can be locally invasive and can result in bone cyst formation and late cartilage and bone loss. Because metalloproteinases have been implicated in the joint destruction occurring in inflammatory arthritis and in the ability of certain tumors to invade adjacent tissues, their presence in PVNS was determined. Synovial tissue samples were collected at surgical synovectomy from the knees of 10 patients with a prior histological diagnosis of PVNS. Pigmented villonodular synovitis synovium was examined for the presence of the metalloproteinases collagenase and stromelysin. Messenger RNA (mRNA) for collagenase and stromelysin was present in all patient samples, although in varying amounts. In situ hybridization studies on synovial tissue sections identified synovial lining cells as the predominant cells expressing these metalloproteinases. Occasional infiltrating mononuclear histiocytic cells also were producing metalloproteinase mRNA. Giant cells did not express mRNA for the metalloproteinases collagenase and stromelysin. These results suggest that collagenase and stromelysin may be among the mediators of cartilage and bone loss that can occur in PVNS.

Adult↗