Search PubMed⌕ Search

Biomedical subjects

E M Shevach

Publications and source records attributed to E M Shevach.

At least 91 records · Page 5Linked to original sources
PubMed1991

The mouse vitronectin receptor is a T cell activation antigen.

In this report, we demonstrate that the T cell activation antigen, recognized by monoclonal antibody H9.2B8, is the murine homologue of the vitronectin receptor (VNR) and, thereby, we provide initial evidence that VNR is expressed on lymphoid cells. VNR is expressed on a variety of T cell lines, tumors, and Con A-activated splenocytes, but not resting T cells, and is capable of binding to the extracellular matrix proteins fibronectin, fibrinogen, and vitronectin, via the tripeptide sequence RGD. There was no evidence of novel beta chains pairing with the VNR alpha chain, as has been demonstrated in some human cells. In view of recent studies demonstrating that this molecule functions as an accessory molecule in T cell activation, the VNR may play an important role in mouse T cell functions.

Amino Acid Sequence↗
PubMed1991

The vitronectin receptor serves as an accessory molecule for the activation of a subset of gamma/delta T cells.

Constitutive production of cytokines was observed in 3 of 12 gamma/delta T cell lines derived from murine epidermis and correlated with the expression of the C gamma 4, V delta 6 T cell receptor (TCR). After adaptation of one of the lines (T195/BW) to serum-free culture conditions, cessation of the "spontaneous" production of interleukin 4 (IL-4) was observed and IL-4 production could then by induced by the addition of RGD-containing extracellular matrix (ECM) proteins to the culture. The response to the ECM proteins could be completely inhibited by a mAb to the murine vitronectin receptor (VNR). However, the induction of IL-4 production could also be inhibited by anti-CD3 and by an anti-clonotypic mAb to the TCR-gamma/delta of T195/BW. As TCR-gamma/delta loss mutants of T195/BW also failed to respond to ECM proteins, these data demonstrate that engagement of the VNR by its ligand is necessary, but not sufficient, for the induction of IL-4 production. Furthermore, the VNR is expressed by many other T cell clones (both gamma/delta and alpha/beta), none of which produce lymphokines constitutively. Taken together, these observations strongly favor the view that not only is coexpression of the VNR and TCR required for the induction of IL-4 production, but that the TCR must also be engaged by its ligand, most likely a cell surface antigen expressed by the hybridoma itself.

Animals↗
PubMed1991

Evidence that cyclosporine treatment during pregnancy predisposes offspring to develop autoantibodies.

Cyclosporine was administered (11 mg/kg/day) to pregnant mice to study the effects of passively transferred CsA on the developing immune system. Placental transfer of CsA was shown by the detection of fetal-tissue levels ranging from 400 to 1500 ng CsA/g tissue. Treatment clearly altered the developing immune system. Thymuses from the day-18 embryos exposed to CsA were partially depleted of CD4+CD8- single positive cells. Eleven of 50 offspring born to CsA-treated mothers developed significant levels of IgG autoantibodies to gastric antigens. In addition, two animals that received CsA in utero developed an extensive mononuclear cell infiltrate in the gastric mucosa resembling autoimmune gastritis. These results raise the possibility that the administration of CsA during pregnancy may result in potential long-term effects on the developing immune system. Offspring of such pregnancies may suffer an increased incidence or severity of autoimmune diseases.

Animals↗
PubMed1990

Chromosomal location of the Ly-49 (A1, YE1/48) multigene family. Genetic association with the NK 1.1 antigen.

The Ly-49 (A1, YE1/48) Ag is a disulfide-linked dimer with 44-kDa subunits, and is expressed on the cell surface of rare T cell tumors of C57BL/6 origin. Although this Ag is undetectable by flow microfluorimetry analysis, normal cells have been shown to express the A1 Ag by immunoprecipitation experiments performed on surface radioiodinated spleen and thymus cells. We (J. Immunol. 143:1379, 1989) and others (J. Immunol. 142:1727, 1989) have recently isolated cDNA encoding the Ly-49 Ag. Southern blots with an Ly-49 cDNA probe revealed multiple bands, consistent with cross-hybridization to other members of a multigene family, and significant RFLP between the C57BL/6 and BALB/c strains. When the RFLP patterns displayed by other common laboratory strains as well as informative recombinant inbred strains were examined, the Ly-49 gene family displayed five RFLP patterns and the entire family was found to reside on a contiguous stretch of the distal portion of mouse chromosome 6, the same region to which the NK1.1 Ag has been mapped. Although tissue distribution studies and transfection analysis ruled out the possibility that Ly-49 was identical to NK1.1 Ag, approximately 20% of NK1.1 cells isolated from normal spleen coexpressed Ly-49 and all Ly-49+ cells were CD3-. Although spleen cells cultured in high doses of rIL-2 demonstrated similar coexpression of NK1.1 and Ly-49, approximately 10% of CD3+ cells coexpressed Ly-49. The chromosomal mapping data and the expression of the Ly-49 and NK1.1 Ag suggest that the NK1.1 Ag may be a member of the Ly-49 multigene family.

Animals↗
PubMed1990

Mechanism of action of cyclosporine A in vivo. II. T cell priming in vivo to alloantigen can be mediated by an IL-2-independent cyclosporine A-resistant pathway.

Cyclosporine A (CsA) inhibits T lymphocyte activation in vitro by blocking at a pretranslational level the production of IL-2 and other cytokines. It is widely assumed that the effectiveness of CsA as an immunosuppressive drug is secondary to a similar mechanism of action in vivo. We have previously demonstrated that certain parameters of T cell activation in the draining popliteal lymph node in response to the injection of alloantigen in the footpad were either completely resistant or enhanced by the administration of CsA. In the present study, we have shown that the mechanism of action of CsA in vivo is identical to that seen in vitro as CsA completely suppressed the induction of IL-2 mRNA as detected in a nuclease protection assay in lymph node cells from alloantigen-primed animals. Nevertheless, T cells from CsA-treated animals appeared to have undergone both priming and differentiation. Thus, upon culture in vitro in the presence of CsA, cells from CsA-treated animals manifested a vigorous proliferative response that could not be inhibited by the addition of a large panel of anti-cytokine mAb. Furthermore, cells from CsA-treated animals demonstrated an enhanced secondary response to the priming alloantigen, which suggests that they had undergone clonal expansion in vivo. Although CTL activity was markedly suppressed in cells from CsA-treated animals, after a 36-h culture in the absence of CsA, CTL activity equivalent to that detected in cells from nontreated animals was present. Collectively, these data support the existence of an alternative IL-2-independent, CsA-resistant pathway of T cell activation/differentiation that may play a prominent role in the generation of certain T effector functions in vivo.

Animals↗
PubMed1990

Role of phosphatidylinositol-anchored proteins in T cell activation.

A novel class of cell surface proteins are attached to the plasma membrane via a phosphatidylinositol (PI)-glycan anchoring structure, and these proteins can be selectively removed from the cell surface by the enzyme PI-specific phospholipase C (PI-PLC). Enzyme treatment led to a prolonged reduction in cell surface expression of several PI-anchored proteins. Activation of T cells led to a marked decrease in the ability of PI-PLC to remove PI-anchored surface proteins from the activated T cells. This decrease in PI-PLC sensitivity may reflect an alteration in the PI-glycan anchoring structures, or in a general membrane property, which renders the PI-anchored proteins inaccessible to the enzyme. When murine T lymphocytes were treated with PI-PLC and then stimulated with either Con A, the calcium ionophore A23187 and PMA, or an anti-CD3 mAb, the response to Con A stimulation was inhibited by 90%, whereas the responses to ionophore and PMA or anti-CD3 were not affected. Removal of PI-anchored proteins inhibited an early event in the activation process in response to Con A because both IL-2 production and IL-2R expression were inhibited by the PI-PLC treatment. Inhibition of the Con A response was secondary to removal of a PI-linked protein from the responder T cell population because PI-PLC treatment of T-depleted spleen cells did not alter their ability to act as a source of accessory cells. It is unlikely that removal of the known PI-linked proteins on murine T cells, Thy-1 and Ly-6, can fully account for the inhibition of Con A response because the cell line M2B3, that lacks these surface proteins, responded normally to Con A stimulation. These studies demonstrate that one or more PI-anchored T cell proteins play an important role in an early step of Con A activation, perhaps involving T cell-accessory cell interactions. In contrast, the ability to stimulate T cells by direct cross-linking of TCR/CD3 complex is not dependent on the presence of these PI-anchored proteins.

Animals↗
PubMed1990

Characterization of agretopes and epitopes involved in the presentation of beef insulin to T cells.

Beef insulin-specific I-Ad-restricted T cell hybridomas were derived from the fusion of antigen-primed (BALB/c X B6)F1 T cells with BW5147 thymoma. Specificity analysis revealed that the A-chain loop region is involved in antigen recognition. Hybridoma A20.2.15 is specific for beef insulin and cross-reacted with sheep insulin, but not with pork insulin. Using synthetic peptides we showed that the A-chain loop containing peptide A1-A14 jointed to the B7-B15 peptide by a disulfide bond can activate this hybridoma. Fragments generated by enzyme digest further suggest that the peptide recognized on beef insulin appears to involve A-chain loop residues A5-A12 and B-chain residues B7-B13 that are linked by the A7-B7 disulfide bridge. We found that beef insulin needs to be processed prior to T cell activation. Glutaraldehyde fixation and chloroquine treatment of presenting cells abolished their capacity to present insulin. Beef insulin denatured by pH changes cannot activate, thus suggesting that simple denaturation is not sufficient for presentation by antigen presenting cells. Finally, the agretope on beef insulin is comprised of two functional regions B7-B13 on the B chain and the A-chain loop in the A-chain, while residues A8 and A10 are probably involved in interaction with the T cell receptor.

Amino Acid Sequence↗
PubMed1989

A murine T lymphocyte antigen belongs to a supergene family of type II integral membrane proteins.

A murine cell surface, disulfide-linked 85kDa dimer, defined with murine mAb A1, is expressed at high levels on EL-4 cells, but at low levels on normal C57BL/6 T cells. A similar structure is recognized by the rat mAbs YE1/32 and YE1/48. We isolated a cDNA clone encoding the antigen recognized by mAb A1 by immunoselection of a cDNA library in the eukaryotic expression vector CDM8. COS 7.2 cells transfected with this cDNA clone expressed an mAb A1-reactive 85 kDa disulfide-linked dimer with 44 kDa subunits, which was also reactive with the mAbs YE1/32 and YE1/48. The A1 gene displayed extensive strain polymorphism, underwent no rearrangement in EL-4, and hybridized with multiple restriction fragments, suggesting that it is a member of a multi-gene family. The deduced polypeptide contained 262 residues with an m.w. of 30,648, multiple cysteines, and three potential N-linked glycosylation sites, consistent with previous observations. In contrast to most integral membrane proteins, the putative A1 protein had features of a type II integral membrane protein structure, with its carboxyl terminus exposed extracellularly and an intracytoplasmic amino terminus. There was significant homology with several type II integral membrane proteins, including the human and chicken asialoglycoprotein receptors, and especially the human low affinity Fc epsilon receptor, in the putative extracellular domains of these proteins. This analysis suggested that the A1 gene belongs to a novel supergene family of type II integral membrane proteins and suggested that the A1 protein itself may be involved in binding a soluble ligand such as carbohydrates or immunoglobulin.

Amino Acid Sequence↗
PubMed1989

Activation of T lymphocytes through the Ly-6 pathway is defective in A strain mice.

The Ly-6 alloantigens have been shown to participate in the process of T cell activation based on the ability of anti-Ly-6 mAb to induce IL-2 production and proliferation of T lymphocytes. In the present investigation we have demonstrated that peripheral T lymphocytes from A strain mice exhibited abnormally low proliferative responses after stimulation through Ly-6A/E and Ly-6C molecules when compared to responses of T cells from numerous other mouse strains. The abnormal activation of the Ly-6 pathway of A strain T cells was not due to ineffective FcR cross-linking of the anti-Ly-6 mAb, to inappropriate cellular expression of the Ly-6A/E alloantigen in A strain T cells, or to an active suppressive phenomenon. T lymphocytes from A strain mice proliferated normally when the cells were activated by mAb to Thy-1 or the CD3/TCR complex suggesting that A strain mice did not exhibit a generalized T cell activation defect. Cell separation studies of T cells and accessory cells demonstrated that this defect was quantitative, rather than qualitative, and that it was complex, residing at both the T cell and accessory cell levels. These results suggest that activation of T lymphocytes via the Ly-6 molecule may involve a signaling pathway and/or cell-cell interactions distinct from those required for optimal activation via CD3/TCR.

Animals↗
PubMed1989

Murine T cells express a cell surface receptor for multiple extracellular matrix proteins. Identification and characterization with monoclonal antibodies.

Cell-cell and cell-extracellular (ECM) protein interactions are mediated through heterodimers termed integrins. We have demonstrated that dendritic epidermal T cell (DETC) lines adhere to the ECM proteins, fibronectin, fibrinogen, and vitronectin but not to collagen, laminin, or control proteins. This adhesion was blocked by the tetrapeptide RGDS, but not the control peptide, RGES. We have derived a hamster mAb H9.2B8, and a rat mAb, 8.18E12, from immunizations with DETC lines. The mAbs in combination, but not individually, specifically inhibited the adhesion of DETC lines to fibronectin, fibrinogen, and vitronectin. Immunoprecipitation analysis revealed that both mAbs reacted with a heterodimer composed of noncovalently linked 140- and 95-kD subunits. The 140-kD subunit can be reduced to 120- and 23-kD fragments. Although the two mAbs did not cross-compete for binding to DETC, sequential immunoprecipitation studies indicated that they react with the same 120-kD fragment. While all DETC cell lines and several T cell clones were reactive with the mAbs, the mAbs were not reactive with normal spleen, lymph node, thymus, or skin. Stimulation of splenic T cells with Con A or allogeneic cells induced mAb reactivity after 1 wk in vitro. These data demonstrate that a single lymphocyte receptor, with biochemical features characteristic of integrins, mediates RGD-dependent binding to the ECM proteins, fibronectin, fibrinogen, and vitronectin. Furthermore, since this integrin is expressed by long-term activated T cells, this receptor may play a physiological role in T cell function.

Animals↗
PubMed1989

Biochemical characterization of the inhibitory effect of CsA on cytolytic T lymphocyte effector functions.

We have examined the effects of cyclosporine A (CsA) on a number of CTL effector functions. CsA partially inhibited the CTL-mediated lysis of Ag-bearing target cells. Both target cell- and anti-TCR mAb-induced granule exocytosis were markedly inhibited by CsA. In addition, marked inhibition of PMA and calcium ionophore (A23187) induced granule exocytosis was produced by CsA suggesting that the inhibitory effects of CsA on granule exocytosis involve biochemical events after protein kinase C activation and increases in intracellular free Ca2+. CsA had no inhibitory effects on TCR-mediated phosphatidylinositol metabolism. The inhibitory effects of CsA were not mediated by the cAMP-dependent protein kinase inhibitory pathway and no effect of CsA on the Ca2+-induced binding of calmodulin to calmodulin-binding proteins could be demonstrated. CsA was also a potent inhibitor of IgE receptor-mediated exocytosis in rat basophil leukemia cells. CsA had no effect on receptor-mediated phosphatidylinositol hydrolysis; 400 ng/ml CsA resulted in a 90% inhibition of serotonin release but had no effect on phosphatidylinositol hydrolysis. These results indicate that CsA may inhibit some common event in Ca2+-dependent secretory cells. Taken together, these results suggest that CsA does not inhibit signal transduction but rather interferes with the biochemical events in the later stages of Ca2+-dependent reactions that follow the binding of calmodulin to cytoskeletal or cytoplasmic calmodulin binding proteins.

Animals↗
PubMed1989

Delta-chains of dendritic epidermal T cell receptors are diverse but pair with gamma-chains in a restricted manner.

To assess the diversity of TCR delta-gene expression in dendritic epidermal T cells (DETC), we characterized the delta-gene used by a panel of cell lines that express the gamma delta TCR on the cell surface. Northern hybridization analyses with a panel of V delta probes representing each of the six known V delta families showed that each of these lines expressed either V delta 1 or V delta 6 containing transcripts. Southern hybridization analysis with the V delta probes gave rearrangement patterns consistent with Northern hybridization results. The correlation of V delta expression with V gamma expression revealed that not only may V delta usage be restricted in DETC cells, but that the pairing of gamma- and delta-chains may not be random. In the DETC cells examined, V delta 1 chains paired exclusively with V gamma 3C gamma 1 chains and V delta 6 chains paired with C gamma 2 and C gamma 4 chains. Sequence analysis of delta-cDNA clones corresponding to the expressed delta-chains from one of the V delta 1 expressing cell lines and two of the V delta 6 expressing cell lines revealed that, although the V delta 1 gene segment sequence and one of the V delta 6 gene segment sequences were identical to previously published V delta sequences, considerable variable region diversity was generated by complex V-D-J rearrangement patterns that utilized various combinations of N-additions, D delta 2 and possibly D delta 1 segments, and J delta 1 or J delta 2 segments. These complex rearrangement patterns distinguish these DETC lines from early thymocytes and suggest that, if DETC are thymic derived, they originate from relatively mature thymic cells.

Animals↗
PubMed1989

Ly-6: a multigene family in search of a function.

Differentiation antigens can be simply defined as cell surface antigens that distinguish one cell type from another. The Ly-6 family of cell surface antigens might easily be regarded as a prototype for the analysis of the function of differentiation antigens on immunocompetent cells. Although a number of studies with antisera, in the late 1970s, established that the Ly-6 antigens exhibit an unusual pattern of expression on T and B lymphocytes, functional studies failed to demonstrate that the expression of Ly-6 defines a unique subpopulation of immunocompetent cells. Paradoxically, the development of anti-Ly-6 monoclonal antibodies led to a further decline in interest in this group of antigens, as the complexity of the results relegated Ly-6 to the realm of the die-hard immunogeneticist. Over the past five years, interest in Ly-6 has been rekindled and in this review Ethan Shevach and Patricia Korty summarize recent studies on the serology, biochemical and molecular structure, and the function of the products of this multigene family.

Amino Acid Sequence↗
PubMed1989

Interleukin 7 is a T-cell growth factor.

Interleukin 7 (IL-7) is a 25-kDa cytokine which was purified and its corresponding cDNA was cloned based upon its ability to stimulate the proliferation of pre-B cells. It has been shown that IL-7 can also function as a costimulator with Con A for the proliferation of T lymphocytes by inducing the production of interleukin 2 (IL-2). We demonstrate here that IL-7 in combination with phorbol 12-myristate 13-acetate can directly drive the proliferation of purified T cells and that this response is not inhibited by cyclosporine A or by antibodies to IL-2 and IL-4. Stimulation of T cells with phorbol myristate acetate and IL-2, IL-4, or IL-7 prepared T cells to respond to any of the three lymphokines. Although T cells activated in vitro by anti-CD3 or allogeneic cells failed to proliferate when challenged with IL-7, T cells primed in vivo to the same stimuli demonstrated a significant proliferative response when restimulated in vitro with IL-7. IL-7 can, therefore, function both as a growth factor for T cells in an IL-2-independent manner and as a competence factor for the induction of lymphokine responsiveness. The ability to induce IL-7 responsiveness via stimulation of the T-cell receptor complex in vivo, but not in vitro, raises the possibility that IL-7 may play a role in T-cell growth and differentiation in vivo.

Animals↗
PubMed1989

Very early (VEA) and very late (VLA) activation antigens have distinct functions in T lymphocyte activation.

We have described in detail the properties of two types of murine T-cell activation antigen. mAbs to these cell surface antigens were initially identified because they demonstrated apparent selective reactivity with a number of cloned cell lines derived from dendritic epidermal T cells. After careful study, both of these antigens were also found to be expressed by normal T cells. The dimer recognized by mAb H1.2F3 which we have termed Very Early Activation (VEA) antigen, is rapidly and transiently expressed during the process of normal T-cell activation. It is likely that the VEA antigen plays a major role in T-cell activation as mAb H1.2F3 is a potent stimulator of T-cell proliferation in the presence of PMA and accessory cells. The second cell surface antigen recognized by mAbs H9.2B8 and 8.18E12 is a Very Late Activation (VLA) antigen that has biochemical, tissue distribution, and functional properties which resemble the human VLA antigens, which have been recently shown to be members of the integrin superfamily of cell surface receptors. While the physiologic ligand for the VEA antigen has yet to be identified, the integrin identified by mAbs H9.2B8 and 8.18E12 mediates cell adhesion to a well-defined group of ligands consisting of the ECM-proteins fibronectin, fibrinogen, and vitronectin. We postulate that the VEA and VLA antigens have distinctly different functions and that further detailed analysis of these dimers will further elucidate the role of these cell surface antigens in T-cell activation and function.

Animals↗
PubMed1988

Expression of C gamma 4 T cell receptors and lack of isotype exclusion by dendritic epidermal T cell lines.

Although four murine C gamma gene segments (C gamma 1, 2, 3, and 4) are known to exist, the large majority of expressed gamma-chains have been shown to be of the C gamma 1 isotype and no evidence exists for the expression of more than one receptor by gamma delta TCR-bearing cells. We investigated the nature of the TCR expressed on a number of murine dendritic epidermal T cell-derived cell lines by using both Northern blot and immunoprecipitation analyses. One of these CD3+ cell lines (T195) expresses C gamma 4, V gamma 1, and delta mRNA, and its CD3-associated TCR complex can be precipitated by both anti-C gamma 4 and anti-delta sera, indicating that this receptor is a C gamma 4/delta heterodimer. Furthermore, we show that two cell lines (Y245, Y93) express two distinct TCR gamma-chains, one derived from the C gamma 4 locus, whereas the second gamma-chain is probably derived from the C gamma 2 locus. Together with the previous demonstration of C gamma 1/delta TCR on a number of dendritic epidermal T cell lines (DETC), these results indicate that such DETC are capable of expressing a variety of gamma delta TCR and that, in some DETC, isotype exclusion of gamma-chain expression does not occur.

Animals↗