Effects of extracellular ATP and adenosine on different thymocyte subsets: possible role of ATP-gated channels and G protein-coupled purinergic receptor.

To explore the possible role of purinergic receptors in thymocyte development and in pathogenesis of adenosine deaminase SCID, we studied effects of extracellular adenosine triphosphate (ATP(ext)) and adenosine on TCR- and steroid hormone-triggered processes in mouse thymocytes. Reverse transcriptase-PCR analysis confirms the mRNA expression of several types of purinergic receptors, while the functioning of ATP receptors in thymocytes is reflected by ATP(ext)-induced intracellular calcium increases and by thymocyte subset-specific sensitivity to the effects of ATP(ext) and adenosine. Only ATP(ext), but not the ATP catabolites, adenosine, dexamethasone, or TCR cross-linking, was efficient in triggering rapid protein synthesis independent lysis of CD4+8- thymocytes and peripheral CD4+ T cells. In contrast, extracellular adenosine specifically induced the apoptosis of CD4+8+ thymocytes. ATP(ext) also induced a slower process of DNA fragmentation and protein synthesis-dependent apoptosis in all thymocyte subsets. ATP(ext) had an additive effect with TCR cross-linking in the induction of thymocyte death, but, unexpectedly, the effects of ATP(ext) at high concentration were antagonistic to steroid-induced apoptosis. Described here, the properties of ATP(ext) and adenosine are consistent with their involvement in the regulation of T cell development due to differential expression and signaling through purinergic receptors in different thymocyte subsets. The possible role of purinergic receptor signaling in T cell differentiation and adenosine deaminase SCID is discussed.

Murine T lymphocytes modulate activity of an ATP-activated P2Z-type purinoceptor during differentiation.

Murine T, but not B, lymphocytes constitutively express a membrane receptor for adenosine nucleotides that opens a nonspecific pore that admits Ca2+ and ethidium (314 Da), but not propidium (415 Da) ions. ATP, ADP, and AMP show decreasing potency; UTP and adenosine are inactive. Nonhydrolyzable ATP analogues are completely ineffective. Oxidized ATP inhibits the response. Activity is detectable at ATP concentrations of 125 microM and peaks at 1 mM. The intracellular free Ca2+ ([Ca2+]i) rise is not reversed by removing ATP by centrifugation or apyrase. The kinetics, agonist and antagonist profiles, and the passage of ions as large as ethidium are the characteristics of a P2z-type purinoceptor. No expression of classical P2x-, P2u-, or P2Y-type purinoceptors can be detected. The [Ca2+]i elevating activity of the ATP receptor is modulated during T cell differentiation. CD4+8+ double-positive thymocytes are the least responsive. CD4-8+ single-positive thymocytes, CD8+ splenic T cells, CD4+8- single-positive thymocytes, and CD4+ splenic T cells show increasing reactivity. Measurement of P2Z expression by the rate of ethidium ion uptake correlates with the [Ca2+]i. The trimodal expression of P2Z by splenic CD4+ T cells correlates with the subsets defined by CD44 and CD45RB, differentiation Ags that distinguish memory cells: P2Zlow cells are CD44brightCD45RBbright; P2Zint are CD44dullCD45RBint; P2Zhigh are CD44brightCD45RBdull. It is suggested that P2Z receptor-mediated signaling could be involved in the regulation of differentiation and cell death in the thymus and peripheral T lymphocytes.

Antigen-specific cell conjugate formation and long-lasting calcium responses in recognition of Mls cellular superantigen by cloned murine T lymphocytes.

T lymphocyte recognition of cell-associated minor lymphocyte stimulation (Mls) superantigen was studied by simultaneous flow cytometric measurement of T cell free ionized intracellular calcium ([Ca2+]i) with the fluorescent probe indo-1 and T cell binding to antigen-presenting cells stained with a long-chained, membrane-fixed cyanine dye. Cloned T cell-B lymphocyte antigen-presenting cell conjugate formation and increased T cell [Ca2+]i were antigen specific and tightly linked in four Mls-reactive T cell clones. The T cell-antigen-presenting cell conjugates were extremely stable and, like T cell [Ca2+]i elevation, were maintained for more than 2 hr. Three ligand-receptor pairs, (i) T cell receptor/antigen+class II, (ii) CD4/class II, and (iii) LFA-1/ICAM-1, were obligatory participants in T cell recognition of cell-bound superantigen since monoclonal antibodies to any of them blocked formation of cell conjugates. Despite previous reports, Mls recognition occurred by the conventional T cell recognition pathway with easily detected phosphoinositide hydrolysis. B cell activation greatly enhances their recognition by Mls-specific T cells.

Two separate mechanisms of T cell clonal anergy to Mls-1.

T cell tolerance to superantigen can be mediated by clonal anergy in which Ag-specific mature T cells are physically present but are not able to mount an immune response. We induced T cell unresponsiveness to minor lymphocyte stimulations locus antigen (Mls)-1a in mice transgenic for TCR V beta 8.1 in three different systems: 1) injection of Mls-1a spleen cells, 2) mating with Mls-1a mice, and 3) bone marrow (BM) chimeras in which Mls-1a is present only on nonhematopoietic cells. CD4+8-V beta 8.1+ cells from all these groups did not proliferate in response to irradiated spleen cells from Mls-1a mice. We compared the response of these cells by T cell/stimulator cell conjugate formation, Ca2+ mobilization, and proliferation assays. The mechanisms underlying the unresponsiveness of these T cells appear to differ. CD4+8-V beta 8.1+ cells from Mls-1a spleen cell-injected mice mobilized cytoplasmic Ca2+ but proliferated at a reduced level in response to cross-linking with anti-TCR mAb. However, these cells formed conjugates, mobilized Ca2+, and proliferated in response to Mls-1a when activated B cells were used as stimulators, although they produced reduced levels of IL-2. In Mls-1a/b V beta 8.1 transgenic mice, a subset in CD4+8-V beta 8.1+ cells did not mobilize cytoplasmic Ca2+ after TCR cross-linking. Their conjugate formation, Ca2+ mobilization, or proliferation in response to Mls-1a on activated B cells was undetectable. Finally, CD4+8-V beta 8.1+ cells from the BM chimeras proliferated to TCR cross-linking at a partially reduced level and formed conjugates, mobilized Ca2+, and proliferated in response to Mls-1a on activated B cells. These features suggest that the mechanisms underlying the maintenance of anergy in Mls-1a spleen cell-injected mice are distinct from those in Mls-1a mice.

Regulation of IgD-receptor expression on murine T cells. II. Upregulation of IgD receptors is obtained after activation of various intracellular second-messenger systems; tyrosine kinase activity is required for the effect of IgD.

The presence of IgD receptors (IgD-R) on T cells during a primary response to antigen causes augmented antibody production and facilitates priming for a secondary response. Cross-linked, but not monomeric IgD leads to a rapid upregulation of these receptors on T cells. As shown in the present study, the rapid upregulation of IgD-specific receptors is also induced by cross-linking of T cell surface molecules known to mediate triggering of T cell activation, such as CD3, CD2, and Thy 1. Furthermore, IgD-R are also upregulated by pharmacologically active compounds that increase intracellular cAMP and by PMA/DiOG plus ionomycin, but not by either PMA or ionomycin alone. The upregulation of IgD-R by anti-CD3 is inhibited by both calphostin C and herbimycin A, while that due to DiOG plus ionomycin is only inhibited by calphostin C. Upregulation of IgD-R by increased cAMP is blocked by HA1004, but not by low concentrations of staurosporine or herbimycin A. IgD itself does not cause an increase in intracellular cAMP, protein kinase C translocation, influx of extracellular Ca2+, or a change in membrane potential. Relatively specific inhibitors of these activation pathways, HA1004, calphostin C, and neomycin, also fail to interfere with IgD-receptor upregulation by IgD itself. However, tyrosine kinase inhibitors, including herbimycin A, tyrphostin C11, and genistein, completely prevent the effect of IgD on IgD-R expression. Although an influx of Ca2+ is apparently not involved, a role for intracellular Ca2+ in the upregulation of IgD-R by IgD on T cells is indicated by the susceptibility to inhibition by BAPTA, W7, and FK520. We conclude that activation of at least three different second-messenger systems can cause IgD-R upregulation, but that the effect of IgD itself requires tyrosine kinase activity, perhaps in an intracellular Ca(2+)-dependent manner.

Lack of voltage sensitive potassium channels and generation of membrane potential by sodium potassium ATPase in murine T lymphocytes.

Voltage sensitive K+ channels, which are responsible for generation of membrane potential in most cells, are functionally absent in about one-third of peripheral murine T cells and greatly reduced in the rest as shown by resistance of their membrane potential to changes in extracellular potassium concentration and failure of K+ channel dependent volume regulation. Despite the absence of voltage- sensitive K+ channels, the membrane potential of peripheral T cells is between -60 and -70 mV, the same as thymocytes. A total of 40 to 70 mV of the membrane potential of peripheral T cells is produced by the direct electrogenic action of the asymmetric Na+K+ ATPase pump because the cells are depolarized by ouabain, an inhibitor of the pump, removal of extracellular potassium or reduction of temperature. The residual, ouabain-resistant membrane potential, is sensitive to the K+ channel blocker, quinine, and thus due to electrodiffusion through K+ channels. Na+ and K+ turnover, and sensitivity to ouabain, are the same in peripheral T cells and thymocytes. The predominant mechanism of membrane potential generation changes during T lymphocyte differentiation from electrodiffusion in the thymus to electrogenic in peripheral T cells and back to electrodiffusion upon peripheral cell activation.

T cell receptor-mediated recognition of self-ligand induces signaling in immature thymocytes before negative selection.

Shaping of the T cell repertoire by selection during intrathymic maturation involves T cell receptor (TCR) recognition of major histocompatibility complex/self-antigen complexes. In this communication, we studied the ability of minor lymphocyte stimulating (Mls) determinants to act as self-tolerogens in the selection of the T cell repertoire. We demonstrate that unprimed T cells from normal as well as TCR transgenic mice form Mls-specific conjugates with antigen-presenting cells, and that this TCR-ligand interaction leads to elevation of intercellular Ca2+ ([Ca2+]i). Peripheral T cells from TCR transgenic mice expressing receptors specific for self-Mls antigen show no reactivities to Mlsa. However, a proportion of immature thymocytes from these mice show specific binding and strong [Ca2+]i elevation in response to self-antigen-presenting cells, although these thymocytes do not proliferate. This self-reactivity of thymocytes is inhibited by antibodies specific for TCR, CD4, CD8, class II molecules, lymphocyte function-associated antigen 1, and intercellular adhesion molecule 1. These results demonstrate for the first time that before thymic negative selection, immature T cells can specifically interact with cells bearing self-antigen, and suggest that the resulting TCR-dependent signal transduction events provide a basis for negative selection of self-reactive T cells.

Failure of signaling through a chimeric class I-immunoglobulin molecule expressed on the surface of transfected B lymphoma cells and cells of transgenic mice.

To test the possibility that the crosslinkage of molecules expressing a transmembrane region derived from the membrane form of the mu immunoglobulin heavy chain would be sufficient for signal transduction in B cells, a chimeric gene (Kk-mu) consisting of extracellular exons of the class I gene H-2Kk and the transmembrane and cytosolic exons of the mu constant region gene was introduced into WEHI-231 B lymphoma cells and into mouse blastocysts. A protein consistent with the predicted product of the Kk-mu gene was expressed in a transfected cell clone (S18) and in transgenic mice. Crosslinkage of Kk-mu protein with soluble, Sepharose-bound, or dextran-conjugated anti-H-2Kk antibodies failed to induce the accumulation of inositol phosphates or to elevate intracellular calcium concentrations in either S18 cells or B lymphocytes from transgenic mice. Furthermore, crosslinkage of Kk-mu did not inhibit growth of S18 cells or stimulate DNA synthesis by transgenic B cells, in the presence or absence of interleukin-4. The failure of crosslinkage of Kk-mu to initiate detectable intracellular biochemical change or to effect cellular growth suggests that simple crosslinkage of molecules expressing the mu transmembrane region is not sufficient to transduce signals in B cells.

Murine splenic null cell compartment contains distinct haemopoietic subpopulations: enlargement of a myeloid and an undifferentiated subset with the development of splenomegaly in New Zealand black mice.

We have previously reported that non-T, non-B 'null' cells increase with age in New Zealand Black (NZB) mice resulting in splenomegaly. Using a panel of monoclonal antibodies recognizing lineage-specific cell surface antigens we demonstrate four distinct subsets within this null cell compartment: (1) undifferentiated; (2) T lineage with undetectable Thy-1.2; (3) myeloid/erythroid; and (4) a pre-B/plasma cell type. All four subsets also occur in non-autoimmune mice. The frequency of these populations are similar in the young mice of all the strains examined, although the total number of null cells is higher in NZB. The elevation of null cells in young NZB mice is controlled by a single dominant gene in the genetic cross with New Zealand White (NZW) mice and does not appear closely related to the subsequent development of autoimmune disease. The proportion of myeloid/erythroid null cells increases with age in NZB as splenomegaly develops.

Induction of proliferation by high molecular weight B cell growth factor or low molecular weight B cell growth factor is associated with increases in intracellular calcium in different subpopulations of human B lymphocytes.

The activation of resting B cells with anti-surface Ig is associated with transient increases in intracellular calcium. In the present study, we demonstrate that stimulation of B cells which have already been activated by Staphylococcus aureus Cowan I (Sac), with high molecular weight B cell growth factor (HMW-BCGF) or low molecular weight B cell growth factor (LMW-BCGF), but not IL-2, IL-4, or interferon-gamma, is associated with an increase in intracellular calcium, which is modest compared to that seen with anti-Ig (approximately 100 nM vs approximately 400 nM). The increases in intracellular calcium induced by HMW-BCGF or LMW-BCGF occur in distinct but overlapping subpopulations of B cells. Thus, increases in intracellular calcium in human B cells occur not only upon activation but also upon the induction of proliferation by certain (but not all) B cell growth factors. Presumably, the effect of increasing intracellular calcium during the induction of proliferation is to modify a different group of intracellular molecules than those induced during activation.

Intracellular signaling events associated with the induction of proliferation of normal human B lymphocytes by two different antigenically related human B cell growth factors (high molecular weight B cell growth factor (HMW-BCGF) and the complement factor Bb).

High molecular weight B cell growth factor (HMW-BCGF) and the complement component, Factor B, are antigenically related. HMW-BCGF and the physiologic Factor B activation fragment Bb, are both mitogenic for B lymphocytes and compete for binding to the B cell plasma membrane (Peters, M., Ambrus, J. L., Jr., Fauci, A., and Brown, E. (1988) J. Exp. Med. 168, 1225-1235). To understand which second messengers that occur after ligand-receptor interaction are associated with mitogenesis, we have examined the early signaling events after stimulation of activated B cells with these related growth factors. HMW-BCGF but not Bb increased [cAMP]i with a maximum between 45 and 60 min after stimulation. The increase in [cAMP]i was inhibited by indomethacin, suggesting that prostaglandin synthesis is involved in this response. Increase in [cAMP]i induced by HMW-BCGF, cholera toxin, or dibutyryl cAMP was associated with increased expression of the HMW-BCGF receptor, but there was no increase in proliferation of activated B cells when they were stimulated with cAMP agonists other than HMW-BCGF. These data suggest that cAMP is associated with regulation of receptor expression but is neither necessary nor sufficient for induction of proliferation. Both HMW-BCGF and Bb increased cellular levels of diacylglycerol and a water-soluble molecule which could be labeled with both [3H]myoinositol and [14C] glucosamine. However, only HMW-BCGF induced increases in intracellular calcium. Thus, two antigenically related B cell growth factors, HMW-BCGF and Bb, produce overlapping but distinct sets of second messengers after incubation with Sac-activated B cells. Since both induced increases in diacylglycerol and water-soluble inositol, one or both of these molecules may be involved in the proliferative signal generated by the related growth factors. In contrast, the increase in [cAMP]i caused by HMW-BCGF but not Bb is involved in the signal to increase HMW-BCGF receptor expression, but is unrelated to proliferation.

Effects of induced anemia in normal and autoimmune mice.

Normal and autoimmune mice were studied with regard to signals eliciting differentiation and division of bone marrow stem cells. The erythropoiesis induced by anemia following serial bleedings was analyzed in young autoimmune New Zealand Black (NZB) mice and non-autoimmune strains. No difference in the response to the stimulus created by anemia was noted between the strains. After serial bleedings as a stimulus to stem cell proliferation, a five-fold increase in numbers of proliferating spleen cells occurred in both NZB and DBA/2 strains; the increased proliferating spleen cells in both strains were non-lymphoid. The bled animals had decreased percentages of B cells. The production of autoantibodies was not significantly altered by the experimentally induced anemia. In contrast, anti-immunoglobulin activation of resting B cells was increased in response to anemia. Young mice which had experimentally induced anemia had several characteristics in common with old autoimmune NZB mice. Both old NZB mice and young anemic animals had splenomegaly, increased numbers of proliferating spleen cells, decrease in splenic Ly 5+ cells and an increase in splenic colony forming units (CFUs). The anemic normal strains of animals lacked other characteristics of old NZB mice such as hyperimmunoglobulinemia or autoantibody production or elevated CD5+B cell numbers. This work supports the concept that the increase in spleen cell number, proliferating spleen cells, CFUs and the increased percentages of non-Ly-5 cells (which include erythroid precursors) found in the spleens of old NZB mice may in part result from their autoimmune hemolytic anemia.

Heterogeneity of lymphocyte calcium metabolism is caused by T cell-specific calcium-sensitive potassium channel and sensitivity of the calcium ATPase pump to membrane potential.

Calcium management differs in T and B lymphocytes. [Ca2+]i elevation in response to calcium ionophores is up to 10 times greater in T cells than B cells. There is no difference between them in ionophore uptake. T cells, but not B cells, possess a calcium-sensitive potassium channel which produces membrane hyperpolarization at [Ca2+]i above 200 nM. This alters T cell density providing a rapid and easy method of cell separation. In contrast, B cells depolarize when [Ca2+]i is increased. Isolated B cell membrane vesicle ATP-dependent calcium pump activity is higher than T cell vesicles. Membrane depolarization reduces the [Ca2+]i response to ionomycin, most dramatically in T cells because they are hyperpolarized by increased [Ca2+]i. The most likely basis of this behavior is an effect of membrane potential on lymphocyte membrane calcium pump activity. This mechanism provides an explanation of the inhibitory effect of membrane depolarization on T lymphocyte responses.

Fixation and long-term storage of human lymphocytes for surface marker analysis by flow cytometry.

A method to preserve stained human lymphocytes for subsequent cell surface analysis by flow cytometry (FCM) is described. Cells stained with fluorescein isothiocyanate (FITC) and phycoerythrin (PE)-conjugated monoclonal antibodies and then fixed in 1% paraformaldehyde, followed by extensive washing and resuspension in 1% BSA medium, could be stored at 4 degrees C for at least 2 weeks prior to FCM analysis without significant alteration in the light scatter or fluorescence properties of the cells. Furthermore, the method was also suitable for analyzing lymphocytes that express T-cell activation markers in certain disease conditions. In addition, we have identified monoclonal antibody combinations that discriminate different lymphocyte subsets that are satisfactory for multiparameter analysis after 2 weeks of storage. This method should prove useful for enumerating lymphocyte subsets in field study sites remote from flow cytometry laboratories.

Aluminum fluoride induces phosphatidylinositol turnover, elevation of cytoplasmic free calcium, and phosphorylation of the T cell antigen receptor in murine T cells.

Antigen activation of murine T lymphocytes leads to phosphorylation of three subunits of the murine T cell antigen receptor (L.E. Samelson, M.D. Patel, A.M. Weissman, J.B. Harford, and R.D. Klausner. 1986. Cell 46:1083). Two kinases are activated in this process: protein kinase C which leads to phosphorylation of the gamma and, to a lesser extent, the epsilon subunits on serine residues and a tyrosine kinase which phosphorylates the p21 subunit (M.D. Patel, L.E. Samelson, and R.D. Klausner. 1987. J. Biol Chem. 262:5831). We sought to determine whether treatment of these cells with NaF could simulate any of these antigen-induced events. Indeed NaF treatment resulted in breakdown of polyphosphoinositides and production of phosphoinositols. This treatment also resulted in a rise in cytosolic free Ca2+. EGTA failed to block this rise suggesting that NaF liberated intracellular stores of Ca2+. Finally NaF treatment resulted in phosphorylation of the gamma and epsilon chains of the T cell receptor indistinguishable from the effects of phorbol esters. The NaF effect was potentiated by addition of A1Cl3 consistent with the view that the active moiety is A1F4-. The A1F4--induced phosphorylations were abolished in cells in which protein kinase C was depleted by prior treatment with phorbol myristate acetate. All of these observations are compatible with the interpretation that the A1F4- phosphorylation is mediated by protein kinase C. Antigen and anti-receptor antibody-induced receptor serine phosphorylation and phophatidylinositol turnover are blocked by raising intracellular levels of cyclic adenosine monophosphate. In contrast, A1F4--induced effects were insensitive to cyclic adenosine monophosphate.

Molecular events in the induction of a nonresponsive state in interleukin 2-producing helper T-lymphocyte clones.

Exposure of normal interleukin 2 (IL-2)-producing helper T-cell clones to antigen and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide-treated antigen-presenting cells results in proliferative unresponsiveness to subsequent stimulation with antigen and normal antigen-presenting cells. In the present study, we have examined the molecular events that accompany the induction of this unresponsive state. T cells stimulated in this manner failed to produce IL-2, but interleukin 3, interferon-gamma, and IL-2 receptors were partially induced and T-cell receptor beta mRNA was fully induced. Although T-cell unresponsiveness correlated with an IL-2 production defect, addition of IL-2 during the induction phase failed to prevent development of the unresponsive state. The critical biochemical event appeared to be an increase in intracellular calcium. Removal of calcium from the medium prevented induction of the unresponsive state, whereas addition of the calcium ionophore ionomycin induced unresponsiveness as well as all of the related partial activation events. Thus, an increase in intracellular calcium under nonmitogenic conditions appears to initiate an alternative activation program that prevents the T cell from producing IL-2 in response to subsequent normal activation signals. The significance of this in vitro model for tolerance induction in vivo is discussed.

Protein kinase C activation blocks anti-IgM-mediated signaling BAL17 B lymphoma cells.

Short term pretreatment of the B lymphoma, BAL17, with phorbol 12-myristate, 13-acetate (PMA) blocks elevation in inositol trisphosphate (InsP3) and increases in intracellular free calcium concentration ([Ca2+]i) in response to anti-IgM. The inhibition of enhanced InsP3 level is detected at 30 sec after the addition of anti-IgM, the earliest point measured, and is reversed by 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride, an inhibitor of protein kinase C (PKC). The blockade of increased [Ca2+]i by PMA is also observed at the earliest time examined (15 sec), is reversed by 1-(5-isoquinoline-sulfonyl)-2-methylpiperazine dihydrochloride, and is mimicked by dioctanoylglycerol, a physiologic activator of PKC. The enhanced production of inositol phosphates in response to NaF is also blocked in BAL17 cells pretreated with PMA. Extended treatment of BAL17 cells with PMA depletes cellular PKC. Such pretreatment with PMA enhances rather than inhibits increased InsP3 levels in response to anti-IgM and leads to more sustained elevations in [Ca2+]i than in normal BAL17 cells. These results lead us to conclude that PMA-blockade of the response of B cells to anti-IgM represents a disruption of the transmembrane signaling process (desensitization of the signaling pathway) as a result of a PKC-mediated phosphorylation event.

Flow cytometric analysis of murine splenic B lymphocyte cytosolic free calcium response to anti-IgM and anti-IgD.

The accurate measurement of ionized intracellular calcium [Ca++]i in single cells by flow cytometry with the use of a new fluorescent calcium chelator, indo-1, is described. We have developed a dependable in situ calibration technique that indicates a resting [Ca++]i in lymphocytes of 100 nM. The enhanced fluorescence of this probe permits its use at sufficiently low cytoplasmic concentrations that buffering of [Ca++]i transients does not occur. The [Ca++]i response of small resting B lymphocytes to crosslinking of surface antigen receptors by anti-immunoglobulin is heterogeneous. With maximal stimulus, the peak [Ca++]i response occurs in 10 to 20 seconds with most cells reaching levels greater than/1 microM. Mean [Ca++]i falls to between 300 and 800 nM by 100 seconds where it remains for more than 10 min. Anti-delta is a more potent stimulus of increased [Ca++]i than anti-mu in terms of both [Ca++]i level and fraction of B cells responding. Whether this is due to the greater density of surface IgD than IgM, a difference in signal transduction efficiency, or both, is not yet known. Surface immunoglobulin receptors are present in great excess. Less than 3% of surface immunoglobulin is crosslinked at the peak of the [Ca++]i response.

HLA-DR antigens in systemic lupus erythematosus: association with specificity of autoantibody responses to nuclear antigens.

HLA-DR antigens and autoantibodies to the nuclear or cytoplasmic antigens Ro/SSA, La/SSB, Sm, and RNP were determined in North American and Austrian patients with systemic lupus erythematosus (SLE). Analysis of the association of antibodies to these ribonucleic acid (RNA)-protein antigens with HLA-DR antigens showed that HLA-DR3 was related to the presence of anti-Ro/SSA or anti-La/SSB, or both. In contrast, anti-Sm or anti-RNP, or both were associated with HLA-DR4. HLA-DR5 was associated with absence of these autoantibodies. The data extend evidence for the complexity and heterogeneity of SLE. Moreover, they indicate that, in SLE, genes linked to those coding for HLA-DR antigens, are related to the specificity of autoantibody responses rather than to the primary immunological abnormalities of this disorder.