MHC class II-bound self peptides from autoimmune MRL/lpr mice reveal potential T cell epitopes for autoantibody production in murine systemic lupus erythematosus

The systemic lupus erythematosus-like syndrome in MRL/lpr mice involves high-titered IgG autoantibodies, particularly antinuclear Abs that target histones, DNA, and RNA particles. Although T cell help is required for the generation of antinuclear Abs, the epitopes recognized by such helper T cells are unknown. To address this question, we isolated and sequenced self peptides bound by MHC class II molecules from MRL/lpr mice. We identified a number of peptides that are not seen in similar preparations from nonautoimmune C3H animals. The "abnormal" peptide donors include histone, a protein component of a small nuclear ribonucleoprotein, ribosomal proteins, and RNA processing enzymes. We postulate that the peptides from these donors are T cell epitopes required for the generation of the most frequent antinuclear Abs specificities seen in MRL/lpr mice.     

Co-dominant restriction by a mixed-haplotype I-A molecule (alpha k beta b) for the lysozyme peptide 52-61 in H-2k x H-2b F1 mice

Helper (CD4+) T lymphocytes recognize protein Ag as peptides associated to MHC class II molecules. The polymorphism of class II alpha- and beta-chains has a major influence on the nature of the peptides presented to CD4+ T lymphocytes. For instance, T cell responses in H-2k and H-2b mice are directed at different epitopes of the hen egg lysozyme (HEL) molecule. The current studies were undertaken with the aim of defining the role of mixed haplotype I-A (alpha k beta b and alpha b beta k) molecules in T cell responses to HEL in (H-2k x H-2b)F1 mice, as well as the nature of the immunogenic peptides of HEL recognized in the context of I-A alpha k beta b and I-A alpha b beta k. A series of HEL-reactive T cell lines and hybridomas derived from MHC class II heterozygous (C57BL/6 x C3H F1) mice were established. Their responsiveness to HEL and synthetic HEL peptides was analyzed with the use of L cells transfected with either I-A alpha k beta b or I-A alpha b beta k as APC. Out of 28 clonal T cell hybridomas tested, 13 (46%) only responded to HEL presented by I-A alpha k beta b, 11 (40%) by I-A alpha b beta k (and to a minor extent I-A alpha k beta k), only 4 (14%) were primarily restricted by I-Ak, and none by I-Ab. All the I-A alpha k beta b-restricted T cell hybridomas responded to the HEL peptide 46-61 and to its shorter fragment 52-61, even at concentrations as low as 0.3 nM. As this determinant has been previously defined as immunodominant for I-Ak but not for I-Ab mice, these results suggest a role for the I-A alpha k chain in the selection and immunodominance of HEL 52-61 in H-2k mice. The fine specificity of I-A alpha k beta b-restricted T cell hybridomas for a series of different HEL peptides around the sequence 52 to 61 suggests that peptide 52-61 binds to I-A alpha k beta b with higher affinity than to I-A alpha k beta k. The peptides recognized in the context of I-A alpha b beta k and I-A alpha k beta k were not identified.     

T cells that recognize peptide sequences of self MHC class II molecules exist in syngeneic mice.

T cell reactivity toward self MHC class II molecules has been recognized in syngeneic MLR in a number of studies, where the T cells are believed to recognize the combination of self/nonself peptide and self MHC molecule. We investigated the stimulation of T cell proliferation by synthetic peptides of sequences corresponding to the first polymorphic amino terminal domain of alpha- and beta-chains of self I-A molecules. Both unprimed and primed T cells responded to a number of peptides of alpha 1 and beta 1 domains of self I-Ad molecules. The response was dependent on the presentation of I-Ad peptides by syngeneic APC and was blocked by anti-class II MHC mAb. Upon further investigation it was observed that I-Ad peptides could inhibit the stimulation of Ag-specific MHC class II-restricted T cell hybridoma due to self presentation of peptides rather than to direct binding of free peptides to the TCR, further supporting their affinity/interaction with intact self MHC class II molecules. The peptide I-A beta d 62-78 showed high affinity toward intact self MHC II molecule as determined by the inhibition of Ag-specific T cell stimulation and yet was nonstimulatory for syngeneic T cells, therefore representing an MHC determinant that may have induced self tolerance. Thus we have shown that strong T cell proliferative responses can be generated in normal mice against the peptides derived from self MHC class II molecules and these cells are part of the normal T cell repertoire. Therefore complete tolerance toward potentially powerful immunodominant but cryptic determinants of self Ag may not be necessary to prevent autoimmune diseases.     

H2-Mbeta 1 and H2-Mbeta 2 heterodimers equally promote clip removal in I-A(q) molecules from autoimmune-prone DBA/1 mice

Antigen-presenting cells degrade endocytosed antigens, e.g. collagen type II, into peptides that are bound and presented to arthritogenic CD4(+) helper T cells by major histocompatibility complex (MHC) class II molecules. Efficient loading of many MHC class II alleles with peptides requires the assistance of H2-M (HLA-DM in humans), a heterodimeric MHC class II-like molecule that facilitates CLIP removal from MHC class II molecules and aids to shape the peptide repertoire presented by MHC class II to CD4(+) T cells. In contrast to the HLA-DM region in humans, the beta-chain locus is duplicated in mice, with the H2-Mb1 beta-chain distal to H2-Mb2 and the H2-Ma alpha-chain gene. H2-M alleles appear to be associated with the development of autoimmune diseases. Recent data showed that Mbeta1 and Mbeta2 isoforms are differentially expressed in isolated macrophages and B cells, respectively. The tissue expression and functional role of these heterodimers in promoting CLIP removal and peptide selection have not been addressed. We utilized the human T2 cell line, which lacks part of chromosome 6 encompassing the MHC class II and DM genes, to construct transgenic cell lines expressing the MHC class II heterodimer I-A(q) alone or in the presence of H2-Malphabeta1 or H2-Malphabeta2 heterodimers. Both H2-M isoforms facilitate the exchange of CLIP for cognate peptides on I-A(q) molecules from arthritis-susceptible DBA/1 mice and induce a conformational change in I-A(q) molecules. Moreover, I-A(q) cell-surface expression is not absolutely dependent on H2-M molecules. These data suggest that I-A(q) exhibits a high affinity for CLIP since virtually all I-A(q) molecules on T2 cells were found to be associated with CLIP in the absence of both H2-M isoforms.     

B and T cell responses to the spliceosomal heterogeneous nuclear ribonucleoproteins A2 and B1 in normal and lupus mice

Autoantibodies directed against spliceosomal heterogeneous nuclear ribonucleoproteins (hnRNPs) are a typical feature of rheumatoid arthritis, systemic lupus erythematosus, and mixed-connective tissue disease. With the aim of investigating a potential pathogenic role of these Abs, we have studied the Ab response to A2/B1 hnRNPs in different murine models of lupus. The specificity of anti-A2/B1 Abs was tested with a series of 14 overlapping synthetic peptides covering the region 1-206 of A2 that contains most of the epitopes recognized by patients' Abs. A major epitope recognized very early during the course of the disease by Abs from most of MRL lpr/lpr mice but not from other lupus mice and from mice of different MHC haplotypes immunized against B1 was identified in residues 50-70. This peptide contains a highly conserved sequence RGFGFVTF also present in other hnRNPs and small nuclear ribonucleoproteins. Abs reacting with a second A2 epitope identified in residues 35-55 were detectable several weeks later, suggesting an intramolecular B cell epitope spreading during the course of the disease. We identified several T cell epitopes within the region 35-175 that generated an effective Th cell response with IL-2 and IFN-gamma secretion in nonautoimmune CBA/J mice sharing the same MHC haplotype H-2k as MRL/lpr mice. None of the peptides stimulated T cells primed in vivo with B1. Because Abs to peptide 50-70 were detected significantly earlier than Abs reacting with other A2 peptides and the protein itself, it is possible that within the protein, this segment contains residues playing an initiator role in the induction of the anti-A2/B1 and antispliceosome Ab response.     

A therapeutic peptide in lupus alters autophagic processes and stability of MHCII molecules in MRL/lpr B cells

The P140 phosphopeptide encompassing residues 131–151 of the spliceosomal U1-70K snRNP protein

displays protective properties in lupus patients and MRL/lpr mice. It increases peripheral blood lymphocyte apoptosis via a mechanism involving γδ T cells. After intravenous administration, P140 accumulates in the lungs and spleen. It binds both the HSC70/Hsp73 chaperone and MHC class II (MHCII) molecules, which colocalize in splenic MRL/lpr B cells. Expression of HSC70 and MHCII, which is increased in MRL/lpr splenic B

cells, is diminished after P140 administration. P140 impairs refolding properties of HSC70 and alters expression of stable MHCII molecules in B lymphocytes. In MRL/lpr B cells, P140 increases the accumulation of the autophagy markers p62/SQSTM1 and LC3-II, consistent with a downregulation of autophagic flux. Our study reveals a very unique property of P140 peptide that alters the autophagy pathway leading to a defect of endogenous (auto)antigen processing in MRL/lpr antigen-presenting B cells and a decrease of T cell priming and signaling.     

Role of MHC-linked genes in autoantigen selection and renal disease in a murine model of systemic lupus erythematosus

We previously described a renal protective effect of factor B deficiency in MRL/lpr mice. Factor B is in the MHC cluster; thus, the deficient mice were H2b, the haplotype on which the knockout was derived, whereas the wild-type littermates were H2k, the H2 of MRL/lpr mice. To determine which protective effects were due to H2 vs factor B deficiency, we derived H2b congenic MRL/lpr mice from the 129/Sv (H2b) strain. Autoantibody profiling using autoantigen microarrays revealed that serum anti-Smith and anti-small nuclear ribonucleoprotein complex autoantibodies, while present in the majority of H2k/k MRL/lpr mice, were absent in the H2b/b MRL/lpr mice. Surprisingly, 70% of MRL/lpr H2b/b mice were found to be serum IgG3 deficient (with few to no IgG3-producing B cells). In addition, H2b/b IgG3-deficient MRL/lpr mice had significantly less proteinuria, decreased glomerular immune complex deposition, and absence of glomerular subepithelial deposits compared with MRL/lpr mice of any H2 type with detectable serum IgG3. Despite these differences, total histopathologic renal scores and survival were similar among the groups. These results indicate that genes encoded within or closely linked to the MHC region regulate autoantigen selection and isotype switching to IgG3 but have minimal effect on end-organ damage or survival in MRL/lpr mice.     

A therapeutic peptide in lupus alters autophagic processes and stability of MHCII molecules in MRL/lpr B cells

The P140 phosphopeptide encompassing residues 131-151 of the spliceosomal U1-70K snRNP protein displays protective properties in lupus patients and MRL/lpr mice. It increases peripheral blood lymphocyte apoptosis via a mechanism involving γδ T cells. After intravenous administration, P140 accumulates in the lungs and spleen. It binds both the HSC70/Hsp73 chaperone and MHC class II (MHCII) molecules, which colocalize in splenic MRL/lpr B cells. Expression of HSC70 and MHCII, which is increased in MRL/lpr splenic B cells, is diminished after P140 administration. P140 impairs refolding properties of HSC70 and alters expression of stable MHCII molecules in B lymphocytes. In MRL/lpr B cells, P140 increases the accumulation of the autophagy markers p62/SQSTM1 and LC3-II, consistent with a downregulation of autophagic flux. Our study reveals a very unique property of P140 peptide that alters the autophagy pathway leading to a defect of endogenous (auto)antigen processing in MRL/lpr antigen-presenting B cells and a decrease of T cell priming and signaling.     

Histone deacetylase 9 deficiency protects against effector T cell-mediated systemic autoimmunity

Co-repressor histone deacetylase 9 (HDAC9) plays a key role in the development and differentiation of many types of cells, including regulatory T cells. However, the biological function of HDAC9 in T effector cells is unknown. Systemic autoimmune diseases like lupus, diabetes, and rheumatoid arthritis have dysfunctional effector T cells. To determine the role of HDAC9 in systemic autoimmunity, we created MRL/lpr mice with HDAC9 deficiency that have aberrant effector T cell function. HDAC9 deficiency led to decreased lympho-proliferation, inflammation, autoantibody production, and increased survival in MRL/lpr mice. HDAC9-deficient mice manifested Th2 polarization, decreased T effector follicular cells positive for inducible co-stimulator, and activated T cells in vivo compared with HDAC9-intact MRL/lpr mice. HDAC9 deficiency also resulted in increased GATA3 and roquin and decreased BCL6 gene expression. HDAC9 deficiency was associated with increased site-specific lysine histone acetylation at H3 (H3K9, H3K14, and H3K18) globally that was localized to IL-4, roquin, and peroxisome proliferator-activated receptor-γ promoters with increased gene expression, respectively. In kidney and spleen, HDAC9 deficiency decreased inflammation and cytokine and chemokine production due to peroxisome proliferator-activated receptor γ overexpression. These findings suggest that HDAC9 acts as an epigenetic switch in effector T cell-mediated systemic autoimmunity.     

The Ia.2 epitope defines a subset of lipid raft-resident MHC class II molecules crucial to effective antigen presentation

Previous work established that binding of the 11-5.2 anti-I-A(k) mAb, which recognizes the Ia.2 epitope on I-A(k) class II molecules, elicits MHC class II signaling, whereas binding of two other anti-I-A(k) mAbs that recognize the Ia.17 epitope fail to elicit signaling. Using a biochemical approach, we establish that the Ia.2 epitope recognized by the widely used 11-5.2 mAb defines a subset of cell surface I-A(k) molecules predominantly found within membrane lipid rafts. Functional studies demonstrate that the Ia.2-bearing subset of I-A(k) class II molecules is critically necessary for effective B cell-T cell interactions, especially at low Ag doses, a finding consistent with published studies on the role of raft-resident class II molecules in CD4 T cell activation. Interestingly, B cells expressing recombinant I-A(k) class II molecules possessing a β-chain-tethered hen egg lysosome peptide lack the Ia.2 epitope and fail to partition into lipid rafts. Moreover, cells expressing Ia.2(-) tethered peptide-class II molecules are severely impaired in their ability to present both tethered peptide or peptide derived from exogenous Ag to CD4 T cells. These results establish the Ia.2 epitope as defining a lipid raft-resident MHC class II conformer vital to the initiation of MHC class II-restricted B cell-T cell interactions.     

Selective modulation of CD4+ T cells from lupus patients by a promiscuous, protective peptide analog

A peptide encompassing residues 131-151 of the spliceosomal U1-70K protein and its analog phosphorylated at Ser140 were synthesized as potential candidates for the treatment of patients with lupus. Studies in the MRL/lpr and (NZB x NZW)F1 lupus models have demonstrated that these sequences contain a CD4+ T cell epitope but administration of the phosphorylated peptide only ameliorates the clinical manifestations of treated MRL/lpr mice. Binding assays with soluble HLA class II molecules and molecular modeling experiments indicate that both peptides behave as promiscuous epitopes and bind to a large panel of human DR molecules. In contrast to normal T cells and T cells from non-lupus autoimmune patients, we found that PBMCs from 40% of lupus patients selected randomly and CFSE-labeled CD4+ T cells proliferate in response to peptide 131-151. Remarkably, however, we observed that phosphorylation of Ser140 prevents CD4+ T cells proliferation but not secretion of regulatory cytokines, suggesting a striking immunomodulatory effect of phosphorylated analog on lupus CD4+ T cells that was unique to patients. The analog might act as an activator of regulatory T cells or as a partial agonist of TCR.     

Roles for asparagine endopeptidase in class II MHC-restricted antigen processing

The adaptive immune response depends on the creation of suitable peptides from foreign antigens for display on MHC molecules to T lymphocytes. Similarly, MHC-restricted display of peptides derived from self proteins results in the elimination of many potentially autoreactive T cells. Different proteolytic systems are used to generate the peptides that are displayed as T cell epitopes on class I compared with class II MHC molecules. In the case of class II MHC molecules, the proteases that reside within the endosome/lysosome system of antigen-presenting cells are responsible; surprisingly, however, there are relatively few data on which enzymes are involved. Recently we have asked whether proteolysis is required simply in a generic sense, or whether the action of particular enzymes is needed to generate specific class II MHC-associated T cell epitopes. Using the recently identified mammalian asparagine endopeptidase as an example, we review recent evidence that individual enzymes can make clear and non-redundant contributions to MHC-restricted peptide display.     

Cutting edge: a single MHC anchor residue alters the conformation of a peptide-MHC complex inducing T cells that survive negative selection

We generated transgenic mice that expressed hen egg-white lysozyme (HEL) under a class II MHC promoter. The A7 line expressed HEL with a point mutation in the Asp(52) residue, the main anchor amino acid responsible for the selection of the chemically dominant family of peptides (52-60) by I-A(k) molecules. Mice expressing HEL with Ala(52) were completely unresponsive when immunized with the same protein, i.e., HEL A52. However, the same mice immunized with wild-type HEL elicited T cells that recognized a conformation of the 52-61 core sequence uniquely different between Asp(52) and Ala(52) containing peptides. Importantly, some T cells also recognized the HEL A52 peptide given exogenously but not the same peptide processed from HEL A52 protein. Thus, a core MHC anchor residue influences markedly the specificity of the T cells. We discuss the relevance of these findings to autoimmunity and vaccination with altered peptides.     

Cutting edge: H-2DM is responsible for the large differences in presentation among peptides selected by I-Ak during antigen processing

We quantitated the amounts of peptides from hen egg-white lysozyme presented by I-A(k) molecules in APC lines. The large chemical gradient of presentation of the four hen egg-white lysozyme epitopes observed in cell lines expressing HLA-DM or H-2DM (referred to in this study as DM) was significantly diminished in the T2.A(k) line lacking DM. Differences in levels of presentation between wild-type and DM-deficient APC were observed for all four epitopes, but differences were most evident for the highest affinity epitope. As a result of these quantitative differences in display, presentation of all four epitopes to T cells was impaired in the line lacking DM. The binding affinity of the pool of naturally processed peptides from DM-expressing lines was higher than that from the DM-deficient line. Thus, using a direct biochemical approach in APC, we demonstrate that DM influences the selection of peptides bound to MHC class II by favoring high affinity peptides.     

Diverse repertoire of the MHC class II-peptide complexes is required for presentation of viral superantigens

Among other features, peptides affect MHC class II molecules, causing changes in the binding of bacterial superantigens (b-Sag). Whether peptides can alter binding of viral superantigens (v-Sag) to MHC class II was not known. Here we addressed the question of whether mutations limiting the diversity of peptides bound by the MHC class II molecules influenced the presentation of v-Sag and, subsequently, the life cycle of the mouse mammary tumor virus (MMTV). T cells reactive to v-Sag were found in mice lacking DM molecules as well as in A(b)Ep-transgenic mice in which MHC class II binding grooves were predominantly occupied by an invariant chain fragment or Ealpha(52-68) peptide, respectively. APCs from the mutant mice failed to present v-Sag, as determined by the lack of Sag-specific T cell activation, Sag-induced T cell deletion, and by the aborted MMTV infection. In contrast, mice that express I-A(b) with a variety of bound peptides presented v-Sag and were susceptible to MMTV infection. Comparison of v-Sag and b-Sag presentation by the same mutant cells suggested that presentation of v-Sag had requirements similar to that for presentation of toxic shock syndrome toxin-1. Thus, MHC class II peptide repertoire is critical for recognition of v-Sag by the T cells and affects the outcome of infection with a retrovirus.     

Defective T cell response to presented antigen in autoimmune mice

The effect of the single autosomal recessive gene lpr on antigen presentation was studied. MRL/Mp-lpr/lpr, C3H/HeJ-lpr/lpr, C57BL/6J-lpr/lpr, and their normal congenic partners were investigated. Mice bearing the lpr gene were unable to respond to TNP-KLH when presented by syngeneic antigen-presenting cells. The congenic normal partners gave a brisk response. Mixing experiments demonstrated that the defect resided with the lpr responding T cell and not with the lpr antigen-presenting cell. Antigen-presenting cells from lpr mice were capable of inducing T cell proliferation in normal congenic partners, whereas antigen-presenting cells from normal mice failed to stimulate lpr T cells. This defect was intrinsic to an Lyt-1+2- cell. Pharmacologic restoration was attempted by in vivo and in vitro administration of interleukin 2. However, cells from lpr mice remained unaffected. The relationship of these findings to autoimmunity is discussed.     

Protection of murine systemic lupus by the Ea transgene without expression of I-E heterodimers

A high-level expression of the Ea transgene encoding the MHC class II I-E alpha-chain is very effective in the protection from systemic lupus erythematosus (SLE) in mice. However, it has not been elucidated whether this protection results from the induction or increased expression of I-E heterodimers or from the generation of I-E alpha-chain-derived peptides displaying high affinity for I-A molecules, because previous studies were conducted in lupus-prone mice expressing I-E beta-chains. To address this question, we assessed the protective effect of the Ea transgene in lupus-prone BXSB mice bearing the H2(q) haplotype (i.e., unable to express I-E heterodimers because of a deficiency in I-E beta-chains). We observed that the Ea transgene expression resulted in a marked suppression of the development of SLE in H2(q) BXSB mice despite the absence of I-E expression. The observed protection was not associated with any detectable levels of T cell depletion and regulatory T cell expansion. Significantly, transgenic I-E alpha-chains were substantially expressed on the surface of B lymphocytes and dendritic cells, but not of macrophages, without apparent formation of mixed-isotype heterodimers with I-A beta-chains. Our results demonstrate for the first time that the Ea transgene is able to prevent the development of SLE without induction of I-E heterodimer expression, indicating a critical role of I-E alpha-chains, but not I-E heterodimers, in the Ea transgene-mediated protection from SLE. Taken together, our data favor a model of autoimmunity prevention based on competition for Ag presentation between I-E alpha-chain-derived peptides and autoantigens.     

Defects in the peripheral taste structure and function in the MRL/lpr mouse model of autoimmune disease

While our understanding of the molecular and cellular aspects of taste reception and signaling continues to improve, the aberrations in these processes that lead to taste dysfunction remain largely unexplored. Abnormalities in taste can develop in a variety of diseases, including infections and autoimmune disorders. In this study, we used a mouse model of autoimmune disease to investigate the underlying mechanisms of taste disorders. MRL/MpJ-Fas^lpr/J (MRL/lpr) mice develop a systemic autoimmunity with phenotypic similarities to human systemic lupus erythematosus and Sjögren''s syndrome. Our results show that the taste tissues of MRL/lpr mice exhibit characteristics of inflammation, including infiltration of T lymphocytes and elevated levels of some inflammatory cytokines. Histological studies reveal that the taste buds of MRL/lpr mice are smaller than those of wild-type congenic control (MRL/+/+) mice. 5-Bromo-2′-deoxyuridine (BrdU) pulse-chase experiments show that fewer BrdU-labeled cells enter the taste buds of MRL/lpr mice, suggesting an inhibition of taste cell renewal. Real-time RT-PCR analyses show that mRNA levels of several type II taste cell markers are lower in MRL/lpr mice. Immunohistochemical analyses confirm a significant reduction in the number of gustducin-positive taste receptor cells in the taste buds of MRL/lpr mice. Furthermore, MRL/lpr mice exhibit reduced gustatory nerve responses to the bitter compound quinine and the sweet compound saccharin and reduced behavioral responses to bitter, sweet, and umami taste substances compared with controls. In contrast, their responses to salty and sour compounds are comparable to those of control mice in both nerve recording and behavioral experiments. Together, our results suggest that type II taste receptor cells, which are essential for bitter, sweet, and umami taste reception and signaling, are selectively affected in MRL/lpr mice, a model for autoimmune disease with chronic inflammation.     

IL-21 receptor is required for the systemic accumulation of activated B and T lymphocytes in MRL/MpJ-Fas(lpr/lpr)/J mice

MRL/MpJ-Fas(lpr/lpr)/J (MRL(lpr)) mice develop lupus-like disease manifestations in an IL-21-dependent manner. IL-21 is a pleiotropic cytokine that can influence the activation, differentiation, and expansion of B and T cell effector subsets. Notably, autoreactive CD4(+) T and B cells spontaneously accumulate in MRL(lpr) mice and mediate disease pathogenesis. We sought to identify the particular lymphocyte effector subsets regulated by IL-21 in the context of systemic autoimmunity and, thus, generated MRL(lpr) mice deficient in IL-21R (MRL(lpr).IL-21R(-/-)). Lymphadenopathy and splenomegaly, which are characteristic traits of the MRL(lpr) model were significantly reduced in the absence of IL-21R, suggesting that immune activation was likewise decreased. Indeed, spontaneous germinal center formation and plasma cell accumulation were absent in IL-21R-deficient MRL(lpr) mice. Correspondingly, we observed a significant reduction in autoantibody titers. Activated CD4(+) CD44(+) CD62L(lo) T cells also failed to accumulate, and CD4(+) Th cell differentiation was impaired, as evidenced by a significant reduction in CD4(+) T cells that produced the pronephritogenic cytokine IFN-γ. T extrafollicular helper cells are a recently described subset of activated CD4(+) T cells that function as the primary inducers of autoantibody production in MRL(lpr) mice. Importantly, we demonstrated that T extrafollicular helper cells are dependent on IL-21R for their generation. Together, our data highlighted the novel observation that IL-21 is a critical regulator of multiple pathogenic B and T cell effector subsets in MRL(lpr) mice.     

Treatment of autoimmune MRL/Ipr mice with monoclonal antibody to Thy-1.2: a single injection has sustained effects on lymphoproliferation and renal disease

MRL/Mp-lpr/lpr (MRL/lpr) mice spontaneously develop an autoimmune disease characterized by anti-DNA antibodies, immune-complex glomerulonephritis, and massive proliferation of a distinct population of T cells. The proliferating T cells have the phenotype Thy-1.2+, T200+, Lyt-1+,2-,3-, but Thy-1.2 and Lyt-1 are expressed in abnormally low density. These cells appear to function as helper cells, and neonatal thymectomy prevents both lymphoproliferation and autoimmunity, which suggests that autoimmunity in MRL/lpr mice is secondary to T cell proliferation. We therefore attempted to reduce lymphoproliferation by treating MRL/lpr mice with a single injection of rat monoclonal antibody (MAb) to Thy-1.2 (30-H12, IgG2b). Mice were treated at 8 wk, before the onset of overt disease. We found that MRL/lpr mice were resistant to depletion of circulating T cells (CTC) by anti-Thy-1.2; 0.6 mg of antibody totally depleted CTC from normal mice, but had little or no effect on CTC in MRL/lpr mice. However, treatment with 6 mg of MAb against Thy-1.2 reduced CTC in MRL/lpr mice by over 70%. Moreover, this single treatment markedly reduced the proliferation of CTC over the ensuing 3 mo, despite clearance of the anti-Thy-1.2 from the circulation within 3 wk. Treated mice maintained better renal function than untreated controls, as assessed by levels of blood urea nitrogen (BUN), although anti-DNA antibodies were not significantly reduced. The effect of anti-Thy-1.2 was specific; treatment with rat MAb to the common leukocyte antigen T200 produced only a transient effect on circulating lymphocytes and did not reduce renal disease. The prolonged effects of a single injection of anti-Thy-1.2 suggest that the MAb produces a sustained alteration in immune regulation. The improvement in renal disease is in accord with evidence that autoimmune disease in MRL/lpr mice is T cell dependent. Monoclonal anti-lymphocyte antibodies may be useful in the treatment of autoimmunity.