Peptides from antibodies to DNA elicit cytokine release from peripheral blood mononuclear cells of patients with systemic lupus erythematosus: relation of cytokine pattern to disease duration.

Peptides from VH regions of antibodies to DNA drive immune responses in systemic lupus erythematosus (SLE). We studied peptide-induced cytokine release by peripheral blood mononuclear cells (PBMC) of patients, the influence of peptide concentration, disease characteristics and HLA-D haplotypes. Cells secreting cytokines (IFNgamma, IL-2, IL-4 and IL-10) were measured by ELISPOT in PBMC from 31 patients with SLE and 20 matched healthy controls in response to seven peptides (A-G) from the CDR1/FR2 to CDR2/FR3 VH regions of human anti-DNA MAbs. Disease activity was assessed by SELENA-SLEDAI. HLA-DR and -DQ alleles were determined by molecular typing techniques. PBMC from significantly higher proportions of SLE patients than controls responded to VH peptides by generating IFNgamma and IL-10. Type of cytokines released in response to at least one peptide (D) depended on antigen concentration. Cytokine release was not associated with clinical features of SLE except for disease duration. A shift occurred from IFNgamma, IL-4 and IL-10 production in early disease to IL-4 and IL-10 in late disease (suggesting increasing TH2-like responses over time). Three peptides (B, D, G) were more stimulatory in the SLE patients than controls. Although none of the peptides was restricted by any particular MHC class II allele, among responders there was increased prevalence of HLA- DQB1*0201 and/or DRB1*0301, alleles known to predispose to SLE. Thus, responses to some VH peptides are more frequent in SLE and vary with disease duration. Increased responses in individuals with HLA class II genotypes that predispose to SLE suggest that peptide presentation by those molecules permits brisker peripheral blood cell responses to autoantibody peptides, thus increasing risk for disease.

Treatment with a consensus peptide based on amino acid sequences in autoantibodies prevents T cell activation by autoantigens and delays disease onset in murine lupus.

OBJECTIVE: To test the hypothesis that an artificial peptide, based on an algorithm describing T cell stimulatory sequences from the VH regions of murine IgG antibodies to DNA, is an effective tolerogen in vivo in the (NZB/NZW)F1 (BWF1) mouse model of systemic lupus erythematosus. METHODS: Using proliferative T cell responses to 439 Ig peptides, an algorithm was constructed that describes the amino acid sequences likely to stimulate BWF1 T cells. Stimulatory (pConsensus [pCONS]) or nonstimulatory (pNegative [pNEG]) peptides were synthesized. Groups of 10-week-old (healthy) or 20-week-old (diseased) BWF1 mice received monthly intravenous injections of 1,000 microg of peptide or saline. Ex vivo splenic T cell responses and in vivo clinical effects were measured. RESULTS: Tolerance was induced by pCONS, but not by pNEG, with respect to ex vivo T cell proliferation and T cell help for antibodies to DNA. T cell help for IgG anti-DNA was impaired not only after T cell stimulation by pCONS but also after stimulation by some peptides from nucleosomal and Ro antigens. Treatment with pCONS significantly delayed the onset of nephritis and inhibited increases in the plasma levels of total IgG, IgG antibodies to DNA, nucleosome, cardiolipin, interferon-gamma, and interleukin-4. In contrast, antibody responses to an exogenous antigen were not impaired. Survival was significantly prolonged in both healthy and diseased mice treated with pCONS. CONCLUSION: Induction of immune tolerance in response to treatment with pCONS in autoreactive T cell helper populations is highly effective in delaying the appearance of multiple autoantibodies, cytokine increases, and nephritis in BWF1 mice, and dramatically prolongs survival. A striking effect is the ability of the peptide to tolerize T cell help for anti-DNA that is induced by multiple, structurally unrelated self antigens.

Evidence for multiple mechanisms of polyclonal T cell activation in murine lupus.

Individuals with systemic autoantibody-mediated diseases such as lupus have polyclonal T and B cell activation. Yet, autoantibody production is restricted to certain autoantigens. The mechanisms underlying this phenomenon remain unclear. We propose three potential mechanisms by which autoreactive helper T cell responses diversify to become polyclonal, yet are restricted to certain antigens. First, using a model where self-Ig peptides spontaneously activate T cells and modulate disease in lupus mice, we demonstrate that the numbers of autoantibody-augmenting T helper peptides increased across the Ig molecule as mice aged ("intramolecular determinant spreading"). Secondly, a single T cell hybridoma established from a (NZB x NZW)F1 mouse immunized with one self-Ig peptide recognized several Ig-derived determinants, which had little sequence homology with the immunizing peptide. Such determinant degeneracy can lead to polyclonality. To explore a mechanism for restriction to certain autoantigens, a protein database search was done for homologies with sequences of selected stimulatory Ig peptides. Identical sequences of such determinants were not found in murine proteins other than Ig. These occurred infrequently in nonautoantibody Ig, but quite commonly in lupus-related autoantibodies such as antibodies to DNA, cardiolipin, and erythrocytes. Thus, determinant spreading and degenerate recognition in T cells coupled with recurring use of T cell determinant sequences among autoantibodies result in polyclonality that is restricted to certain autoantigens.

Self Ig peptides that help anti-DNA antibody production: importance of charged residues.

Young NZB/NZW F1 (BWF1) mice develop T cell repertoires that are spontaneously stimulated by peptides derived from the VH regions of BWF1 J558-encoded autoantibodies (autoAb) to DNA, but not to VH region peptides of a J558-encoded antibody to an exogenous antigen. Immunization of young BWF1 mice with selected Ig-derived peptides accelerates anti-DNA production and nephritis, and immune tolerance induction to a combination of these determinants delays anti-DNA production and disease onset. To further characterize this immunoregulatory circuitry, we asked whether this phenomenon of spontaneous T cell activation by VH region peptides is restricted to anti-DNA Ab of the VH J558 family, and what are the charge and structural attributes of these T cell determinants? We studied spontaneous T cell proliferative responses to peptides derived from an autoAb to DNA constructed from VH 7183 and found that it contains several T cell determinants. Both charge and size of certain amino acids (AA) within each peptide seemed to be important. Peptides containing arginine (R) or glutamic acid (E) were more likely to be T cell determinants than peptides without those AA; replacement of charged AA with uncharged AA abolished T cell recognition of a peptide. We previously reported that some Abs to DNA are enriched in R in their VH; pathogenic BWF1 IgG anti-DNA are enriched in positively and negatively charged AA in VH regions. Therefore, we speculate that peptides from natural IgM autoAb may initially activate BWF1 T cells, and as somatic mutations of Ig occur, charged AA introduced into V regions increase the number of T cell determinants, thus favoring upregulation of pathogenic Ab subsets. Therefore, in predisposed individuals, the ability of T cells to recognize more charged T cell determinants in autoAb may be one mechanism promoting development of disease.

Immune tolerance to autoantibody-derived peptides delays development of autoimmunity in murine lupus.

Mechanisms that initiate and maintain autoantibody (autoAb) production in individuals with autoimmune diseases like SLE are poorly understood. Inadequate suppression of autoreactive T cells and/or unusual activation of T and B cells may underlie the persistence of pathogenic autoAbs in lupus. Here, we examine the possibility that in mice with lupus, autoAb molecules may be upregulating their own production by activating self-reactive T cells via their own processed peptides; downregulation of this circuit may decrease autoAb production and delay the development of lupus. We found that before the onset of clinical disease, lupus-prone (NZB/NZW) F1 [BWF1] (but not MHC-matched nonautoimmune mice) developed spontaneous T cell autoimmunity to peptides from variable regions of heavy chains (VH) of syngeneic anti-DNA mAbs but not to peptides from the VH region of an mAb to an exogenous antigen. Tolerizing young BWF1 mice with intravenous injections of autoAb-derived determinants substantially delayed development of anti-DNA antibodies and nephritis and prolonged survival. Thus, in such an autoAb-mediated disease, the presence of autoreactive T cells against VH region determinants of autoAbs may represent an important mechanism involved in the regulation of autoimmunity. Our findings show that tolerizing such autoreactive T cells can postpone the development of an autoimmune disease like SLE.

T cell determinants from autoantibodies to DNA can upregulate autoimmunity in murine systemic lupus erythematosus.

(NZB x NZW) F1 (BWF1) mice develop spontaneous T cell autoimmunity to VH region determinants of syngeneic anti-DNA before the onset of clinical disease. In this study, we characterized the immunogenicity, MHC binding, and lymphokine secretion patterns induced by T cell determinants from the VH region of one such anti-DNA mAb (A6.1) and examined their role in the regulation of autoimmunity. Determinants were identified by proliferation of syngeneic splenic T cells from young, unprimed BWF1 mice in response to overlapping 12-mer peptides representing the entire VH region sequence. Immunization of young BWF1 mice with any of three determinants (A6H 34-45 [p34], A6H 58-69 [p58], and A6H 84-95 [p84]) elicited proliferative responses upon in vitro recall. Upon immunization with the whole A6.1 molecule, however, proliferative responses could be recalled only to the p58 peptide, defining this as immunodominant. The other two peptides (p34 and p84) elicited minimal or no proliferation and could be termed cryptic. Proliferative responses elicited by the cryptic determinants were restricted by a single class II (I-Ed for p34 and I-Au for p84), whereas the immunodominant p58 determinant was restricted by both I-Ed and I-Eu. The cryptic p34 and p84 bound strongly to I-Ed and I-Au, respectively, whereas the immunodominant p58 peptide bound poorly to I-Ed. A6H p84 elicited T cells that secreted lymphokines in a pattern consistent with a Th1-like phenotype, whereas p58 induced a Th2-like cytokine pattern. Immunization with p34 or p84, or adoptive transfer of a p84-reactive T cell line to young BWF1 mice significantly increased IgG anti-DNA levels, accelerated nephritis, and decreased survival. In conclusion, in BWF1 mice, autoreactive T cells recognizing both cryptic and dominant self-determinants on anti-DNA autoantibodies escape deletion or anergy induction. Furthermore, since these cells are spontaneously activated before the onset of clinical disease, they may be involved in the development of the autoimmune process.

Murine and human antibodies to native DNA that cross-react with the A and D SnRNP polypeptides cause direct injury of cultured kidney cells.

Murine monoclonal and human affinity-purified Abs to native DNA (anti-nDNA) that cross-react with the A and D SnRNP polypeptides were analyzed for direct injurious effects against cultured pig kidney (PK15) cells under ordinary cell-culture conditions. Of the two murine nephritogenic Abs derived from NZB/NZW F1 mice (BWds1 and BWds3), BWds1 initially bound to the cell surface and subsequently penetrated into cells localizing in nuclei and cytoplasm. BWds3 was consistently and abundantly associated with the surface of live cells without penetration. In the presence of rabbit C, BWds3 caused massive cell lysis (85% dead cells) whereas BWds1 had only a modest lytic effect (24% dead cells). One of the nonpathogenic murine anti-nDNA Abs (5GD5) that did not cross-react with the A and D polypeptides showed no interaction with PK-15 cells and had no injurious effects. Affinity-purified autoantibodies to nDNA isolated from two SLE patients with high anti-nDNA titers and clinically active lupus nephritis showed properties similar to the murine mAbs. They both strongly cross-reacted with the A and D SnRNP polypeptides and interacted with live PK-15 cells. One of them (Cr) penetrated into live cells and localized within cytoplasm and nuclei whereas the other (Pe) bound mostly to the cell surface and caused significant cell lysis in the presence of C. Results of this study suggest that the nephritogenic murine anti-nDNA as well as subpopulations of human anti-nDNA Abs could exert their injurious influence through direct interactions with kidney cells using two different pathogenic mechanisms (i.e., C-mediated cytotoxicity and potential cell cycle dysfunctions. Interestingly, cross-reactivity of anti-nDNA Abs with the A and D SnRNP polypeptides appears to be a prerequisite for their direct pathogenicity.

Comparison of pathogenic and non-pathogenic murine antibodies to DNA: antigen binding and structural characteristics.

Three pathogenic and two non-pathogenic NZB/NZW F1 mAbs to DNA were compared. Pathogenicity was defined as the ability to induce nephritis in BALB/c mice. All mAbs were IgG2a or 2b, had high avidity for double-stranded DNA and fixed complement well. All three pathogens expressed idiotype IdGN2. Mice receiving pathogenic mAbs (compared with non-pathogenic) had more glomerular IgG deposits. The unique properties of two of the pathogens were: strong homogeneous staining of Hep-2 nuclei and the ability to bind (i) nucleosomes, (ii) histone (after mAb complexed with DNA), (iii) heparan sulfate in renal basement membranes (after complexing with DNA/histone) and (iv) nuclei in vivo. Comparison of nucleotide and amino acid sequences of the V regions of heavy and light Ig chains showed use of multiple VHDJH and V kappa J kappa gene families, with representation of several anti-DNA 'families' described by others. Arginine (R) occurred in the CDR2 or CDR3 of VH chains in all pathogens; R was absent in the CDRs of VH chains of non-pathogens. Positively and negatively charged AA were more frequent in VH CDR of pathogens than of non-pathogens. We hypothesize that the tertiary structure of mAbs determined by VH CDR regions permits stronger binding to negatively charged antigens (DNA and heparan sulfate) and to positively charged molecules (histone) in pathogens compared with non-pathogens.

Lupus autoantibodies to native DNA cross-react with the A and D SnRNP polypeptides.

Antibodies to native DNA (nDNA) in sera from patients with systemic lupus erythematosus have been found to frequently correlate with antibodies to the A and D SnRNP proteins measured in Western blot assays. 40 of 54 SLE (74.1%) sera with anti-nDNA bound to A and D proteins, while 9 of 113 sera (8%) without anti-nDNA bound the A and D proteins, P < 10(-8) by Fisher's exact test. Antibodies to nDNA correlated closely with anti-A and anti-D in seven of eight patients followed sequentially, r = 0.7865. Nine human polyclonal anti-nDNA populations were isolated from DNA cellulose columns. Seven reacted equally with A and D, and two reacted predominantly with D. Two of three murine monoclonal anti-DNA antibodies isolated from NZB/NZW F1 hybrid mice bound A and D equally in Western blot with a titer > 1/40,000. These reactions were directed to the unfolded A and D proteins measurable in Western blot since these monoclonals (and several of the human anti-nDNA populations) failed to react with native U1RNP in ELISA or in RNA immunoprecipitation experiments. These newly recognized cross reactions of anti-nDNA may amplify the immune response to DNA and be part of the original immunogenic drive.

A peptide derived from an autoantibody can stimulate T cells in the (NZB x NZW)F1 mouse model of systemic lupus erythematosus.

OBJECTIVE: To assess the ability of peptides derived from anti-DNA to stimulate syngeneic T lymphocytes and influence lupus in (NZB x NZW)F1 (NZB/W) mice. METHODS: We synthesized (by Geysen pin method) overlapping 12-mer peptides recapitulating the amino acid sequence of the VH region of a nephritogenic monoclonal IgG2a anti-DNA antibody (A6.1) from an NZB/W mouse. Splenic T cells were cultured with the peptides; multiple experiments assayed 12-mer and 16-mer peptides which contained a triple-base motif (KFKGK). We immunized 20-week-old NZB/W mice with the 12-mer and evaluated them for evidence of nephritis and for quantities of antibodies in plasma and glomeruli. RESULTS: Three clusters of peptides caused proliferation of spleen cells from young, nonimmunized mice. Both the 12-mer FYNQKFKGKATL and the 16-mer GDTFYNQKFKGKATLT peptides stimulated purified T cells. The KXKXK motif occurs in 15% of murine Ig VH (NBRF protein database), compared with 100% (6 of 6) of NZB/W anti-DNA monoclonal antibody. Immunization with the 12-mer peptide increased plasma levels of IgG, anti-DNA, and immune complexes, and levels of anti-DNA in glomeruli; nephritis was accelerated. CONCLUSION: NZB/W anti-DNA contain peptide sequences in their VH regions that stimulate self-T cells. At least one motif is frequent in NZB/W anti-DNA. If some activated T cells provide help, this mechanism may contribute to sustained up-regulation of autoantibodies in murine lupus.

Idiotypic characteristics of immunoglobulins associated with human systemic lupus erythematosus. Association of high serum levels of IdGN2 with nephritis but not with HLA class II genes predisposing to nephritis.

Serum levels of IdGN2 (an idiotype enriched in nephritogenic antibodies), IdX (an idiotype not enriched in nephritogenic antibodies), IgG, and anti-DNA were measured in 23 Caucasian patients with lupus nephritis, in age- and sex-matched lupus patients without nephritis, and in similarly matched healthy individuals. Serum levels of IdGN2 were significantly higher in the patients with lupus nephritis than in those without, and they were higher in all lupus patients compared with the healthy control subjects. However, the same observations were true for serum levels of IdX. There were significant positive correlations between the serum levels of IgG, IdGN2, IdX, and anti-DNA. HLA typing at the DR and DQ loci was performed in 105 lupus patients of different races (Caucasian, black, and Asian/Polynesian/Filipino). Serum levels of IdGN2 in 83 of these individuals did not correlate with any of the HLA class II haplotypes currently known to predispose to lupus nephritis. We conclude that the high serum levels of IdGN2, which are characteristic of some patients with lupus nephritis, may often result from polyclonal B cell activation rather than from idiotype-specific upregulation associated with one or more of the class II genes that predispose to nephritis in this disease.

Structural characteristics of the variable regions of immunoglobulin genes encoding a pathogenic autoantibody in murine lupus.

We have studied several monoclonal anti-double-stranded (ds) DNA antibodies for their ability to accelerate lupus nephritis in young NZB X NZW F1 female mice and to induce it in BALB/c mice. Two identified as pathogens in both strains have characteristics previously associated with nephritogenicity: expression of IgG2a isotype and IdGN2 idiotype. Both pathogenic antibodies used the combination of genes from the VHJ558 and VK9 subfamilies. Two weak pathogens failed to accelerate nephritis in young BW mice, but induced lupus nephritis in BALB/c mice. They both express IdGN2; one is cationic and an IgG3, the other is an IgG2a. Additional MAbs (some IgG2a, one IdGN2-positive) did not accelerate or induce nephritis. We have cloned and sequenced the variable regions of the immunoglobulin genes of one pathogenic autoantibody. No unique V, D, or J gene segments and no evidence of unusual mechanisms in generating diversity were used to construct this antibody. These data argue against use of unique abnormal Ig genes by systemic lupus erythematosus individuals to construct pathogenic autoantibody subsets. Instead, the major abnormality may be immunoregulatory.

Idiotypic characteristics of immunoglobulins associated with systemic lupus erythematosus. Studies of antibodies deposited in glomeruli of humans.

Indirect immunofluorescence with monoclonal antibodies to 6 different idiotypes was used to characterize immunoglobulins deposited in the glomeruli of renal biopsy samples from 32 patients with systemic lupus erythematosus (SLE) and 19 patients with nonlupus immune glomerulonephritis. IdGN2 was present in 75% of the biopsy specimens from SLE patients and in 6% of those from patients with non-lupus nephritis; IdGN1 occurred in 38% and 6%, respectively. The other idiotypes were not increased in biopsy samples from patients with SLE. Deposition of IdGN2 was associated with a subendothelial location of Ig and proliferative changes in the glomeruli. In studies of glomerular eluates from 4 immunosuppressed SLE patients, an average of 26% of total Ig and 37% of anti-DNA was composed of IdGN2. Compared with IdGN2- immunoglobulin, IdGN2+ immunoglobulin was enriched in IgG1 in all 4 eluates, and was enriched in high-avidity anti-DNA in 2 eluates. IdGN2 is a marker of antibody subsets that are characteristic of SLE and are associated with severe lupus nephritis.

Pathogenic subsets of antibodies to DNA.

Among the autoantibodies that are known to play a role in the pathogenesis of autoimmune diseases, antibodies to DNA (anti-DNA) have been the subject of much study. Several interesting observations have resulted. The ability to make antibodies that bind DNA is not abnormal. Normal mice and humans can produce antibodies that bind DNA. On the other hand, large quantities of antibodies to DNA are found in the sera of patients with systemic lupus erythematosus (SLE), and complement-fixing antibodies to double-stranded (ds) DNA cause some of the tissue lesions, especially glomerulonephritis (GN). Why, then, do some individuals make anti-DNA that deposits in glomeruli, skin, and other tissue, resulting in organ damage? It is likely that disease results from a combination of several factors--ability to make pathogenic antibody subsets, inability to downregulate those subsets, and "tissue susceptibility" to injury from those antibodies and their immune complexes. This chapter will focus on the characteristics of pathogenic antibody subsets and their regulation.

T cell up-regulation of B cells via their idiotypes contributing to the development of systemic lupus erythematosus. A hypothesis.

A mechanism for sustained production of pathogenic autoantibody subsets in patients and mice with systemic lupus erythematosus may be centered on selective up-regulation of B cells bearing certain idiotypes. Public idiotypes are characteristic of some autoantibodies, including anti-DNA. Evidence is reviewed that suggests that immunoglobulins bearing certain public idiotypes, such as IdGN2, contain autoantibody subsets that are nephritogenic in human systemic lupus erythematosus and in New Zealand black/New Zealand white F1 mice. Up-regulation of such cells could promote development of nephritis. Work from several laboratories has shown that production of immunoglobulin G antibodies to DNA depends upon T cell help. In New Zealand black/New Zealand white F1 mice, cloned T cells are dominated by autoreactive cells that produce B cell growth factors. We suggest that this sustained release of B cell growth factors combined with selection by T helper cells for B cells bearing IdGN2 are a major mechanism for sustained up-regulation of nephritogenic subsets of autoantibodies.

Allogeneic MHC antigen requirements for lupus-like autoantibody production and nephritis in murine graft-vs-host disease.

A graft-vs-host (GVH) reaction of parental T cells in allogeneic F1 mice can lead to an autoimmune disease resembling human SLE. We analyzed the contribution of MHC genes to the development of IgG antinuclear antibody production and immune complex glomerulonephritis in MHC-congenic F1 recipients. DBA/2 T cells elicited IgG antibodies to histone, ssDNA, and dsDNA in all histoincompatible F1 recipients that were studied. The anti-DNA antibody responses were quantitatively similar among the F1 combinations and displayed comparable IgG2a subclass and cationic charge characteristics. In contrast, severe renal disease was manifested only in F1 mice that expressed H-2b encoded class II gene products. Disease susceptibility was associated with a decrease in circulating anti-DNA antibodies and a characteristic localization of immune complexes in the glomeruli. The data suggest that the production of potentially pathogenic IgG anti-nuclear antibodies is not sufficient for the development of renal disease in GVH-induced lupus. Thus, another event separate from autoantibody production is MHC dependent and appears to be critical for the formation and/or deposition of pathologic immune complexes.

Idiotypic spreading promotes the production of pathogenic autoantibodies.

We propose that a major immunoregulatory abnormality in murine and human autoantibody-mediated disease is idiotypic spreading. By this mechanism, B cells with the genetic information to produce immunoglobulin (Ig) bearing certain public idiotypes (Ids) are selectively upregulated, probably by Id-recognizing helper T cells. The model in which we are testing the hypothesis is systemic lupus erythematosus (SLE) in humans and NZB/NZW F1 (BW) female mice. Recent experiments have shown that the number of public Ids expressed on the Ig of nephritic BW mice is quite restricted. IdX is the dominant Id on serum Ig; IdGN1 and IdGN2 are also common. All three Ids were initially derived from spontaneous antibodies to DNA. Together the three are present on 85% of the total Ig repertoire. Such restriction suggests idiotypic spreading. In glomerular Ig deposits from nephritic BW mice, IdGN1 and IdGN2 are found on 45% of the total Ig: IdX is present in minute amounts. Furthermore, suppression of IdGNs by administration of anti-IdGN1 to BW mice resulted in significant delay in the onset of nephritis, but the IdGNs escaped from control and eventually caused a fatal nephritis. Finally, studies of glomerular Ig deposits in renal biopsies of patients with SLE have shown that IdGN2 dominates such Ig, being present in 76% of renal biopsies from SLE patients and in 6% from patients with non-lupus immune nephritis. Therefore, we have concluded that IdGN1 and IdGN2 are markers of nephritogenic subsets of autoantibodies and are probably the products of idiotypic spreading most likely to cause disease. Finally, after a review of recent experiments suggesting the dominance of autoreactive, Mossman Type 2 T helper cells in nephritic BW mice, it is hypothesized that autoreactive, IdGN-recognizing helper T cells may be central to the sustained upregulation of pathogenic autoantibodies in murine and human SLE.

Idiotype regulatory networks promote autoantibody formation.

There is good evidence that the idiotypic network of the immune system can be implicated in the synthesis of pathogenic subsets of autoantibodies. The individual with systemic lupus erythematosus must have the immunoglobulin gene information which permits synthesis of those idiotypes, helper T populations which drive or select for the B cells producing them, and inadequate mechanisms to suppress those activated effector cells.

Idiotype restriction in murine lupus; high frequency of three public idiotypes on serum IgG in nephritic NZB/NZW F1 mice.

Antibodies to self-antigens are characteristic of several human and murine autoimmune diseases. Subsets of those autoantibodies cause organ damage in some instances, such as IgG antibodies to DNA in human and murine systemic lupus erythematosus (SLE). Our experiments in the NZB/NZW F1 (BW) female mouse model of SLE were designed to define idiotypic (Id) structures on antibodies to DNA in attempts to distinguish pathogens from nonpathogens within the anti-DNA population. Two important findings emerged. First, the number of public Id expressed became relatively restricted as the mice aged, with three such Id (IdX, IdGN1 and IdGN2) dominating and accounting for 30 to 95% of the total serum IgG in all individual nephritic mice studied, and 81 to 86% of the total IgG in serum pools from 30-wk-old nephritic mice. Second, IdGN1 and IdGN2 constituted approximately 50% of the IgG deposited in glomeruli of nephritic mice; IdX was present in negligible quantities in glomeruli, whereas it was usually the most frequent Id in BW serum. These latter findings suggested that pathogens and nonpathogens can be distinguished by their idiotypy in this animal model. The finding of relative Id restriction suggests the occurrence of an idiotypic "spreading" phenomenon, in which a regulatory process appears as BW mice age that results in repeated selection and expansion of this small number of Id, one group of which, the IdGN, is pathogenic. This process was further suggested in experiments in which IdX was suppressed by administration of anti-IdX; the "escape" antibodies to DNA appearing after suppression of IdX were composed largely of IdGN1 and IdGN2, without a major contribution from Id-negative mutants. Defining the basis of this Id spreading or restriction phenomenon may provide important information regarding the pathogenesis of this autoimmune disease.