Severe hydralazine-induced lupus presenting as systemic lupus erythematosus.

Despite its long history of untoward side effects of a systemic autoimmune disease, drug-induced lupus can be difficult to recognize because of the disconnect between chronic drug usage and onset of symptoms. In this case, the patient was treated with hydralazine for two years when symptoms were initially reported, but a diagnosis of hydralazine-induced lupus was not considered for another half year. Despite treatment with steroidal and nonsteroidal anti-inflammatory medications during this period, rheumatologic symptoms and signs continued to deteriorate, consistent with the diagnosis of systemic lupus erythematosus. Not until the patient voluntarily discontinued hydralazine did symptoms begin to improve, fully resolving over the subsequent 6-12 months largely in the absence of anti-inflammatory medication. This patient demonstrates that failure to recognize a drug-induced disease etiology can result in substantial worsening of rheumatologic symptoms over the subsequent six months, ultimately satisfying criteria for systemic lupus erythematosus. While symptoms and signs largely normalized, some laboratory abnormalities and occasional arthralgia remained two years after discontinuing hydralazine, suggesting smoldering inflammatory disease.

Complement-fixing properties of antinuclear antibodies distinguish drug-induced lupus from systemic lupus erythematosus.

The immunofluorescence antinuclear antibody (ANA) test has been widely used to monitor autoimmune disease, but its value for diagnostic purposes is compromised by low specificity and high prevalence in disease-free individuals. The capacity of autoantibodies to fix serum complement proteins when bound to antigen is an important effector function because this property is associated with acute and chronic inflammatory processes. The current study evaluates the complement-fixing properties of antinuclear antibodies (CANA) in three well-defined and clinically-related patient groups: systemic lupus erythematosus (SLE), drug-induced lupus (DIL) and drug-induced autoimmunity (DIA). Of 20 patients diagnosed with SLE, 90% displayed complement-fixing ANA while this feature was present in only two of 18 patients with DIL and no patients with DIA without associated disease even though the mean ANA titres were similar among these patient groups. CANA was significantly correlated with anti-Sm activity. Because SLE but not DIL or DIA can be a life-threatening disease associated with complement consumption in vivo, these results demonstrate that measurement of CANA is a diagnostically useful tool and may have immunopathologic implications.

Binding to histone of an anomalous IgG from patients with SLE and drug-induced lupus.

The mechanism of attachment of circulating immune complexes (CIC) to glomerular basement membranes (GBM) in systemic lupus erythematosus (SLE) has not yet been elucidated. One difficulty is that CIC must be strongly cationic for such deposition to occur, which is opposite to the anionic nature of putative DNA-anti-DNA immune complexes (DNA-IC). The strongly cationic histone has been proposed as a potential "planted antigen"; it would decorate the GBM to function as a ligand for DNA in the DNA-IC. However, DNA-IC, aggregated IgG and most of the IgG "anti-histone antibodies" in SLE patient sera bind to histone on a solid phase not through DNA, but through the Fcgamma. Here, we investigated the nature of the anti-histone "antibody" in sera of 18 patients with SLE and 57 with drug-induced lupus (DIL). The binding to nucleosomes of IgG from these patients was mainly pepsin-resistant and F(ab')(2)-dependent, whereas the binding to histone was mainly pepsin-sensitive and Fcgamma-dependent. Surprisingly, after molecular sieving of 12 of these sera, the pepsin-sensitive histone-binding IgG was located mainly in the 150-kDa monomeric IgG peak. The binding to nucleosomes was only in the 150-kDa peak. These findings are consistent with the existence of an anomalous IgG in SLE and DIL sera, capable, like aggregated IgG, DNA-IC and other CIC, of binding to histone-decorated structures. We propose that this anomalous IgG plays an essential role in the pathogenesis of lupus nephritis and other related inflammatory conditions. These observations also explain the large discrepancies in the reports on anti-histone autoantibodies in autoimmune conditions.

The autoimmune response to chromatin antigens in systemic lupus erythematosus: autoantibodies against histone H1 are a highly specific marker for SLE associated with increased disease activity.

This study investigates specificity, sensitivity and concomitant presence of antibodies against histone H1 (H1), nucleosomes (NUC), chromatin (CHR) and dsDNA in patients with systemic lupus erythematosus (SLE), analyses their association with SLE disease activity and characterizes the immunodominant epitope reactivity of anti-H1 antibodies and its relation to SLE disease activity. In a cross-sectional study 394 sera of patients with various rheumatic diseases and healthy subjects were analysed by ELISA for antibodies against H1, NUC, CHR and dsDNA. In addition, a longitudinal analysis was performed that included 121 sequential serum samples derived from 16 SLE patients to assess the relation of these antibodies as well as antibodies to histone H2B to SLE disease activity. To assess epitope reactivity of anti-H1 antibodies overlapping synthetic peptides covering the entire H1 sequence were used. Anti-H1 antibodies yielded a sensitivity of approximately 45% and a specificity of over 98% for SLE, which was comparable to that found for anti-dsDNA antibodies. Anti-CHR and anti-NUC antibodies were of similar sensitivity but slightly (anti-CHR) or considerably (anti-NUC) less specific for SLE (95 and 85%, respectively). The sequential analysis revealed a strong correlation of anti-H1 antibodies with SLE disease activity that was better than the correlation of anti-dsDNA and anti-NUC antibodies, while only weak correlation was found for anti-CHR and anti-H2B antibodies. The immunodominant epitope for anti-HI was localised between amino acids 204 and 218 (pp204-218) and immune reactivity to this epitope also correlated with disease activity. Anti-H1 is a highly specific marker for SLE with a diagnostic value comparable to anti-dsDNA. A positive testing for anti-H1 indicates increased disease activity, as does the appearance of antibodies to its immunodominant epitope pp204-218.

A nondeletional mechanism for central T-cell tolerance.

To be positively selected, immature thymocytes must receive signaling through their T-cell receptor (TCR), and engagement of relatively low-affinity self-peptides permits further T-cell maturation. However, mature T cells no longer overtly respond to such low-affinity antigens, indicating that T cells acquire a higher threshold for activation during thymopoiesis. We wondered whether partial interference in positive selection could produce T cells that respond to the selecting self-peptide. This possibility was tested by injecting procainamide-hydroxylamine (PAHA), a lupus-inducing drug, into the thymus of adult normal mice. Three weeks after the second injection, IgG antichromatin antibodies appeared in the circulation and remained for several months. The murine antichromatin antibodies reacted with the (H2A-H2B)-DNA subnucleosome complex, the predominant specificity in patients with procainamide-induced lupus. In thymus organ and reaggregate cultures, PAHA had no effect on negative selection of T cells with high affinity for a co-present antigen, but acted on CD4+ CD8+ immature T cells as they underwent positive selection. TCR transgenic T cells specific to cytochrome c peptide 88-104 acquired the capacity to respond to the low-affinity analogue at position 99 (lys-->ala) if PAHA was present during their development. PAHA also blocked the capacity of a T-cell line to become anergic after anti-CD3 treatment, suggesting that PAHA prevents the production of negative regulators that accumulate in response to partial signaling through the TCR. These results are consistent with the view that T cells acquire self-tolerance during positive selection, and disruption of this process can result in autoreactive T cells and systemic autoimmunity.

Early cellular events in systemic autoimmunity driven by chromatin-reactive T cells.

In vivo exposure of the thymus of normal mice to procainamide-hydroxylamine, a lupus-inducing drug, causes development of chromatin-reactive T cells. Autoantibodies subsequently appear, but their origin and significance are unknown. The current studies were undertaken to determine the specificities of B cells that respond to chromatin-reactive T cells at the initiation of this autoimmune process. Three days after adoptive transfer of 6 x 10(6) chromatin-reactive T cells, B cells with the capacity to secrete IgM anti-chromatin antibodies were detected in 1/10(6) splenocytes, and these became 10- to 50-fold more numerous if either the donor T cells or the recipient had defective Fas due to the lpr allele. Five days later these mice developed IgG anti-chromatin-secreting B cells at a precursor frequency of 3-6 x 10(-5). B cells with dDNA-binding activity isolated from mice primed in vivo to a complex of methylated pigeon cytochrome c and dDNA could stimulate naive, cytochrome c-reactive T cells in vitro, demonstrating that B cells can internalize dDNA-bound proteins through their dDNA immunoblobulin receptor and can functionally present a T cell epitope. However, no capacity of chromatin for binding anti-dDNA antibodies was detected, and IgM dDNA-specific B cells did not expand when challenged with chromatin-reactive T cells in vivo. The rapid and robust expansion of anti-chromatin-secreting B cells indicates that the normal immune repertoire includes nontolerant autoreactive B cells that respond to strong T cell drive and are readily manifested if Fas-mediated activation-induced cell death is inhibited.

Phagocyte-mediated oxidation in idiosyncratic adverse drug reactions.

The drug-metabolizing capacity of the liver is well known but cannot account for most idiosyncratic adverse drug reactions. Of the extrahepatic sources of reactive drug metabolites, the neutrophil has received the most attention because of its vast numbers and robust oxidizing machinery. Many drugs associated with autoimmunity are susceptible to oxidative transformation by the enzymatic action of myeloperoxidase, a protein released into the extracellular environment when neutrophils are activated. Production of the resulting drug metabolites within lymphoid organs maximizes their immune-perturbing effects. Mechanisms proposed for the initiation of drug-induced blood dyscrasias, hypersensitivity reactions, or lupus-like symptoms center around three views: (1) presentation of the implicated compound in the major histocompatibility complex of antigen-presenting cells via direct binding or after processing as a hapten bound to self-macromolecules, (2) direct cytotoxicity, or (3) interference in the development of T-cell tolerance in the thymus. How participation of reactive drug metabolites in these processes might lead to symptomatic disease is discussed.

Thymus function in drug-induced lupus.

Autoimmunity develops when a lupus-inducing drug is introduced into the thymus of normal mice, but the relevance of this model to the human disorder is unclear in part because it is widely assumed that the thymus is non-functional in the adult. We compared thymus function in 10 patients with symptomatic procainamide-induced lupus to that in 13 asymptomatic patients who only developed drug-induced autoantibodies. T cell output from the thymus was quantified using a competitive polymerase chain reaction that detects T cell receptor DNA excision circles in peripheral blood lymphocytes. Despite the advanced age of the patient population under study, newly generated T cells were detected in all subjects. Although there was no overall quantitative difference between the symptomatic and asymptomatic patients, we found a positive correlation between the level of T cell receptor excision circles in peripheral lymphocytes and serum IgG anti-chromatin antibody activity in patients with drug-induced lupus. The association between autoantibodies and nascent peripheral T cells supports the requirement for T cells in autoantibody production. Our observations are consistent with findings in mice in which autoreactive T cells derived from drug-induced abnormalities in T cell development in the thymus.

Disruption of positive selection of thymocytes causes autoimmunity.

To differentiate into T cells, immature thymocytes must engage, through their antigen-specific T-cell receptor, peptides derived from self proteins presented by cortical epithelial cells in the thymus, a process called positive selection. Despite this requirement for self-recognition during development, mature T cells do not normally show autoreactivity. Mice injected in the thymus with procainamide-hydroxylamine, a metabolite of procainamide, develop autoimmune features resembling drug-induced lupus. Here, we show that when thymocytes undergo positive selection in the presence of procainamide-hydroxylamine, they fail to establish unresponsiveness to low affinity selecting self antigens, resulting in systemic autoimmunity.

The lupus erythematosus cell phenomenon: comparative analysis of antichromatin antibody specificity in lupus erythematosus cell-positive and -negative sera.

OBJECTIVE: To compare and investigate antihistone and antichromatin antibody responses as well as clinical variables in patients with systemic lupus erythematosus (SLE) who were either positive (LEC+) or negative (LEC-) for the lupus erythematosus (LE) cell phenomenon. METHODS: The binding properties of LEC+ and LEC- SLE sera to chromatin-associated nuclear antigens (histones H1, H2A, H2B, H3, H4; complexes of H2A-H2B, [H2A-H2B]-DNA, H1-DNA; total and H1-stripped chromatin; native and denatured DNA) were investigated. In addition, sera from patients with drug-induced lupus (by procainamide, hydralazine, or quinidine), as well as from patients with rheumatoid arthritis and osteoarthritis, were assessed. Enzyme-linked immunosorbent assay was used to detect specific antibody binding. RESULTS: Mirroring the important role of histone H1 in the formation of LE cells, anti-histone H1 reactivity was 8-fold higher in LEC+ sera than in LEC- sera. In addition, reactivities to most of the other antigens tested, i.e., other histones and histone-DNA complexes as well as chromatin and DNA, were significantly higher in LEC+ sera than in LEC- sera. All but 1 serum sample from the patients with drug-induced lupus were negative for LE cell formation as well as for anti-histone H1 reactivity, but displayed high antibody reactivities to histone-DNA complexes, including chromatin. Sera from patients with rheumatoid arthritis and osteoarthritis did not show significant binding to these antigens. When comparing the clinical features of LEC+ and LEC- SLE patients, severe organ involvement, including nephritis and central nervous system involvement, was common in the LEC+ group, but rare in the LEC- group. CONCLUSION: A positive LE cell phenomenon not only correlated with the presence of high anti-histone H1 antibody levels in SLE, but also indicated serologically and clinically active disease with major organ involvement.

Etiology and mechanisms of drug-induced lupus.

Systemic rheumatic symptoms occur with widely different frequencies as a side effect of long-term therapy with some 39 medications currently in use. Because symptoms are nonspecific, subjective, and protean, diagnosis of drug-induced lupus (DIL) requires awareness of this risk of chronic medication. However, laboratory features and the characteristic of full recovery after discontinuing treatment are helpful in differentiating drug-induced from spontaneous lupus or other syndromes. Drug-induced lupus is probably mediated by reactive drug metabolites, not the ingested medications, and susceptibility to neutrophil-mediated oxidative transformation is a property of ten lupus-inducing drugs reported so far. Mechanisms for DIL modeled after drug hypersensitivity reactions are unsupported experimentally and inconsistent with the features of DIL. However, several new lines of investigation using mouse models have opened up promising leads into the origin of autoreactive T cells and disease development in DIL.

Initiation of autoimmunity by a reactive metabolite of a lupus-inducing drug in the thymus.

Drug-induced lupus is a side effect of deliberate ingestion of various medications, but its etiology, underlying mechanisms, and pathogenesis are puzzling. In vivo metabolic transformation of lupus-inducing drugs to reactive products explains how a heterogeneous set of drugs can mediate the same disease syndrome. Evidence has accumulated that drugs are transformed by extracellular oxidation from reactive oxygen species and myeloperoxidase produced when neutrophils are activated, maximizing the in situ accumulation of reactive drug metabolites within lymphoid compartments. The metabolite of procainamide, procainamide hydroxylamine, displays diverse biologic properties, but no apparent autoimmune effect has been observed. However, when procainamide hydroxylamine was introduced into the thymus of young adult normal mice, a delayed but robust autoimmune response developed. Disruption of central T-cell tolerance by intrathymic procainamide hydroxylamine resulted in the production of chromatin-reactive T cells that apparently drove the autoantibody response in the periphery. Drug-induced autoantibodies in this mouse model were remarkably similar to those in patients with procainamide-induced lupus. Therefore, this system has considerable promise to provide insight into the initiating events in drug-induced lupus and may provide a paradigm for how other xenobiotics might induce systemic autoimmunity.

Linkage of immune self-tolerance with the positive selection of T cells.

Development and maturation of antigen-specific T cells take place in the thymus in a process dependent on recognition by the T cell antigen receptor (TCR) of endogenous self-peptides presented by several types of specialized stromal cells. Paradoxically, emerging T cells are not self-reactive, and it is commonly believed that deletion of high avidity autoreactive T cells is the principal mechanism for establishing self-tolerance. However, there is increasing evidence that the positive selection of T cells on self-peptides presented by thymic cortical epithelial cells must be linked with a process that prevents their subsequent activation when the same self-peptides are encountered in the periphery. Consequently, a higher activation threshold is established that can be overcome only with ligands of higher affinity, which would normally be foreign peptides. The molecular basis for this increase in activation threshold is unknown, but observations on differential signaling by peptide analogs, on increased TCR expression during T cell maturation and on energy induction in the absence of costimulation provide promising leads. Linkage of self-tolerance with positive selection is a simple and evolutionary sound explanation for self/non-self discrimination and offers a framework for understanding systemic autoimmunity.

Persistence of autoreactive T cell drive is required to elicit anti-chromatin antibodies in a murine model of drug-induced lupus.

Long-term treatment with procainamide and numerous other medications is occasionally associated with the development of drug-induced lupus. We recently established a murine model for this syndrome by disrupting central T cell tolerance. Two intrathymic injections of procainamide-hydroxylamine (PAHA), a reactive metabolite of procainamide, into (C57BL/6 x DBA/2)F1 mice resulted in the appearance of chromatin-reactive T cells and anti-chromatin autoantibodies. The current study explores in this model the role of autoreactive T cells in autoantibody production and examines why autoantibodies after a single intrathymic drug injection were much more limited in isotype and specificity. Injection of as few as 5000 chromatin-reactive T cells into naive, syngeneic mice induced a rapid IgM anti-denatured DNA response, while injection of at least 100-fold greater number of activated T cells was required for induction of IgG anti-chromatin Abs, suggesting that small numbers of autoreactive T cells can be homeostatically controlled. Mice subjected to a single intrathymic PAHA injection after receiving splenic B cells from an intrathymic PAHA-injected syngeneic donor also developed anti-chromatin Abs, but adoptive transfer of similarly primed T cells or of B cells without intrathymic PAHA injection of the recipient failed to produce an anti-chromatin response. However, anti-chromatin Abs developed after a single intrathymic PAHA injection in Fas-deficient C57BL/6-lpr/lpr mice, suggesting that activation-induced cell death limited autoimmunity in normal mice. Taken together, these results imply that chromatin-reactive T cells produced by intrathymic PAHA created a B cell population primed to somatically mutate and Ig class switch when subjected to a heavy load or second wave of autoreactive T cells.

Antinuclear antibodies (ANA) and complement fixing ANA in systemic connective tissue diseases.

Screening for antinuclear antibodies (ANA) with parallel tests for complement fixing ANA (C-ANA) reveal that C-ANA react either as strongly as or more strongly than ANA in most cases of systemic lupus erythematosus (SLE) and related disorders including CREST syndrome. But sera of drug induced LE and other ANA positive subjects have weak or no C-ANA. (P < 0.0005). Titrations with parallel C-ANA/ANA tests of two cases reveal primarily ANA and less C-ANA reactions in a case of drug induced LE but in CREST syndrome both ANA and C-ANA tests yield elevated titers with stronger C-ANA reactions. These findings point to distinct immunochemical mechanisms in C-ANA and ANA reactions.

Bridging of neutrophils to target cells by opsonized zymosan enhances the cytotoxicity of neutrophil-produced H2O2.

Hydrogen peroxide (H2O2) is a well-established cytotoxic agent released by activated neutrophils into the extracellular environment. However, a maximum of only 5 microM H2O2 was detected in the medium when 10(6) neutrophils/ml were activated with opsonized zymosan (OZ), more than 50-fold lower than the concentration of exogenous H2O2 required to produce equivalent killing of a cell line. In addition PMA-activated neutrophils were noncytotoxic, despite the capacity of PMA to generate two- to fourfold as much H2O2 for five times longer. The basis for this discrepancy was explored. NaN3 increased cytotoxicity to >90% only when neutrophils were activated with OZ due in part to inhibition of myeloperoxidase-mediated hydrolysis of H2O2, while catalase completely prevented cytotoxicity of OZ-activated neutrophils. These results indicate that H2O2 was solely responsible for the observed cytotoxicity. OZ-mediated cytotoxicity was prevented by intermittent agitation of the cultures or by the addition of soluble complement receptor type 1, suggesting that a physical association between neutrophils and target cells mediated by OZ was required to generate a cytotoxic environment. Significant numbers of neutrophil-target cell aggregates were observed by microscopic examination only under low hydrodynamic shear conditions. We conclude that the cytotoxic potency of H2O2 produced by neutrophils activated with OZ was due to a localized high concentration of H2O2 to which the target cells were exposed as a result of their labile adherence to OZ. This phenomenon may reflect a mechanism that neutrophils have acquired for maximizing the antimicrobial power of extracellular oxidants toward microbes that escape phagocytotosis.

A metabolite of the lupus-inducing drug procainamide prevents anergy induction in T cell clones.

Therapeutic treatment with procainamide is occasionally associated with the development of drug-induced lupus. This syndrome has become the prototype for an aseptic systemic autoimmune disease caused by a known environmental agent, but the underlying mechanisms remain puzzling. We explored the possibility that lupus-inducing drugs affect processes involved in T cell tolerance to self-Ag. An in vitro model of anergy using established T cell clones was used to determine whether procainamide or one of its metabolites could prevent development of T cell nonresponsiveness to cognate Ag. Addition of procainamide-hydroxylamine, but not procainamide or its further oxidation products during anergy induction by CD3 engagement, caused a dose-dependent recovery of the capacity of T cells to proliferate and secrete IFN-gamma upon subsequent Ag challenge. Resistance to anergy induction required 2 h of exposure to procainamide-hydroxylamine, and this state remained for 8 h, suggesting that uptake of the drug caused a reversible interference in signaling pathways involved in establishing anergy. We suggest that prevention of anergy induction by procainamide-hydroxylamine may also take place in vivo during establishment of T cell tolerance to self-Ag, thereby allowing the production of autoreactive T cells.

Autoimmunity caused by disruption of central T cell tolerance. A murine model of drug-induced lupus.

A side effect of therapy with procainamide and numerous other medications is a lupus-like syndrome characterized by autoantibodies directed against denatured DNA and the (H2A-H2B)-DNA subunit of chromatin. We tested the possibility that an effect of lupus-inducing drugs on central T cell tolerance underlies these phenomena. Two intrathymic injections of procainamide-hydroxylamine (PAHA), a reactive metabolite of procainamide, resulted in prompt production of IgM antidenatured DNA antibodies in C57BL/6xDBA/2 F1 mice. Subsequently, IgG antichromatin antibodies began to appear in the serum 3 wk after the second injection and were sustained for several months. Specificity, inhibition and blocking studies demonstrated that the PAHA-induced antibodies showed remarkable specificity to the (H2A-H2B)-DNA complex. No evidence for polyclonal B cell activation could be detected based on enumeration of Ig-secreting B cells and serum Ig levels, suggesting that a clonally restricted autoimmune response was induced by intrathymic PAHA. The IgG isotype of the antichromatin antibodies indicated involvement of T cell help, and proliferative responses of splenocytes to oligonucleosomes increased up to 100-fold. As little as 5 microM PAHA led to a 10-fold T cell proliferative response to chromatin in short term organ culture of neonatal thymi. We suggest that PAHA interferes with self-tolerance mechanisms accompanying T cell maturation in the thymus, resulting in the emergence of chromatin-reactive T cells followed by humoral autoimmunity.

Heterogeneity and clinical significance of glomerular-binding antibodies in systemic lupus erythematosus.

We used an ELISA employing extracts of human glomerular basement membrane (GBM) to detect, characterize, and evaluate the clinical significance of glomerular-binding IgG in patients with SLE nephritis. Most patients with SLE nephritis exhibited GBM-binding IgG, although many patients with active nonrenal SLE or symptomatic, drug-induced lupus had similar reactivity, albeit at lower levels. IgG binding to GBM in SLE nephritis patients was decreased by DNase pretreatment of GBM, restored after DNase with nuclear antigens (most notably with nucleosomes), inhibited by exogenous nuclear antigens (particularly nucleosomes), but unaffected by exposure of serum to DNase/high ionic strength. The characteristics of IgG binding to GBM largely paralleled the patients' underlying autoimmune response, which was dominated either by antibodies to DNA/nucleosomes or to nucleosomes alone. Binding of lupus sera to nonrenal extracellular matrix (even with nucleosomes) was not equivalent to GBM. Collagenase pretreatment of GBM variably decreased IgG binding, depending on the level and type of binding. SLE nephritis patients with high levels of GBM-binding IgG exhibited more severe disease clinically, but the same renal histopathology, as patients with lower levels. The level of GBM-binding IgG at presentation did not predict the therapeutic response, but decreased in responders to therapy. In sum, glomerular-binding IgG in lupus nephritis binds to epitopes on chromatin, which adheres to GBM in part via collagen. These autoantibodies appear necessary, but not sufficient, for the development of nephritis, and correlate with clinical rather than histopathologic parameters of disease activity.

Intrathymic injection of polynucleosomes delays autoantibody production in BXSB mice.

T-cell dependent autoimmunization with nucleosomes appears to be an early event in the induction of lupus anti-chromatin antibodies. We investigated this phenomenon by injecting H1-stripped chromatin polynucleosomes into the thymuses of BXSB male lupus-prone mice. In comparison to uninjected controls, the production of IgG antichromatin, anti-native DNA, and anti-denatured DNA were significantly reduced among the injected animals for a period of 8 to 10 weeks. Peripheral T-cells from intrathymic (i.t.)-treated animals showed decreased proliferative responses to polynucleosomes compared to those from uninjected controls. Treatment did not affect T-cell antigen receptor V beta profiles, excluding the possibility that results were due to superantigen-imposed deletions. In situ staining using the TUNEL method demonstrated that generation and phagocytosis of apoptotic material in thymuses of unmanipulated BXSB mice were similar to normal controls. These findings show that polynucleosomes likely comprise the antigens for helper T-cell engagement and induction of lupus-associated anti-chromatin antibodies. Bypassing the underlying defect of T-cell tolerance for polynucleosomal antigens among BXSB mice, by i.t. administration of exogenous polynucleosomes, results in abrogation of autoantibody production.