Production Of Antinuclear Antibodies Research Articles

自己抗体の産生機序とその病原性 こう原病における抗核抗体の産生機序とその病原性

Genesis of antinuclear antibodies is one of the most important immunological abnormalities that are closely associated with clinical manifestations, disease activity, and prognosis of systemic autoimmune diseases. Previous reports indicated that autoreactive T cells have critical roles not only in antinuclear antibody production but also in organ damages. Recently, the possible molecules that are involved in the mechanism of antinuclear antibody production have been determined by using lupus-prone mice. Target treatment against the key molecules associated with autoreactive T cells and B cells may be the useful therapeutic strategy for systemic autoimmune diseases along with the suppression of disease-specific antinuclear antibody production.

Therapeutic effect of CpG-enriched plasmid administration on the tight-skin mouse model of scleroderma

Immunostimulatory CpG motifs can preferentially induce Th1 immune responses and have been applied to treat Th2-dominant disease. In this study, we investigated whether a plasmid with the addition of 20 copies of an immunostimulatory CpG motif (pB-CpG20) might prevent the development of scleroderma-like syndrome in tight-skin (Tsk/+) mice. Administration of pB-CpG20 to Tsk/+mice every 3 weeks starting at the age of 1 week reduced skin thickness and collagen content compared to that of pB or saline. The reduction was long lasting even after halting the treatment. Furthermore, this treatment partially reduced the production of anti-nuclear antibodies although it did not decrease the incidence of lung emphysema. pB-CpG20 increased the number of spleen cells secreting IFN-γ and reduced that of the cells secreting IL-4 in vivo and in vitro compared to saline. These results suggest that repeated administration of a CpG-enriched plasmid can ameliorate scleroderma-like syndrome by biasing Th1 immunity in young Tsk/+mice.

Exposure to mercuric chloride during the induction phase and after the onset of collagen‐induced arthritis enhances immune/autoimmune responses and exacerbates the disease in DBA/1 mice

In susceptible mice, mercuric chloride induces a systemic autoimmune response that is characterized by elevated serum levels of immunoglobulin G1 (IgG1) and immunoglobulin E (IgE), production of anti-nucleolar antibodies (ANolAs) and the formation of renal IgG deposits. We have previously shown that mercury can also enhance immune/autoimmune responses in mouse strains genetically prone to develop spontaneous autoimmune disease. Here, we investigated whether mercury can enhance the severity of murine collagen-induced arthritis (CIA), an inducible (acquired) autoimmune disease that cannot be induced by mercury itself. While mercury administered prior to the induction phase of CIA exerted little, if any, influence, administration of mercury during the induction phase and following onset aggravated the symptoms of this disease and increased the serum levels of IgE and IgG2a antibodies directed against collagen type II (CII). Furthermore, while animals injected with mercury alone exhibited a significant decrease in the ratio of the levels of interferon-gamma (IFN-gamma) to interleukin-4 (IL-4) mRNA in their spleens, this ratio was increased in mice with CIA, with or without administration of mercury. Finally, the production of anti-nuclear antibodies, a hallmark of autoimmunity in response to mercury, was observed in all mice with CIA treated with this heavy metal. Our findings suggest that exposure to mercury during the development of CIA may influence immunological factors in such a way as to synergistically promote disease development.

T- and B-Cell Abnormalities in Systemic Lupus Erythematosus

Systemic lupus erythematosus (SLE) is a complex autoimmune disease characterized by the production of antinuclear autoantibodies and clinical involvement in multiple organ systems. T cells from patients with SLE have been shown to be activated in vivo and provide help to autoreactive B cells. Abnormal expression of key signaling molecules, defective signal transduction pathways, and permanent mitochondrial dysfunction--associated with a significantly increased mitochondrial mass--appear to be the axis of T-lymphocyte dysfunction. Lupus T cells exhibit persistent mitochondrial hyperpolarization (MHP), cytoplasmic alkalinization, increased ROI production, and ATP depletion that mediate enhanced spontaneous and diminished activation-induced apoptosis and sensitize lupus T cells to necrosis. Necrotic, but not apoptotic, cell lysates activate dendritic cells and may account for increased interferon-alpha production, inflammation, and antinuclear antibody production. Recent data indicate that B cells are not merely the passive producers of autoantibodies, but also play a central role in autoimmunity via nonconventional mechanisms, including autoantigen presentation and modulation of other immune cells. This article reviews recent advancements in the understanding of the molecular mechanisms involved in the pathogenesis of lupus autoimmunity and highlights the development of novel therapies in SLE.

Effect of the B cell superantigen protein A from S. aureus on the early lupus disease of (NZB×NZW) F 1 mice

The (NZB×NZW) F 1 mouse develops a spontaneous autoimmune disease process with striking similarities to human systemic lupus erythematosus (SLE). In female (NZB×NZW) F 1 mice, the production of IgG antinuclear antibodies, including antibodies to double-stranded DNA (dsDNA), is associated with the development of a severe immune complex-mediated glomerulonephritis that results in death from renal failure in virtually all animals by 12 months of age. Since B-1 and marginal zone (MZ) cells represent a potential source of pathogenic antibodies and because B cell superantigens have been demonstrated to reduce B-1 and MZ cells in vivo, we tested the effect of repeated injections of the superantigen protein A (SpA) from S. aureus on the disease of this lupus model. We found that weekly intraperitoneal injections of SpA delay the progression of serum anti-DNA IgG and reduce proteinuria early in young female (NZB×NZW) F 1 mice. This superantigen also induced a specific depression in the numbers of peritoneal B-1 cells, as compared to mice treated with a control protein. These results support the role of B-1 cells in the development of the autoimmune disease in this mouse model and suggest that B cell superantigens may be useful in the management of autoimmune conditions.

IgM monomers accelerate disease manifestations in autoimmune-prone Fas-deficient mice

The monomeric form of IgM, also known as low molecular weight IgM, is found in increased concentrations in patients chronically infected with a variety of viral and bacterial pathogens or suffering from various autoimmune diseases. Whether monomeric IgM contributes to the disease process, however, is not known. To address this question, transgenic mice were created that secreted elevated levels of IgM monomers. In normal mice (C57BL/6), the presence of IgM monomers did not alter the composition of the immune system significantly: lymphocyte subsets and serum antibody levels were normal, with the exception of increased levels of IgM due to the presence of the monomers. Immune responses also appeared to be normal. Transgenic mice did develop antinuclear antibodies (ANA) earlier than non-transgenic littermates, but did not develop further indications of autoimmune disease. When the transgene was expressed in the autoimmune-prone strain of mice, B6.MRL- Tnfrsf6 lpr (B6/lpr), these mice developed autoimmune manifestations more rapidly than non-transgenic littermates, including hypergammaglobulinemia, splenomegaly, and ANA production. Transgenic mice also displayed earlier evidence of immune complex deposition in the kidneys. From these results, we conclude that monomeric IgM does not induce autoimmune disease, but its presence can accelerate the onset of autoimmune manifestations in otherwise autoimmune prone animals.

Expression of the Epstein-Barr virus nuclear antigen-1 (EBNA-1) in the mouse can elicit the production of anti-dsDNA and anti-Sm antibodies

Systemic lupus erythematosus (SLE) is a chronic autoimmune disorder characterized by the production of anti-nuclear antibodies. The etiology of SLE is unknown, although several viruses including the Epstein-Barr virus (EBV) have been implicated. An increase in the frequency of EBV infection has been observed in SLE patients relative to normal individuals. Some patients with SLE develop antibodies that recognize a proline rich epitope in the ribonucleoprotein Sm B/B that is similar to an epitope in EBNA-1, a major nuclear antigen of EBV. In the present study we have cloned the EBNA-1 gene under the control of the CMV promoter in the vector pcDNA3. We now report for the first time that expression of the entire EBNA-1 protein in the mouse can elicit the production of IgG antibodies to Sm and to double-stranded DNA (dsDNA). Our data suggest that the anti-Sm response arises as a consequence of antigenic cross-reactivity by anti-EBNA-1 antibodies. These results support a possible association between EBV infection and SLE.

Multiple loci are linked with anti-red blood cell antibody production in NZB mice -- comparison with other phenotypes implies complex modes of action.

The New Zealand Black (NZB) mouse strain is a model of autoimmune haemolytic anaemia (AHA) and systemic lupus erythematosus (SLE), characterized by the production of anti-red blood cell (RBC) antibodies and anti-nuclear antibodies (ANA), respectively. A linkage analysis was carried out in an (NZB x BALB/c) F(2) cross in order to identify loci involved in the production of both anti-RBC IgM and IgG antibodies. These regions of linkage were compared with linkage data to ANA from the same cohort and other linkage analyses involving New Zealand mice. Four previously described NZB loci linked to anti-RBC antibodies were confirmed, and eight novel loci linked to this trait were also mapped: five of which were of NZB origin, and three derived from the non-autoimmune BALB/c background. A comparison between loci linked with anti-RBC antibodies and ANA demonstrated many that co-localize, suggesting the presence of genes that result in the general breaking of tolerance to self-antigen. Furthermore, the observation that some loci were associated only with the anti-RBC response suggests an antigen specific mechanism in addition to a general breaking of tolerance. A locus linked with anti-RBC antibodies and ANA on distal chromosome 7 in this cohort is orthologous to one on the q arm of human chromosome 11, a region linked to AHA and ANA in human SLE.

Molecular mimicry of mitochondrial and nuclear autoantigens in primary biliary cirrhosis

Background & Aims: The mechanism for development of primary biliary cirrhosis (PBC) remains enigmatic, but molecular mimicry has been implicated because of well-known cross-reactivity of human mitochondrial autoantigens and equivalent bacterial antigens. Virtually all patients with PBC have antimitochondrial autoantibodies (AMA), but, interestingly, approximately 50% also manifest antinuclear antibodies (ANA). Methods: To determine whether generation of ANA are due to molecular mimicry of mitochondrial peptides, we established 6 T-cell clones selected by a peptide corresponding to the E2 subunit of mitochondrial pyruvate dehydrogenase complex and analyzed for reactivity to mimicry peptides derived from mitochondrial and nuclear autoantigens, including control sequences. Results: For mitochondrial autoantigens, 1 peptide from the E2 subunit of the pyruvate dehydrogenase complex, 1 peptide from the E2 subunit of the oxo-glutarate dehydrogenase complex, 1 peptide from the E2 subunit of the branched-chain 2-oxoacid dehydrogenase complex, and 1 peptide from the E3-binding protein cross-reacted with these T-cell clones. For the nuclear autoantigens, 5 peptides from gp210 and 1 from Sp100 cross-reacted with these clones. Furthermore, 1 of 3 T-cell clones selected by recombinant gp210 protein reacted with a mimicry peptide corresponding to amino acids 188-201 of gp210, indicating that this part of the protein is a naturally processed immunodominant T-cell epitope. Conclusions: These results demonstrate molecular mimicry between mitochondrial and nuclear autoantigens in PBC and that a mimicry peptide may become an immunodominant T-cell epitope. These data have significance not only for PBC but also for the production of ANA in other disease processes.

EFFECT OF MURINE RECOMBINANT IL-2 ON THE COURSE OF LUPUS-LIKE DISEASE IN (NZB×NZW) F1 Female Mice

ABSTRACTThe exacerbation of pre-existing autoimmune diseases is a potential toxic effect of immunoactive drugs. An increase in the incidence of autoimmune thyroiditis has been noted in patients treated with human recombinant interleukin-2 (rIL-2). In contrast, human rIL-2 tends to protect mice from autoimmunity. As the effects of murine rIL-2 on autoimmunity have not been reported in mice, lupus-prone female (NZB×NZW) F1 mice were treated with 20,000 IU murine rIL-2 intraperitoneally, twice weekly for 13 weeks, beginning at 15 weeks of age. No evidence of an exacerbating effect of murine IL-2 on the lupus disease of (NZB×NZW) F1 mice was observed as no change in the following parameters were seen, namely mean survival time, mean body weight, anti-DNA and antinuclear antibody production. These results show that: 1) like human rIL-2, murine rIL-2 does not exacerbate autoimmunity in mice; 2) the biological effects of human as well as murine rIL-2 in mice differ from those seen with human rIL-2 in man. These latter findings suggest that the selection of the relevant animal species for immunotoxicity studies with recombinant cytokines and derivatives may be less straightforward than previously thought.

Autoimmune uveitis in children: clinical correlation between antinuclear antibody positivity and ocular recurrences

The aim of this study was to identify the correlation between antinuclear antibody (ANA) titre and the onset and clinical course of uveitis in children with juvenile idiopathic arthritis (JIA) or without any other systemic autoimmune disease, i.e., idiopathic uveitis (IU). Twenty-two patients affected by uveitis were examined. Ten had JIA-associated uveitis, 12 had IU. Follow-up ranged from 7 to 101 months. The ANA were titrated three times per year and additionally in case of ocular recurrences. All patients were treated with immunosuppressive drug combination therapy (IDCT). JIA-associated uveitis: ocular recurrences were noted in three ANA-positive patients and in one ANA-negative patient. IU uveitis: ocular recurrences were noted in one ANA-positive and in one ANA-negative patient. No significant rise in ANA titre was noted in either group during uveitis recurrence. (1) ANA had no value in predicting the recurrence of uveitis. (2) IDCT does not influence ANA production.

Differences in the reactivity of CD4+ T-cell lines generated against free versus nucleosome-bound SV40 large T antigen.

Previous results have revealed a strong correlation between polyomavirus BK reactivation and disease activity and antinuclear auto-antibody production in the human autoimmune disease systemic lupus erythematosus. BK virus establishes a latent infection in most humans, and reactivation requires the production of the DNA-binding large T antigen. Experimentally induced expression of the polyomavirus SV40 large T antigen in mice induces both an immune response to large T antigen and autoimmune response to nuclear antigens and antinuclear antibody production. Previous results have indicated that human T-antigen-specific CD4+ T-cell lines are stimulated equally by free, soluble and nucleosome-bound T antigen. This study was designed to determine how antigen processing of nucleosomes containing bound SV40 large T antigen may affect the specificity and response characteristics of experimentally induced T-antigen-specific CD4+ T cells. The results indicated that CD4+ T-cell lines generated from mice immunized with soluble, free T antigen responded very poorly in response to stimulation with T antigen bound to nucleosomes. CD4+ T-cell lines generated from mice immunized with nucleosomes that had bound T antigen in situ responded to both free and nucleosome-bound T antigen. The T-antigen-specific, CD4+ memory T cells induced by latent polyomavirus infections in humans may be uniquely suited to initiate autoimmunity to nuclear antigens upon virus reactivation.

Increased Frequency and Specific Reactivity of Serum Antinuclear Antibodies in Patients with Nonimmune Chronic Idiopathic Neutropenia of Adults

This study describes the frequency of serum organ-specific and organ-nonspecific autoantibodies in 157 patients with nonimmune chronic idiopathic neutropenia of adults (NI-CINA). Forty-two age- and gender-matched volunteers were used as controls. We found that patients with NI-CINA had increased frequency of antinuclear antibodies (ANA) compared to controls (33.1 vs. 9.5%, p = 0.0025), and that ANA positivity inversely correlated with the number of circulating neutrophils (r = –0.2765, p < 0.0001). Speckled pattern of reactivity was seen in 84.6% of ANA-positive patients, and diffuse pattern in the remaining 15.4%. Patients had also increased levels of circulating immune complexes compared to controls (3.30 ± 2.41 vs. 1.70 ± 1.19 µg/ml, p = 0.0042), which inversely correlated with the number of circulating neutrophils (r = –0.2405, p = 0.0154) but not with the titer of ANA positivity. No significant differences were found between the patients and the normal controls in the frequency of positive tests for antibodies to dsDNA, Sm, nRNP, SSA, SSB and Scl-70 antigens, or for parietal cell antibodies, anti-neutrophil cytoplasmic antibodies (ANCA), anti-cardiolipin and anti-thyroid antibodies. Serum levels of rheumatoid factor, C-reactive protein (CRP) and complement factors C3 and C4 ranged within normal limits in the patients studied, but a highly significant correlation was noted between the levels of CRP and ANA positivity (r = 0.3936, p < 0.0001). These findings are suggestive of a chronic inflammation in NI-CINA patients which provides the antigenic stimulus for ANA production, and they further support our previously reported suggestion for the possible involvement of such a low-grade chronic inflammatory process in the pathogenesis of neutropenia in the affected subjects.

Interleukin-13 (IL-13)in autoimmune rheumatic diseases: relationship with autoantibody profile

The production of rheumatoid factor (RF) and antinuclear antibody by B-cells could depend on different cytokines action. We evaluated IL-13 serum levels in 230 patients with autoimmune rheumatic diseases including rheumatoid arthritis (RA) [M/F=22/62; mean age=55.2 (25-76) yrs; mean disease duration = 116 (5-605) months], SLE [M/F=17/97; mean age=38.3 (15-70) yrs; mean disease duration = 77 (1-456) months], Sjogren's syndrome (SS) [M/F=2/50; mean age = 55.2 (26-81) yrs; mean disease duration = 82 (3-540) months], and systemic sclerosis (ScS) [M/F = 1/31; mean age=50.6 (20-73) yrs; mean disease duration = 113 (12-276) months], in order to investigate the relationship of this cytokine with the autoantibody profile. Serum levels of IL-13 (pg/ml) were significantly increased in patients with RA (P < 0.00003), with SLE (P < 0.03), with SS (P < 0.0007), with ScS (P < 0.025) as compared to controls. IL-13 serum levels correlated with those of RF in RA (P < 0.00001), SLE (P < 0.003) and ScS (P < 0.03). IL-13 levels were higher in RA (P < 0.0002), SLE (P < 0.004) and ScS (P < 0.05) patients with RF than patients without RF. SS patients with anti-SSA/Ro antibodies had significantly higher IL-13 levels than SS patients without this autoantibody (P < 0.036). No statistically significative correlation was found between IL-13 levels and any other antinuclear autoantibody or total immunoglobulin levels or main clinical features of each disease. This study suggests that IL-13 may be involved in the pathogenesis of autoimmune rheumatic diseases, with a relevant role on RF production. In SS, the lack of correlation between IL-13 and RF is probably due to the peculiar characteristics of this antibody in the disease. We can conclude that the mechanisms involved in RF synthesis recognise different pathways depending on the underlying autoimmune disease.

Anti-DNA and autoantibodies.

Systemic lupus erythematosus is a prototypic autoimmune disease characterized by antinuclear antibodies (ANAs), including pathogenic specificities to DNA. As shown by recent research, ANA production is a genetically determined process in which self antigens drive B and T cells that have escaped the normal mechanisms of tolerance. Although antibodies can bind isolated protein or nucleic acid species, the in vivo driving antigens likely exist as complexes that have been released from apoptotic cells. The clinical measurement of ANAs, although valuable in assessing diagnosis and prognosis, must nevertheless be interpreted with caution because ANAs, despite their disease associations, can occur in healthy individuals.

Peritoneal and peripheral B-1-cell populations in patients with endometriosis.

The purpose of this study was to investigate the frequency of B-1 cells in the peritoneal cavity and peripheral blood of patients with endometriosis. We examined 31 patients with endometriosis and 14 normal nonpregnant women. Peripheral blood cells and peritoneal exudate cells (PECs) were stained with FITC or PE-labeled anti-CD5/CD19 monoclonal antibodies. Immunofluorescence analysis was performed using a flow cytometer. The significance of differences between the patient and control groups was determined by the non-parametric Mann-Whitney test. There was no significant difference in the percentages of B-1 cells in the peripheral blood of women with and without endometriosis (median, 22.7%; range, 4.7-92.3% vs median, 20.05%; range, 11.1-12.6%, respectively). Endometriosis patients with antinuclear antibodies (ANAs) demonstrated significantly elevated B-1 cells compared to both endometriosis patients without ANAs and normal controls (p < 0.005 and p < 0.05, respectively). Endometriosis patients demonstrated significantly higher B-1 cell populations (B-1 cells/total B-cell ratio) in PECs than did non-endometriosis patients (p < 0.05). The peripheral B-1-cell population in patients with endometriosis is related to ANA production. B-1 cells might play important roles in the development of endometriosis through autoantibody production.

Identification of intervals on chromosomes 1, 3, and 13 linked to the development of lupus in BXSB mice.

To identify intervals containing systemic lupus erythematosus (SLE) susceptibility alleles in the BXSB strain of mice. We analyzed 286 (B10 x [B10 x BXSB]F1) backcross mice for a range of phenotypic traits associated with the development of SLE in BXSB mice. The mice were genotyped using 93 microsatellite markers, and the linkage of these markers to disease was studied by extreme-phenotype and quantitative trait locus analysis. The disease phenotype in these backcross mice was less severe than that in BXSB mice. However, antinuclear antibody production was increased compared with the parental strain. We identified 4 areas of genetic linkage to disease on chromosome 1 (Bxs1-4), 1 on chromosome 3 (Bxs5), and another interval on chromosome 13 which were associated with various aspects of the phenotype. Bxs4 and Bxs5 are located in regions not previously linked to disease in other models of SLE. SLE in the BXSB mouse model has a complex genetic basis and involves at least 5 distinct intervals located on chromosomes 1 and 3. There is evidence that different intervals affect particular aspects of the SLE phenotype.

84 Active DNA or active histone triggering the production of antinuclear antibodies: Inducing the generation of kidney injury in mice

84 Active DNA or active histone triggering the production of antinuclear antibodies: Inducing the generation of kidney injury in mice

Silicone Gel Enhances the Development of Autoimmune Disease in New Zealand Black Mice but Fails To Induce It in BALB/cAnPt Mice

Anecdotal evidence links silicone gel breast implants with the development of autoimmune connective tissue disease in women. To investigate whether silicone gel is capable of directly inducing and/or enhancing the development of autoimmune disease, female BALB/cAnPt (BALB/c) and New Zealand Black (NZB) mice were injected subcutaneously with silicone gel, pristane, a nonmetabolizable substance that can cause plasmacytomas in BALB/c and NZB mice, or saline and monitored for the development of glomerulonephritis and autoantibody production. NZB, but not BALB/c, mice spontaneously develop autoantibodies and an autoimmune hemolytic anemia by 12 months of age. Over a period of 10 months, biweekly screening for proteinuria revealed increases in urinary protein in NZB mice that received multiple injections of either silicone gel or pristane. In contrast, urinary protein was unaffected in identically treated BALB/c mice. Although, silicone gel had no effect on serum titers of anti-erythrocyte antibodies in NZB mice, the hematocrits were significantly decreased. Moreover, silicone gel both increased the concentration of IgM anti-type I collagen antibodies and skewed the immunofluorescent staining pattern of serum autoantibodies on HEp-2 cells. In contrast, silicone gel failed to induce the production of anti-erythrocyte or antinuclear antibodies in BALB/c mice and induced only slight increases in IgG anti-type I collagen antibodies. These results suggest that silicone gel can exacerbate the development of autoimmune disease in autoimmune NZB mice, but fails to induce disease in normal BALB/c mice. This is consistent with several epidemiological studies failing to demonstrate an increase in the incidence of autoimmune disease in women with breast implants. However, because silicone gel was able to exacerbate autoimmune disease in NZB mice, it may play a similar role in the development of autoimmune disease in a small percentage of women who are genetically susceptible to such diseases.

Systemic lupus: basic mechanisms and prospects for specific immunotherapy

Nucleosomes are the primary immunogens that initiate the pathogenic autoimmune response in SLE. The production of pathogenic anti-nuclear antibodies in murine as well as human lupus is mediated by a MHC class-II restricted, cognate interaction between select populations of autoimmune T helper (Th) cells and autoimmune B cells that recognize epitopes in the different molecular components of the nucleosome particle. In the SNF1 model of lupus, we have localized the critical peptide autoepitopes for lupus nephritis-inducing Th cells in the core histones of nucleosomes, at amino acid positions 10–33 of H2B, and 16–39 and 71–94 of H4. Remarkably, the nephritogenic epitopes are located in the regions of histones that are also targeted by lupus autoantibodies, as well as the sites where the histones contact with DNA in the nucleosome, indicating that they are specially protected during antigen processing. The localization of the critical peptide autoepitopes in nucleosomes is a basic step towards defining how the pathogenic Th cells emerge in lupus. In addition, the pathogenic Th cells and B cells of lupus have a regulatory defect in the expression of CD40 ligand (CD40L or gp39), which mediates abnormal co-stimulatory signals sustaining the production of pathogenic autoantibodies. Immunotherapy that specifically blocks the pathogenic T and B cell interaction in lupus can be developed based on these studies.