Association between polyclonal B cell activation and the presence of autoantibodies in mice infected with Yersinia enterocolitica O:3

Eight-week old conventional female Swiss mice were inoculated intravenously with Yersinia enterocolitica O:3. A second group of normal mice was used as control. Five mice from each group were bled by heart puncture and their spleens were removed for spleen cell collection on the 3rd, 5th, 7th, 10th, 14th and 21st day after infection. Immunoglobulin-secreting spleen cells were detected by the isotype-specific protein A plaque assay. Total immunoglobulin levels were determined in mouse serum by single radial immunodiffusion and the presence of autoantibodies was determined by ELISA. We observed a marked increase in the total number of cells secreting immunoglobulins of all isotypes as early as on the 3rd day post-infection and the peak of secretion occurred on the 7th day. At the peak of the immunoglobulin response, the total number of secreting cells was 19 times higher than that of control mice and most immunoglobulin-secreting cells were of the IgG2a isotype. On the 10th day post-infection, total serum immunoglobul in values were 2 times higher in infected animals when compared to the control group, and continued at this level up to the 21st day post-infection. Serum absorption with viable Y. enterocolitica cells had little effect on antibody levels detected by single radial immunodiffusion. Analysis of serum autoantibody levels revealed that Y. enterocolitica infection induced an increase of anti-myosin and anti-myelin immunoglobulins. The sera did not react with collagen. The present study demonstrates that Y. enterocolitica O:3 infection induces polyclonal activation of murine B cells which is correlated with the activation of some autoreactive lymphocyte clones.

Presence of autoantibodies against HeLa small nuclear ribonucleoproteins in chagasic and non-chagasic cardiac patients

We detected anti-human small nuclear ribonucleoprotein (snRNP) autoantibodies in chagasic patients by different immunological methods using HeLa snRNPs. ELISA with Trypanosoma cruzi total lysate antigen or HeLa human U small nuclear ribonucleoproteins (UsnRNPs) followed by incubation with sera from chronic chagasic and non-chagasic cardiac patients was used to screen and compare serum reactivity. Western blot analysis using a T. cruzi total cell extract was also performed in order to select some sera for Western blot and immunoprecipitation assays with HeLa nuclear extract. ELISA showed that 73 and 95% of chronic chagasic sera reacted with HeLa UsnRNPs and T. cruzi antigens, respectively. The Western blot assay demonstrated that non-chagasic cardiac sera reacted with high molecular weight proteins present in T. cruzi total extract, probably explaining the 31% reactivity found by ELISA. However, these sera reacted weakly with HeLa UsnRNPs, in contrast to the chagasic sera, which showed autoantibodies with human Sm (from Stefanie Smith, the first patient in whom this activity was identified) proteins (B/B', D1, D2, D3, E, F, and G UsnRNP). Immunoprecipitation reactions using HeLa nuclear extracts confirmed the reactivity of chagasic sera and human UsnRNA/RNPs, while the other sera reacted weakly only with U1snRNP. These findings agree with previously reported data, thus supporting the idea of the presence of autoimmune antibodies in chagasic patients. Interestingly, non-chagasic cardiac sera also showed reactivity with T. cruzi antigen and HeLa UsnRNPs, which suggests that individuals with heart disease of unknown etiology may develop autoimmune antibodies at any time. The detection of UsnRNP autoantibodies in chagasic patients might contribute to our understanding of how they develop upon initial T. cruzi infection.

Serum titres of anti-glutamic acid decarboxylase-65 and anti-IA-2 autoantibodies are associated with different immunoregulatory milieu in newly diagnosed type 1 diabetes patients

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Production of autoantibodies associated with polyclonal activation in Yersinia enterocolitica 0:8-infected mice

Polyclonal lymphocyte stimulation is one of the immunomodulatory mechanisms induced by arthritogenic pathogens. In this study we examined the polyclonal activation potential of a virulent strain of Y. enterocolitica serotype O:8 (WA 2707+) and its plasmidless isogenic pair (WA 2707-). SPF Swiss mice were infected intragastrically and spleen cells were obtained on days 7, 14, 21, 28, 35 and 42 after infection. The number of cells secreting nonspecific immunoglobulins of IgG, IgM and IgA isotypes was determined by the ELISPOT technique. The presence of serum-specific antibodies was investigated by ELISA and the presence of autoantibodies by dot-blot assay. Although the patterns of infection of the two bacterial strains were almost the same, only the animals infected with the virulent strain presented clinical anomalies. Neither arthritic nor inflammatory signs were observed in the joints of the infected animals. The greatest activation observed was that of the nonspecific IgM-secreting cells, and their peak of secretion occurred between the 28th and the 42nd day after infection, for both strains of Y. enterocolitica O:8. Only the animals infected with the virulent strain (WA 2707+) produced IgG-specific antibodies in the serum, from the 28th day after infection. The serum of animals infected with either strain showed reactivity to all the autologous constituents tested, mainly on the 28th and 42nd day after infection. It was concluded that infection of mice with either the virulent strain of Y. enterocolitica O:8 or with its plasmidless isogenic pair resulted in the polyclonal activation of the splenic B lymphocytes including some autoreactive clones.