Complement subcomponent C1q stimulates Ig production by human B lymphocytes

The regulation of Ig production by human B lymphocytes is a complex process involving interactions among B cells, APC, T lymphocytes and soluble factors including activation, growth, and differentiation factors. Components of the complement system, including C3a, C3b, C3d, and C5a, have been shown to influence various stages in this process. In this study, we demonstrate that the C1q subcomponent of complement binds to both small resting and large activated B cells and stimulates immunoglobulin production by Staphylococcus aureus Cowan-activated tonsillar B lymphocytes. This effect is present whether C1q is added to the B cells either at the beginning or near the end of a 7-day culture period and is not associated with enhancement of proliferation. The C1q stimulation of Ig production is, however, associated with increased steady state levels of mRNA for the mu Ig H chain. Furthermore, C1q stimulated IgM production by the human B cell line SKW 6.4, which is capable of secreting IgM in response to B cell differentiation factors (BCDF). SLE is a disorder frequently associated with polyclonal activation of B lymphocytes. We studied the effect of C1q on B cells from two patients with this disorder and one with an SLE-like illness, all selected for the predominance of either IgM or IgG in serum. Spontaneous or BCDF-stimulated Ig secretion was of the isotype predominant in vivo, whereas C1q selectively stimulated B cells to produce the other isotype (IgG vs IgM). Thus, C1q interacts with B lymphocytes in a manner distinct from that of BCDF found in mixed lymphocyte supernatants. C1q may be an important factor influencing the production of Ig by B lymphocytes in normal individuals and in patients with abnormalities of B cell activity.     

Cutting edge: a role for B lymphocyte stimulator in systemic lupus erythematosus

Increased levels of B lymphocyte stimulator (BLyS) are associated with systemic autoimmunity in animal models of spontaneous autoimmune disease, and transgenic animals expressing BLyS develop typical autoimmune disease. Here, we demonstrate significant elevations of BLyS in the patients with systemic lupus erythematosus (SLE). The BLyS isolated from the sera of SLE patients had the same m.w. as the natural soluble form and was able to stimulate B cell activation in vitro. Increased BLyS in SLE patients was partially associated with higher levels of anti-dsDNA Ab of the IgG, IgM, and IgA classes, but not associated with the disease activity. Our results suggest that BLyS may be a useful marker for early activation of an autoimmune diathesis and likely plays a critical role in triggering activation of self-Ag-driven autoimmune B cells in human SLE. BLyS may provide an effective therapeutic target in systemic autoimmunity.     

Effect of B cell differentiation factor (BCDF) on biosynthesis and secretion of immunoglobulin molecules in human B cell lines

Stimulation of human B lymphoblastoid cell lines, CESS and SKW6-CL4 with BCDF induced an increase in IgG- and IgM-secreting cells, as well as a slight increase in IgG and IgM expression on their respective surfaces. Biosynthetic labeling demonstrated that BCDF stimulation induced an increase in synthesis of secretory gamma- and mu-chains, as well as their precursors. A slight but significant increase in synthesis of membrane-bound gamma- and mu-chains and their precursors was also observed by BCDF stimulation. However, an increase in synthesis of secretory heavy chains was much higher than that in membrane-bound heavy chains in both cell lines. Pulse-chase experiments showed that increased synthesis of secretory heavy chains was not due to a decrease in degradation. BCDF stimulation induced a preferential increase in mRNA specific for secretory gamma- and mu-chains in CESS and SKW6-CL4 cells, respectively. These results suggest that BCDF induces an increase in the level of mRNA specific for secretory heavy chains, and then induces final maturation of B cells into immunoglobulin-secreting cells.     

Probing the normal and autoimmune B cell repertoire with Epstein-Barr virus. Frequency of B cells producing monoreactive high affinity autoantibodies in patients with Hashimoto's disease and systemic lupus erythematosus

The frequency of cell precursors producing Ig of different classes and Ag-binding activities were determined, using EBV-infection and limiting dilution assays, in healthy subjects and patients with autoimmune disease. A large proportion of circulating B cells from healthy subjects were committed to the production of IgM antibodies that were polyreactive and bound a variety of self- and exogenous Ag, i.e., IgG Fc fragment, ssDNA, thyroglobulin, thyroid microsomal Ag, insulin, and tetanus toxoid. Similar frequencies of these polyreactive antibody-producing cells were found in patients with Hashimoto's disease and SLE. In contrast, significantly higher frequencies of cell precursors producing monoreactive IgG autoantibodies to thyroid Ag (thyroglobulin and thyroid microsomal Ag) and ssDNA were found in Hashimoto's disease and SLE patients, respectively. Calculation of the Kd revealed that monoclonal polyreactive antibodies were in general low affinity (Kd, 10(-3) to 10(-7) mol/liter), whereas monoclonal monoreactive autoantibodies were high affinity (Kd, 10(-9) to 10(-11) mol/liter). The detected frequency and high affinity of the monoreactive autoantibodies in Hashimoto's disease and SLE patients were comparable to those of anti-tetanus toxoid and anti-insulin IgG mAb produced by B cell clones from vaccinated healthy subjects and insulin-treated patients with insulin-dependent diabetes mellitus, respectively. These findings support the hypothesis that the autoimmune B cell repertoire in patients with organ-specific and systemic autoimmunity is shaped by Ag-driven responses rather than merely reflecting a polyclonal B cell activation.     

Screening of an endothelial cDNA library identifies the C-terminal region of Nedd5 as a novel autoantigen in systemic lupus erythematosus with psychiatric manifestations

Anti-endothelial-cell antibodies are associated with psychiatric manifestations in systemic lupus erythematosus (SLE). Our primary aim in this study was to seek and characterize molecules that behave as endothelial autoantigens in SLE patients with psychiatric manifestations. By screening a cDNA library from human umbilical artery endothelial cells with serum from an SLE patient with psychosis, we identified one positive strongly reactive clone encoding the C-terminal region (C-ter) of Nedd5, an intracytoplasmatic protein of the septin family. To evaluate anti-Nedd5 serum immunoreactivity, we analyzed by ELISA specific IgG responses in 17 patients with SLE and psychiatric manifestations (group A), 34 patients with SLE without psychiatric manifestations (group B), 20 patients with systemic sclerosis, 20 patients with infectious mononucleosis, and 35 healthy subjects. IgG specific to Nedd5 C-ter was present in 14 (27%) of the 51 SLE patients. The mean optical density value for IgG immunoreactivity to Nedd5 C-ter was significantly higher in patients of group A than in those of group B, those with infectious mononucleosis, or healthy subjects (0.17 +/- 0.14 vs, respectively, 0.11 +/- 0.07, P = 0.04; 0.11 +/- 0.06, P = 0.034; and 0.09 +/- 0.045, P = 0.003, on Student's t-test). Moreover, IgG immunoreactivity to Nedd5 C-ter was significantly higher in patients with systemic sclerosis than in patients of group B or healthy subjects (0.18 +/- 0.18 vs, respectively, 0.11 +/- 0.07, P = 0.046; and 0.09 +/- 0.045, P = 0.003). The percentage of patients with anti-Nedd5 C-ter serum IgG was higher in group A than in group B (8 (47%) of 17, vs 6 (17%) of 34, P = 0.045, on Fisher's exact test). In order to clarify a possible mechanism by which Nedd5 might be autoantigenic, we observed that Nedd5 relocated from cytoplasm to the plasma membrane of EAhy926 endothelial cells after apoptotic stimuli. In conclusion, Nedd5 is a novel autoantigen of potential clinical importance that could be successfully used for a more thorough investigation of the pathogenesis of psychiatric manifestations in SLE. Although anti-Nedd5 autoantibodies are not specific to SLE, they are significantly associated with neuropsychiatric SLE and may represent immunological markers of psychiatric manifestations in this pathology.     

Anti-dsDNA Antibody Isotypes in Systemic Lupus Erythematosus: IgA in Addition to IgG Anti-dsDNA Help to Identify Glomerulonephritis and Active Disease

Objectives

To evaluate the role of serum IgG, IgM and IgA anti-dsDNA antibody isotypes in the diagnosis of systemic lupus erythematosus (SLE), and their association with clinical features and disease activity, in a large cohort of SLE patients.

Methods

Sera of 200 SLE patients (mean age 34±10.3 years; 26 male and 174 female; median duration of disease 115 months, range 7–378), and of 206 controls, including 19 Sjögren''s syndrome, 27 rheumatoid arthritis, 26 psoriatic arthritis, 15 idiopathic inflammatory myopathies (IIM), 13 systemic sclerosis, 49 infectious diseases and 57 healthy subjects, were tested for anti-dsDNA IgG, IgM and IgA isotypes.

Results

Selecting a cutoff corresponding to 95% specificity, the sensitivity of IgG, IgM and IgA anti-dsDNA antibodies in SLE was 55%, 30% and 49%, respectively; 12.5%, 1% and 7.5% of SLE patients had positive IgG, IgM or IgA isotype alone, respectively. SLE patients with glomerulonephritis showed higher levels of IgA anti-dsDNA (p = 0.0002), anti-dsDNA IgG/IgM (p = 0.001) and IgA/IgM (p<0.0001) ratios than patients without renal disease. No significant associations have been found between anti-dsDNA isotypes and other clinical features. IgA anti-dsDNA (p = 0.01) (but not IgG or IgM) and IgG/IgM ratio (p = 0.005) were significantly higher in patients with more active disease (ECLAM score >4).

Conclusions

The detection of IgA anti-dsDNA autoantibodies seems to improve our ability to diagnose SLE and to define lupus nephritis phenotype and active disease. By contrast, IgM anti-dsDNA antibodies might be protective for renal involvement. These data support the hypothesis that anti-dsDNA antibody class clustering may help to refine SLE diagnosis and prognosis.     

Human helper T cell factor(s). IV. Demonstration of a human late-acting B cell differentiation factor acting on Staphylococcus aureus Cowan I-stimulated B cells

At least two distinct B cell stimulatory factors (BSF) were found to be involved in the differentiation of Staphylococcus aureus Cowan I (SAC)-stimulated human B cells to IgG-producing cells. A factor tentatively called B cell differentiation factor I (BCDF I) was found in one fraction, and a second factor, BCDF II was found in another fraction. The BCDF I fraction alone induces IgG-production in SAC-stimulated B cells, but the BCDF II fraction does not. The BCDF II fraction enhances IgG production in SAC-stimulated B cells in the presence of the BCDF I fraction. Studies concerning the time-course of the action of the BCDF II fraction revealed that it contains a late-acting differentiation factor that acts on B cells most effectively when it is added to the SAC-stimulated B cell culture after the addition of BCDF I fraction; it induces IgG plaque-forming cells within 1 day. The pI value of a late-acting BCDF was in the range of 5 to 6; this pI range is different from that of BCDF I but similar to that of BCDF II, which was shown in our previous studies to be able to induce IgG production in Epstein Barr Virus-transformed B lymphoblastoid cell lines. In addition, the m.w. of a late-acting BCDF were about 35,000 and 20,000, which are the same as those of BCDF II, and thus its identity with BCDF II was suggested.     

Lupus IgG VH4.34 antibodies bind to a 220-kDa glycoform of CD45/B220 on the surface of human B lymphocytes

Anti-lymphocyte autoantibodies are a well-recognized component of the autoimmune repertoire in human systemic lupus erythematosus (SLE) and have been postulated to have pathogenic consequences. Early studies indicated that IgM anti-lymphocyte autoantibodies mainly recognized T cells and identified CD45, a protein tyrosine phosphatase of central significance in the modulation of lymphocyte function, as the main antigenic target on T cells. However, more recent work indicates that lupus autoantibodies can also recognize B cells and that CD45 may also represent their antigenic target. In particular, IgM Abs encoded by V(H)4.34 appear to have special tropism for B cells, and strong, but indirect evidence suggests that they may recognize a B cell-specific CD45 isoform. Because V(H)4.34 Abs are greatly expanded in SLE, in the present study we investigated the antigenic reactivity of lupus sera V(H)4.34 IgG Abs and addressed their contribution to the anti-lymphocyte autoantibody repertoire in this disease. Our biochemical studies conclusively demonstrate that lupus IgG V(H)4.34 Abs target a developmentally regulated B220-specific glycoform of CD45, and more specifically, an N-linked N-acetyllactosamine determinant preferentially expressed on naive B cells that is sterically masked by sialic acid on B220-positive memory B cells. Strikingly, our data also indicate that this reactivity in SLE sera is restricted to V(H)4.34 Abs and can be eliminated by depleting these Abs. Overall, our data indicate that V(H)4.34 Abs represent a major component of the lupus IgG autoantibody repertoire and suggest that the carbohydrate moiety they recognize may act as a selecting Ag in SLE.     

Ophiopogonin D attenuates the progression of murine systemic lupus erythematosus by reducing B cell numbers

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease characterized by immune abnormalities leading to multi-organ damage. The activation of autoreactive B cell differentiation will lead to the production of pathogenic autoantibodies, contributing to the development of SLE. However, the effects of Ophiopogonin D (OP-D) on B cell activation and autoantibody production as well as renal injury in the pathogenesis of SLE remain unclear. MRL/lpr mice, one of the most commonly used animal models of SLE, were intragastrically administered with 5 mg/kg/d OP-D at 17 weeks of age for 3 weeks. The survival rates of mice in each group were monitored for 6 weeks until 23 weeks of age. Proteinuria and serum creatinine levels were measured. Serum levels of immunoglobulin (Ig)G, IgM, and anti-dsDNA autoantibodies were detected by enzyme-linked immunosorbent assay. Numbers of CD19⁺ B cells in the blood, spleen and bone marrow and numbers of splenic germinal center (GC) B cells were calculated by using flow cytometry. OP-D treatment prolonged survival in MRL/lpr mice. OP-D treatment reduced proteinuria and serum creatinine levels as well as mitigated renal pathological alternation in MRL/lpr mice. Furthermore, serum levels of IgG, IgM, and anti-dsDNA autoantibodies were reduced by OP-D treatment. OP-D lessened not only CD19⁺ B cells in the spleen and bone marrow but also plasma cells that secreted anti-dsDNA autoantibodies, IgG and IgM in the spleen and bone marrow. OP-D ameliorated the progression of SLE by inhibiting the secretion of autoantibodies though reducing B cell numbers.     

Isotypes of spontaneous and mitogen-induced autoantibodies in SLE-prone mice

A common cellular abnormality of all murine strains prone to systemic lupus erythematosus (SLE) is an increased spontaneous polyclonal expansion of B cells. Our findings support the existence of this SLE-associated abnormality because the numbers of B lymphocytes secreting all the different IgG subclasses and IgM in spleens of all lupus-prone mice are elevated, compared to levels of normal splenic immunoglobulin-producing cells. We also report that 1) spontaneous polyclonal stimulation of immunoglobulin in autoimmune mice is preferential for subclass, and that the preferentially stimulated isotypes in each SLE strain consistently dominate both circulating and kidney-deposited immune complexes; 2) distinct patterns of isotype preference exist among the autoimmune strains determined by inherent B cell proliferative abnormalities or by B cell proliferation affected by thymus-derived lymphocytes; and 3) chronic administration of the TI B cell mitogen Lipid A in late-life SLE-prone mice induces an early-life glomerulonephritis with auto-antibodies of an isotype composition characteristic of those spontaneously produced by inherently abnormal B cells of early-life lupus mice.     

Characterization of lymphokines mediating B cell growth and differentiation from monoclonal anti-CD3 antibody-stimulated T cells

Addition of anti-CD3 mAb 147 (IgG1), 446 (IgG1), or 454 (IgG2a) to cultures of T plus non-T cells can result in both B cell growth and differentiation. To determine whether lymphokines mediating these activities were similar to those described from conventional mitogen-induced T cell activation, normal peripheral blood T cells were stimulated with anti-CD3 mAb for 48 h. The supernatants were assayed for factors inducing B cell growth or differentiation (BCDF). A marked increase in Ig secretion was observed when either EBV-transformed B cell lines or normal B cells, pre-activated with Staphylococcus aureus Cowan I strain, were cultured in the presence of mAb 446 (anti-CD3) stimulated T cell supernatant whereas no significant increase in Ig secretion was noted with either mAb 454- or 147-induced T cell supernatant despite equivalent T cell proliferative responses to these antibodies. In contrast, IL-2 secretion was detectable in T cell supernatants from T cells stimulated with either mAb 454 or 147 but not 446. Factors promoting B cell proliferation were detected in all antibody-stimulated T cell supernatants but, contrary to BCDF, appear to act only on non-activated B cells. To determine whether these effector activities were due to distinct lymphokines, supernatants were pooled and concentrated by ammonium sulfate precipitation. Superose 12 permeation chromatography revealed BCDF activity with an apparent Mr of approximately 30,000 Da. The growth factor activity eluted over a wider and higher molecular weight range which overlapped the differentiation factor activity. Fractions containing BCDF activity were pooled, dialyzed, applied to a Mono Q anion-exchange column, and eluted with a linear NaCl gradient. The growth factor activity came off in a single-peak while BCDF was found divided into two major areas. The growth factor eluted at an ionic strength between the two BCDF activities. BCDF has an apparent isoelectric point (pI) of 6, in contrast to the reported pI 5 for IL-6 and more acidic than the documented basic pI of IFN-gamma. Lastly, peaks with BCDF activity were not active in assays for either IL-2 or IL-4. In addition, a rabbit anti-IL-6 heteroantiserum failed to inhibit the pI 6 BCDF, suggesting non-identity between IL-6 and anti-CD3 induced BCDF. Thus, anti-CD3 activated T cells generate both growth factor activity and BCDF as separate molecular entities distinct from IFN-gamma, IL-2, IL-4, and conventional IL-6.     

Serological and Genetic Evidence for Altered Complement System Functionality in Systemic Lupus Erythematosus: Findings of the GAPAID Consortium

Systemic lupus erythematosus is a chronic autoimmune disease with multifactorial ethiopathogenesis. The complement system is involved in both the early and late stages of disease development and organ damage. To better understand autoantibody mediated complement consumption we examined ex vivo immune complex formation on autoantigen arrays. We recruited patients with SLE (n = 211), with other systemic autoimmune diseases (n = 65) and non-autoimmune control subjects (n = 149). Standard clinical and laboratory data were collected and serum complement levels were determined. The genotype of SNP rs1143679 in the ITGAM gene was also determined. Ex vivo formation of immune complexes, with respect to IgM, IgG, complement C4 and C3 binding, was examined using a functional immunoassay on autoantigen microarray comprising nucleic acids, proteins and lipids. Complement consumption of nucleic acids increased upon binding of IgM and IgG even when serum complement levels were decreased due to consumption in SLE patients. A negative correlation between serum complement levels and ex vivo complement deposition on nucleic acid autoantigens is demonstrated. On the contrary, complement deposition on tested protein and lipid autoantigens showed positive correlation with C4 levels. Genetic analysis revealed that the non-synonymous variant rs1143679 in complement receptor type 3 is associated with an increased production of anti-dsDNA IgG antibodies. Notwithstanding, homozygous carriers of the previously reported susceptible allele (AA) had lower levels of dsDNA specific IgM among SLE patients. Both the non-synonymous variant rs1143679 and the high ratio of nucleic acid specific IgG/IgM were associated with multiple organ involvement. In summary, secondary complement deficiency in SLE does not impair opsonization of nucleic-acid-containing autoantigens but does affect other antigens and potentially other complement dependent processes. Dysfunction of the receptor recognizing complement opsonized immune complexes promotes the development of class-switched autoantibodies targeting nucleic acids.     

Comparison of autoantibodies to the collagen-like region of C1q in hypocomplementemic urticarial vasculitis syndrome and systemic lupus erythematosus.

Hypocomplementemic urticarial vasculitis syndrome (HUVS) is an apparent autoimmune disorder that resembles SLE. We previously showed that C1q precipitins in HUVS sera are IgG autoantibody to human C1q. We have compared HUVS anti-C1q autoantibody to a similar autoantibody in the serum of some patients with SLE. As with anti-C1q autoantibody in SLE sera, the HUVS autoantibody binds only to the collagen-like region (CLR) of C1q. In both HUVS and SLE, IgG2 is the predominant subclass of IgG autoantibody and IgM autoantibody to C1q is uncommon. In both diseases, anti-C1q autoantibodies bind preferentially to surface-adsorbed C1q or CLR fragments compared to these antigens in solution. Finally, when HUVS or SLE autoantibodies were added to CLR-coated wells already bound, respectively, by SLE or HUVS autoantibodies, no increases in CLR binding were observed, suggesting that HUVS and SLE autoantibodies to C1q bind to the same CLR epitope(s).     

Cell cycle-related expression of the receptor for a B cell differentiation factor

Cloned, neoplastic B cells (BCL1) have been used to evaluate the expression of the receptor for the B cell differentiation factor, BCDF mu. These cells do not secrete IgM before stimulation with BCDF mu-containing T cell supernatants (SN). By inducing cell cycle synchrony in this homogeneous population, the expression of the BCDF mu receptor could be evaluated as a function of the cell cycle. Responsiveness to BCDF mu-containing SN is maximal when the cells are in S and G2 phases of the cell cycle, and a 2-hr exposure of cells to BCDF mu-containing SN during S/G2 results in optimal IgM secretion 5 days later. Cells in S/G2 are also maximally effective in absorbing BCDF mu activity from SN. These data support the hypothesis that B cells do not respond to differentiative signals until after they are committed to at least one round of cell division.     

Human double-negative T cells in systemic lupus erythematosus provide help for IgG and are restricted by CD1c

To understand the mechanism of T cell help for IgG production in systemic lupus erythematosus (SLE) we investigated the response of CD4- and CD8-negative (double-negative (DN)) T cells because 1) DN T cells are present at unusually high frequency in patients with SLE and can induce pathogenic autoantibodies; 2) the DN T cell repertoire includes cells restricted by CD1 Ag-presenting molecules; and 3) CD1c is expressed on a population of circulating B cells. We derived DN T cell lines from SLE patients and healthy individuals. In the presence of CD1(+) APCs, DN T cell lines from SLE patients produced both IL-4 and IFN-gamma, whereas DN T cells from healthy donors produced IFN-gamma, but no IL-4. In general, cells from patients with highly active disease produced high levels of IFN-gamma; cells from those with little activity produced high IL-4. Coculture of CD1c-directly reactive T cells from healthy donors with CD1c(+) B cells elicited IgM Abs, but little or no IgG. In contrast, CD1c-directly reactive T cells from SLE patients induced isotype switching, with a striking increase in IgG production. Neutralizing Abs to CD1c inhibited the ability of DN T cells to induce IgG production from CD1c(+) B cells, further indicating that CD1c mediated the T and B cell interaction. IgG production was also inhibited by neutralizing Abs to IL-4, correlating with the cytokine pattern of DN T cells derived from these patients. The data suggest that CD1c-restricted T cells from SLE patients can provide help to CD1c(+) B cells for IgG production and could therefore promote pathogenic autoantibody responses in SLE.     

9G4 autoreactivity is increased in HIV-infected patients and correlates with HIV broadly neutralizing serum activity

The induction of a broadly neutralizing antibody (BNAb) response against HIV-1 would be a desirable feature of a protective vaccine. Vaccine strategies thus far have failed to elicit broadly neutralizing antibody responses; however a minority of HIV-infected patients do develop circulating BNAbs, from which several potent broadly neutralizing monoclonal antibodies (mAbs) have been isolated. The findings that several BNmAbs exhibit autoreactivity and that autoreactive serum antibodies are observed in some HIV patients have advanced the possibility that enforcement of self-tolerance may contribute to the rarity of BNAbs. To examine the possible breakdown of tolerance in HIV patients, we utilized the 9G4 anti-idiotype antibody system, enabling resolution of both autoreactive VH4-34 gene-expressing B cells and serum antibodies. Compared with healthy controls, HIV patients had significantly elevated 9G4+ serum IgG antibody concentrations and frequencies of 9G4+ B cells, a finding characteristic of systemic lupus erythematosus (SLE) patients, both of which positively correlated with HIV viral load. Compared to the global 9G4-IgD--memory B cell population, the 9G4+IgD--memory fraction in HIV patients was dominated by isotype switched IgG+ B cells, but had a more prominent bias toward "IgM only" memory. HIV envelope reactivity was observed both in the 9G4+ serum antibody and 9G4+ B cell population. 9G4+ IgG serum antibody levels positively correlated (r = 0.403, p = 0.0019) with the serum HIV BNAbs. Interestingly, other serum autoantibodies commonly found in SLE (anti-dsDNA, ANA, anti-CL) did not correlate with serum HIV BNAbs. 9G4-associated autoreactivity is preferentially expanded in chronic HIV infection as compared to other SLE autoreactivities. Therefore, the 9G4 system provides an effective tool to examine autoreactivity in HIV patients. Our results suggest that the development of HIV BNAbs is not merely a consequence of a general breakdown in tolerance, but rather a more intricate expansion of selective autoreactive B cells and antibodies.     

Lymphokines regulate immunoglobulin isotype expression in an antigen-specific immune response

The control of immunoglobulin class switching appears to involve T cell-derived lymphokines. Such lymphokines have been shown to affect isotype expression in polyclonally activated B cells. We show in this paper that the same lymphokines similarly control isotype expression in an antigen-specific response acting in concert with a "T cell independent" antigen. In this situation, B cell growth factor II (BCGF II) enhances the production of antigen-specific IgM antibodies, whereas the production of antigen-specific IgG1 antibodies is only observed in the presence of B cell differentiation factor gamma (BCDF gamma). These results suggest that these lymphokines (and perhaps additional ones) are involved in the control of isotype expression in antigen-specific responses.     

Altered recruitment of Lyn,Syk and ZAP-70 into lipid rafts of activated B cells in Systemic Lupus Erythematosus

There is evidence that B cells from patients with Systemic Lupus Erythematosus (SLE) could be hyperactivated due to changes in their lipid rafts (LR) composition, leading to altered BCR-dependent signals. This study aimed to characterize possible alterations in the recruitment of protein tyrosine kinases (PTK) into B cells LR from SLE patients. Fifteen patients with SLE and ten healthy controls were included. Circulating B cells were isolated by negative selection and stimulated with goat Fab´2 anti-human IgM/IgG. LR were isolated with a non-ionic detergent and ultracentrifuged on 5–45% discontinuous sucrose gradients. Proteins from each fraction were analyzed by Western Blot. Total levels of Lyn, Syk, and ZAP-70 in resting B cells were similar in SLE patients and healthy controls. Upon BCR activation, Lyn, Syk and ZAP-70 recruitment into LR increased significantly in B cells of healthy controls and patients with inactive SLE. In contrast, in active SLE patients there was a great heterogeneity in the recruitment of signaling molecules and the recruitment of ZAP-70 was mainly observed in patients with decreased Syk recruitment into LR of activated B cells. The reduction in Flotilin-1 and Lyn recruitment in SLE patients seem to be associated with disease activity. These findings suggest that in SLE patients the PTK recruitment into B cell LR is dysregulated and that B cells are under constant activation through BCR signaling. The decrease of Lyn and Syk, the expression of ZAP-70 by B cells and the increase in Calcium fluxes in response to BCR stimulation in active SLE patients, further support that B cells from SLE patients are under constant activation through BCR signaling, as has been proposed.     

Simultaneous Assessment of Rotavirus-Specific Memory B Cells and Serological Memory after B Cell Depletion Therapy with Rituximab

The mechanisms that contribute to the maintenance of serological memory are still unclear. Rotavirus (RV) memory B cells (mBc) are enriched in IgM⁺ and CD27^- subpopulations, which are associated with autoimmune diseases pathogenesis. In patients with autoimmune diseases treated with Rituximab (RTX), some autoantibodies (auto-Abs) decrease after treatment, but other auto-Abs and pathogen-specific IgG Abs remain unchanged. Thus, maintenance of autoimmune and pathogen-specific serological memory may depend on the type of antigen and/or Ab isotype evaluated. Antigen-specific mBc and antigen-specific Abs of different isotypes have not been simultaneously assessed in patients after RTX treatment. To study the relationship between mBc subpopulations and serological memory we characterized total, RV- and tetanus toxoid (TT)-specific mBc by flow cytometry in patients with autoimmune diseases before and after treatment with RTX. We also measured total, RV- and TT-Abs, and some auto-Abs by kinetic nephelometry, ELISA, and EliA tests, respectively. Minor differences were observed between the relative frequencies of RV-mBc in healthy controls and patients with autoimmune disease. After RTX treatment, naïve Bc and total, RV- and TT-specific mBc [IgM⁺, switched (IgA⁺/IgG⁺), IgM⁺ only, IgD⁺ only, and CD27^- (IgA⁺/IgG⁺/IgM⁺)] were significantly diminished. An important decrease in total plasma IgM and minor decreases in total IgG and IgA levels were also observed. IgM rheumatoid factor, IgG anti-CCP, and IgG anti-dsDNA were significantly diminished. In contrast, RV-IgA, RV-IgG and RV-IgG₁, and TT-IgG titers remained stable. In conclusion, in patients with autoimmunity, serological memory against RV and TT seem to be maintained by long-lived plasma cells, unaffected by RTX, and an important proportion of total IgM and serological memory against some auto-antigens seem to be maintained by short-lived plasma cells, dependent on mBc precursors depleted by RTX.     

Associations between the FAS ?670?A/G and ?1,377?G/A polymorphisms and susceptibility to autoimmune rheumatic diseases: a meta-analysis

The aim of this study was to explore whether FAS ?670?A/G and ?1,377?G/A polymorphisms confer susceptibility to autoimmune rheumatic diseases. A meta-analysis was conducted on the associations between the FAS ?670?A/G and ?1,377?G/A polymorphisms and autoimmune rheumatic diseases using allele contrast, a recessive model, a dominant model, and an additive model. Thirteen articles with 21 comparison studies (16 on FAS ?670?A/G and 5 on ?1,377?G/A polymorphisms) including systemic lupus erythematosus (SLE), four systemic sclerosis, four Sjogren’s syndrome, three rheumatoid arthritis (RA), one juvenile idiopathic arthritis, and one spondyloarthropathy were available for the meta-analysis. Meta-analysis revealed an association between rheumatic diseases and the FAS ?670?A/G polymorphism in the dominant model (odds ratio [OR]?=?0.761, 95?% confidence interval [CI]?=?0.621–0.932, p?=?0.008]. Stratification by ethnicity indicated an association between the FAS ?670?G allele carrier and rheumatic diseases in Asian (OR?=?0.569, 95?% CI?=?0.409–0.791, p?=?0.001). Furthermore, stratification by disease indicated an association between the FAS ?670?G allele carrier and SLE and RA (OR?=?0.578, 95?% CI?=?0.358–0.934, p?=?0.025; OR?=?0.609, 95?% CI?=?0.398–0.934, p?=?0.023, respectively). The FAS ?670?G allele was negatively associated with SLE susceptibility. Meta-analysis of the FAS ?1,377?G/A polymorphism stratified by disease showed an association between the FAS ?1,377 A allele and SLE (OR?=?0.783, 95?% CI?=?0.613–0.997, p?=?0.047). Meta-analyses using the dominant model also showed a significant association in SLE (OR?=?0.712, 95?% CI?=?0.528–0.961, p?=?0.027). This meta-analysis demonstrates that the FAS ?670?A/G polymorphism confers susceptibility to rheumatic diseases in Asians and SLE and RA, and the FAS ?1,377?G/A polymorphism is associated with SLE susceptibility.