The Role Of B Lymphocyte In Sle-like Syndrome Induced By Syngeneic Apoptotic Dna

Systemic lupus erythematosus (SLE) is a severe multi-systemic autoimmune disease serologically characterized by the production of a variety of antinuclear antibodies (ANA) and the tissue damage resulting from the local deposition of immune complexes. However, the pathogenic mechanisms triggering the onset of SLE have not yet been fully defined, which has been brought to the fore work at multiple laboratories in both mice and humans.Our previous study about the SLE pathogenesis has established a lupus model in BALB/c mice induced by syngeneic apoptotic DNA derived from ConA stimulation induced apoptotic splenocytes. Immunization of naive mice with apoptotic DNA evoked high levels of anti-ds-DNA antibody production. The pathogenecity of the anti-ds-DNA antibodies was demonstrated by the renal pathological changes, including high level of urine proteins, cellular hyperplasia and tubular dilation together with renal glomerular IgG deposits in the apoptotic DNA treated mice. However, genomic DNA from normal splenocytes failed to induce the anti-ds-DNA antibodies and lupus nephritis.In fact, the underlying mechanism for the induction of anti-ds-DNA antibodies by apoptotic DNA remains undefined. The reactivation of CD4+T cell was responsible for the disease manifestations besides the autoantibody formation and what's the motive for T cell activation?In the last several years, a growing body of evidence shows that B cells contribute importantly to the pathogenesis of SLE disease. In the peripheral blood of SLE patients, up-regulation of the inhibitory Fc receptor, FcyRIIb, was decreased in B cells and reduced the threshold for auto-reactive B activation. The importance of BAFF (B cell activator of the TNF family) in human SLE, a key cytokine that promotes B cell survival, has been demonstrated by the finding of elevated serum levels in SLE patients and the correlation with serum IgG and autoantibody levels. MRL/lpr lupus-prone mice genetically B cell-deficient have strikingly attenuated disease. In special, the efficacy of B cell depletion therapy in clinical supported the key role of the B cell in the disease. B cells are important as APC as well as autoantibody secreting cells. Whether B cells played a role for both the antibody production and activators of T cell in our SLE-like syndrome? If so, what's the mechanism?We firstly set up the SLE-mice model to study the role of B lymphocyte in the induction course of disease by apoptotic DNA. It can provide the theory bases for the underlying of SLE pathogenesis, which may facilitate the disease therapy.Partâ… :TLR9 signaling in B lymphocytes enhance its ability of anti-ds-DNA antibody production induced by apoptotic DNAIn the period of our work about SLE, we found that genomic DNA derived from ConA-stimulated syngeneic splenocytes, but not normal lymphocyte-derived DNA, could induce high levels of anti-ds-DNA antibodies and subsequent end organ damage in BALB/c mice. We further investigated that the increased anti-ds-DNA level was associated with apoptotic level of the splenocytes after ConA stimulation and the apoptotic characteristic of genomic DNA was responsible for its immunogenicity. Normal BALB/c mice were immunized with apoptotic DNA derived from the Annexin-V+ cells after ConA stimulation 6 days. Finally, high levels of anti-ds-DNA antibodies and urine proteins were detected in the apoptotic DNA treated mice. The pathological examination of the kidney tissue showed cellular hyperplasia and tubular dilation together with renal glomerular IgG deposits and inflammatory infiltrates, which are important manifestations of lupus nephritis in the SLE-like syndrome. These results demonstrated that we successfully conducted the apoptotic DNA-induced SLE-like syndrome in normal BALB/c mice.B cell as anti-ds-DNA antibody secreting cell has been the focus subject in the investigation of the underlying mechanism of SLE-mice model induction. Of note, the nuclei acid receptor TLR9 or TLR7 was recognized as an essential role of ANA production. It is important to be aware that generation of specific antibody responses requires direct activation of TLRs in B cells in addition to antigen recognition and CD4+T cell help. We detected the percentage and absolute number of B cell from apoptotic DNA or normal DNA immunized mice, and found that the percentage and absolute number of B cell in SLE mice compared with the control group. Further we sorted CD19+B cell from different group of mice to analyze TLR9 expression of it by RT-PCR, Real-time PCR and FACS. All the results showed a dynamic up-expression of TLR9 in B cells from apoptotic DNA immunized mice. Next, we immunized mice with apoptotic DNA in the presence of chloroquine and found that the induction of anti-ds-DNA antibody was decreased while the inducible level of TLR9 expression in B cell was lower. To confirm the results, purified B cells were pre-treated with choloqiune in vitro and consequently the ability to secrete antibodies was impaired. It's also demonstrated that down-regulation of transcriptional regulators for PC differentiation after chloroqiune treatment may be the reason for the decreased antibody. All of the results suggested apoptotic DNA could induce the TLR9 up-expression in B cells to facilitate the production of anti-ds-DNA antibody, which was meaningful for the initiation of SLE.Partâ…¡:The accessory function of B cell to activate CD4+ T cell is beneficial for its generation of anti-ds-DNA antibody via a B-cell/T-cell self-regulatory loopIn recent years, the accumulating evidence implicated that both antibody-dependent and antibody-independent mechanisms of B cells were important in SLE. Autoantibody-independent B cell functions have been postulated to include auto-antigen presentation, T-cell activation and polarization, and immuno-regulatory cytokines production. Given that a relatively stable level of specific auto-antibodies is crucial for the ongoing disease, remain at least in the short-term, the role of B cell to induce proliferation and activation of T cell is highlighted. Once generated, these activated CD4+ cells also can reengage the activated B-cell, leading to more auto-antibody production and more T effector-cell generation. B-cell/T-cell interactions constitute a positive feedback loop that enables continuous amplification of the autoimmune response.We further explored whether B cell was involved in the apoptotic DNA stimulation of the CD4+ T cell. The apoptotic DNA induced the B cell production of anti-ds-DNA antibodies in a CD4+ T cell-dependent manner, while B cells from apoptotic DNA immunization contributed to the reactivation of CD4+ T cell in turn. Then a trans-well assay was performed to find that close cell-cell contact was required for the activation of CD4+ T cell by apoptotic DNA primed B cells. The expression of co-stimulatory molecules CD80/86 on B cells were detected to up-regulate after the apoptotic DNA immunization. Anti-CD86 blocking Ab significantly abrogated the effect of B cells on the proliferation of CD4+ T cells. Chloroquine treatment of B cell could suppress the expression of CD80/86, which suggested that the efficient APC function of B cells was mediated by the inducible TLR9 signaling by apoptotic DNA and the subsequent CD80/CD86 pathway.Based on the findings from the aforementioned studies, the following conclusions may be drawn: SLE-like syndrome was successfully established by syngeneic apoptotic DNA and the TLR9 of B cell was inducible up-expressed. It's defined that TLR9 signaling is essential for the B cell secretion of anti-ds-DNA antibody and activation of CD4+ T cell. These findings would not only facilitate our further understanding of the SLE pathogenesis, but also provide novel insights and experimental bases into clinical application of B cell therapy for SLE.