Study On Disturbance Of Helper T Cells And Its Effect In Human IgA Nephropathy

IgA nephropathy (IgAN) is the most common form of glomerulonephritis defined by the predominant deposition of IgA in the glomerular mesangium, which was first reported by Berger in 1968. About 30–40% of IgAN patients will develop progressive renal failure and eventually require the renal replace therapy. It follows that the elucidation of the pathogenesis of IgAN is a key research goal that, if followed by the design of successful specific treatment interventions, will have a substantial positive health impact.The initiating event in the pathogenesis of IgAN is the mesangial deposition of IgA, which is predominantly polymeric IgA of the IgA1 subclass (pIgA1). With or without the additional deposition of IgG and C3 complement. Arising evidences showed IgAN patients produce a pool of circulating IgA1 molecules with abnormal O-glycosylation structure, which particularly form pIgA1 or IgA-IgG complex, and promote mesangial deposition, then provoke glomerular inflammation and injury. Therefore the increased IgA1 with abnormal glycosylation structure could be regard as the important pathogenetic factor in IgAN. The mechanisms of circulating pIgA1 overproduction and abnormal O-glycosylation IgA1 formation in IgAN have not been fully elucidated.IgA class antibodies are released from plasma cells, which are developed successively from cytoplasmicμ-chain positive pre-B cells via IgM-, IgG-, and IgA-bearing B lymphocytes. During these development stages, regulation by T cells is necessary. Previous studies have found the disbalance of Th1/Th2 in IgAN, but there was controversy on which one is predominant. Furthermore the role of Th1/Th2 in IgAN has not been elucidated. Th2 cytokines (IL-4, 5, and 6) are proved to promote B cell class switching to IgA, and subsequent proliferation and differentiation of IgA-producing cells. Recently Th2 cytokines have been reported to influence IgA N-glycosylation in murine system, although the effect on human IgA O-glycosylation need further studies.The aim of the present study is to investigate the frequency of helper T subsets in the peripheral blood of IgAN patients, and explore the correlation with the pathological phenotype as well as clinical parameters. In addition, we observe the effect of Th2 on the abnormal O-glycosylation of human IgA1 in vitro and explore the potential role of Th2 in the immunopathogenesis of IgAN. Chapter 1 The expression of helper T subsets in the peripheral blood of IgA nephropathy patientsObjective:To detect the frequency of Th1 and Th2 in the peripheral blood of IgAN patients, and observe the bias of Th1/Th2, explore the correlation with clinical and pathological parameters.Methods:20 IgAN patients and 22 healthy normal controls with comparable age and gender distribution were enrolled. All patients were pathologic diagnosed primary IgA nephropathies excluding secondary reasons, such as systemic lupus erythematosus chronic alcoholic liver disease, and Henoch-Sch?nlein purpura. None of the patients have been treated with glucocorticoid or immunosuppressants in one month. Serum samples from patients were obtained at the time of renal biopsy. The clinical and pathologica parameters were collected.The frequency of Th1 and Th2 in the peripheral blood of IgAN and healthy control was examined by intracellular cytokine detection method with flow cytometry, and calculated the ratio of Th1/Th2. The association with clinical and pathological parameters of IgAN was analyzed by Spearman's or Pearson rank correlation test. Results:Compared to healthy controls, the percentage of Th1 in CD3+ lymphocytes were decreased in IgAN, but no significant difference. While the percentage of Th2 were significantly increased in IgAN patients compared to healthy controls, respectively was 2.57±0.72% vs 1.81±1.10%. Accordingly the ratio of Th1/Th2 reduced. The frequency of Th2 was positively correlated with serum IgA level (r=0.47, P < 0.05), negatively correlated with eGFR (r=-0.50, P < 0.05). No significant correlation between Th1, Th1/Th2 ratio and clinical or pathological parameters was detected.Conclusions:The disbalance of Th1/Th2 was found towards Th2 in IgAN patients. The frequency of Th2 was positively correlated with serum IgA level, negatively correlated with eGFR. Chapter 2 The effect of Th2 on abnormal glycosylation of IgA1 in human IgA nephropathyObjective To observe the effect of Th2 on IgA1 glycosylation secreted by PBMC of IgAN patients, explore the role of Th2 in the immunopathogenesis of IgAN. Methods The PBMC of IgAN patients and normal controls were separated by Ficoll method, and the Th2 was removed by MACS. The equal number cells of whole PBMC and PBMC without Th2 were cultured in vitro with pokeweed mitogen, and collected the supernatant after 7 days. The concentration of IgA1 of supernatant and its capability to combined with VVL, PNA were detected by ELISA. The difference between two groups were analysis by t test.Results The concentration of IgA1 produced by PBMC without Th2 of IgAN patients were significantly lower than the whole PBMC, respectively were 1.87±1.31μg/ml vs 0.61±0.40μg/ml. While there was no significant difference between the IgA1 concentration produced by PBMC of normal control with or without Th2. Both in IgAN patients and normal controls, the capacity to combine with VVL, PNA of IgA1 secreted by PBMC with or without Th2 were similar. Conclusions Th2 was found to contribute to IgA1 production in IgAN patients, but no direct effect on IgA1 glycosylation.Conclusions:1.The disbalance of Th1/Th2 was found towards Th2 in IgAN patients, Th2 could contribute to development of IgAN by producing exceed IgA1.2. In vitro experiment result confirmed the effect of Th2 on quantity of IgA1, but no effect on the IgA1 glycosylation.