IGA CELLULAR RECEPTOR INTERACTIONS IN IGA NEPHROPATHY

IgA肾病（IgAN）是糖尿病患者肾小球肾炎的主要原因。 世界IgA 1在系膜中的显著沉积是糖尿病的标志。 疾病尽管经过了25年的调查， 定位于系膜，这种沉积与 肾功能不全的发展缓慢，但在一些患者进行性 Igan仍然未知。在本提案中，我们将检验一般假设 在lgAN中，循环lgA 1结构的异常导致 不成比例的IgA绕过正常的分解代谢 途径和/或选择性地结合到肾小球系膜细胞， 肾脏该项目将重点关注O-连接的寡糖结合 IgA 1的铰链区肽，因为这些结构 区分lgA 1与lgA 2和其他主要Ig同种型。的结合 lgA 1与两种细胞受体，即培养细胞上的去唾液酸糖蛋白受体 肝癌细胞和系膜细胞原代培养物的FcalphaR受体 将详细检查细胞。这些受体选自 因为它们最有可能参与免疫球蛋白A受体， 分解代谢缺陷（ASGPR）或异常IgA靶向肾脏 （系膜FcalphaR）。这两种受体都需要寡糖在其 配体结合。三个具体的假设将被测试：1）， 来自一些IgAN患者的lgA 1将相对更好地结合FeaR 2）部分IgAN患者的IgA 1结合率高于ASGPR， 诱导更多FcalphaR的表达和细胞因子的产生， 生长因子从培养的系膜细胞，和3），一些 在1和2中定义的功能异常可以归因于异常的 IgA 1铰链区的糖基化。此中要使用的方法 研究包括通过凝集素亲和层析法从 检测了20例IgA肾病患者、10例正常人和10例 其他形式的肾小球肾炎。结合的详细研究， 选择的IgA 1的内化和细胞内命运 将进行准备工作。不同的lgA 1制剂的作用 将通过北方印迹和FcalphaR表达的释放来评估 转化生长因子、血小板衍生生长因子和IL-6， 免疫测定。碳水化合物结构分析将由β 消除和HPLC，凝集素结合和酶降解。铰链 用IgA特异性蛋白酶和胰蛋白酶产生的糖蛋白将被 提交用于AA序列分析。成功完成这些研究 应鉴别出一种或多种IgA结构和功能异常 使一些lgAN患者易发生IgA的系膜沉积。这些 研究最终应该能够识别出有风险的患者， lgAN的发展或进展。特定治疗部位 这些研究也可能建议采取干预措施。

lgA nephropathy (IgAN) is the leading cause of glomerulonephrits in the world. Prominent deposition of IgA1 in the mesangium is the hallmark of the disease. Despite over 25 years of investigation, the reason why lgA localizes to the mesangium and the relationship between this deposition and slow but progressive development of renal dysfunction in some patients with IgAN remains unknown. In this proposal, we will test the general hypothesis that in lgAN, abnormalities in the structure of the circulating lgA1 cause a disproportionate fraction of the lgA to bypass the normal catabolic pathway in the liver and/or to bind selectively to the mesangial cells in the kidney. The project will focus on the O-linked oligosaccharides bound the hinge region peptide of lgA1, since these are the structures that distinguish lgA1 from lgA2 and the other major Igs isotypes. The binding of lgA1 to two cellular receptors, the asialoglycoprotein receptor on cultured hepatoma cells and the FcalphaR receptor of primary cultures of mesangial cells will be examined in detail. These receptors were chosen from a number of lgA receptors, since they are most likely to be involved in an lgA catabolic defect (ASGPR) or in targeting of the abnormal lgA to the kidney (mesangial FcalphaR). Both receptors require oligosaccharides on their ligands for binding. Three specific hypotheses will be tested: 1) that the lgA1 from some patients with IgAN will bind relatively better to the FeaR than the ASGPR, 2) that binding of the lgA1 from some patients with lgAN induces the expression of more FcalphaR and the production of cytokines and growth factors from cultured mesangial cells, and 3) that some of the functional abnormalities defined in l and 2 can be attributed to aberrant glycosylation of the hinge region of lgA1. The methods to be used in this study include isolation by lectin affinity chromatography of lgA1 from the serum of 20 patients with lgAN, 10 normals subjects and 10 patients with other forms of glomerulonephritis. Detailed studies of the binding, internalization and the intracellular fate of the selected lgA1 preparations will be undertaken. The effect of various preparations of lgA1 on FcalphaR expression will be assessed by Northern blotting and release of transforming growth factor, platelet derived growth factor and lL-6 by immunoassays. Carbohydrated structural analysis will be performed by beta elimination and HPLC, lectin binding and enzymatic degradation. The hinge glycoproteins produced with lgA-specific proteases and trypsin will be submitted for aa sequence analysis. Successful completion of these studies should identify one or more abnormalities of lgA structure and function predispose some patients with lgAN to mesangial deposition of lgA. These studies should eventually allow identification of patients at risk for development or progression of lgAN. Specific sites for therapeutic interventions may also be suggested by these studies.