GM130-C1GalT1在IgA肾病低糖基化IgA1生成中的作用及机制研究

IgA nephropathy(IgAN) is the most common type of primary glomerulonephritis, and its most critical pathogenesis is that elevated circulating levels of galactose-deficient HR O-linked glycans of IgA1 (Gd-IgA1) and deposited in glomerular. The decreased expression and activity of Core 1 β1,3-galactosyltransferase(C1GalT1) could lead to increase the production of Gd-IgA1 in patients with IgA nephropathy, are related to the occurring development of IgAN, but the mechanism has not been elucidated. GM130 facilitates C1GalT1 vesicle to the Golgi docking as a vesicle “tethering factor”. We found that GM130 decreased significantly in IgAN patients, and it negative with Gd-IgA1 production. What’s more, we used siRNA degraded GM130 expression in PBMC, the Gd-IgA1 production increased significantly, and C1GalT1 expression decreased, which indicated that GM130 play an important role in IgA1 glycosylation. We speculated that GM130 involved in the IgA1 glycosylation defection by regulating C1GalT1. GM130 decreasing prevents C1GalT1 vesicle docking to the Golgi, resulting in IgA1 O-glycosylation defects in IgAN patients. In this study, we focus on the CD19(+) B cells of peripheral blood from IgAN patients, investigated the mechanism of GM130 on Gd-IgA1 production in IgAN by immunofluorescence, Western blot, PCR and gene silencing and overexpression. The goal of this proposal is to dissect the role and underlying mechanisms of the production of Gd-IgA1, in order to provide scientific and experimental direction for the treatment of IgAN in future.

IgA肾病（IgAN）是最常见原发性肾小球疾病，其关键始动环节是过多低糖基化IgA1(Gd -IgA1)产生并沉积于肾小球。IgAN患者β1,3半乳糖转移酶（C1GalT1）表达下降致Gd-IgA1增加，但机制尚不明。GM130为C1GalT1囊泡进入高尔基体关键衔接蛋白。我们前期研究发现IgAN患者GM130表达显著下降，并与Gd-IgA1分泌呈负相关，下调人PBMC GM130表达，Gd-IgA1分泌显著增加，C1GalT1表达下降，提示GM130通过C1GalT1在IgA1糖基化过程中起重要作用。故推测：IgAN患者GM130下降，使C1GalT1进入高尔基体障碍，导致IgA1低糖基化发生。本研究以IgAN患者外周血CD19(+)B细胞为研究对象，采用细胞分子生物学技术，探索GM130对IgA1糖基化调控机制。旨在揭示IgAN患者Gd-IgA1产生机制，为IgAN防治提供新思路。

IgA肾病（IgAN）是最常见原发性肾小球疾病，其关键始动环节是过多低糖基化IgA1(Gd-IgA1)产生并沉积于肾小球。IgAN患者β1,3半乳糖转移酶（C1GalT1）表达下降致Gd-IgA1增加，但机制尚不明。我们研究发现IgAN患者外周血单个核细胞GM130表达显著下降，并与Gd-IgA1分泌呈负相关。下调人外周血单个核细胞 GM130表达，C1GalT1表达下降，Gd-IgA1分泌显著增加。若上调IgAN患者外周血单个核细胞GM130表达，C1GalT1蛋白表达增加，Gd-IgA1分泌显著下降。这提示GM130在IgAN患者Gd-IgA1产生过程中起到重要作用。另外，我们通过分子生物技术，通过分别下调GRASP65和Giantin两个高尔基体锚点蛋白，发现下调Giantin蛋白后，可导致C1GalT1表达下降，细胞分泌Gd-IgA1显著增加，从而探明GM130通过Giantin蛋白协助C1GalT1囊泡在高尔基体上的锚定及正常转位 参与 IgA1 糖基化调控的分子机制。揭示了在IgA肾病患者中，GM130表达下降导致C1GalT不能锚钉并进入高尔基体内，而在细胞内快速降解，从而导致IgA1 糖基化水平下降，产生大量低糖基化IgA1，这为IgAN防治提供新思路。我们在进行本研究同时，建立了IgA肾病生物样本库和随访队列，为后续大规模临床研究提供基础。.IgA肾病患者低糖基化IgA目前普遍认为来源于粘膜免疫，肠道作为体内最大粘膜系统，其在IgAN中的作用日益受到关注。我们构建了IgAN小鼠模型，初步检测了IgAN小鼠肠道粘膜中GM130表达较正常对照组显著降低。证实了GM130同样参与了IgAN肠道粘膜免疫，为后续深入研究提供了那就基础。.另外，在新冠疫情期间，通过统计分析湖南省新冠患者的临床资料，发现输入性病例较本土病例更容易发展为重型肺炎以及需要糖皮质激素及丙种球蛋白治疗，这可能与病毒毒力在传代过程中减弱以及患者在疾病过程中的心理等综合因素有关。论文已发表。在临床慢性炎症性肾脏疾病病例中发现外周血淋巴细胞中TIM-3的表达变化对与患者的免疫状态具有重要作用，经过系统的文献复习与临床观察研究，我们撰写论文已发表。