蛋白质核心岩藻糖基化修饰在肾间质纤维化中的特征性改变及作用机制

Pericyte activation is the key event in the process of renal fibrosis, but the underlying mechanism remains obscure. Glycosylation modification is an indispensable episode regulating the function of glycoprotein. With the development of proteomics, the research of glycosylation has become a new promising area. But the findings of glycosylation in the field of nephrology are almost barren. Our previous studies have suggested that excessive activation of α1,6-fucosyltransferase(fucosyltransferase 8, FUT8) regulated-core fucosylation may play an important role in the process of renal interstitial fibrosis. Moreover, our preliminary screen has found out several candidate key proteins relevant to fibrosis. Based on our previous creative work, bionic microfluidic chip model consisting of renal tubular epithelial cell, microvascular uint and interstitial substance, which is highly superior to routine 2-dimensional static cell culture in exsistance, will be established to imitate the pathological changes of renal interstitial fibrosisin vitro. FUT8 gene knockout and knockdown mice models will be used for in vivo study, the dynamic changes of core fucosylation will be analyzed by glycomics technique. The effects and the molecular mechanism of core fucosylation on pericyte activation will be explored. Finally, the comparisons between the suppression of mutiple signaling pathways and the suppression of single signaling pathway will be made to evaluate the efficacy and the superiority of core fucosylation intervention on pericyte activation and renal interstitial fibrosis. We will propose and then verify the academic idea “core fucosylation modification of protein is the keypoint in the regulation of renal interstitial fibrosis”. We believe that our findings will promote the communication between glycobiology and nephrology. We will also provide a novel therapeutic strategy and researching idea for the treatment of renal interstitial fibrosis.

周细胞活化是肾间质纤维化的重要事件，其机制不清。糖基化修饰是糖蛋白发挥功能的重要环节，是蛋白质组学发展后新开拓的重要领域，但在肾脏病领域的研究几近空白。我们前期发现α-1,6岩藻糖转移酶FUT8调控的核心岩藻糖基化在肾间质纤维化中起重要作用，并初步筛选出关键差异蛋白。本研究拟在上述创新性工作基础上，利用微流控芯片组织工程构建仿生“肾小管-微血管-间质微环境”模型行体外研究，该技术远远优于常规二维静态细胞培养；利用FUT8敲除、敲低小鼠行体内研究，糖蛋白质组学观察肾间质纤维化中核心岩藻糖基化特征性改变，研究核心岩藻糖基化在周细胞活化中的作用及机制，比较多信号通路活性阻抑、单信号通路活性阻抑对周细胞活化及肾间质纤维化的干预效果。提出并验证“蛋白质核心岩藻糖基化修饰是调控肾间质纤维化发生和进展的关键环节”学术观点，促进糖生物学和肾脏病学交叉渗透，为肾间质纤维化防治提供新思路和治疗策略。

周细胞活化是肾间质纤维化的重要事件，其机制不清。糖基化修饰是糖蛋白发挥功能的重要环节，是蛋白质组学发展后新开拓的重要领域，但在肾脏病领域的研究几近空白。我们前期发现α-1,6岩藻糖转移酶FUT8调控的核心岩藻糖基化在肾间质纤维化中起重要作用，并初步筛选出关键差异蛋白。拟利用微流控芯片组织工程构建仿生“肾小管-微血管-间质微环境”模型行体外研究，利用FUT8敲除、敲低小鼠行体内研究，糖蛋白质组学观察肾间质纤维化中核心岩藻糖基化特征性改变，研究核心岩藻糖基化在周细胞活化中的作用及机制，比较多信号通路活性阻抑、单信号通路活性阻抑对周细胞活化及肾间质纤维化的干预效果。本研究成功建立研究所需的模式动物、仿生肾脏微流控芯片以及核心岩藻糖组学分析等技术平台：首先制作了fut8转基因动物资源库及PDGFRβ-GFP荧光报告鼠；构建了高通量集成“肾小管-间质-微血管”微流控芯片模型；在建立位点特异性核心岩藻糖修饰定量分析方法基础上，建立位点特异性的糖链结构解析与定量研究策略，实现了复杂结构的融合蛋白质药物糖修饰定量分析。在此基础上，在多种肾病模型中开展系列研究，系统解析了CF修饰参与肾脏损伤、肾间质纤维化的调控机制，并探究以CF修饰为干预靶点的治疗与机制。验证了“蛋白质核心岩藻糖基化修饰是调控肾间质纤维化的关键环节”观点，为肾间质纤维化防治提供了基于CF修饰的新思路与干预策略。研究成果发表SCI收录论文19篇，参编专著2部，授权专利2项，申请2项。