Niban通过Fas/FasL凋亡途径影响肾间质纤维化的机制研究

Renal interstitial fibrosis（RIF）is closely correlated with end stage renal disease. Renal tubular epithelial cells can promote RIF through apoptosis pathway of Fas/FasL. We screened out differential protein, Niban, in proteomics study of UUO rats. In turn, we found that expression of Niban decreased obviously in patients, animal and cell model of RIF, contrary to increasing apoptosis of renal tubular epithelial cells and caspase8 which was the marker protein of Fas/FasL. Transfecting siRNA of Niban increased phenomenon above. So we put forward the hypothesis that Niban can regulate RIF through apoptosis pathway of Fas/FasL. In order to confirm Niban can regulate pathway of Fas/FasL, and make sure the binding protein interact with Niban, we plan to observe impact of Niban on Fas, FADD and caspase8 through vivo and vitro experiments with methods of immune coprecipitation, mass spectrum, transfection plasmid and renal vein injection Niban overexpression intervention to gene knockout mice. This topic will provide experimental basis for Niban being the possible and effective therapeutic target in RIF.

肾间质纤维化(RIF)是各种慢性肾脏病进展至终末期肾病的共同通路和主要病理基础。已证实肾小管上皮细胞可通过Fas/FasL凋亡途径促进RIF。课题组前期在UUO大鼠模型蛋白质组学中筛选出显著差异蛋白Niban，并首次发现RIF患者、动物及细胞模型中，随着Niban表达降低，肾小管上皮细胞凋亡增加，Fas/FasL途径标志蛋白caspase8表达增多；Niban siRNA干预后，上述现象更显著。因此我们提出假说：Niban能通过Fas/FasL凋亡途径调控RIF。本课题组拟采用免疫共沉淀、质谱、转染质粒、肾静脉注射Niban过表达干预基因敲除小鼠等方法，从体内外实验两个层面分别观察Niban对Fas、FADD、caspase8的影响，证实Niban能调控Fas/FasL途径，并进一步寻找与Niban相互作用的结合蛋白。本课题将为Niban成为可能的RIF有效治疗靶点提供实验依据。

目的：探讨Niban蛋白是否通过调节肾小管上皮细胞凋亡在肾间质纤维化中起作用，并探讨其机制。.方法：采用单侧输尿管梗阻（UUO）模型对C57B/6J小鼠进行手术，分为假手术组、第3天、第7天和第14天。免疫组化和免疫印迹检测Niban表达.采用TUNE L法检测细胞凋亡。分别在HK-2细胞中转染Nibansi RNA和过表达Niban质粒，探讨Niban在血管紧张素II（AngII）和内质网（ER）应激损伤中的凋亡相关机制。.结果：随着梗阻的发展，Niban的表达逐渐减少，而凋亡增加。沉默Niban不仅增加了AngII和ER应激诱导的细胞凋亡，而且促进了caspase8、caspase9、Bip和Chop的表达。过表达Niban可降低AngII诱导的细胞凋亡，降低caspase8和caspase9的表达。.结论：Niban蛋白可能通过caspase依赖的途径参与HK-2细胞凋亡调控。