TNFR1/NF-κB通路介导的炎症反应在肾小管上皮细胞损伤及肾草酸钙结石形成中的机制研究

Renal calculus affect around 6.0% of total population, among them 80% are calcium oxalate stones. Despite great patient population, the underlying mechanisms that cause the disease are still poorly understood. Recent research shows tubular epithelial injury is the key for disease onset and inflammation plays dominant role in it. TNFR1 and downstream NF-κB signaling pathway plays major role in the onset of many inflammatory diseases. Our result shows that high oxalate could induce the expression of TNFR1 and activate downstream NF-κB signaling pathway. High oxalate could also induce apoptosis. In mouse kidney calcium oxalate stone model, serum level of proinflammatory cytokines which are target genes of NF-κB signaling pathway increased dramatically, severe apoptosis is also detected in tubular epithelial, kidney damage marker is also increased. Based on that, we hypothesis that TNFR1 might regulate inflammatory response through its downstream NF-κB signaling to promote the formation of kidney stones. A combined approach including siRNA, inhibitors will be used together on cell culture, animal model and human tissue specimen to uncover the detailed mechanism of high oxalate induced activation of TNFR1 and downstream NF-κB pathway, to explore the role of activated NF-κB signaling in the damage of tubular epithelial and understand the molecular mechanism behind kidney oxalate stone disease, so to provide novel treatment and marker for kidney oxalate stone disease.

肾结石患病率达6.0%，其中约80%为草酸钙结石，其具体发病机制不明。目前研究认为肾小管上皮细胞损伤是结石形成的关键，炎症反应在其中起重要作用，而TNFR1及下游NF-κB信号通路参与了多种炎症反应相关疾病的发生。课题组预实验发现：高草酸刺激可上调TNFR1蛋白表达，并可激活细胞内TNFR1下游NF-κB通路；同时，高草酸还可诱导细胞发生凋亡；在小鼠肾草酸钙结晶模型中，血清NF-κB信号通路下游促炎性因子显著上调，肾小管上皮细胞出现明显凋亡。我们猜想，TNFR1复合物可能通过下游NF-κB信号通路调控炎症反应从而促进肾结石形成。课题组拟应用siRNA敲低、抑制剂处理等方法，分别在细胞、小鼠模型及人组织标本上进行检测和验证，揭示高草酸激活TNFR1下游NF-κB信号通路的机制，探索其在肾小管上皮细胞损伤中的作用，阐明肾草酸钙结石形成的分子机制，为临床预防和治疗肾草酸钙结石提供新靶点。

草酸钙结石是肾结石中的主要类型，不但发病率高，且易复发，其具体发病机制尚不明确，目前临床上除手术治疗外，尚无有效的预防措施。既往研究认为肾小管上皮细胞损伤是草酸钙结石形成的关键步骤。本项目探讨了内质网应激和NF-κB信号通路在草酸诱导的肾小管上皮细胞损伤中的作用及其可能的分子机制。结果发现，草酸刺激后，小鼠肾组织肿胀，肾小管上皮细胞凋亡增多，并可激活内质网信号通路和NF-κB信号通路。细胞实验表明，草酸可诱导HK-2细胞自噬、凋亡和线粒体损伤，激活ERS和NF-κB通路；干预NF-κB表达可抑制NF-κB信号通路，HK-2细胞仍处于较高水平的ERS，但凋亡率明显下降。实验结果为草酸钙结石的预防和治疗提供了一个新思路和方向。