RNA结合蛋白NOVA1调控p53稳定性在脓毒症致肾小管上皮细胞凋亡中的分子机制研究

Apoptosis of renal tubular epithelial cells (RTEC) plays a key role in acute renal injury (AKI) induced by sepsis, but the specific molecular mechanism is unclear. Researches at home and abroad in recent years and our early study indicate that p53 is an important mediator of RTEC apoptosis. In our preliminary study, it was observed that NOVA1, a RNA-binding protein, was significantly increased in the RTEC of sepsis-AKI rats; silencing of NOVA1 decreased the apoptosis of RTEC induced by lipopolysaccharide (LPS). Moreover, NOVA1 was proved to increase the stability of p53 mRNA, and NOVA1 protein level was positively regulated by lncRNA MALAT1. We therefore inferred that up-regulated MALAT1 increases the expression of NOVA1 protein and then stabilizes p53 mRNA, thereby transcriptional activating its downstream apoptotic signaling pathway to induce apoptosis of RTEC. The aim of this study is to determine the role of NOVA1 in RTEC apoptosis in vivo and in vitro by means of cell biology and molecular biology techniques. This study will elucidate the probable mechanism of NOVA1 in the sepsis-induced RTEC apoptosis and also provide a key target for the treatment of AKI.

肾小管上皮细胞（RTEC）凋亡在脓毒症致急性肾损伤（AKI）中起着关键性作用，但具体分子机制不清楚。近年国内外研究和我们的前期工作表明p53是RTEC凋亡的重要介导分子。本课题前期研究显示，脓毒症致AKI大鼠RTEC中RNA结合蛋白NOVA1表达显著升高；干扰NOVA1降低脂多糖诱导的RTEC凋亡；此外，NOVA1能够增加p53 mRNA稳定性，而NOVA1蛋白水平受lncRNA MALAT1正向调节。由此，我们推测，肾小管上皮细胞中，高表达MALAT1上调NOVA1蛋白水平，稳定p53 mRNA，增加其蛋白表达，转录活化其下游凋亡信号通路，诱导肾小管上皮细胞凋亡。该项目旨在细胞和模式动物水平上，借助于分子生物学、细胞生物学等技术手段，研究NOVA1在RTEC凋亡中的作用。本项目的完成不仅有助于阐明NOVA1在脓毒症致RTEC凋亡中的内在机制，并为AKI治疗提供一个直接的作用靶点。

越来越多的证据证实，肾小管上皮细胞（RTEC）的凋亡是脓毒症急性肾损伤（AKI）发病和发展的关键因素，但脓毒症急性肾损伤（AKI）期间RTEC凋亡上调的病理机制尚不完全清楚。在前期的实验中，我们已经发现lncRNA NONRATG019935.2 (9935)的表达水平在败血症AKI大鼠和LPS处理的NRK-52E细胞（大鼠RTEC系）中均表现出最显著的降低。在本研究中，我们发现9935通过限制RTEC中人类抗原R（HuR）和Tp53 mRNA的3′UTR区的结合，降低了人类抗原R（HuR）介导的Tp53 mRNA稳定性。HuR的过度表达消除了pcDNA-9935对LPS诱导的NRK-52E和大鼠原代RTEC凋亡的抑制作用。总之，9935通过抑制p53介导的RTEC凋亡在败血症性AKI中发挥作用，而9935的这一重要作用依赖于其对HuR介导的Tp53 mRNA稳定性的破坏作用。这提示外源性补充9953可以减轻LPS诱导的人RTEC的损伤，为临床脓毒症AKI的治疗提供了有益的参考。