EVI-1/NM IIA/MG53通路在急性肾小管损伤中的作用研究

AKI is a common clinical syndrome with elusive molecular mechanisms. Non-muscle myosin IIA (NM IIA) is a molecular motor protein. It can carry MG53 to the cell membrane to contribute to the membrane repair. Our preliminary data showed that NM IIA and MG53 were significantly reduced in kidney tissues of AKI mice and cisplatin-treated RTECs. Silencing NM IIA significantly reduced the expression of MG53, while NM IIA agonist could increase MG53 expression. Meanwhile, Overexpression of NM IIA or MG53 inhibited RTEC apoptosis induced by cisplatin, suggesting that NM IIA might be of importance in AKI by modulating MG53. In addition, bioinformatics analysis showed multiple binding sites of transcription factor EVI-1 in NM IIA promoter region. We also found EVI-1 was significantly reduced in kidney tissues of cisplatin-induced AKI mice and cisplatin-treated RTECs. More importantly, silencing EVI-1 reduced NM IIA expression in RTECs. Thus, we hypothesized that the insults of AKI, such as toxins, ischemia and ischemia, et al. could reduce EVI-1 expression to block the transcription of NM IIA transcription, leading to the renal tubular injury via reducing the MG53 levels. In the present proposal, employing the models of animal, cell and molecule, we will fully investigate the role of EVI-1/NM IIA/MG53 pathway in the pathogenesis of AKI. The results from current study will provide new insights into the understanding and therapy of AKI.

AKI是临床多发病，但具体分子机制尚需研究。非肌肉肌球蛋白NM IIA被报道可以通过运输E3泛素连接酶MG53参与细胞膜修复。预实验发现AKI小鼠及顺铂处理的肾小管上皮细胞（RTEC）的NM IIA和MG53表达下调；NM IIA siRNA能下调RTEC中MG53表达，而NM IIA激动剂则上调MG53；同时，过表达NM IIA及MG53能明显抑制顺铂诱导的细胞凋亡，提示NM IIA可能通过调控MG53参与了急性肾小管损伤的发生。生物信息学分析发现转录因子EVI-1在NM IIA启动子区有2个结合位点，在顺铂处理的AKI小鼠及RTEC中都下调，敲低EVI-1降低了NM IIA。由此推测：致AKI因素通过下调EVI-1导致了NM IIA表达降低，进而通过抑制MG53导致RTEC损伤和AKI。本课题将在动物、细胞和分子水平深入研究这一通路在急性肾小管损伤中的作用，为临床防治AKI提供新视点。

AKI是临床多发病，但具体分子机制尚需研究。非肌肉肌球蛋白NM IIA被报道可以通过运输E3泛素连接酶MG53参与细胞膜修复。实验发现在AKI患者肾活检组织及顺铂诱导的体内外模型中，NM IIA表达下调；肾小管特异性敲除NM IIA会加重顺铂及缺血再灌注导致的AKI；体内外过表达NM IIA及MG53或给予NM IIA激动剂CA均能明显抑制顺铂诱导的AKI；通过基因调控和免疫共沉淀发现NM IIA可以调控MG53；通过转录组学分析发现肾小管特异性敲除NM IIA会显著影响PI3K-AKT通路；通过基因干预和双荧光素酶报告基因发现EVI-1可能是NM IIA的转录调节因子，EVI-1在顺铂模型中表达下调，体外过表达EVI-1可以改善顺铂诱导的细胞凋亡。由此推测：致AKI因素通过下调EVI-1导致了NM IIA表达降低，进而通过抑制MG53及PI3K通路的激活导致RTEC损伤和AKI发生。此外证实，中药单体DT-13作为NM IIA激动剂也能改善AKI。本研究在动物、细胞和分子水平深入研究这一通路在急性肾小管损伤中的作用，为临床防治AKI提供新视点和潜在靶向治疗药物。