miR-451调控糖尿病肾病细胞外基质增生的LMP7/NF-κB信号通路研究

Extracellular matrix accumulation of glomerular mesangial cells plays an important role in the development of diabetic nephropathy (DN). NF-κB signals abnormality is closely associated with the accumulation of extracellular matrix. However, the mechanism of NF-κB activity in DN is not fully clear. Here, we emphasized the key role of miRNA in the aberrant activation of NF-κB. Our previous studies found that the expression of miRNA-451 was significantly decreased in the kidney of DN mice by miRNA Microarray Assay. Moreover, we indicated that immunosubunit LMP7 (encoded by psmb8) was a target for miRNA-451 by Dual Glo Luciferase assay and Western blot. More importantly, reduced expression of miRNA-451 and concomitant enhanced expression of LMP7 were observed in the kidney of DN mice..In this project, how miRNA-451 activated NF-κB pathway by targeting of LMP7, and thus, induced an enhanced secretion of cytokine and chemotactic factors, will be discussed. The effect of miR-451 on the proliferation of mesangial cells and deposition of extracellular matrix will be examined by upregulating and downregulating the expression of miR-451. Further, the influence of miRNA-451 on the expression of LMP7 and key signaling molecules in NF-κB pathway in vitro and in vivo will be investigated. The present study may help to elucidate the physiopathological mechanism of DN progression.

肾小球系膜细胞外基质聚积是糖尿病肾病(DN)进展的重要原因之一，核因子-κB（NF-κB）信号异常与细胞外基质增生密切相关，但DN发生发展中NF-κB活化的机制尚不清楚。本项目提出miRNA在NF-κB异常活化中起关键作用。我们前期工作发现miRNA-451在DN小鼠显著下调，双荧光素酶报告系统和Western blot证实psmb8基因（编码LMP7）是miR-451的靶基因，观察到DN小鼠肾脏组织miR-451表达降低与LMP7高表达同时存在。.我们将研究miR-451通过调节靶基因LMP7的表达，从而影响NF-κB活化，引发一系列细胞因子和趋化因子分泌增强的机理。采用分别上调和下调肾小球系膜细胞中miR-451的表达，研究其对系膜细胞增殖和细胞外基质增生的影响，体外和体内实验研究miR-451对LMP7及NF-κB信号途径中关键分子表达的影响。本研究有助于阐明DN发生的病理生理机制。

核因子-kB（NF-kB）信号异常与肾小球外基质增生和炎症密切相关，而miR-451与糖尿病肾损害密切相关。大型多功能蛋白酶7（LMP7），一种免疫蛋白酶体亚基，可以激活NF-kB。然而，目前仍不清楚是否miR-451 通过调节LMP7 影响NF-kB引起DN炎症。在这项研究中，我们分别使用了原位杂交，实时定量PCR，双荧光素酶报告基因检测，Western blot和染色质免疫沉淀技术。结果发现，在db/db小鼠肾脏，DN患者的外周血单个核细胞和在高糖条件下培养的肾小球系膜细胞中发现miR-451明显下调。此外，miR-451直接通过降低LMP7表达抑制NF-kB活性，下调系膜细胞中促炎症分子的转录。同时，在db/db糖尿病小鼠肾脏中通过尾静脉增加miR-451水平后，明显抑制LMP7 / NF-kB活性，减少尿微量白蛋白、血糖、肾小球外基质增生和肾小球损伤。总之，这些结果证实DN中，miR-451可以通过调节LMP7 / NF-kB炎症通路缓解糖尿病肾病发病过程。