Liang Xinyue, Xu Wen
Department of Geriatrics, Renji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200127, P.R. China.
State Key Laboratory of Bioreactor Engineering and Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, Shanghai 200127, P.R. China.
Exp Ther Med. 2020 Aug;20(2):1121-1128. doi: 10.3892/etm.2020.8780. Epub 2020 May 21.
Diabetic nephropathy (DN) is a chronic loss of kidney function that frequently occurs in patients with diabetes mellitus and is characterized by abnormal glomerular mesangial cell (GMC) proliferation and apoptosis. By using microarray analysis, microRNA (miR)-181a-5p has previously been identified to be dysregulated in DN. The present study aimed to determine the underlying molecular mechanisms and function of miR-181a-5p in GMCs under DN conditions. First, reverse transcription-quantitative PCR was performed to detect miR-181a-5p and kruppel-like factor 6 (KLF6) expression in GMCs following high-glucose treatment. Subsequently, MTT and flow cytometric assays were performed in order to determine the effect of miR-181a-5p and KLF6 on high-glucose-driven GMC proliferation and apoptosis. After confirming that KLF6 was a target gene of miR-181a-5p via a bioinformatics analysis and luciferase reporter assay, the mRNA and protein expression levels of associated factors in different treatment groups were measured. The results demonstrated that miR-181a-5p was significantly downregulated, while KLF6 was significantly upregulated in GMCs following treatment with high glucose. Furthermore, overexpression of miR-181a led to suppression of cell proliferation and promoted apoptosis of GMCs induced by high glucose, while these effects were inhibited by co-transfection with KLF6. Finally, miR-181-5p was demonstrated to inhibit the expression of KLF6, Bcl-2, Wnt1 and β-catenin, while increasing the expression levels of Bax and caspase-3. In conclusion, the expression levels of miR-181a-5p were downregulated in GMCs following treatment with high glucose and overexpression of miR-181a-5p may inhibit GMC proliferation and promote apoptosis, at least partially through targeting KLF6 via the Wnt/β-catenin signaling pathway. Overall, the results of the present study suggest that miR-181a-5p may have a crucial role in the occurrence and development of DN and may be a valuable diagnostic marker and therapeutic target for DN.
糖尿病肾病（DN）是一种常见于糖尿病患者的慢性肾功能丧失疾病，其特征为肾小球系膜细胞（GMC）异常增殖和凋亡。通过微阵列分析，先前已确定微小RNA（miR）-181a-5p在DN中表达失调。本研究旨在确定DN条件下miR-181a-5p在GMC中的潜在分子机制和功能。首先，进行逆转录定量PCR以检测高糖处理后GMC中miR-181a-5p和 kruppel样因子6（KLF6）的表达。随后，进行MTT和流式细胞术检测，以确定miR-181a-5p和KLF6对高糖驱动的GMC增殖和凋亡的影响。通过生物信息学分析和荧光素酶报告基因检测确认KLF6是miR-181a-5p的靶基因后，测量不同处理组中相关因子的mRNA和蛋白质表达水平。结果表明，高糖处理后GMC中miR-181a-5p显著下调，而KLF6显著上调。此外，miR-181a的过表达导致细胞增殖受到抑制，并促进高糖诱导的GMC凋亡，而与KLF6共转染可抑制这些作用。最后，证明miR-181-5p抑制KLF6、Bcl-2、Wnt1和β-连环蛋白的表达，同时增加Bax和caspase-3的表达水平。总之，高糖处理后GMC中miR-181a-5p的表达水平下调，miR-181a-5p的过表达可能至少部分通过Wnt/β-连环蛋白信号通路靶向KLF6来抑制GMC增殖并促进凋亡。总体而言，本研究结果表明miR-181a-5p可能在DN的发生和发展中起关键作用，并且可能是DN的有价值的诊断标志物和治疗靶点。
