Department of Nephrology, Qingdao Municipal Hospital, No.5, Middle Donghai Road, Qingdao, 266071, Shandong, China.
Department of Nephrology, Qingdao Haici Med Ctr, Qingdao, China.
Mol Cell Biochem. 2021 Aug;476(8):3051-3064. doi: 10.1007/s11010-021-04140-2. Epub 2021 Apr 2.
MiR-17 is found upregulated in diabetic mice; however, its effect(s) on renal fibrosis of diabetic nephropathy remain(s) unknown. This study aimed to explore the mechanism underlying the downregulation of miR-17 in renal fibrosis of diabetic nephropathy (DN). Patients with diabetes mellitus (DM) and DN and normal healthy individual controls, mice (db/db, db/m), and human mesangial cells (HMCs) and human proximal tubule epithelial cells (HK-2) were used as research subjects in the study. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to measure the expression of miR-17 in the serum samples, renal tissues and cells. Acid-Schiff (PAS) and Masson staining experiments were performed to detect glomerular mesangial matrix and collagen deposition. Levels of fibrosis-related proteins (E-Cadherin (E-cad), vimentin, fibronectin and collagen I) were measured by Western blot (WB). The target gene of miR-17 was predicted by TargetScan 7.2 and confirmed by dual-luciferase reporter analysis. The study found that miR-17 expression was elevated in the serums of DN patients as well as in the serums and kidney tissues of db/db mice. db/db mice showed a severe renal fibrosis condition. The levels of E-cad in db/db mice, HMC and HK-2 cells were increased by downregulating miR-17 expression, while expressions of vimentin, fibronectin and collagen I were reduced. Smad7 was predicted to be the target gene of miR-17, and its expression was promoted by downregulation of miR-17. Moreover, the reduced Smad7 expression could inhibit the expressions of fibrosis-related proteins, which, however, can be ameliorated by the downregulation of miR-17. In addition, downregulation of miR-17 could suppress renal fibrosis mediated by TGF-β1 through targeting Smad7, which might be a clinical therapeutic target for patients with DN.
miR-17 在糖尿病小鼠中上调；然而，其对糖尿病肾病（DN）肾纤维化的影响尚不清楚。本研究旨在探讨 miR-17 在糖尿病肾病肾纤维化中下调的机制。糖尿病患者和糖尿病肾病患者以及正常健康个体对照、小鼠（db/db、db/m）以及人肾小球系膜细胞（HMC）和人近端肾小管上皮细胞（HK-2）被用作研究对象。定量实时聚合酶链反应（qRT-PCR）用于测量血清样本、肾组织和细胞中 miR-17 的表达。酸-Schiff（PAS）和 Masson 染色实验用于检测肾小球系膜基质和胶原沉积。Western blot（WB）用于测量纤维化相关蛋白（E-钙粘蛋白（E-cad）、波形蛋白、纤连蛋白和胶原 I）的水平。miR-17 的靶基因通过 TargetScan 7.2 预测，并通过双荧光素酶报告分析确认。研究发现，DN 患者血清以及 db/db 小鼠血清和肾组织中 miR-17 表达升高。db/db 小鼠表现出严重的肾纤维化。下调 miR-17 表达可增加 db/db 小鼠、HMC 和 HK-2 细胞中的 E-cad 水平，同时降低波形蛋白、纤连蛋白和胶原 I 的表达。Smad7 被预测为 miR-17 的靶基因，其表达受 miR-17 下调的促进。此外，下调 miR-17 可通过下调 Smad7 抑制纤维化相关蛋白的表达，但可通过下调 miR-17 得到改善。此外，下调 miR-17 通过靶向 Smad7 抑制 TGF-β1 介导的肾纤维化，这可能是 DN 患者的临床治疗靶点。
