Feng Fen, Yang Jie, Wang Gang, Huang Ping, Li Yongjie, Zhou Bin
School of Pharmacy, Shaoyang University, Shaoyang, China.
Department of Endocrinology, The Central Hospital of Shaoyang, Shaoyang, China.
Nephron. 2023;147(3-4):212-222. doi: 10.1159/000525440. Epub 2022 Jul 22.
Many studies have confirmed that circular RNA (circRNA) is an important target for regulating human disease progression. This study aimed to explore the role of circ_0068087 in diabetic nephropathy (DN) progression.
High glucose (HG)-induced renal tubular cells (HK2) were used to mimic DN cell models in vitro. The expression levels of circ_0068087, microRNA (miR)-106a-5p, and Rho-associated coiled-coil-containing kinase 2 (ROCK2) were detected by quantitative real-time PCR. Cell proliferation and apoptosis were examined by cell counting kit-8 assay, 5-ethynyl-2'-deoxyuridine assay, colony formation assay, and flow cytometry. The protein levels were examined by Western blot analysis. Cell oxidative stress was assessed by measuring MDA level and SOD activity, and cell inflammation was evaluated by detecting the concentrations of inflammatory factors. RNA interaction was verified by dual-luciferase reporter assay and RNA pull-down assay.
The present study showed that circ_0068087 was highly expressed in the serum of DN patients and HG-induced HK2 cells. Interference of circ_0068087 alleviated HG-induced apoptosis, oxidative stress, inflammation, and fibrosis in HK2 cells, while accelerated cell proliferation. miR-106a-5p could be sponged by circ_0068087, and its inhibitor eliminated the regulation of circ_0068087 knockdown on HG-induced HK2 cell injury. ROCK2 was a target of miR-106a-5p, and its expression was suppressed by circ_0068087 knockdown. miR-106a-5p overexpression suppressed HG-induced HK2 cell injury, and this effect was reversed by ROCK2 upregulation.
Our data indicated that circ_0068087 downregulation mitigated HG-induced HK2 cell injury through the miR-106a-5p/ROCK2 axis, providing a potential circRNA-targeted therapy for DN.
许多研究证实，环状RNA（circRNA）是调节人类疾病进展的重要靶点。本研究旨在探讨circ_0068087在糖尿病肾病（DN）进展中的作用。
使用高糖（HG）诱导的肾小管细胞（HK2）在体外模拟DN细胞模型。通过定量实时PCR检测circ_0068087、微小RNA（miR）-106a-5p和Rho相关卷曲螺旋蛋白激酶2（ROCK2）的表达水平。通过细胞计数试剂盒-8检测、5-乙炔基-2'-脱氧尿苷检测、集落形成检测和流式细胞术检测细胞增殖和凋亡。通过蛋白质印迹分析检测蛋白质水平。通过测量丙二醛水平和超氧化物歧化酶活性评估细胞氧化应激，并通过检测炎症因子浓度评估细胞炎症。通过双荧光素酶报告基因检测和RNA下拉检测验证RNA相互作用。
本研究表明，circ_0068087在DN患者血清和HG诱导的HK2细胞中高表达。干扰circ_0068087可减轻HG诱导的HK2细胞凋亡、氧化应激、炎症和纤维化，同时促进细胞增殖。miR-106a-5p可被circ_0068087吸附，其抑制剂消除了circ_0068087敲低对HG诱导的HK2细胞损伤的调节作用。ROCK2是miR-106a-5p的靶标，其表达受到circ_0068087敲低的抑制。miR-106a-5p过表达抑制了HG诱导的HK2细胞损伤，而ROCK2上调可逆转这种作用。
我们的数据表明，circ_0068087下调通过miR-106a-5p/ROCK2轴减轻HG诱导的HK2细胞损伤，为DN提供了一种潜在的circRNA靶向治疗方法。
