Cao Yaochen, Hu Jialin, Sui Jianying, Jiang Limei, Cong Yakun, Ren Guoqing
Department of Nephrology, Daqingshi No. 4 Hospital, Daqing, Heilongjiang 163712, P.R. China.
Exp Ther Med. 2018 Sep;16(3):2442-2448. doi: 10.3892/etm.2018.6489. Epub 2018 Jul 20.
Patients with chronic kidney disease (CKD) are characterized by a gradual loss of kidney function over time. A number of studies have indicated that tubule interstitial fibrosis (TIF) is associated with the occurrence and development of CKD. The aim of the present study was to investigate the effect of quercetin treatment on the fibrosis of renal tubular epithelial cells and to determine whether the anti-fibrotic effects of quercetin are achieved via microRNA (miR)-21. Human tubular epithelial HK-2 cells were cultured with transforming growth factor (TGF)-β to induce fibrosis and the expression of fibrotic markers collagen I, fibronectin, α-smooth muscle actin (SMA) and epithelial-cadherin were measured using reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blotting. Cells were treated with 7.5, 15 or 30 mg/ml quercetin, following which fibrosis and miR-21 expression were evaluated. Quercetin-treated cells were transfected with miR-21 mimics and the expression of fibrotic markers was examined using RT-qPCR. Finally, the expression of fibrosis-associated miR-21 target genes, phosphatase and tensin homolog (PTEN) and TIMP Metallopeptidase Inhibitor 3 (TIMP3), was measured in cells treated with quercetin with or without miR-21 mimics using RT-qPCR, western blotting and immunocytochemistry. The results revealed that TGF-β treatment induced a significant increase in the expression of fibrotic markers in HK-2 cells, while quercetin treatment partially inhibited the fibrosis of HK-2 cells. Furthermore, quercetin treatment significantly inhibited TGF-β-induced miR-21 upregulation and transfection with miR-21 mimics reversed the anti-fibrotic effects of quercetin. Quercetin treatment markedly upregulated PTEN and TIMP3 expression, whereas transfection with miR-21 mimics reversed this effect. The results of the present study suggest that quercetin is able to alleviate TGF-β-induced fibrosis in HK-2 cells via suppressing the miR-21 and upregulating PTEN and TIMP3. Quercetin may have potential as an anti-fibrotic treatment for patients with renal fibrosis.
慢性肾脏病(CKD)患者的特征是肾功能随时间逐渐丧失。多项研究表明,肾小管间质纤维化(TIF)与CKD的发生和发展相关。本研究的目的是探讨槲皮素治疗对肾小管上皮细胞纤维化的影响,并确定槲皮素的抗纤维化作用是否通过微小RNA(miR)-21实现。用转化生长因子(TGF)-β培养人肾小管上皮HK-2细胞以诱导纤维化,并用逆转录-定量聚合酶链反应(RT-qPCR)和蛋白质印迹法检测纤维化标志物I型胶原、纤连蛋白、α-平滑肌肌动蛋白(SMA)和上皮钙黏蛋白的表达。用7.5、15或30mg/ml槲皮素处理细胞,然后评估纤维化和miR-21表达。用miR-21模拟物转染经槲皮素处理的细胞,并用RT-qPCR检测纤维化标志物的表达。最后,用RT-qPCR、蛋白质印迹法和免疫细胞化学法检测在用或不用miR-21模拟物处理的经槲皮素处理的细胞中纤维化相关miR-21靶基因、磷酸酶和张力蛋白同源物(PTEN)以及金属蛋白酶组织抑制因子3(TIMP3)的表达。结果显示,TGF-β处理可显著增加HK-2细胞中纤维化标志物的表达,而槲皮素处理可部分抑制HK-2细胞的纤维化。此外,槲皮素处理可显著抑制TGF-β诱导的miR-21上调,用miR-21模拟物转染可逆转槲皮素的抗纤维化作用。槲皮素处理可显著上调PTEN和TIMP3表达,而用miR-21模拟物转染可逆转此作用。本研究结果表明,槲皮素能够通过抑制miR-21并上调PTEN和TIMP3来减轻TGF-β诱导的HK-2细胞纤维化。槲皮素可能具有作为肾纤维化患者抗纤维化治疗药物的潜力。