Qingshen granule-medicated serum attenuates high glucose-induced epithelial-mesenchymal transition of HK-2 cells via inhibiting oxidative stress-mediated NF-κB signaling pathway
10.3969/j.issn.1000-4718.2022.07.011
- VernacularTitle:清肾颗粒含药血清通过抑制氧化应激介导的NF-κB信号通路减轻高糖诱导的肾小管上皮细胞转分化
- Author:
Lei ZHANG
1
;
Hua JIN
;
Dong WANG
;
Meng CHENG
;
Zhuo-Ya LI
;
Yi-Ping WANG
Author Information
1. 安徽中医药大学第一附属医院肾内科,安徽合肥230031
- Keywords:
Qingshen granules;
NF-κB signaling pathway;
HK-2 cells;
Epithelial-mesenchymal transition;
Oxidative stress
- From:
Chinese Journal of Pathophysiology
2022;38(7):1234-1245
- CountryChina
- Language:Chinese
-
Abstract:
AIM:To investigate whether Qingshen granules(QSG)-medicated serum inhibits oxidative stress-mediated NF-κB signaling pathway and attenuates epithelial-mesenchymal transition(EMT)of human proximal tubule epi?thelial HK-2 cells induced by high glucose. METHODS:The active components in QSG were analyzed by HPLC. The HK-2 cells were randomly divided into control group,mannitol group,high glucose group,low-dose QSG group,medium-dose QSG group,high-dose QSG group and pyrrolidine dithiocarbamate(PDTC)group. The morphological changes of the cells were observed by inverted phase contrast microscopy. MTT assay was used to detect the cell viability. Flow cytometry was used to detect the content of reactive oxygen species(ROS)in HK-2 cells. ELISA was used to detect the content of malondi?al dehyde(MDA)and the activity of superoxide dismutase(SOD). The DNA binding activity of nuclear factor-κB(NF-κB) p65 in HK-2 cells was detected by electrophoretic mobility-shift assay(EMSA). The protein expression of NF-κB p65, phosphorylated inhibitor of kappa B alpha(p-IκBα),inhibitor of kappa B kinase alpha(IKKα),monocyte chemoattractant protein-1(MCP-1)and intercellular adhesion molecule-1(ICAM-1)in HK-2 cells was detected by Western blot. Immuno?fluorescence staining was used to detect NF-κB p65 andα-smooth mucle actin(α-SMA)protein expression in HK-2 cells. RESULTS:Chlorogenic acid,berberine hydrochloride,plantamajoside,6,7-dimethoxycoumarin,epiberberine,copti?sine,lithospermicacid B,palmatine,leonurine hydrochloride,rheic acid and tanshinone ⅡA in QSG were preliminarily de?termined by HPLC. Compared with control group,the levels of ROS and MDA in HK-2 cells induced by high glucose in?creased(P<0. 05),while the activity of SOD decreased(P<0. 05). The protein levels of NF-κB p65,p-IκBα,IKKα, MCP-1,ICAM-1 andα-SMA were increased(P<0. 05). After intervened by QSG-medicated serum,the levels of ROS and MDA were decreased(P<0. 05),while the activity of SOD was increased(P<0. 05). The protein levels of NF-κB p65,p-IκBα,IKKα,MCP-1,ICAM-1 andα-SMA were decreased(P<0. 05). CONCLUSION:QSG-medicated serum inhibited oxidative stress-mediated NF-κB signaling pathway,thus attenuating the EMT of HK-2 cells induced by high glucose.
