Abstract
Aim. The purpose of this study was to investigate the effects of pioglitazone on oxidative stress and the expressions of p22phox and p47phox, subunits of NADPH oxidase, in mesangial cells (MCs). Method. Rat mesangial cells were cultured and randomly divided into normal glucose (NG) group, high glucose (HG) group, and pioglitazone group. After 48 h exposure, the supernatants and cells were collected. The expressions of p22phox and p47phox in MCs were detected by RT-PCR and western blot. The levels of intracellular ROS were determined by flow cytometry. Coloimetry method was used to detect malondialdehyde (MDA) concentrations and superoxide dismutase (SOD) activities. Results. Compared with the NG group, the expression levels of p22phox, p47phox and ROS significantly increased, the activity of SOD decreased in HG group, while the concentration of MDA greatly increased (P < 0.01). Pioglitazone significantly suppressed HG-induced p22phox and p47phox expressions and oxidative stress. The protein and gene expressions of p22phox and p47phox were markedly reduced after pioglitazone treatment, so did the ROS generation. The activities of SOD in MCs increased, while the concentrations of MDA in the supernatant decreased greatly by pioglitazone. Conclusions. Pioglitazone can inhibit HG-induced oxidative stress in MCs through suppressing p22phox and p47phox expressions.
Highlights
Diabetic nephropathy (DN) is the most common cause of end stage renal failure and is a chronic disease characterized by proteinuria, glomerular hypertrophy, decreased glomerular filtration, and renal fibrosis with loss of renal function
HyClone (UT, USA); fetal bovine serum (FBS) was purchased from Hangzhou Sijiqing Biological Engineering Materials Co., Ltd. (Hangzhou, China); trypsin, BCA protein assay kit, and the ECL Plus kit were from Beyotime Institute of Biotechnology (Nantong, China); pioglitazone was a gift from Jiang Su Heng Rui Medicine Co., Ltd. (Lianyungang, China); 2,7-dichlorofluorescein diacetate (DCFH-DA) and ethidium bromide (EB) were from Sigma (MO, USA); rabbit p22phox, p47phox antibody and goat anti-rabbit IgG were purchased from Santa Cruz Biotechnology Inc. (CA, USA); β-actin was from Sangon Biological Engineering Technology Corporation (Shanghai, China); TRIzol was obtained from Invitrogen (CA, USA); reverse transcription reagents and pcr kit were from TaKaRa Biotechnology Co., Ltd. (Dalian, China); superoxide dismutase (SOD) and malonaldehyde (MDA) assay were from Nanjing Jiancheng Bioengineering Institute (Nanjing, China)
high glucose (HG) is the key factor contributing to long term complications of diabetes mellitus, including DN, which is characterized by mesangial cells proliferation, extracellular matrix accumulation, and tubulointerstitial fibrosis in the glomerular [9], but the molecular mechanisms of chronic complications caused by the exposure to elevated levels of glucose remain to be completely defined
Summary
Diabetic nephropathy (DN) is the most common cause of end stage renal failure and is a chronic disease characterized by proteinuria, glomerular hypertrophy, decreased glomerular filtration, and renal fibrosis with loss of renal function. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase is the predominant enzyme source for ROS generation, which is composed of five subunits comprising a membrane-associated p22phox, a gp91phox subunit and at least four cytosolic subunits: p47phox, p67phox, p40phox, and rac-1/2. Subunits such as p22phox and p47phox were mainly expressed in the kidney. Mesangial cells (MCs), an inherent cell of kidney, play an important role during the development and progression of chronic kidney disease, including DN [3]. The present study was undertaken to observe the effects of pioglitazone on expressions of p22phox and p47phox in rat glomerular mesangial cells in vitro and explore its potential antioxidative mechanisms
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
