Protective effects of gliclazide on high glucose and AGEs-induced damage of glomerular mesangial cells and renal tubular epithelial cells via inhibiting RAGE-p22phox-NF-kB pathway.

Abstract

Gliclazide is one of the most widely used therapeutic drugs for diabetes. As a second-generation sulfonylurea oral hypoglycemic drug, it can lower blood glucose level and delay the occurrence and development of diabetic nephropathy (DN). However, the underlying mechanism remains unclear. Therefore, the aim of this study was to explore whether gliclazide had protective effects on high glucose and advanced glycation end products (AGEs)-induced injury of human mesangial cells (HMCs) and renal tubular epithelial cells. HMC and renal tubular epithelial cell lines [human kidney 2 (HK-2)] were cultured in vitro. All cells were then divided into the follow groups: 1) blank control group (5.6 mmol/L glucose), 2) AGEs group [400 μg/mL AGE-bovine serum albumin (AGE-BSA)], 3) high glucose group (25 mmol/L glucose), 4) gliclazide + AGEs group (400 μg/mL AGE-BSA + 20 μmol/L gliclazide) and 5) gliclazide + high glucose group (25 mmol/L glucose + 20 μmol/L gliclazide). Cell counting kit-8 (CCK-8) assay was adopted to determine cell viability. Flow cytometry was used to detect cell apoptosis. The levels of malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) were measured as well. Furthermore, the mRNA expressions of receptor for AGE (RAGE), p22phox and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) were measured via fluorescence quantitative Real-time polymerase chain reaction (qRT-PCR). Compared with control group, significantly accelerated apoptosis of HMCs and HK-2, increased MDA level, decreased SOD and GSH-Px levels, and up-regulated mRNA expressions of RAGE, p22phox and NF-κB were observed in HMCs and HK-2 of high glucose group and AGEs group. Meanwhile, there were obviously alleviated apoptosis of HMCs and HK-2, decreased MDA level, increased SOD and GSH-Px levels, as well as down-regulated mRNA expressions of RAGE, p22phox and NF-κB in HMCs and HK-2 of gliclazide group compared with high glucose and AGEs group. Furthermore, significant correlations were found between the mRNA expression of RAGE and the apoptosis rate of HMCs and HK-2 (HMCs: r=0.701, p=0.004 and HK-2: r=0.633, p=0.011). Gliclazide has protective effects on high glucose and AGEs-induced damage of glomerular mesangial cells and renal tubular epithelial cells via inhibiting RAGE-NADPH oxidase-NF-kB pathway.