Resveratrol retards progression of diabetic nephropathy through modulations of oxidative stress, proinflammatory cytokines, and AMP-activated protein kinase

Abstract

Background: Diabetic nephropathy (DN) has been recognized as the leading cause of end-stage renal disease. Resveratrol (RSV), a polyphenolic compound, has been indicated to possess an insulin-like property in diabetes. In the present study, we aimed to investigate the renoprotective effects of RSV and delineate its underlying mechanism in early-stage DN.

Methods: The protective effects of RSV on DN were evaluated in streptozotocin (STZ)-induced diabetic rats.

Results: The plasma glucose, creatinine, and blood urea nitrogen were significantly elevated in STZ-induced diabetic rats. RSV treatment markedly ameliorated hyperglycemia and renal dysfunction in STZ-induced diabetic rats. The diabetes-induced superoxide anion and protein carbonyl levels were also significantly attenuated in RSV-treated diabetic kidney. The AMPK protein phosphorylation and expression levels were remarkably reduced in diabetic renal tissues. In contrast, RSV treatment significantly rescued the AMPK protein expression and phosphorylation compared to non-treated diabetic group. Additionally, hyperglycemia markedly enhanced renal production of proinflammatory cytokine IL-1β. RSV reduced IL-1β but increased TNF-α and IL-6 levels in the diabetic kidneys.

Conclusions: Our findings suggest that RSV protects against oxidative stress, exhibits concurrent proinflammation and anti-inflammation, and up-regulates AMPK expression and activation, which may contribute to its beneficial effects on the early stage of DN.

Figure 1

The effects of RSV on mesangial expansion in DN. Photomicrographs of rodent glomeruli sections of CON (A), STZ-DM (B), DM-R0.1 (C) and DM-R1 (D) groups were represented at × 400 magnification from periodic acid-Schiff-stained kidney. CON: non-diabetic control, STZ-DM: streptozotocin-induced diabetes, DM-R0.1: DM treated with RSV (0.1 mg/kg/day) for 7 days, DM-R1: DM treated with RSV (1 mg/kg/day) for 7 days.

Figure 2

The effects of RSV on superoxide anion production, lipid peroxidation, protein carbonyl level, MnSOD and CuZnSOD protein expressions in the renal tissues of STZ-DM rats. (A) Superoxide anion content was measured by lucigenin-enhanced chemiluminescence. (B) Lipid peroxidation was determined using the modified thiobarbituric acid reactive substances method. (C) Protein carbonyl level was evaluated by 2,4-dinitrophenylhydrazine method. (D) Equal amounts of proteins were resolved on 10 and 15% SDS-PAGE and blotted with MnSOD and CuZnSOD antibodies, respectively. The blots were shown at the top and the quantified ratios were shown at the bottom. Results were expressed as means ± standard error (n = 5 per group). *: P < 0.05 vs. control, †: P < 0.05 vs. STZ-DM, ‡: P < 0.05 vs. DM-R0.1, RLU: relative light unit, LPO: lipid peroxidation, PCL: protein carbonyl level, SOD: superoxide dismutase, CON: non-diabetic control, STZ-DM: streptozotocin-induced diabetes, DM-R0.1: DM treated with RSV (0.1 mg/kg/day) for 7 days, DM-R1: DM treated with RSV (1 mg/kg/day) for 7 days.

Figure 3

The effects of RSV on IL-1β, TNF-α, IL-6, and NF-κB protein expression in the renal tissues of STZ-DM rats. Samples were processed to measure IL-1β (A), TNF-α (B), or IL-6 (C) using a competitive ELISA. (D) Equal amounts of proteins were resolved on 10% SDS-PAGE and blotted with NF-κB antibody. The blots were shown at the top and the quantified ratios were shown at the bottom. Results were expressed as means ± standard error (n = 5 per group). *: P < 0.05 vs. control, †: P < 0.05 vs. STZ-DM, ‡: P < 0.05 vs. DM-R0.1, IL-1β: interleukin 1β, TNF-α: tumor necrosis factor α, IL-6: interleukin 6, NF-κB: nuclear factor kappa B, CON: non-diabetic control, STZ-DM: streptozotocin-induced diabetes, DM-R0.1: DM treated with RSV (0.1 mg/kg/day) for 7 days, DM-R1: DM treated with RSV (1 mg/kg/day) for 7 days.

Figure 4

The effects of RSV on Erk, p38, JNK, Akt, and AMPK phosphorylation and total protein expressions in the renal tissues of STZ-DM rats. Equal amounts of proteins were resolved on 10% SDS-PAGE and blotted with Erk, p-Thr202/Tyr204-Erk, p38, p-Thr180/Tyr182-p38, JNK, p-Thr183/Tyr185-JNK (A), and Akt, p-Thr308-Akt, AMPK, p-Thr172-AMPK (B) antibodies. The blots were shown at the top and the quantified ratios were shown at the bottom. Results were expressed as means ± standard error (n = 5 per group). *: P < 0.05 vs. control, †: P < 0.05 vs. STZ-DM, ‡: P < 0.05 vs. DM-R0.1, CON: non-diabetic control, STZ-DM: streptozotocin-induced diabetes, DM-R0.1: DM treated with RSV (0.1 mg/kg/day) for 7 days, DM-R1: DM treated with RSV (1 mg/kg/day) for 7 days.