Mechanisms of resveratrol in alleviating diabetic nephropathy: focus on tumor necrosis factor receptor-related factor expression and toll-like reeptor 4/nuclear factor-kappaB signaling pathway.

Abstract

The role of inflammation and oxidative stress (OS) is significant in the progression and development of diabetic nephropathy (DN). Resveratrol (Res) has various pharmacological effects including anti-inflammatory and anti-OS. Tumor necrosis factor receptor-related factor (TRAF3) has been shown to have anti-inflammatory and anti-OS effects in a variety of diseases. This study investigated the potential mechanisms underlying the renoprotective effects of Res, with a special focus on the regulation of TRAF3 in the Toll-like receptor 4 (TLR4)-mediated nuclear factor kappaB (NF-κB) inflammatory signaling pathway. A high-fat diet combined with intraperitoneal injection of streptozotocin (35 mg/kg) was used to induce type 2 diabetes mellitus (T2DM) in a rat model, and a high glucose (HG, 30 mmol/L)-treated glomerular thylakoid cell model was established in HBZY-1 rats. Res (40 mg/kg/day) was studied in vivo by gavage for 8 weeks and in vitro by treatment with Res (25, 50, and 100 μmol/L) for 48 hours. The degree of renal injury was evaluated by blood urea nitrogen (BUN), blood creatinine (Cr) and urine protein. Renal structure and glycogen changes were observed by hematoxylin and eosin and periodic acid Schiff staining (PAS). Inflammatory factors, including interleukin (IL)-1βt IL-6, monocyte chemotactic protein (MCP)-1, tumor necrosis factor-alpha (TNF-α), and oxidative stress markers, including superoxide dismutase (SOD), catalase (CAT), and malondialdehyde (MDA), were measured by enzyme-linked immunosorbent assay. Immunoblotting was performed to detect TRAF3, TLR4, MyD88, phosphorylated (p)-IκBβ, p-p65, and p65. Protein blotting was applied for final mechanistic validation using lentiviral transfection of diabetes mellitus (DM) rats and high glucose-induced cells against TRAF3. As a result Res effectively ameliorated renal loss and renal histopathologic changes in DM rats, as evidenced by decreased BUN (P<0.01), Cr (P<0.01), urine protein (P<0.01), and renal structural lesions and reduced basement membrane glycogen (P<0.015). In terms of molecular mechanisms, Res was able to significantly promote the expression of TRAF3, which in turn inhibited the activation of the TLR4/NF-κB signaling pathway and reduced the release of pro-inflammatory cytokines such as IL-1β, IL-6, MCP-1, and TNF-α (all P<0.01), thereby attenuating inflammatory responses in the kidney. In addition, Res significantly reduced OS in renal tissues and in the HG cell model, such as increasing SOD and CAT activities and decreasing MDA levels (all P<0.05). Silencing of TRAF3, on the other hand, partially reduced the anti-inflammatory and anti-OS effects of Res and activated the TLR4-mediated NF-κB signaling pathway. To sum up Res can alleviate DN by attenuating inflammation and OS, and the underlying mechanisms are related to TRAF3 in the TLR4/NF-κB-mediated anti-inflammatory pathway.