Resveratrol ameliorated endothelial injury of thoracic aorta in diabetic mice and Gly-LDL-induced HUVECs through inhibiting TLR4/HIF-1α.

Abstract

To explore the effects of resveratrol on the levels of inflammatory cytokines and Toll‐like receptor‐4/ hypoxia‐inducible transcription factors‐1α (TLR4/HIF‐1α) signalling pathway in diabetes mellitus. C57BL/6 mice received intraperitoneal injection of streptozocin for constructing diabetic mice models. Human umbilical vein endothelial cells (HUVECs) were treated with 50 µg/mL Gly‐LDL for inducing injury models. 10, 100 and 1000 mmol/L resveratrol were obtained and added into each group. Haematoxylin‐eosin (H&E) staining was used for histological evaluation. CCK8 assay was performed for determination of cell viability, and Transwell assay was implemented for detecting cell migration ability. Cell apoptosis was analysed using flow cytometry. The content of inflammatory factors including interleukin‐6 (IL‐6), tumour necrosis factor‐α (TNF‐α), vascular adhesion molecule‐1 (VCAM‐1) and vascular endothelial growth factor (VEGF) were measured by ELISA. GST pull‐down assay was employed for determining interactions between TLR4 and HIF‐1α. The protein expression of TLR4 and HIF‐1α was detected using Western blotting and immunohistochemistry, while relative mRNA expression was measured by RT‐qRCR. Resveratrol could reduce bodyweight and ameliorate endothelial injury of thoracic aorta in diabetic mice. Both in vivo and in vitro results revealed that the level of IL‐6, TNF‐α, VCAM‐1 and VEGF was significantly down‐regulated after being treated with resveratrol. Resveratrol inhibited the increase of MDA and ROS and increased the level of SOD in diabetic mice. Western blotting, IHC and RT‐qPCR results showed that the levels of TLR4 and HIF‐1α were significantly down‐regulated in resveratrol group. Overexpression of TLR4 or HIF‐1α could reverse the effect of resveratrol. GST pull‐down elucidated that there might be a close interaction between TLR4 and HIF‐1α. Resveratrol ameliorated endothelial injury of thoracic aorta in diabetic mice and Gly‐LDL‐induced HUVECs through inhibiting TLR4/HIF‐1α signalling pathway.