Ma Yuhong, Li Weizu, Yin Yanyan, Li Weiping
Department of Pharmacology, College of Basic Medicine, Anhui Medical University, Hefei, Anhui 230032, P.R. China.
Int J Mol Med. 2015 Jun;35(6):1667-74. doi: 10.3892/ijmm.2015.2188. Epub 2015 Apr 17.
Endothelial cell apoptosis plays an important role in the pathophysiological mechanisms of vascular complications in diabetes mellitus (DM). NADPH oxidase 4 (Nox4)-dependent reactive oxygen species (ROS) aggregation is the main cause of vascular endothelial cell apoptosis. The transforming growth factor-β1 (TGF-β1)/Smad2 signaling pathway is involved in the apoptosis of several types of cells. However, the association between vascular endothelial cell apoptosis and Nox4, and the involvement of the TGF-β1/Smad2 signaling pathway in vascular endothelial cell apoptosis remain unclear. In the present study, we aimed to investigate the role of Nox4-dependent ROS production and to determine the involvement of the TGF-β1/Smad2 signaling pathway in endothelial cell apoptosis induced by oxidative stress which causes vascular injury in DM. We demonstrated that hydrogen peroxide (H2O2) increased Nox4-dependent-ROS aggregation, as well as the expression of TGF-β1, Smad2, Bax and caspase-3, decreased Bcl-2 expression and increased the apoptosis of human umbilical vein endothelial cells (HUVECs). Treatment with diphenyliodonium (DPI), a specific inhibitor of Nox4 or astragaloside IV (AST IV), a monomer located in an extract of astragaloside, decreased Nox4 expression and the levels of ROS, decreased TGF-β1 and Smad2 expression, altered the expression of apoptosis-related genes and decreased the apoptosis of HUVECs. Treatment with LY2109761, a selective inhibitor of the TGF-β1/Smad2 pathway, produced results similar to those of DPI; however, LY2109761 had no effect on Nox4 expression and ROS levels. Taken together, the findings of the present study suggest that H2O2 contributes to HUVEC apoptosis by inducing Nox4-dependent ROS aggregation and activating the TGF-β1/Smad2 signaling pathway. Our data indicate that the protective effects of AST IV against vascular endothelial cell apoptosis in DM are mainly associated with the decrease in Nox4 expression through the TGF-β1/Smad2 signaling pathway. Furthermore, the inhibition of the activation of the TGF-β1/Smad2 signaling pathway may be another potential therapeutic strategy in the treatment of DM.
内皮细胞凋亡在糖尿病（DM）血管并发症的病理生理机制中起重要作用。NADPH氧化酶4（Nox4）依赖性活性氧（ROS）聚集是血管内皮细胞凋亡的主要原因。转化生长因子-β1（TGF-β1）/Smad2信号通路参与多种类型细胞的凋亡。然而，血管内皮细胞凋亡与Nox4之间的关联以及TGF-β1/Smad2信号通路在血管内皮细胞凋亡中的作用仍不清楚。在本研究中，我们旨在探讨Nox4依赖性ROS产生的作用，并确定TGF-β1/Smad2信号通路在由氧化应激诱导的内皮细胞凋亡中的作用，氧化应激会导致DM中的血管损伤。我们证明，过氧化氢（H2O2）增加了Nox4依赖性ROS聚集，以及TGF-β1、Smad2、Bax和caspase-3的表达，降低了Bcl-2表达并增加了人脐静脉内皮细胞（HUVECs）的凋亡。用Nox4的特异性抑制剂二苯基碘鎓（DPI）或黄芪甲苷提取物中的单体黄芪甲苷IV（AST IV）处理，降低了Nox4表达和ROS水平，降低了TGF-β1和Smad2表达，改变了凋亡相关基因的表达并降低了HUVECs的凋亡。用TGF-β1/Smad2通路的选择性抑制剂LY2109761处理产生了与DPI相似的结果；然而，LY2109761对Nox4表达和ROS水平没有影响。综上所述，本研究结果表明，H2O2通过诱导Nox4依赖性ROS聚集和激活TGF-β1/Smad2信号通路促进HUVEC凋亡。我们的数据表明，AST IV对DM中血管内皮细胞凋亡的保护作用主要与通过TGF-β1/Smad2信号通路降低Nox4表达有关。此外，抑制TGF-β1/Smad2信号通路的激活可能是治疗DM的另一种潜在治疗策略。
