Hosseinipour Mahsa, Rashidi Mojtaba, Seif Faezeh, Babaahmadi-Rezaei Hossein
Hyperlipidemia Research Center, Department of Clinical Biochemistry, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran.
Department of Basic sciences , Shoushtar Faculty of Medical sciences , Shoushtar , Iran.
Int J Mol Cell Med. 2022;11(4):297-305. doi: 10.22088/IJMCM.BUMS.11.4.297.
Caveolin-1(Cav-1) is one of the most important components of caveolae in the cell membrane, which plays an important role in cell signaling transduction, such as EGFR and TGF-β receptor transactivation. The purpose of this study was to evaluate the effect of c-Abl and NAD(P)H oxidases (NOX) on phosphorylation of Cav-1 and consequently their effect on phosphorylation of Smad2C induced by Endothelin-1 in human vascular smooth muscle cells (VSMCs). In this study, all experiments were performed using human VSMCs. The phosphorylation level of the Caveolin-1 and Smad2C proteins were assessed by western blotting using Phospho-Caveolin-1 (Tyr14) antibody and phospho-Smad2 (Ser465/467) antibody. The data were reported as mean ± SEM. The VSMCs treated with endothelin-1(ET-1) (100 nanomolar (nmol)) demonstrated a time-dependent increase in the pCav-1 level (p<0.05). The inhibitors of NOX (diphenyleneiodonium) (p<0.05), cholesterol depleting agent (beta-cyclodextrin) (p<0.05) and c-Abl inhibitor (PP1) (p<0.01) were able to reduce the level of the phospho-Cav-1 and phospho-Smad2C induced by Et-1 (p<0.05). Our results proposed that caveolae structure, NOX, c-Abl played an important role in the phosphorylation of Cav-1 induced by ET-1 in the human VSMCs. Furthermore, our findings showed that phosphoCav-1 involved in TGFR transactivation. Thus, Et-1 via a transactivation-dependent mechanism can cause phosphorylation of Smad2C.
小窝蛋白-1（Cav-1）是细胞膜中小窝的最重要组成部分之一，在细胞信号转导中起重要作用，如表皮生长因子受体（EGFR）和转化生长因子-β（TGF-β）受体的反式激活。本研究旨在评估c-Abl和烟酰胺腺嘌呤二核苷酸磷酸（NAD(P)H）氧化酶（NOX）对Cav-1磷酸化的影响，以及它们对内皮素-1在人血管平滑肌细胞（VSMC）中诱导的Smad2C磷酸化的影响。在本研究中，所有实验均使用人VSMC进行。使用磷酸化小窝蛋白-1（酪氨酸14）抗体和磷酸化Smad2（丝氨酸465/467）抗体，通过蛋白质免疫印迹法评估小窝蛋白-1和Smad2C蛋白的磷酸化水平。数据以平均值±标准误表示。用内皮素-1（ET-1）（100纳摩尔（nmol））处理的VSMC显示pCav-1水平呈时间依赖性增加（p<0.05）。NOX抑制剂（二亚苯基碘鎓）（p<0.05）、胆固醇耗竭剂（β-环糊精）（p<0.05）和c-Abl抑制剂（PP1）（p<0.01）能够降低Et-1诱导的磷酸化小窝蛋白-1和磷酸化Smad2C的水平（p<0.05）。我们的结果表明，小窝结构、NOX、c-Abl在ET-1诱导的人VSMC中Cav-1磷酸化中起重要作用。此外，我们的研究结果表明，磷酸化小窝蛋白-1参与TGF受体反式激活。因此，Et-1通过反式激活依赖性机制可导致Smad2C磷酸化。
