Testosterone Induces Vascular Smooth Muscle Cell Migration by NADPH Oxidase and c-Src-Dependent Pathways
Testosterone Induces Vascular Smooth Muscle Cell Migration by NADPH Oxidase and c-Src-Dependent Pathways
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DOI：
10.1161/hypertensionaha.111.180620
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发表时间：
2012-06-01
期刊：
影响因子：
8.3
通讯作者：
Tostes, Rita C.
中科院分区：
文献类型：
--
作者：
Chignalia, Andreia Z.;Schuldt, Elke Z.;Tostes, Rita C.
睾酮被认为与高血压相关的血管重塑有关。这背后的分子机制尚不清楚，但氧化应激可能是重要的。我们假设睾酮可刺激血管平滑肌细胞(VSMCs)产生活性氧(ROS)和迁移，从而增强自发性高血压大鼠(SHR)细胞的作用。确定了睾酮诱导ROS产生的机制(基因组和非基因组)以及c-Src的作用，c-Src是氧化还原敏感迁移的调节因子。用睾酮(10(-7)mol/L，0~120min)刺激雄性Wistar-京都大鼠和SHR的VSMCs。通过二氢乙锭荧光和光敏素增强的化学发光(30分钟[SHR]和60分钟[两个菌株])来评估，睾酮增加了ROS的产生。雄激素受体拮抗剂氟他胺和基因转录抑制剂放线菌素D可抑制ROS的产生(60min)。实时定量聚合酶链式反应和免疫印迹检测显示，睾酮可增加NOX1和NOX4的mRNA水平和p47Phox蛋白的表达。氟他胺、放线菌素D和放线菌亚胺(蛋白质合成抑制剂)可减弱睾酮对p47Phox的影响。在30分钟(SHR)和120分钟(Wistar-京都大鼠)观察到c-Src的磷酸化。在SHR中，3-(4-氯苯基)1-(1，1-dimethylethyl)-1H-pyrazolo[3，4-day]pyrimidin-4-amine(c-Src抑制剂)可抑制睾酮诱导的ROS生成，而抗氧化剂/NADPH氧化酶抑制剂apocynin可降低两株SHR的ROS生成。通过伤口愈合技术评估，睾酮刺激VSMCs迁移，对SHR的影响更大。氟他胺、apocynin和3-(4-氯苯基)1-(1，1-二甲乙基)-1H-吡唑并[3，4天]嘧啶-4-胺均可阻断睾酮诱导的VSMCs迁移。我们的研究表明，睾酮通过NADPH氧化酶衍生的ROS和c-Src依赖的途径，通过基因组和非基因组机制诱导VSMCs迁移，这两种机制在Wistar-京都大鼠和SHR的VSMCs中受到不同的调控。(高血压。2012年；59：1263-1271。)。在线数据补充
Testosterone has been implicated in vascular remodeling associated with hypertension. Molecular mechanisms underlying this are elusive, but oxidative stress may be important. We hypothesized that testosterone stimulates generation of reactive oxygen species (ROS) and migration of vascular smooth muscle cells (VSMCs), with enhanced effects in cells from spontaneously hypertensive rats (SHRs). The mechanisms (genomic and nongenomic) whereby testosterone induces ROS generation and the role of c-Src, a regulator of redox-sensitive migration, were determined. VSMCs from male Wistar-Kyoto rats and SHRs were stimulated with testosterone (10(-7) mol/L, 0-120 minutes). Testosterone increased ROS generation, assessed by dihydroethidium fluorescence and lucigenin-enhanced chemiluminescence (30 minutes [SHR] and 60 minutes [both strains]). Flutamide (androgen receptor antagonist) and actinomycin D (gene transcription inhibitor) diminished ROS production (60 minutes). Testosterone increased Nox1 and Nox4 mRNA levels and p47phox protein expression, determined by real-time PCR and immunoblotting, respectively. Flutamide, actinomycin D, and cycloheximide (protein synthesis inhibitor) diminished testosterone effects on p47phox. c-Src phosphorylation was observed at 30 minutes (SHR) and 120 minutes (Wistar-Kyoto rat). Testosterone-induced ROS generation was repressed by 3-(4-chlorophenyl) 1-(1,1-dimethylethyl)-1H-pyrazolo[3,4-day]pyrimidin-4-amine (c-Src inhibitor) in SHRs and reduced by apocynin (antioxidant/NADPH oxidase inhibitor) in both strains. Testosterone stimulated VSMCs migration, assessed by the wound healing technique, with greater effects in SHRs. Flutamide, apocynin, and 3-(4-chlorophenyl) 1-(1,1-dimethylethyl)-1H-pyrazolo[3,4-day] pyrimidin-4-amine blocked testosterone-induced VSMCs migration in both strains. Our study demonstrates that testosterone induces VSMCs migration via NADPH oxidase-derived ROS and c-Src-dependent pathways by genomic and nongenomic mechanisms, which are differentially regulated in VSMCs from Wistar-Kyoto rats and SHRs. (Hypertension. 2012; 59: 1263-1271.). Online Data Supplement