Expression of a Functionally Active gp91phox-Containing Neutrophil-Type NAD(P)H Oxidase in Smooth Muscle Cells From Human Resistance Arteries: Regulation by Angiotensin II
Expression of a Functionally Active gp91phox-Containing Neutrophil-Type NAD(P)H Oxidase in Smooth Muscle Cells From Human Resistance Arteries: Regulation by Angiotensin II
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DOI：
10.1161/01.res.0000020404.01971.2f
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发表时间：
2002-06
期刊：
影响因子：
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通讯作者：
R. Touyz;Xin Chen;F. Tabet;Guoying Yao;G. He;M. Quinn;P. Pagano;E. Schiffrin
中科院分区：
文献类型：
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作者：
R. Touyz;Xin Chen;F. Tabet;Guoying Yao;G. He;M. Quinn;P. Pagano;E. Schiffrin
血管平滑肌细胞（VSMC）超氧化物的主要来源是NAD（P）H氧化酶。然而，这种酶的分子特征和调控尚不清楚。我们研究了来自人阻力动脉（HVSMC）的VSMC是否具有功能活性、血管紧张素II（Ang II）调节的NAD（P）H氧化酶，该氧化酶含有中性粒细胞氧化酶亚基，包括p22 phox、gp 91 phox、p40 phox、p47 phox和p67 phox。gp 91 phox同源物nox 1和nox 4的mRNA表达也在HVSMCs、人主动脉平滑肌细胞和大鼠VSMCs中进行了评估。HVSMC获自健康受试者臀肌活检的阻力动脉。HVSMCs中检测到gp 91 phox和nox 4，而未检测到nox 1。人主动脉平滑肌细胞和大鼠血管平滑肌细胞表达nox 1和nox 4，但不表达gp 91 phox。逆转录-聚合酶链反应和免疫印迹法检测到HVSMC中存在所有NAD（P）H氧化酶亚基。Ang Ⅱ增加NAD（P）H氧化酶亚单位的丰度。放线菌酮可抑制上述作用。急性血管紧张素II刺激（10至15分钟）增加p47 phox丝氨酸磷酸化，并诱导p47 phox和p67 phox易位。这与NAD（P）H氧化酶激活有关。在转染gp 91 phox反义寡核苷酸的细胞中，Ang II介导的作用被废除。NADPH诱导的超氧化物生成减少gp 91 ds-tat和夹竹桃苷，抑制剂p47 phox-gp 91 phox相互作用。我们的研究结果表明，HVSMCs具有功能活性的gp 91 phox含嗜中性粒细胞样NAD（P）H氧化酶。血管紧张素II通过诱导p47 phox的磷酸化、胞质亚基的易位和从头蛋白质合成来调节该酶。这些新的发现提供了深入了解NAD（P）H氧化酶的分子调控血管紧张素II在HVSMCs。此外，我们确定了大鼠和人的小动脉和大动脉的VSMCs中gp 91 phox同源物表达的差异。
A major source of vascular smooth muscle cell (VSMC) superoxide is NAD(P)H oxidase. However, the molecular characteristics and regulation of this enzyme are unclear. We investigated whether VSMCs from human resistance arteries (HVSMCs) possess a functionally active, angiotensin II (Ang II)–regulated NAD(P)H oxidase that contains neutrophil oxidase subunits, including p22phox, gp91phox, p40phox, p47phox, and p67phox. mRNA expression of gp91phox homologues, nox1 and nox4, was also assessed in HVSMCs, human aortic smooth muscle cells, and rat VSMCs. HVSMCs were obtained from resistance arteries from gluteal biopsies of healthy subjects. gp91phox and nox4, but not nox1, were detected in HVSMCs. Nox1 and nox4, but not gp91phox, were expressed in human aortic smooth muscle cells and rat VSMCs. All NAD(P)H oxidase subunits were present in HVSMCs as detected by reverse transcriptase–polymerase chain reaction and immunoblotting. Ang II increased NAD(P)H oxidase subunit abundance. These effects were inhibited by cycloheximide. Acute Ang II stimulation (10 to 15 minutes) increased p47phox serine phosphorylation and induced p47phox and p67phox translocation. This was associated with NAD(P)H oxidase activation. In cells transfected with gp91phox antisense oligonucleotides, Ang II–mediated actions were abrogated. NADPH-induced superoxide generation was reduced by gp91ds-tat and apocynin, inhibitors of p47phox-gp91phox interactions. Our results suggest that HVSMCs possess a functionally active gp91phox-containing neutrophil-like NAD(P)H oxidase. Ang II regulates the enzyme by inducing phosphorylation of p47phox, translocation of cytosolic subunits, and de novo protein synthesis. These novel findings provide insight into the molecular regulation of NAD(P)H oxidase by Ang II in HVSMCs. Furthermore, we identify differences in gp91phox homologue expression in VSMCs from rats and human small and large arteries.