Department of Biochemistry and Biophysics, University of Kalyani, Kalyani, West Bengal 741235, India.
Cell Biol Int. 2019 Jun;43(6):678-694. doi: 10.1002/cbin.11145. Epub 2019 Apr 30.
We sought to determine the mechanism by which angiotensin II (ANGII) stimulates NADPH oxidase-mediated superoxide (O ) production in bovine pulmonary artery smooth muscle cells (BPASMCs). ANGII-induced increase in phospholipase D (PLD) and NADPH oxidase activities were inhibited upon pretreatment of the cells with chemical and genetic inhibitors of PLD2, but not PLD1. Immunoblot study revealed that ANGII treatment of the cells markedly increases protein kinase C-α (PKC-α), -δ, -ε, and -ζ levels in the cell membrane. Pretreatment of the cells with chemical and genetic inhibitors of PKC-ζ, but not PKC-α, -δ, and -ε, attenuated ANGII-induced increase in NADPH oxidase activity without a discernible change in PLD activity. Transfection of the cells with p47phox small interfering RNA inhibited ANGII-induced increase in NADPH oxidase activity without a significant change in PLD activity. Pretreatment of the cells with the chemical and genetic inhibitors of PLD2 and PKC-ζ inhibited ANGII-induced p47phox phosphorylation and subsequently translocation from cytosol to the cell membrane, and also inhibited its association with p22phox (a component of membrane-associated NADPH oxidase). Overall, PLD-PKCζ-p47phox signaling axis plays a crucial role in ANGII-induced increase in NADPH oxidase-mediated O production in the cells.
我们试图确定血管紧张素 II（ANGII）刺激牛肺动脉平滑肌细胞（BPASMC）中 NADPH 氧化酶介导的超氧化物（O ）产生的机制。在用 PLD2 的化学和遗传抑制剂预处理细胞后，ANGII 诱导的 PLD 和 NADPH 氧化酶活性的增加被抑制，但 PLD1 则不然。免疫印迹研究表明，ANGII 处理细胞可显著增加细胞膜中蛋白激酶 C-α（PKC-α）、-δ、-ε 和 -ζ 水平。用 PKC-ζ 的化学和遗传抑制剂预处理细胞，但不是 PKC-α、-δ 和 -ε，可减弱 ANGII 诱导的 NADPH 氧化酶活性的增加，而 PLD 活性没有明显变化。用 p47phox 小干扰 RNA 转染细胞可抑制 ANGII 诱导的 NADPH 氧化酶活性增加，而 PLD 活性无明显变化。用 PLD2 和 PKC-ζ 的化学和遗传抑制剂预处理细胞可抑制 ANGII 诱导的 p47phox 磷酸化及其随后从细胞质向细胞膜的易位，还可抑制其与 p22phox（膜结合 NADPH 氧化酶的一个组成部分）的结合。总的来说，PLD-PKCζ-p47phox 信号轴在 ANGII 诱导的细胞中 NADPH 氧化酶介导的 O 产生增加中起关键作用。
