Nanduri Jayasri, Vaddi Damodara Reddy, Khan Shakil A, Wang Ning, Makarenko Vladislav, Semenza Gregg L, Prabhakar Nanduri R
Institute for Integrative Physiology and Center for Systems Biology of O2 Sensing, Biological Sciences Division, University of Chicago, Chicago, Illinois, United States of America.
Vascular Program, Institute for Cell Engineering; Department of Pediatrics, Medicine, Oncology, Radiation Oncology and Biological Chemistry; and Mckusick-Nathans Institute of Genetic Medicine, The Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America.
PLoS One. 2015 Mar 9;10(3):e0119762. doi: 10.1371/journal.pone.0119762. eCollection 2015.
Hypoxia-inducible factor 1 (HIF-1) mediates many of the systemic and cellular responses to intermittent hypoxia (IH), which is an experimental model that simulates O2 saturation profiles occurring with recurrent apnea. IH-evoked HIF-1α synthesis and stability are due to increased reactive oxygen species (ROS) generated by NADPH oxidases, especially Nox2. However, the mechanisms by which IH activates Nox2 are not known. We recently reported that IH activates xanthine oxidase (XO) and the resulting increase in ROS elevates intracellular calcium levels. Since Nox2 activation requires increased intracellular calcium levels, we hypothesized XO-mediated calcium signaling contributes to Nox activation by IH. We tested this possibility in rat pheochromocytoma PC12 cells subjected to IH consisting alternating cycles of hypoxia (1.5% O2 for 30 sec) and normoxia (21% O2 for 5 min). Kinetic analysis revealed that IH-induced XO preceded Nox activation. Inhibition of XO activity either by allopurinol or by siRNA prevented IH-induced Nox activation, translocation of the cytosolic subunits p47phox and p67phox to the plasma membrane and their interaction with gp91phox. ROS generated by XO also contribute to IH-evoked Nox activation via calcium-dependent protein kinase C stimulation. More importantly, silencing XO blocked IH-induced upregulation of HIF-1α demonstrating that HIF-1α activation by IH requires Nox2 activation by XO.
缺氧诱导因子1（HIF-1）介导了许多对间歇性缺氧（IH）的全身和细胞反应，IH是一种模拟反复呼吸暂停时发生的氧饱和度曲线的实验模型。IH诱发的HIF-1α合成和稳定性归因于烟酰胺腺嘌呤二核苷酸磷酸（NADPH）氧化酶，尤其是Nox2产生的活性氧（ROS）增加。然而，IH激活Nox2的机制尚不清楚。我们最近报道，IH激活黄嘌呤氧化酶（XO），由此产生的ROS增加会提高细胞内钙水平。由于Nox2的激活需要细胞内钙水平升高，我们推测XO介导的钙信号传导有助于IH激活Nox。我们在经历由缺氧（1.5% O2，持续30秒）和常氧（21% O2，持续5分钟）交替循环组成的IH的大鼠嗜铬细胞瘤PC12细胞中测试了这种可能性。动力学分析表明，IH诱导的XO先于Nox激活。用别嘌呤醇或小干扰RNA（siRNA）抑制XO活性可防止IH诱导的Nox激活、胞质亚基p47phox和p67phox向质膜的转位及其与gp91phox的相互作用。XO产生的ROS还通过钙依赖性蛋白激酶C刺激促进IH诱发的Nox激活。更重要的是，沉默XO可阻断IH诱导的HIF-1α上调，表明IH激活HIF-1α需要XO激活Nox2。
