Nicorandil inhibits hypoxia-induced apoptosis in human pulmonary artery endothelial cells through activation of mitoKATP and regulation of eNOS and the NF-κB pathway

Wang,Hui; Zuo,Xiangrong; Wang,Qiang; Yu,Yanzhe; Xie,Lijun; Wang,Hong; Wu,Hongcheng; Xie,Weiping

doi:10.3892/ijmm.2013.1379

Nicorandil inhibits hypoxia-induced apoptosis in human pulmonary artery endothelial cells through activation of mitoKATP and regulation of eNOS and the NF-κB pathway

Authors:
- Hui Wang
- Xiangrong Zuo
- Qiang Wang
- Yanzhe Yu
- Lijun Xie
- Hong Wang
- Hongcheng Wu
- Weiping Xie
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Affiliations: Department of Respiratory Medicine, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China, Department of Rheumatism, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China, Department of Respiratory Medicine, Lihuili Hospital, Medical Center of Ningbo, Ningbo, Zhejiang 315041, P.R. China
Published online on: May 13, 2013 https://doi.org/10.3892/ijmm.2013.1379
Pages: 187-194

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Abstract

Apoptosis of human pulmonary artery endothelial cells (HPAECs) is the initial step and triggering event for pulmonary hypertension (PH). However, little is known about the actions of nicorandil on HPAECs in vitro. In the present study, we investigated the anti-apoptotic effect of nicorandil on HPAECs exposed to hypoxia, and explored the underlying mechanism(s) of action. Cell viability was assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Annexin V and propidium iodide staining, and Hoechst 33342 staining assay were employed to detect apoptosis. In addition, the protein expression of Bax, Bcl-2, caspase-9 and -3, endothelial nitric oxide synthase (eNOS), nuclear factor-κB (NF-κB) and IκBα were determined by western blotting to investigate the possible mechanisms. We found that exposure to hypoxia for 24 h significantly decreased cell viability and increased cell apoptosis. Pretreatment with nicorandil (100 µM) effectively abolished the influence of hypoxia on HPAECs. However, these protective effects of nicorandil were significantly inhibited by an antagonist of mitochondrial adenosine triphosphate-sensitive potassium (mitoKATP) channels, 5-hydroxydecanoate (5-HD, 500 µM), and by an eNOS inhibitor, NG-nitro-L-arginine methyl ester (L-NAME, 300 µM). We further observed that nicorandil could upregulate the decreased protein expression of eNOS and IκBα, and downregulate the increased protein expression of NF-κB, induced by hypoxia. In addition, nicorandil inhibited the enhancement of caspase-3 and -9 expression, and the increase in the Bax/Bcl-2 expression ratio, induced by hypoxia. However, these effects were also abolished by 5-HD and L-NAME. Collectively, these findings suggest that nicorandil inhibits hypoxia-induced apoptosis of HPAECs through activation of mitoKATP channels and increased eNOS expression, which in turn inhibits the NF-κB pathway and the mitochondrial apoptotic pathway.

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