title: Iptakalim induces mitochondria-dependent apoptosis in hypoxic rat pulmonary arterial smooth muscle cells.
abstract: OBJECTIVE
  Pulmonary vascular medial hypertrophy in hypoxic pulmonary arterial hypertension (HPH) is caused in part by decreased apoptosis in pulmonary artery smooth muscle cells (PASMCs). Iptakalim (Ipt), an ATP sensitive potassium channel opener, ameliorates HPH in animal models. Here we investigated the effects of Ipt on proliferation and apoptosis of hypoxic rat PASMCs, and to determine the possible underlying mechanisms.
  
  
  METHODS
  Primary rat PASMCs were isolated and cultured. PASMCs were cultured for 24h in normoxia or hypoxia (5% O2) conditions with and without Ipt. Cell proliferation and cycle were determined by MTT assay and flow cytometry, respectively. Mitochondrial membrane potential (Δym) was detected by fluorescence microscope Western blot assays were used to examine the expression of cyclin D, CDK4, endothelin-1 (ET-1), hypoxia-inducible factor-1 (HIF-1), platelet-derived growth factor-BB (PDGF-BB), Bax, Bcl-2, cytochrome c (Cyt c), caspase-9, and caspase-3 in PASMCs.
  
  
  RESULTS
  We found that hypoxia significantly stimulated proliferation and rendered resistance to apoptosis in PASMCs. Ipt suppressed proliferation and induced cell cycle arrest in hypoxia PASMCs. Ipt decreased the expression of cyclin D, CDK4, HIF-1, ET-1, and PDGF-BB in hypoxia PASMCs. It reversed the depolarization of Δψm in hypoxia PASMCs too. Ipt significantly upregulated Bax expression and downregulated Bcl-2 expression, and promoted the release of Cyt c from mitochondria to cytoplasm in hypoxia PASMCs. Furthermore, Ipt significantly activated the caspase cascades evidenced by increased expression of caspase-9 and caspase-3 in hypoxia PASMCs.
  
  
  CONCLUSIONS
  Ipt could inhibit cell proliferation and induce apoptosis associated with cell cycle arrest, decreased ET-1, HIF-1, cyclin D, CDK4, PDGF-BB and Δψm, increased Bax/Bcl-2 ratio, enhanced Cyt c release, and activation of caspases in PASMCs under hypoxia status. Our data indicated that Ipt could be a therapeutic candidate for treatment of HPH.
authors.affiliation: Department of Respiratory Medicine, The first Affiliated Hospital of Nanjing Medical University Nanjing 210029, China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Xu
authors.affiliation: School of Pharmacy, Nanjing University of Chinese Medicine, Nanjing 210023, China.
authors.forename: Xiang
authors.initials: X
authors.lastname: Wu
authors.affiliation: School of Basic Medicine, Nanjing University of Chinese Medicine, Nanjing 210023, China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Li
authors.affiliation: Department of Respiratory Medicine, The first Affiliated Hospital of Nanjing Medical University Nanjing 210029, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Kong
authors.affiliation: Department of Respiratory Medicine, The first Affiliated Hospital of Nanjing Medical University Nanjing 210029, China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Jin
authors.affiliation: Department of Respiratory Medicine, The first Affiliated Hospital of Nanjing Medical University Nanjing 210029, China.
authors.forename: Weiping
authors.initials: W
authors.lastname: Xie
authors.affiliation: Department of Respiratory Medicine, The first Affiliated Hospital of Nanjing Medical University Nanjing 210029, China. Electronic address: 1319420655@qq.com.
authors.forename: Hong
authors.initials: H
authors.lastname: Wang
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D015687:Cell Hypoxia / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D008297:Male; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000187:drug effects* / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000187:drug effects* / Q000378:metabolism; D011437:Propylamines / Q000494:pharmacology*; D011651:Pulmonary Artery / Q000187:drug effects* / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: C502260:N-(1-methylethyl)-1,1,2-trimethylpropylamine; D011437:Propylamines
keywords: Apoptosis; Caspase; Cytochrome c; Hypoxic pulmonary hypertension; Iptakalim; Pulmonary artery smooth muscle cell