AMPK and FoxO1 Regulate Catalase Expression in Hypoxic Pulmonary Arterial Smooth Muscle
AMPK and FoxO1 Regulate Catalase Expression in Hypoxic Pulmonary Arterial Smooth Muscle
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DOI：
10.1002/ppul.22919
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发表时间：
2014-09-01
影响因子：
3.1
通讯作者：
Dakshinamurti, Shyamala
中科院分区：
文献类型：
--
作者：
Awad, Hanan;Nolette, Nora;Dakshinamurti, Shyamala
背景资料：低氧和活性氧（reactive oxygen species，ROS）（包括H2 O2）在持续性肺动脉高压肺血管重构的发生和发展中起重要作用。过氧化氢酶（CAT）是植物体内清除H2 O2的主要酶，其活性受到组织和环境特异性的调控。目的：探讨缺氧和H2 O2对新生猪肺动脉平滑肌过氧化氢酶表达的调控机制及AMPK-FoxO通路的作用。设计/方法：将PASMC在缺氧（10%O-2）或常氧（21%O-2）条件下培养72小时，检测过氧化氢酶活性和脂质过氧化反应，用qPCR检测CAT、FoxO 1和FoxO 3a的表达，用免疫印迹法检测完整裂解物中CAT、FoxO s、p-AMPK、p-AKT、p-JNK、p-ERK 1/2的蛋白含量，以及核提取物中FoxO s的蛋白含量;和FoxO-1核定位通过免疫细胞化学，通过激光扫描细胞术定量。结果如下：缺氧通过增加CAT转录因子FoxO 1和FoxO 3a的mRNA表达，促进FoxO 1的核转位，上调CAT的转录、含量和活性。而缺氧时PASMC脂质过氧化反应增强。在候选FoxO调节激酶中，缺氧激活AMPK，并降低p-Akt和ERK 1/2。AMPK激活增加FoxO 1（总和核）和CAT，而AMPK抑制抑制FoxO 1和CAT，但不FoxO 3a。外源性H_2O_2使缺氧PASMC p-AMPK降低，p-AKT升高。这降低了活性FoxO 1，并减少CAT的mRNA和蛋白质含量。CAT，AKT抑制（LY 294002），或添加PEG-过氧化氢酶部分改善H2 O2介导的核FoxO 1的损失的hypocalinity诱导。结论：低氧诱导过氧化氢酶表达，但这种适应不足以保护PASMC免受低氧诱导的脂质过氧化作用。这是通过缺氧激活AMPK而发生的，AMPK促进核FoxO 1并从而促进过氧化氢酶表达。外源性ROS可能下调细胞的抗氧化防御; H2 O2激活生存因子Akt，减少核FoxO 1，从而减少过氧化氢酶。(C)2013 Wiley Periodicals，Inc.
Background: Hypoxia and reactive oxygen species (ROS) including H2O2 play major roles in triggering and progression of pulmonary vascular remodeling in persistent pulmonary hypertension. Catalase (CAT), the major endogenous enzyme scavenging H2O2, is regulated in a tissue-and context-specific manner. Objective: To investigate mechanisms by which hypoxia and H2O2 regulate catalase expression, and the role of AMPK-FoxO pathway, in neonatal porcine pulmonary artery smooth muscle (PASMC). Design/Methods: PASMC were grown in hypoxia (10% O-2) or normoxia (21% O-2) for 72 hr. We measured catalase activity and lipid peroxidation; CAT, FoxO1, and FoxO3a expression by qPCR; protein contents of CAT, FoxOs, p-AMPK, p-AKT, p-JNK, p-ERK1/2 in whole lysates, and FoxOs in nuclear extracts, by immunoblot; and FoxO-1 nuclear localization by immunocytochemistry, quantified by laser scanning cytometry. Results: Hypoxia upregulated CAT transcription, content and activity, by increasing CAT transcription factors FoxO1 and FoxO3a mRNA, and promoting nuclear translocation of FoxO1. However, lipid peroxidation increased in hypoxic PASMC. Among candidate FoxO regulatory kinases, hypoxia activated AMPK, and decreased p-Akt and ERK1/2. AMPK activation increased FoxO1 (total and nuclear) and CAT, while AMPK inhibition inhibited FoxO1 and CAT, but not FoxO3a. Exogenous H2O2 decreased p-AMPK and increased p-AKT in hypoxic PASMC. This decreased active FoxO1, and reduced mRNA and protein content of CAT. Hypoxic induction of CAT, AKT inhibition (LY294002), or addition of PEG-catalase partly ameliorated the H2O2-mediated loss of nuclear FoxO1. Conclusions: Hypoxia induces catalase expression, though this adaptation is insufficient to protect PASMC from hypoxia-induced lipid peroxidation. This occurs via hypoxic activation of AMPK, which promotes nuclear FoxO1 and thus catalase expression. Exogenous ROS may downregulate cellular antioxidant defenses; H2O2 activates survival factor Akt, decreasing nuclear FoxO1 and thus catalase. (C) 2013 Wiley Periodicals, Inc.