Yao Yansheng, Wang Yanbing, Zhang Yibo, Liu Chang
Jinzhou Medical University, Jinzhou, Liaoning, China.
Department of Pathogenic Biology, Jinzhou Medical University, Jinzhou, Liaoning, China.
Lipids Health Dis. 2017 Apr 13;16(1):77. doi: 10.1186/s12944-017-0447-0.
Atherosclerosis is a common cardiovascular disease that causes myocardial infarction, heart failure, and stroke. Increased oxidized low density lipoprotein (ox-LDL) in the sub-endothelium is the characteristic origin of atherogenesis. Klotho, an anti-aging protein, has been reported to protect against atherosclerosis and ameliorate endothelial dysfunction in vivo. The aim of this study is to investigatethe anti-oxidative activity of Klothoin ox-LDL-treated human umbilical vein endothelial cells (HUVECs).
After pre-treatment with 200 pMKlotho for 1 h, HUVECs were stimulated with 50 μg/ml ox-LDL for 24 h. Reactive oxygen species (ROS) and superoxide dismutase (SOD) levels were analyzed in the cells. Nitric oxide (NO) concertation was measured in the medium supernatant. Related proteins or genes were detected with Western blot or real time PCR, respectively, in the cell lysates.
Initially, oxidative damage in HUVECs was established by adding 50 μg/mL ox-LDL, which resulted in decreased cellular viability, SOD/Cu/Zn-SOD and endothelial NO synthase (eNOS) expression and NO production, as well as increased malondialdehyde (MDA) levels, ROS production, inducible NO synthase (iNOS), phosphatidyl inositol-3 kinase (PI3K), protein kinase B (Akt), gp91 phox, and lectin-like ox-LDL receptor (LOX-1) expression in HUVECs. Pre-incubation with recombinant Klotho (200 pM) significantly prevented all of these alterations. These results suggest that Klotho can attenuate ox-LDL-induced oxidative stress in HUVECs through upregulating oxidative scavengers (SOD and NO) viaactivating the PI3K/Akt/eNOS pathway and depressing LOX-1expression.
These results suggest that Klotho has a potential therapeutic effect on attenuating endothelial dysfunction and ameliorating atherosclerosis.
动脉粥样硬化是一种常见的心血管疾病，可导致心肌梗死、心力衰竭和中风。内皮下氧化型低密度脂蛋白（ox-LDL）增加是动脉粥样硬化发生的典型起始因素。据报道，抗老化蛋白Klotho可在体内预防动脉粥样硬化并改善内皮功能障碍。本研究旨在探讨Klotho在ox-LDL处理的人脐静脉内皮细胞（HUVECs）中的抗氧化活性。
用200 pM Klotho预处理HUVECs 1小时后，用50 μg/ml ox-LDL刺激24小时。分析细胞中的活性氧（ROS）和超氧化物歧化酶（SOD）水平。测定培养基上清液中的一氧化氮（NO）浓度。分别用蛋白质印迹法或实时PCR检测细胞裂解物中的相关蛋白质或基因。
最初，通过添加50 μg/mL ox-LDL建立HUVECs中的氧化损伤，这导致细胞活力、SOD/Cu/Zn-SOD和内皮型一氧化氮合酶（eNOS）表达以及NO生成降低，同时丙二醛（MDA）水平、ROS生成、诱导型一氧化氮合酶（iNOS）、磷脂酰肌醇-3激酶（PI3K）、蛋白激酶B（Akt）、gp91 phox和凝集素样ox-LDL受体（LOX-1）在HUVECs中的表达增加。用重组Klotho（200 pM）预孵育可显著预防所有这些改变。这些结果表明，Klotho可通过激活PI3K/Akt/eNOS途径上调氧化清除剂（SOD和NO）并抑制LOX-1表达，从而减轻ox-LDL诱导的HUVECs氧化应激。
这些结果表明，Klotho在减轻内皮功能障碍和改善动脉粥样硬化方面具有潜在的治疗作用。
