Suppression of Apoptosis in Human Umbilical Vein Endothelial Cells (HUVECs) by Klotho Protein is Associated with Reduced Endoplasmic Reticulum Oxidative Stress and Activation of the PI3K/AKT Pathway

• Published in: Medical science monitor 2018-11, Vol.24, p.8489-8499
• Main Authors: Cui, Wei, Leng, Bin, Liu, Wei, Wang, GaoPin
• Format: Article
• Language: English
• Subjects: Apoptosis - drug effects; Cell Survival - drug effects; Endoplasmic Reticulum - metabolism; Endoplasmic Reticulum Chaperone BiP; Endoplasmic Reticulum Stress - drug effects; Glucuronidase - physiology; Human Umbilical Vein Endothelial Cells - cytology; Human Umbilical Vein Endothelial Cells - drug effects; Human Umbilical Vein Endothelial Cells - metabolism; Humans; Hydrogen Peroxide - pharmacology; Interleukin-6 - metabolism; Klotho Proteins; Lab/In Vitro Research; Nitric Oxide - metabolism; Oxidative Stress - drug effects; Phosphatidylinositol 3-Kinases - metabolism; Phosphorylation - drug effects; Proto-Oncogene Proteins c-akt - metabolism; Reactive Oxygen Species - metabolism; Signal Transduction - drug effects; Tumor Necrosis Factor-alpha - metabolism
• ISSN: 1643-3750; 1234-1010; 1643-3750
• DOI: 10.12659/MSM.911202

BACKGROUND Klotho protein has been shown to act as a hormone on the cardiovascular system, and to have specific protective effects on vascular endothelial cells. The aim of this study was to investigate the mechanisms of the anti-oxidative and anti-apoptotic effects of klotho protein on hydrogen peroxide (H₂O₂)-induced apoptosis and endoplasmic reticulum oxidative stress in human umbilical vein endothelial cells (HUVECs). MATERIAL AND METHODS HUVECs were cultured in vitro and treated with H₂O₂. The MTT assay evaluated cell viability of H₂O₂-treated HUVECs, and flow cytometry measured cell apoptosis. An enzyme-linked immunosorbent assay (ELISA) measured the levels of nitric oxide (NO), tumor necrosis factor-alpha (TNF-α), and interleukin (IL)-6. Western blot was used to detect the expression of the proteins, 78 kD glucose-regulated protein (GRP78), CCAAT-enhancer-binding protein homologous protein (CHOP), caspase-3, caspase-9, caspase-12, and AKT. The effects of LY294002, a pharmacological inhibitor of PI3K, were evaluated. RESULTS Klotho protein increased the viability of H2O2-treated HUVECs and reduced the expression of NO, TNF-α, and IL-6. Klotho protein reduced the rate of apoptosis of H₂O₂-treated HUVECs and downregulated the expression of proteins associated with endoplasmic reticulum oxidative stress, GRP78 and CHOP, and the expression of the apoptotic proteins, caspase-3, caspase-9, and caspase-12, and activated the phosphorylation of AKT. The addition of LY294002 inhibited klotho protein downregulation of GRP78, CHOP, caspase-3, caspase-9, and caspase-12 expression. CONCLUSIONS In HUVECs, klotho protein suppressed apoptosis mediated by endoplasmic reticulum oxidative stress by activation of the PI3K/AKT pathway.