Overexpression of survivin affects cell proliferation and apoptosis by regulating the expression of EPAS1 and CHOP-10 in vascular endothelial cells with hypoxic treatment

Abstract:

Objective：To investigate the effect of survivin（SVV） gene overexpression on the proliferation and migration of rat vascular endothelial cells with hypoxic treatment and the protein expression of EPAS1 and CHOP-10，as well as the possible mechanisms. Methods：Rat arterial endothelial cells were given hypoxic treatment with 500 μmol/L CoCl2 and were then divided into SVV inter－vention group，negative control group，and blank control group. The cells in the SVV intervention group were transfected with aden－ovirus SVV-EGFP，those in the negative control group were transfected with empty virus，and those in the blank control group were not given any treatment. Transwell assay was used to evaluate migration ability；ELISA was used to measure the changes in the levels of matrix metallopeptidase-2（MMP-2） and matrix metallopeptidase-9（MMP-9）；the WST-1 test was used to assess cell proliferation；flow cytometry was used to determine the cell cycle；Western blot was used to measure the changes in the protein expression of survivin，hypoxia-inducible factor EPAS1，endoplasmic reticulum stress protein CHOP-10，and apoptosis protein caspase-8. U0126 and LY294002 were used to block the EPAS1 upstream signaling pathways P44/42 MAPK and P13K/AKT，and then Western blot was used to measure the expression of EPAS1 to investigate the mechanism of the effect of survivin on EPAS1. Results：SVV overexpression promoted the migration ability of vascular endothelial cells and increased the secretion of MMP-2 and MMP-9. The proliferation ability of endothelial cells was enhanced after survivin trans－fection，with an increase in the number of cells in S phase，and there was an increase in the expression of survivin after transfection（F=326.137，P=0.000）. The relative protein expression of EPAS1 was 1.29±0.11 in the overexpression group，0.97±0.06 in the neg－ative control group，and 0.99±0.06 in the blank control group，suggesting that survivin significantly upregulated the expression of EPAS1（F=17.765，P=0.000）. SVV overexpression significantly inhibited the expression of CHOP-10（F=78.908，P=0.000）. There was a significant reduction in the relative expression of caspase-8 after survivin overexpression（F=53.823，P=0.000）. There were no significant differences in the expression of these three proteins between the negative control group and the blank control group（P=0.841，0.891，and 0.955）. The upregulated expression of EPAS1 due to survivin was inhibited by blocking the P13K/AKT pathway（0.78±0.04 vs. 1.32±0.28，P=0.002），and after U0126 was used to block the P42/44 MAPK pathway，there was a slight reduction in the expression of EPAS1（1.36±0.12 vs. 1.32±0.28，P=0.451）. Conclusion：Under the hypoxic condition，survivin overexpression can promote cell proliferation and migration abilities and increase the secretion of MMP-2 and MMP-9. SVVexerts an anti-apoptosis effect and promotes angiogenesis by upregulating the expression of EPAS1 in endothelial cells via the P13K/AKT pathway，inhibiting the expression of CHOP-10 and caspase-8，and protecting the cells against the hypoxic environment.