Insulin‑receptor substrate 1 protects against injury inendothelial cell models of ox‑LDL‑induced atherosclerosis by inhibiting ER stress/oxidative stress‑mediated apoptosis and activating the Akt/FoxO1 signaling pathway.

Abstract

Oxidized low-density lipoprotein (ox-LDL)-induced endothelial cell (EC) injury is a risk factor for atherosclerosis. Therefore, the present study aimed to investigate the effects of insulin-receptor substrate 1 (IRS-1) on injury to ox-LDL-exposed ECs. For this purpose, thoracic aorta tissues were isolated from rats and cultured to obtain ECs, which were then identified using immunohistochemical staining. IRS-1 overexpression plasmid (pcDNA3.1-IRS-1) and IRS-1-small interfering RNA were synthesized and transfected into ECs pre-exposed to ox-LDL. MTT and TUNEL assays were performed to evaluate the cell proliferative activity and apoptosis. Intracellular reactive oxygen species (ROS) production was determined by a flow cytometry assay. Reverse transcription-quantitative PCR was conducted to measure the peroxisome proliferator-activated receptor gamma co-activator 1 alpha (Ppargcla), phosphoenol-pyruvate carboxykinase 1 (Pck1) and glucose-6-phosphatase catalytic subunit (G6pc) gene transcription levels. Western blot analysis was then performed to determine the IRS-1, forkhead box O1 (FoxO1), phosphorylated (p-)FoxO1, 78-kDa glucose-regulated protein (GRP78), p-eukaryotic translation initiation factor 2A (eIF2α), CHOP, Akt and p-Akt expression levels. Immunofluorescence staining was used to evaluate p-FoxO1 nuclear localization. The results indicated that IRS-1 significantly enhanced the proliferative activity, whereas it inhibited the apoptosis of ECs in a model of ox-LDL-induced atherosclerosis compared with ECs without IRS-1 treatment (P<0.05). IRS-1 significantly decreased the p-FoxO1/FoxO1 ratio compared with ECs without ox-LDL treatment (P<0.05). IRS-1 significantly downregulated the expression of ER stress biomarkers, including GRP78, CHOP and the p-eIF2α/eIF2α ratio in ox-LDL-exposed ECs compared with ECs without ISR-1 treatment (P<0.05). IRS-1 significantly reduced the intracellular ROS levels in the EC models of ox-LDL-induced atherosclerosis compared with ECs without IRS-1 treatment (P<0.05). Moreover, IRS-1 promoted the phosphorylation of Akt in the EC models of ox-LDL-induced atherosclerosis. IRS-1 also significantly suppressed the transcription of atherosclerosis-associated genes in ox-LDL-exposed ECs compared with ECs without IRS-1 treatment (P<0.05). Furthermore, IRS-1 significantly increased the cytoplasmic localization of p-FoxO1 in EC models of ox-LDL-induced atherosclerosis. On the whole, the findings of the present study demonstrate that IRS-1 exerts protective effects in an EC model of ox-LDL-induced atherosclerosis by inhibiting ER stress/oxidative stress-mediated apoptosis and activating the Akt/FoxO1 signaling pathway.