Association between endoplasmic reticulum stress pathway mediated by inositolrequiring kinase 1 and AECⅡ apoptosis in preterm rats induced by hyperoxia

Abstract:

Objective To study the association between endoplasmic reticulum stress (ERS) pathway mediated by inositol-requiring kinase 1 (IRE1) and the apoptosis of type Ⅱ alveolar epithelial cells (AECⅡs) exposed to hyperoxia. Methods The primarily cultured AECⅡs from preterm rats were devided into an air group and a hyperoxia group. The model of hyperoxia-induced cell injury was established. The cells were harvested at 24, 48, and 72 hours after hyperoxia exposure. An inverted phase-contrast microscope was used to observe morphological changes of the cells. Annexin V/PI double staining flow cytometry was performed to measure cell apoptosis. RT-PCR and Western blot were used to measure the mRNA and protein expression of glucose-regulated protein 78 (GRP78), IRE1, X-box binding protein-1 (XBP-1), and C/EBP homologous protein (CHOP). An immunofluorescence assay was performed to measure the expression of CHOP. Results Over the time of hyperoxia exposure, the hyperoxia group showed irregular spreading and vacuolization of AECⅡs. Compared with the air group, the hyperoxia group showed a significantly increased apoptosis rate of AECⅡs and significantly increased mRNA and protein expression of GRP78, IRE1, XBP1, and CHOP compared at all time points (P < 0.05). The hyperoxia group had significantly greater fluorescence intensity of CHOP than the air group at all time points. In the hyperoxia group, the protein expression of CHOP was positively correlated with the apoptosis rate of AECⅡs and the protein expression of IRE1 and XBP1 (r = 0.97, 0.85, and 0.88 respectively; P < 0.05). Conclusions Hyperoxia induces apoptosis of AECⅡs possibly through activating the IRE1-XBP1-CHOP pathway.