PERK Overexpression-Mediated Nrf2/HO-1 Pathway Alleviates Hypoxia/Reoxygenation-Induced Injury in Neonatal Murine Cardiomyocytes via Improving Endoplasmic Reticulum Stress.

Reperfusion processes following acute myocardial infarction (AMI) have been reported to induce additional cardiomyocyte death, known as ischemia-reperfusion (I/R) injury. Endoplasmic reticulum (ER) stress is reported to be involved in the development of I/R injury. There is evidence that PERK exerts beneficial roles in alleviating ER stress. Here, we investigated whether upregulation of PERK improved cardiomyocytes injury induced by I/R. Specific siRNAs or adenovirus vectors were incubated with isolated neonatal cardiomyocytes (NCMs) to regulate expression levels of target genes including PERK, Nrf2, and HO-1. Afterwards, hypoxia and subsequent reoxygenation (H/R) administration was performed as the in vitro model of I/R injury. MTT assay showed that H/R intervention decreased the viability of cells, yet PERK overexpression increased the cellular proliferative rate. Moreover, the upregulation of Nrf2 or HO-1 elevated the growth rate of cells, while gene silencing of Nrf2 or HO-1 reduced the viability of NCMs treated with PERK-rAAV9. In addition, we observed that the apoptotic index of cells with H/R stimulation was reduced when NCMs were pretreated with PERK-rAAV9, Nrf2-rAAV9, or HO-1-rAAV9. After cells were incubated with Nrf2-siRNA or HO-1-siRNA, the upregulation of PERK had no roles in affecting the apoptosis rate of NCMs damaged by H/R. Then, our findings indicated that there was a level decrease of GRP78, CRT, CHOP, and Caspase-12 in NCMs of the PERK-rAAV9 group compared to that of the H/R group. Both Nrf2 overexpression and HO-1 upregulation reduced the expression of ER stress-related proapoptotic factors, yet the expression suppression of Nrf2 and HO-1 increased levels of GRP78, CRT, CHOP, and Caspase-12 in NCMs treated with PERK-rAAV9. Taken together, our results suggested that the effects of PERK against H/R injury might be attributed to the upregulation of Nrf2/HO-1 cascade, followed by the inhibition of ER stress-related apoptotic pathway.