Effect And Mechanism Of HO-1 On Endoplasmic Reticulum Stress-mediated Apoptosis In Acute Lung Injury Induced By Endotoxin

Objective:Previous studies have found that HO-1 plays an important role in acute lung injury induced by endotoxin,but the mechanism is far from clear.Endoplasmic reticulum(ER)stress may be a important factor in the pathogenesis of lung injury,and the regulatory effect and mechanism of HO-1 on ERS are not clear in lung injury.In this study,we mainly discussed the effect and mechanism of HO-1 on ER stress-mediated apoptosis in acute lung injury induced by endotoxin.Methods:Using a mice model of acute lung injury induced by LPS and the LPS attacking cell model,we investigated the role and mechanism of HO-1 in ER stress-mediated apoptosis of acute lung injury induced by endotoxin,which are divided into three parts.In the first part,to verify the role of ER stress-mediated apoptosis in acute lung injury induced by endotoxin,we observed the changes of protein expression of GRP78,ATF6,IREl?,PERK,CHOP and caspase-12 in lung tissue and the degree of lung injury.The second part mainly used the acute lung injury model of HO-1 knockout mice to explore the effect and mechanism of HO-1 on ER stress-mediated apoptosis.The protein expression levels of GRP78,ATF6,IREl?,PERK,CHOP,and caspase-12 in lung tissue were observed,and the expression of HO-1 gene and protein,the content of CO in lung tissue,and the degree of lung injury were detected.In the third part,we used the alveolar type II epithelial cell model,further clarify the possible mechanism of HO-1 regulating ER stress-mediated apoptosis.LPS was used to establish the cell endotoxin model,the main indicators were the same as animal experiments.In addition,to explore the possible pathway of HO-1 regulating ER stress-mediated apoptosis,we pretreated alveolar type II epithelial cells using the inhibitors of IREl?and PERK,and observed the expression of ER stress related proteins and apoptotic proteins.Results:In the first experiment,we used tail intravenous injection of LPS to establish acute lung injury model in C57BL/6 mice,and observed the changes of lung injury,inflammatory factor index and oxidative stress index in mice at different doses and induction time points.Finally,we selected C57BL/6 mice tail intravenous injection of LPS 15 mg/kg to establish acute lung injury model for 12 hours.In addition,we used the ER stress inhibitor TUDCA and the agonist tunicamycin to successfully verify the role of ER stress-mediated apoptosis in the pathogenesis of acute lung injury induced by endotoxin.In experiment 2,we found that compared with wild C57BL/6 mice,HO-1^-/-mice treated with LPS 15 mg/kg for 12 hours can aggravate the degree of lung injury,which is manifested as lung injury score,W/D ratio,total protein content in BALF,oxygenation index.In addition,the expression levels of GRP78,p-IREl?,and p-PERK were significantly increased.Furthermore,AI and the expression levels of CHOP and caspase-12 m RNA and protein were significantly increased.These results may suggest that HO-1,which has endogenous protective effect,may reduce LPS-induced lung injury by inhibiting ER stress-mediated apoptosis.In experiment 3,we also found that HO-1 can protect the alveolar type II epithelial cells from endotoxin attack,which may reduce the injury of cells induced by endotoxin by inhibiting ER stress-mediated apoptosis.Further mechanism studies showed that the PERK inhibitor GSK2606414 pretreatment reduced the protein expression levels of p-PERK,p-e IF2?and CHOP,and the pretreatment with IRE1?inhibitor GSK2850163 decreased p-IREl?,TRAF2 and caspase-12 protein expression levels under LPS conditions in alveolar type II epithelial cells.These results suggest that HO-1 may inhibit ER stress-mediated cell damage through PERK/e IF2?/CHOP and IREl?/TRAF2/caspase-12 pathways.Conclusion:ER stress-mediated apoptosis is involved in the pathogenesis of acute lung injury induced by endotoxin.HO-1 may regulate ER stress-mediated apoptosis through PERK/e IF2?/CHOP and IREl?/TRAF2/caspase-12,so as to reduce lung injury induced by endotoxin.Other mechanisms of HO-1 regulating ER stress-mediated apoptosis needs further study.