HSF1 Protects Sepsis-induced Acute Lung Injury By Inhibiting Ferroptosis

Background and Objective:Sepsis is a life-threatening multi-organ dysfunction caused by dysregulation of host’s response to infection.Septic shock and multiple organ dysfunction were the main causes of death.During sepsis,lung is the most vulnerable to injury.And sepsis is the main risk factor for the most patients to develop acute lung injury.The pathogenesis of sepsis is complex,involving inflammatory response,oxidative stress,coagulation dysfunction,and immune cell death.Ferroptosis is a new type of programmed cell death caused by accumulation of iron dependent lipid peroxides.Ferroptosis is involved in the development of many diseases,such as cancer,nervous system diseases and ischemia-reperfusion injury.Recent studies suggest that ferroptosis is involved in the pathogenesis of sepsis,ferroptosis can aggravates inflammation and acute lung injury caused by sepsis.Therefore,clarifing the mechanism of ferroptosis in the pathogenesis of sepsis is important.Maybe it can provide a potential target for the treatment of sepsis.HSF1 is an endogenous protective factor against environmental changes in the process of biological evolution.A series of previous studies have shown that HSF1 can improve the survival rate of endotoxemia mice and protect LPS-induced acute lung injury by inhibiting macrophage infiltration and NLRP3 inflammasome activation.So it has not been reported whether HSF1 can protect sepsis acute lung injury by inhibiting ferroptosis.Therefore,this study aims to investigate the effect of HSF1 on ferroptosis in sepsis-acute lung injury at the animal and cell levels,and to explore the molecular mechanism of HSF1 regulation of ferroptosis.Methods:（1）.Cecal ligation and puncture（CLP）was used to replicate sepsis model,and LPS induced ferroptosis model of MLE12 cells.The changes of ferroptosis related genes GPX4 was detected by q RT-PCR and western blot,biochemical indexes(GSH,ROS,MDA and Fe²⁺)are detected by kit,finally,the changes of mitochondrial morphological in alveolar epithelial cells of mice were observed by transmission electron microscopy.（2）.After intraperitoneal injection of ferrostatin-1（Fer-1）,the survival rate of mice was observed.Changes of ferroptotic gene GPX4 in lung tissues of mice were detected by q RT-PCR and western Blot.The biochemical indices of iron death(GSH,ROS,MDA and Fe²⁺)were detected by kit.（3）.To construct CLP moels of HSF1^+/+and HSF1^-/-,HE staining was used to observe alveolar interstitial hyperemia and inflammatory cell infiltration to verify the protective effect of HSF1 on acute lung injury in sepsis mice.q RT-PCR and western blot were used to detect the changes of GPX4 in lung tissues of HSF1^-/-and HSF1^+/+mice in CLP model,as well as in LPS-induced ferroptosis model of MLE12 cells after HSF1 overexpression and HSF1 si RNA interference.And the changes of biochemical indexes(GSH,ROS,MDA and Fe²⁺)were detected by kit.（4）.Bioinformatics methods were used to predict the binding of HSF1 to the GPX4 promoter region,and Electrophoretic mobility shift assay（EMSA）and dual luciferase reporter gene assay were used to verify the regulation of HSF1 on the transcription level of GPX4.Results:（1）.Ferroptosis-related genes and biochemical indexes were changed in lung tissue and MLE12 cells of sepsis mice,with decreased expression of GPX4,decreased content of GSH,increased content of MDA and Fe²⁺,changes in mitochondrial morphology,crest reduction or disappearance,increased density of mitochondrial outer membrane and membrane damaged.After the addition of Fer-1,the changes of the above indicators can be reversed.Animal and cell models demonstrate that ferroptosis occurs in sepsis,and Fer-1 can alleviate acute lung injury in sepsis.（2）.Compared with HSF1 wild-type mice,lung injury was more severe in HSF1 knockout mice,and the expression level of GPX4decreased,GSH content decreased,MDA content increased,and Fe²⁺content increased.The same results were obtained after HSF1 si RNA interference in LPS-induced ferroptosis in MLE12 cells.However,after HSF1 overexpression,the opposite result occurred in MLE12 cells,GPX4expression increased,MDA content decreased,and GSH content increased.（3）.Electrophoretic mobility shift assay（EMSA）and dual luciferase reporter gene assay verified that HSF1 could regulate GPX4 expression at the transcriptional level.Conclusion（1）Ferroptosis is involved in sepsis-induced acute lung injury,and inhibition of ferroptosis can alleviate septic lung injury.（2）HSF1 play a protective tole through inhibiting ferroptosis on lung tissues of septic mice and LPS induced ferroptosis in MLE12 cells.（3）HSF1 can promote the expression of GPX4 through binding HSE,and ultimately inhibit sepsis-induced acute lung injury.Figure:20,table:7,reference:62...