Study On The Protective Effect Of Resveratrol On Myocardial Injury In Sepsis And Its Mechanism

Objective: Investigating the protective effect of resveratrol(Res)on myocardial injury in sepsis and its mechanismMehtods: Animal experiment: 24 male SD rats were randomly divided into 3 groups:(1)sham-operated group(i.e.,control group,Con),(2)sepsis group(CLP),and(3)sepsis + resveratrol group(CLP + Res).Sepsis model preparation was performed using cecum ligation perforation surgery(CLP).The rats in the sham-operated group were identical to those in the CLP group except that they were not subjected to ligation and perforation;the sepsis group was subjected to sepsis modeling according to the CLP surgical method;the rats in the resveratrol group were injected with resveratrol(50 mg/kg)intraperitoneally 0.5 h after the CLP procedure.Echocardiography was used to observe the cardiac function of rats in each group,HE staining was used to observe the morphological changes of myocardial tissues,CK-MB and LDH markers in serum of rats,ELISA kit was used to detect IL-6,TNF-α and IL-10 in serum,TUNEL kit was used to detect apoptosis of myocardial cells,and transmission electron microscopy was used to observe the mitochondrial changes in myocardial tissues.Cell experiments: H9c2 cardiomyocytes in the same culture condition were randomly divided into 10groups:(1)control group(Con),(2)sepsis group(LPS),(3)Res group(Con+Res),(4)Res-activated LPS group(LPS+Res),(5)DMSO-activated LPS group(LPS+DMSO),(6)JNK agonist group(Con+JNK agonist),(7)LPS group with JNK agonist action(LPS+JNK agonist),(8)LPS group with Res and JNK agonist action(LPS+JNK agonist+Res),(9)LPS group with Res and JNK inhibitor action(LPS+Res+JNK inhibitor),(10)JNK inhibitor action LPS group(LPS+JNK inhibitor).The CCK-8 kit was used to detect the cell viability of each group,the kits to detect the markers of myocardial injury CK-MB and LDH,ELISA to detect the levels of cellular inflammatory factors IL-6,IL-10 and TNF-α,the kits to detect the levels of redox-related substances MDA,SOD and GSH-Px in cells,the fluorescent probes to detect the expression of reactive oxygen species(ROS)in each group of cells,and Western Blot was used to detect the expression of JNK,p-JNK and apoptosis-related proteins Bax,Bcl-2,Cyt-C,Caspase-3 and Cleaved-Caspase-3.Statistical analysis: Graph Pad Prism 9 software was used for statistical analysis.Numerical variable data were expressed as mean ± standard deviation;one-way ANOVA was used for comparison between groups,and P<0.05 was statistically significant;Pearson correlation analysis was used to analyze the correlation between the data obtained from animal and cell experiments,and P<0.05 was statistically significant,0.5 <|r |≤0.8 was significant correlation,and 0.8 <| r |<1 was highly correlated.Results: Animal experiments:(1)Compared with the sham-operated group,rats in the CLP group had significantly decreased cardiac function EF and FS;myocardial fiber disorder and irregular cardiomyocyte morphology were more obvious;CK-MB and LDH content were increased;inflammatory factors IL-6 and TNF-α were elevated and IL-10 was decreased;apoptotic TUNEL staining fluorescence was enhanced;mitochondrial swelling and deformation of cardiomyocytes were obvious,disorderly arrangement and The internal ridge was significantly reduced,and mitochondrial damage was obvious.(P<0.05)(2)Compared with the CLP group,the CLP+Res group: cardiac function EF and FS increased;HE staining of myocardial fibers was more regular and inflammatory cell infiltration decreased;CK-MB and LDH content decreased;inflammatory factors IL-6 and TNF-α decreased,anti-inflammatory factor IL-10 increased;apoptotic TUNEL staining fluorescence decreased;mitochondrial damage was(P < 0.05).(P < 0.05)Cellular experiments:(1)Compared with the control group,cell viability was significantly decreased in the LPS group;myocardial injury markers CK-MB and LDH content was increased;pro-inflammatory factors IL-6 and TNF-α were significantly increased and anti-inflammatory factor IL-10 content was significantly decreased;increase in cellular reactive oxygen species(ROS);elevated redox-related substances MDA,SOD and GSH-Px were decreased;apoptosis related proteins p-JNK,Bax,Cyt-C,Cleaved-Caspase-3 were increased and the expression of anti-apoptotic protein Bcl-2 was decreased.(P < 0.05)(2)Compared with the LPS group,cell viability was significantly higher in the LPS+Res group;CK-M and LDH levels were decreased;pro-inflammatory factors IL-6 and TNF-α were significantly decreased and IL-10 was increased;ROS fluorescence was decreased;redox substances MDA was decreased and SOD and GSH-Px were increased;apoptosis-related proteins p-JNK,Bax,Cyt-C and Cleaved-Caspase-3 were increased,and the expression of anti-apoptotic protein Bcl-2 was decreased.(P < 0.05)(3)With JNK inhibitor intervention,the alteration of cellular indices was coterminous with the effect of Res.Conversely,the use of JNK agonists acting on H9c2 cardiomyocytes had the same damaging effect on the cells as LPS,and this effect was partially reversed by Res.(P < 0.05)Conclusion: Resveratrol(Res)acts to ameliorate myocardial injury in sepsis by inhibiting JNK/Bax/Cyt-C pathway-mediated mitochondrial apoptosis,suppressing inflammatory responses and oxidative stress.