Mechanism Of Pioglitazone Against High Glucose-Induced Inflammatory Injury In H9c2 Cardiomyocytes

Objective:H9c2 cardiomyocytes were cultured with high glucose to simulate the microenvironment of diabetic myocardium and observe the changes of cell activity.To observe the effect of pioglitazone on the activity of H9c2 cardiomyocytes and the expression of inflammatory markers in high glucose microenvironment.To observe the role of p38MAPK/NF-κB signaling pathway in cardiomyocyte injury caused by high glucose.Methods:1.Simulated cardiomyocytes injury induced by high glucose.H9c2 cells derived from rat embryonic myocardial tissue were cultured in vitro with 5.5m M(mmol/L)low glucose DMEM.Myocardial cells were incubated with 35 m M high glucose DMEM medium for 24 h and 48 h respectively,and the high glucose injury model was established.2.Detection of the activity of H9c2 cardiomyocytes.The H9c2 cells were grouped as follows: NG,HG,HG + PGZ(5,10,15,20 μM)were detected by CCK8 method after treatment for different time(24h,48h).3.Changes in the expression of inflammatory cytokines in H9c2 cardiomyocytes.The cells were divided into four groups:NG,HG,HG + Low dose PGZ(HG +PGZ 10μM,LOW group)and HG + high dose PGZ(HG + PGZ 20μM,HIGH group)according to the cell viability test results.After 48 h of intervention,the cell medium supernatant of each group was collected,and the levels of TNF-α and IL-1β were determined by enzymy-linked immunosorbent assay(ELISA).4.Changes of p-p38 MAPK and NF-κBp-p65 protein expression in H9c2 cardiomyocytes.Cell grouping was the same as "3".After 48 hours of treatment,we detected the expression of phosphorylated p38 and p65 protein in each cell groups by Western-blot.Results:1.Changes of H9c2 cell viability after intervention for different time in different treatment groups.(1)The activity of the HG group was significantly lower than NG group after 24 h intervention(P < 0.01),and the activity of the HG group was more significantly decreased after 48 h intervention(P < 0.01).(2)Compared with the HG group,the cell viability of the HG+ low-dose PGZ group increased after 24 h and 48 h intervention(P > 0.05,P < 0.05),and the cell viability of the HG+ high-dose PGZ group increased more obviously after 24 h and 48 h intervention(P < 0.05,P < 0.01).2.The expression levels of inflammatory cytokines TNF-α and IL-1β were detected by ELISA.After 48 h intervention,compared with NG group,the content of inflammatory cytokines TNF-α and Interleukin-1β in HG group were increased(P < 0.01;P < 0.01);Compared with HG group,the expression levels of TNF-α and Interleukin-1β in HG+ low concentration pioglitazone group decreased(P > 0.05,P < 0.05),and the expression levels of TNF-α and Interleukin-1β in HG + high concentration pioglitazone group decreased more significantly(P < 0.05,P < 0.01).3.The protein expression levels of p-p38 and p-p65 were detected by Western-blot.Compared with the NG group,the expression levels of phosphorylated p38 and p65 protein in HG group were increased after 48 h treatment(P < 0.01;P < 0.01);Compared with the HG group,the protein expression levels of p-p38 and p-p65 in the high glucose + low dose PGZ group were decreased(P < 0.05,P < 0.05),and the protein expression levels of p-p38 and p-p65 in the high glucose + high dose PGZ group were reduced more obviously than those in the low dose group(P < 0.01,P <0.01).Conclusions:The PPAR-γ agonist pioglitazone may reduce the inflammatory injury of H9C2 cardiomyocytes induced by high glucose by restraining the p38MAPK/NF-κB signal transduction pathway.