Protective Effects Of Paeoniflorin On Hypoxic Reoxygenation Injury In H9C2 Cardiomyocytes And The Effect Of MAPK Signaling Pathway

MIRI(Myocardial ischemia-reperfusion injury)is a condition in which the myocardium is continuously ischaemic due to blockage of coronary vessels caused by plaque dislodgement,vasoconstriction and other triggers,and after a period of time blood flow is restored,additional myocardial damage occurs,which has an impact on myocardial structure and in severe cases may result in myocardial fibrosis.Studies have shown that PF(Paeoniflorin)has protective effects against cardiovascular diseases,such as vasodilatation,improved blood flow,inhibition of calcium imbalance in mitochondria and anti-free radicals.In this experiment,an in vitro H/RI(Hypoxiareoxygenation injury)model was used to simulate MIRI at the cellular level to investigate the protective effect of PF on H/RI in H9C2 cells and its effect on the MAPK pathway.The details of the study are as follows.1.The protective effect of PF on H9C2 cell H/RIThis experiment investigated the optimal concentration of PF administration as well as the time of reoxygenation.Toxicity experiments were grouped into Control group,PF group(50 μM,100 μM,200 μM,300 μM),and cell viability was detected using CCK-8 after 24 h treatment with PF;reoxygenation time mapping was grouped into Control group,3 h group,6 h group,9 h group,3 h hypoxia,3 h,6 h and 9 h after reoxygenation Cell viability was detected using CCK-8;effectiveness experiments were grouped into Control group,H/R group,PF+H/R group(50 μM,100 μM,200μM),PF+H/R group was pretreated with PF for 24 h followed by hypoxia for 3h,and cell viability was detected using CCK-8 after reoxygenation for 3h;Anti-oxidative stress capacity tests AST,LDH,MDA,CK-MB.The results showed that the PF concentration of 200 μM had a better protective effect on H/RI of H9C2 cells,reduced the levels of AST,LDH,MDA and CK-MB,and improved the antioxidative stress ability.2.Effect of PF on H/RI apoptosis in H9C2 cellsThis experiment investigated the effect of PF on the antioxidant stress ability,apoptosis rate,ROS production and Caspase-3 protein and m RNA of H9C2 cells.The experiments were grouped into Control group,PF group,H/R group and PF+H/R group(200 μM).the apoptosis rate and ROS production were detected by flow cytometry,and the expression of Caspase-3 m RNA were detected by Western Blot was used to detect the expression of Caspase-3 and Bcl-2 protein,and RT-PCR was used to detect the transcription level of Caspase-3 m RNA.Observe the effect of PF on H/RI apoptosis in H9C2 cells.The results showed that PF decreased the apoptosis rate and ROS production in H9C2 cells;increased the expression of Bcl-2 and decreased the expression of Caspase-3.3.Effect of PF on MAPK signalling pathway in H9C2 cell H/RIIn this experiment,we investigated the effect of PF on MAPK pathway.The expression of p38 MAPK,JNK,ERK1/2 protein and m RNA were examined by Western Blot and RT-PCR.The results showed that PF could reduce the expression of p38 MAPK,JNK,ERK1/2 protein and m RNA.In summary,the protective effect of PF on H9C2 cell H/RI is multifaceted,and its mechanism of action may be related to the fact that PF increases the resistance of H9C2 cells to oxidative stress,decreases intracellular ROS levels,increases Bcl-2expression,decreases Caspase-3 expression,and decreases apoptosis rate,and its mechanism may be related to the regulation of MAPK pathway.