Effects Of Xinmaitong On The Expression Of Akt/PKB And ENOS During Oxidative Injury On Cardiac Myocytes In Rats

Objective:To explore effects of Xinmaitong on the expression of PKB and eNOS under oxidative stress in cardiomyocyte injury model induced by H₂O₂.Methods:1:Single cardiac myocyte from neonatal rats was got by the enzyme digestion way and identified it.A cardiomyocyte injury model induced by H₂O₂ was used in this experiment.2:The cells were seeded into 24 well culture plates and were cultured in 37℃,5% CO₂,and 95%humidity condition,then the cells were randomly divided into 4 groups: Control group(15%non-medicated serum+DMEM),model group(15%non-medicated serum +H₂O₂+DM EM),and Xinmaiong group(15%serum containing Xinmaiong +H₂O₂+DME M), DAng-I(15%non-medicated serum+H₂O₂+DMEM+DAng-I).6 wells/each group.On the third day,interventions were given as following:Control group:Cardiomyocytes were cultured in DMEM containing 15%normal mouse serum for 24 hours,then the cells were cultured in DMEM without serum for 6 hours.Model group:Cardiomyocytes were cultured in DMEM containing 15%normal mouse serum for 24 hours,then the cells were cultured in DMEM and added H₂O₂(200μmol/L) into wells for 6 hours.Xinmaitong group: Cardiomyocytes were cultured in DMEM containing 15%serum containing Xinmaitong for 24 hous,then the cells were cultured in DMEM and added H₂O₂(200μmol/L) into wells for 6 hours.DAng-I group:Cardiomyocytes were cultured in DMEM containing 15%normal mouse serum and DAng-I(5μmol/L)for 24hours,then the cells were cultured in DMEM and added H₂O₂(200μmol/L) into wells for 6 hours.At last,the plates were taken from the culturing box.The Cells were collected for ICC,total RNA extracted and the expression of PKB/eNOS was examined by RT-PCR method.Results:1.A model of cardiomyocyte damaged due to H₂O₂ was made and identified successfully.2.The level of phosphorylation of PKB was low in control group,but it was up-regulated in the model group and other groups.And it was higer in Xinmaitong plus DAng-I group(P＜0.05) compared with model group.There was no significant difference between the DAng-I group and Xinmaitong group(P＞0.05).Similar changes was found in expression of PKB detected by RT-PCR method.3.Expression of eNOS was higher in the control group,but the level was down-regulated in other groups.Expression of eNOS was higher in drugs groups than that in model group(P＜0.05),but there was no significant difference in Xinmaitong group and DAng-I group(P＞0.05).Conclusion:1.The cardiacmyocytes can be damaged in H₂O₂ 200μmol/L.2.Xinmaitong and DAng-I play a protective role on cardiomyocytes injury H₂O₂-induced,the protective role is relation with the activation of PKB and eNOS in cardiomyocytes.