The Study That Reactive Oxygen Species Activates Mitochondrion Apoptosic Pathway In Cardiac Myocytes Apoptosis By High Glucose

ObjectiveTo investigate the effect of different concentration of glucose as well as reactive oxygen species and p53 on cardiac myocytes apoptosis induced by high glucose.Reveal oxidative stress and the transposition of p53 to mitochondrion may be involved in the development of diabetic cardiomyopathy,so provide a direction to cure diabetic cardiomyopthy.Methods1. Cultivate cardiac myocytes of neonatal rat, devided into four groups as follows: group A(control group,glucose concentration is 5.6mmol/L),group B(glucose concentration is 16.7mmol/L),group C (glucose concentration is 25mmol/L)andgroup D(glucose concentration is 33.3mmol/L).The viability of cardiac myocytes was measured by MTT assay,the reactive oxygen species of cardiac myocytes was detected by fluorospectrophotometer in order to get an optimal glucose concentration.2. High glucose-induced myocardial cell as a model,using different intervention factors ,devided into four groups as follows:control group(Mannitol 25 mmol/L),high glucose (25 mmol/L),high glucose+ antioxidant(10mmol/L),high glucose+ Pifithrin-α(50μmol/L).The ROS of cardiac myocytes was measured by fluorospectrophotometer,the SOD of supernatant was measured by spectrophotometer,apoptosis was detected by flow cytometry, intracellular expression of caspase-9 and the level of p53 were analyzed by Western blotting.Results1. The cardiac myocytes were harvested by extraction of mixed enzymatic digestion, and highly purified by different adhesion time, They showedα-smooth muscle actin(+) by cell immunohistochemistry technique.2. Compared with group A(control myocytes),the viability of cardiac myocytes from group B to group D was gradually degraded,while ROS was increased by degrees.Among the total,compared with group C,the viability of cardiac myocytes and ROS of group D had no difference.3. Compared with the control group, in the high glucose group, the results suggested that the cardiomyocyte apoptosis rate, ROS as well as intracellular expression of caspase-9 and p53 increased , while SOD activity decreased significantly. However, in the antioxidant group,the cardiomyocyte apoptosis rate, ROS as well as intracellular expression of caspase-9 and p53 decreased , SOD activity increased remarkably compared with the high glucose group. The cardiomyocyte apoptosis rate,ROS as well as intracellular expression of caspase-9 and p53 decreased , SOD activity increased in the Pifithrin-αgroup in comparison with that in the high glucose group.Conclusion1. High glucose can induce neonatal rat cardiomyocyte apoptosis in vitro.2.The transposition of p53 to mitochondrion induced by active oxygen activates mitochondrial apoptosic pathway in cardiac myocytes apoptosis induced by high glucose.3. P53 had retroaction to ROS,each other that influence cardiomyocyte apoptosis.