The Effect Of CyclinA2on The Hypoxia Myocardial Cell

Background:Cardiovascular disease has becoming the focus of global health, one of the most common is myocardial infarction, treatment of the disease is very uncertain, CyclinA2expression allows myocardial cell cycle gene into differentiation activation, and increases the criticalarea myofilament density, improves cardiac function. CyclinA2gene transfection methods is expected to becoming a highly effective treatment of myocardial infarction.Objectives:Making hypoxia model of myocardial cells, to investigate the role of myocardial cells transfected by CyclinA2.Methods:1. cell culture:(1) Taking twelve SD rats’heart, then digestion,centrifugal and mark, put cells in Carbon dioxide incubator.(2) Immunohistochemical staining detect myocardial cells which fixed by4%paraformaldehyde.2. Transfecting adenovirus(1) When the cells culture36h, transfecting the adenovirus or empty vector virus into cells.(2)culturing cells12h, fixed and detected by Immunofluorescence technique.3. cell hypoxia-ischemia(1) Culturing the myocardial cells with three days in the sub-time periods of hypoxia.(2) Culturing the myocardial cells with sub-time periods of hypoxia which were transfected with adenovirus.4. Western blottingExtracting proteins from Myocardial cells cultured for three days, hypoxia myocardial cells,myocardial cells after transfecting adenovirus and hypoxia myocardial cells after transfecting adenovirus stored at-20℃. western blotting detecting. Results:1. Routine immunohistochemical staining showed that neonatal rat cardiomyocytes cyclinA2present in the cytoplasm and reduces with the increase of culture time.2. Neonatal rat myocardial cells were observed under the inverted microscope after differential velocity adherent for60mins and spot BrdU48h. Cells, most are small fusiform, were beating in different frequencies. After4d, cells had three main forms: spindle-shaped, oval-shaped, irregularly-shaped, pulsation frequency is approximately (80±7)/min.3. The immunofluorescence results showed that under the fluorescence microscope observation, the highest cyclinA2expression of hypoxia cardiomyocytes at6h, the lowest at12h, or even no expression.4. Expression of cyclinA2in myocardial cells was significantly higher after transfecting adenovirus.5. Expression of cyclinA2was changing after transfecting adenovirus, Group1: the cells were transfected by adenovirus, expression of cyclinA2was higher. Group2:the cells were transfected only by empty vector, expression of cyclinA2was no change. Group3:the cells were tranfected by adenovirus and cultured under ischemia and hypoxia, expression of cyclinA2was relatively higher.Group4:the cells were transfected only by empty vector and cultured under ischemia and hypoxia, expression of cyclinA2was lower.Group5:the control group, expression of cyclinA2was no change.Conclusions:1. CyclinA2reduced as time increases.2. CyclinA2expression of ischemia and hypoxia (6h) is into a rising trend, and then into a downward trend, up to12h to a minimum or even disappear.3. Expression of cyclinA2in myocardial cells was significantly higher and also increased in ischemia and hypoxia conditions.