Implanting The HAECs As Well As Its Exosome Into Rat With Acute Myocardial Infarction Would Have A Positive Impact On The Cardiac Function

PartⅠ Human amniotic epithelial cells were cultured and the exosomes were extracted.Objectives:We cultured the Human amniotic epithelial cells(hAECs)and extracted the exosome culture supernatant for next exploration.Methods:We extracted exosomes according to the proposal of exosomes extraction kit,the concentration and characteristic of exosomes was identified.Results:About 300ug extractives were extracted via BCA kit in every T75 cell culture bottle.The electron microscope showed that the extractives were oval and round,with variable size lipid cystic vesicles.The average value(128.5±8.2nm)and main distribution(136.2±.7 nm)of extractive is within the range of exosomes,and the results of surface marker antigen CD63 and CD81 showed that the positive labeling rate was 86.2±1.4(%)and 94.9±1.1(%)respectively.Conclusions:It was efficient and convenient way to extract exosome via the exosomes extraction kit.The characteristic of extractives matched with the exosomes in electron microscope,FCM and expression of surface marker antigen CD81 and CD63.Part Ⅱ The experiment about the cardiac function could be improved by implanting Human amniotic epithelial cells and its exosomes in rat acute myocardial infarction model.Objectives:Establish the stable model of acute myocardial infarction in rat,and observation whether cardiac function could be improved by implanting the Human amniotic epithelial cells(hAECs)and its exosomes in it.Methods:1.Establish the stable model of acute myocardial infarction in rat,and random divide into 3 groups:treat with hAECs;treat with exosome and control.2.We conducted electrocardiogram during coronary artery ligation;and ultrasound cardiogram was performed before surgery and after the surgery 7,14 and 28days to detect the change of cardiac function in each group.3.We random selected one of rat in each group to see the distribution of hAECs and exosome,after the surgery 7 days;and 28 days after the surgery the rest heart tissues were collected and analyzed with Masson’s Trichrome stain to explore the fibrosis of tissues.Results:1.After ligated the coronary artery,the electrocardiogram showed typical dynamic changes.2.ultrasonic cardiogram showed 7,14 and 28 days after surgery,the EF and FS were lower in control group than those in therapy group(P<0.0001),and 14 and 28 days after surgery,LVEDD and LVESD were higher in control group than those in therapy group(P<0.05).3.7 days after surgery,the hAECs and exosomes was scattered in the myocardial tissue;28days after surgery,the area of tissue fibrosis in the control group was larger,and the difference between groups was significant(P<0.0001)。Conclusions:After acute myocardial infarction in rats,implant the hAECs and exosomes could improve the cardiac function。Part Ⅲ The protected mechanism would be explored by vitro experiment.Objectives:Establish the lumen formation ability and apoptosis detection of myocardial cells to explore the potential mechanisms how the hAECs and exosomes could improve the cardiac function.Methods:1.we co-cultured the human umbilical vein endothelial cell(HUVECs)with hAECs and its exosomes,set a blank control,and detected the lumen formation ability of HUVECs.2.we co-cultured the myocardial cell(H9C2)with hAECs and its exosomes,set a blank control,after 5 hours in simulated anaerobic environment,and we detected the apoptosis rate of myocardial cells during AnnexinV-FITC/PI of flow cytometry.Results:1.The lumen formation ability in hAECs group and exosome group were better than in control group(P<0.0001).2.The apoptosis rate of myocardial cells in hAECs group and exosome group were significant decrease than in control group(P<0.0001).Conclusions:The hAECs and exosomes could improve the cardiac function by promoting angiogenesis and reducing apoptosis of cardiac myocytes,which was one of the potential mechanism.