The Study of Human Cardiac Stem Cells Differentiated into Cardiomyocytes in vitro

HOU Hong; Lü Anlin; DA Jing; HOU Zhaolei

Abstract：To explore the differentiation of cardiac stem cells into cardiomyocytes and the methods of improving the differentiation rate of 5 -AZA、II Ang and coinduced cardiac stem cells.Myocardial tissues obtained from the right atrial appendage of patients during surgery were digested with collagenaseⅡwith informed concent.The isolated cells were inoculated in culture flask and passaged,P2 to P8 cells were collected and incubated with monoclonal anti -human PE -CD117 specific antibody,then sterile sorting with flow cytom-etry.Then purified CSCs were collected for cultivation.After fusion around 90%,purified c -kit +CSCs were collected for differentia-tion and divided into four groups:(1)Control group(cells culture medium),(2)5 -AZA group,(3)Ang -Ⅱgroup,(4)5 -AZA com-bined Ang II group.The expression of protein of connexin 43 (Cx43)and cardiac Troponin I (cTn I)by Western Blot in 4 week and the cardiomyocyte differentiation rates with flow cytometry were determined.Identification of induced human CSCs cTn I,Cx43 protein expression:the expression of cTn I and Cx43 protein of each group by immunofluorescence staining was determined and the results showed that each group expressed protein of cTn I and Cx43,and weak expression had been found in the control group,strong expres-sion in the 5 -AZA combined Ang II group.Identification of induced human CSCs differentiation rates:the results detected by flow cy-tometry showed the cardiac differentiation rate of each group as follows:the control group was (27.86 ±4.52)%;the 5 -AZA group was (52.71 ±7.40)%;the Ang II group was (40.48 ±7.02)%;the 5 -AZA combined Ang II group was (64.74 ±6.11)%.Sta-tistics of the differentiation rates between other groups had significant differences (P <0.05).5 -AZA,Ang II can induce cardiac stem cells to differentiate into cardiomyocyte,and the 5 -AZA combined Ang II group is superior to single induced group.