Study Of The Bone Marrow Stem Cells Differentiating To Cardiac-like Myocytes

IntroductionIschemic heart disease, including myocardial infarction, is the major cause of congestive heart failure, accounting for 1/2 cause of death in cardiovascular diseases. Up to now, treatment strategy for ischemic heart disease are medication , interventional therapy and surgery, but they don' t solve foundamental question of cardiac myocytes regeneration. Present, homogeneity heart transplantation is the only radical cure for heart failure. In American, 2500 cases of heart failure received homogeneity heart transplantation, but patients who really need are 30000. The cause is lack of donators. Other methods, such as left ventricular assistor and artificial heart, are very expensive, most of patients don t accept it. In the situation, it is very necessary to find a novel method to change basic questions. Mass animals trials showed that stem cells differentiated to cardiac myocytes in vitro and in vivo. Especial for bone marrow stem cells, they prior to other candidate cells ( embryonic stem cell, cord blood stem cells, satellite cells,cardiac myocytes,et al. ) and apply for clinics. Bone marrow stem cells can be harvested by routine way and no moral problems. The cells from bone marrow stem cells can form electric - mechanical couple with near host myocytes. In addition, acceptors avoid immunological rejection reaction. Dates from clinical trials showed that autological bone marrow stem cells transplantation improved heart function , reperfuse, and decreased infarct size. In view of above mentioned theories, we selected Wistar rats as objects and investigated recovery of situ transplantation for damaged myocardium. At the same time, we studied if bone marrow mesenchymal stem cells could be induced into cardiac -like myocytes and their electrophysiology properties. We divided experiment intothree parts, state as follow:(—) Mobilize autologous bone marrow stem cells to repair infarcted myocardiumObjective: Autologous bone marrow stem cells were mobilized and released by cytokines. On the basis of homing and injured microenvironment theories , we investigated the effective reparation of situ transplantation for acute my-ocardial infarction in rats.Methods.-(1) We divided 60 Wistar rats into situ transplantation group and control group; To duplicate rat' acute myocardial infarction model by injection of drug; CK^LDH level were checked by automatic biochemistry analyzer;(2)30 Wistar rats were injected rhG - CSF 50 jig/kg, day;( 3 ) We did a control analysis on pathological section between two groups by histological staining technique and computer graphic analysis;(4) Heart function were checked by polygraph system after AMI model 4 weeks.Results:(1) Serum cardiac enzymes were higher after AMI model 48 hours than it before AMI model. We saw local necrosis region with grid and trabs shape in endocardium of left ventricular apex and papillary muscle;(2)1116 infarction size in situ transplantation group was smaller than one in control group;(3)The heart function parameters improved significantly in situ transplantation group compared with control group.Discussion:Stem cells are daughter cells that differentiation potency gradually grow down. They can amplify and remain undifferentiation in vitro. According to their differentiation potency, they are divided into totipotential stem cell, multipoten-tial stem cell, multipotent stem cell (adult stem cell). Bone marrow stem cells are multipotent stem cell ( adult stem cell). Bone marrow stem cells have abilityto transdifferentiation, which differentiate into another type of cells in cytokine and microenvironment. It is important for cardiomyocytes regeneration. Therefore , we mobilize rats bone marrow stem cells by granulocyte colony - stimulating factors, migrating and homing to zone of necrosis. It was found that area of myocardial infarction was smaller in situ transplantation group than in control group and heart function improved obviously in situ transplantation group. This method was simple and no invasive compared with interventional means, but further studies and evaluations will be necessary.Conclusions:(l)Rat' s acute myocardial infarction model could be completed by injecting 10 mg/kg isoprenaline interaperitoneally;(2)infarcted myocardium were repaired and heart function improved by u-sing cytokines.Key words: acute myocardial infarction; cytokine; situ transplantation(H)Bone marrow mesenchymal stem cells can be induced to differentiate to cardiac - like myocytes in vitroObjective:To investigate the bone marrow mesenchymal stem cells induced and differentiated to cardiac - like myocytes in Wistar rats.Methods;(1) We selected Wistar rats to isolate and culture bone marrow mesenchymal stem cells;(2)The first generation of bone marrow mesenchymal stem cells were induced with lOujnol/L concentrations 5 - azacytidine agent;(3 ) We identified and evaluated expression of surface markers, including CD34,CD44,CDw90;(4) After bone marrow mesenchymal stem cells were treated with lOjimol/L concentrations 5 -azacytidine agent, they were checked by immunohistochemis-try staining of antibodies against a - sarcomeric actin and troponin T in 2^4 week.Results;(1) The first generation of bone marrow mesenchymal stem cells expressed CD^O arid CD34 was negtive;(2)The cardiac - like myocytes expressed a - sarcomeric actin and tropo-nin T.Discussion:Now, specific surface markers in bone marrow mesenchymal stem cells have not been discovered, which is difficult to identify and purify bone marrow mesenchymal stem cells. It is well known that bone marrow mesenchymal stem cells expresse SH2, SH3> CD29, CD44, CD71, CD90, CD120a, CD124, CD14, CD34, and CD45 is negtive. We cultured and purified bone marrow mesenchymal stem cells by adherence method. These stem cells were in good condition and easy to be induced in vitro.Conclusions: We verified that the first generation of bone marrow mesenchymal stem cells could be induced to differentiate to cardiac - like myocytes. This provided theory foundation and technique method for cardiomyocytes induced in vitro and applied in clinic.Key words: bone marrow mesenchymal stem cells; induce; cardiac - like myocytes(H) Changes of intraceUular calcium concentration in cardiac - like myocytesObjective; To study the effects of verapamil, endothelin on [ Ca2+ ]{ in cardiac - like myocytes derived of bone marrow mesenchymal stem cells. Methods:(1) Bone marrow mesenchymal stem cells and cardiac myocytes were cultured by primary method;(2) Bone marrow mesenchymal stem cells differentiated to cardiac - like myocytes by 5 - azacytidine induction agent;(3) We divided our test into three groups,including the first generation of bone marrow mesenchymal stem cells, cardiac - like myocytes and cardiac myocytes;(4) Hie alteration of [ Ca2+ ], affected by verapamil, endothelin was ob-...