| Myocardial infarction (MI)is one of the major health killers for human being , the morbidity of which is increasingly soaring in concomitance with the improvement of people's life . Due to the myocardial ischemia and anoxia in the infarctional areas, some pathological changes take place, such as the massive necrosis of cardiomyocytes, the proliferation of fibrous tissues, the formation of scars, the remodeling and dysfunction of cardiac ventricle, and often end in refractory heart failure. The present treatments for MI mainly aim at saving the dying cardiomyocytes , improving the function of survival cardiomyocytes , blocking the neuroendocrine system and inversing ventricular remodeling . Although the unregenerability of cardiomyocytes makes it impossible to restore myocardial tissues from the scar tissues , fortunately , cell transplantation can increase the amount of cardiomyocytes in lesion and therefore provide a new therapeutic strategy for MI repair . In recent years, many progresses have been made by foreign and domestic research workers in the field of cellular cardiomyoplasty, especially regarding the embryonic stem cells, the fetal cardiomyocytes, the skeletal muscle myoblasts, the autologous cardiomyocytes, and so on, engrafted into the myocardium. However, the optimal cell type available for transplantation has not been confirmed yet, owing to the problems of the sources of donor cells, the immunological rejection caused by the xenogenic cell transplantation, and the junctions between the grafted cells and the hostcardiomyocytes. As a sort of multipotential stem cell, the bone marrow stromal stem cells have been found to be able to differentiate into myogenic cells or cardiomyocytes after induction in vitro. The objective of the studies is to observe the in vitro culture of the bone marrow stromal stem cells, to observe their influences on the growth, differentiation, and function of postinfarction myocardium after selectively implanted them into the myocardial tissues via the coronary artery, and to provide experimental data of clinical applications to myocardial regeneration with the bone marrow stromal stem cells .Methods: 1.The bone marrow stromal stem cells were isolated by their plastic adherence, then induced by 5-azacytidine to facilitate their differentiation into myogenic cells. The phase-contrast microscope, electron microscope, and immunocytochemical stain were applied to identify the induced cells. 2. An animal model of canine MI was created by means of clipping the anterior descending branch of the left coronary artery. Color Doppler ultrasonic examinations were performed to evaluated the changes of ventricular function before and after clipping left coronary. 3. With the help of coronary artery angiography, the induced bone marrow stromal stem cells labeled with DAPI were selectively engrafted into myocardium through the left coronary artery. The growth, differentiation, and proliferation of these cells in the myocardial tissue were detected. Again, Color Doppler ultrasonic examination was performed to evaluated the changes of ventricular function before and after cell implantation.Results: 1. 2ml bone marrow was extracted and primary cultured in 25cm2 culture flask containing 10ml 20%FBS DMEM. The scattering clone fusiform cell clusters were observed 3 days later, and were induced by 5-azacytidine for 24 hours. The amount of cells amplified to 1×106 and 2.7×108 respectively at the end of 2 and 4 weeks .The cultured cells showedstick-like morphology with striated structure after 2 weeks observed by phase-contrast microscope, appeared plenty of cytoplasmic ribosome and mitochondria, much myofilament-like texture, mononuclear locating in the central of the cells after 4 weeks observed by electron microscope, and displayed positive reaction to anti-desmin, anti-smooth muscle actin, and anti-troponin I antibodies with immunocytochemical stain , all of which confirmed that the cloned bone marrow stromal stem cells differentiated into the myogenic cells with...
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