| Research surroundingsAcute myocardial infarction (AMI) is a familiar and common disease thatseverely threatens the elder's health. At the present time, the ultimate treatmentto regenerate the infarcted myocardium and the pathological coronary arteries islacking. Although dissolving the thrombus and catheterizing in time in 4-6 hoursafter disease breaks out can return the blood of the infarct-associated vesselsand save the dying cardiomyocytes, the reperfusion therapy only can limit theinfarcted size and have no effect on the infarcted myocardium and the ischemiccardiomyocytes of the majority patients have infarcted when they go to see thedoctor. Lacking the ability of differentiation, the infarcted myocardium can onlybe replaced by the fibroblasts to form the scar tissue and have no systolicfunction. Thus the patients are easy to suffer heart failure, even sudden death,and their life quality is also very poor. The cellular transplantation techniquegives us a good prevision to re gnerate the injured myocardium. Fetalcardiomyocytes, embryonic stem cells, skeletal myoblasts, bone marrow stemcells became the focus of this field in present times. To gain the donor of thefetal cardiomyocytes is difficult and the embryonic stem cells also have theethical, legal problems and immune rejection. So it's difficult for them to use inclinic. Skeletal myocyte is also transversus as cardiomyocytes, using the methodof skeletal myoblasts transplantation to regenerate the injured cardiomyocyteshave made some progress. But the quantity of skeletal myoblasts is very limited,just about 4%-8% of the skeletal myocytes, and also decrease with age. At thesame time, the location of skeletal myoblasts is separate and difficult to gatherthe plenty of quantity of myocytes. It's indicated recently that mesenchymalstem cells (MSCs) are multipotent tissue stem cells that can be induced in vitro to differentiate into osteoblasts, chondroblasts, tendon, ligament, fibroblasts, nerve cells and muscles, et al. In addition, MSCs are easy to be separated and cultivated, have the potential ability to expansion in vitro, do not express the costimulation molecule B7, etc. All these characters make MSCs become the focus of the cardiology and may be become the ideal therapeutic method to regenerate the cardiomyocytes after the myocardial infarction.Objectives( 1) To establish a method for isolation, purification and cultivation of MSCs.( 2) To observe whether the human MSCs can differentiate into cardiomyocytes in vitro. ( 3) To observe the migration and differentiation of rat MSCs in normal and infarcted heart tissue. ( 4) To observe the therapeutic effect of the homogeneously exogenous transplantation of rat MSCs on treatment of the rat myocardial infarction. ( 5 ) To observe the therapeutic effect of the heterogeneously exogenous transplantation of human MSCs on treatment of rat myocardial infarction. ( 6) To observe the therapeutic effect of transplanting the myoblasts derived from human MSCs and infected VEGF gene on treatment of the rat myocardial infarction. Above all, our purpose is to provide new ideal, new approach and new method for treatment of myocardial infarction, and to lay a good foundation for the use of stem cells in clinic ultimately.Methods and resultsPart I : We examined the isolation, purification, expansion and differentiation of human MSCs into cardiomyocytes in vitro. Bone marrow mononuclear cells were isolated with 1.073g/mL Percoll. MSCs were obtained by removing the non-adherent cells. Then the MSCs were purified and expanded through passaging in time. To prevent the MSCs from differentiating or slowing their rate of division, each primary culture was replated (first passage) to 2-3 new plates when the cell density within colonies became 70%-80% confluent approximately 2-3 days after replating. 5.0X 105 primary MSCs were expanded for twelve passages, and 2.3 X 109 MSCs were obtained, and increased about 4.6 X 103 folds, and still maintain the s...
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