| Background: Stem cell transplantation promises an effective way in treatment for heart diseases. Many kinds of stem cells have been used in animal models. Among them, mesenchymal stem cells (MSCs) become the most fascinating scenes because they are easily to be obtained and proliferated. Although many problems have yet to be determined, the good results in treating myocardial infarction and cardiomyopathies of animals paved its ways into clinical trials. At the same time, basic research must provide more details and knowledge so that we can use it more safely, effectively and rationally. The mechanisms underlying MSCs improvement of cardiac functions still need more analyzing, especially whether MSCs can differentiated into contractile cells in host myocardium or not. As fortechnical development now, it is hard to observe their contraction directly in beating hearts. This would impede evaluating and improving its functions if this problem was unclear.Objective: to observe structural development of MSCs in host myocardium and to analyze if they hold the integral structural basis for contraction. And to uncover the environmental factors which play major roles in this differentiation process.Materials and Methods:1. Titin, myosin heavy chain (MHC) and Connexin43 (Cx43) of normal myocardium and neonatal cardiomyocytes were detected by immunofluorescence. And sarcomeric structure was also observed through the detection of titin.2. MSCs were labeled with BrdU and DAPI and were injected directly into rats' left ventricle myocardium. Survival animals after operation were randomized into several groups as follows: instant groups, lweek, 2, 4, 6, 8, 10 and 12weeks groups respectively (n=3~4). Their hearts were harvested at each time point. Cryosections were obtained and HE staining was investigated. Z part and M part of titin, MHC as well as Cx43 were detected through immunofluorescence under common immunomicroscope and laser scan confocal microscope. The changes of host myocardium and fibrosis after injection were also observed.3. Culture supernatant of neonatal cardiomyocytes was harvested. MSCs were either cultured in conditioned media, the combination of supernatant and common media, or cocultured with neonatal cardiomyocytes at 1:1 ratio. After 7 days of induction, immunofluorescence was employed to detect titin, MHC and Cx43 expression in MSCs.Results:1. Through detecting titin, regular sarcomeric structure was easily to be observed in normal myocardium and cardiomyocytes. Cx43 was expressed at interface of adjacent cells.2. MSCs could survive in normal host myocardium for at least 12 weeks. Some of transplanted cells expressed titin and the assembled titin could be detected in the cells adjacent to host myocardium. Cx43 could be observed between these cells. But the size and structure of transplanted cells were different from normal contractile myocardium.3. Most of the grafted cells were isolated by fibrous tissue. Part of these cells differentiated into fibrocytes. The structure of host myocardium was slightly disturbed. Neovascularization could be observed. Tissue rejection was not found.4. Cx43 and titin were expressed in few of normal MSCs. Conditioned medium and coculture could activate MSCs' cardiomyocyte-like differentiation. It was more differentiated in coculture group than inconditioned group. MSCs and myocytes could form gap junction when cocultured in vitro.Conclusions: MSCs could survive and develop in normal myocardium. But they have not the integral structural basis for normal contraction because they were less differentiated and premature. Forming gap junctions with cardiomyocytes is necessity for MSCs' high level of development.
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