Differentiation Of Rabbit Bone Marrow Mesenchymal Stem Cells Into Smooth Muscle Cells: An In Vitro Study

Objective: To find an efficient method to induce rabbit bone marrow mesenchymal stem cells different towards smooth muscle cells in vitro, so as to establish a stabile cells source for further study.Methods: Rabbit bone marrow mesenchymal stem cells were isolated from bone marrow by the method of density gradient centrifuge. By adhering culture, a further purification was carried out. Morphology observation and Flow cytometry were performed to identify the stem cells. In order to induce them different towards smooth muscle cells, the cells were treated with 5-azacytidine, supernatant and direct cocultured with smooth muscle cells, respectively. Morphology observation and immunohistochemistry were performed to compare the inductivity of each means. Results: The isolated bone marrow mesenchymal stem cells comprised a single phenotypic population and displayed a fibroblast-like morphology with active prolification. Subcultivate cells grow well. The cells have very low expression of CD34, CD45 and high expression of CD90,indicates a high purity. In the induce experiment, treated the cells with 5-azacytidine and then cocultured them with smooth muscle cells resulted in the highest inductivity, just cocultured them with smooth muscle cells caused a much lower inductivity, cells which were induced by 5-azacytidine and supernatant also presented a low inductivity, but there were no apparent difference between the two groups. Only treated the cells with supernatant resulted in the lowest inductivity.Conclusion: By density gradient centrifuge and adhering culture, we can obtain a high purity bone marrow stem cell population. 5-Azacytidine induction in addition to coculture is an efficient way to induce bone marrow mesenchymal stem cells different towards smooth muscle cells in vitro.