Isolation And Amplification In Vitro Of Rabbit Endometrium And Smooth Muscle Cells

Isolating and culturing endometrial epithelial cells, stroma cells, and smooth muscle cells can provid ideal model for studying on physiology and pathology such as pregnancy and disease of uterus. Obtaining a great deal of puried uterine cells is one of the critical steps as well as domestic and foreign scholars are concerned about issues of the world. To explore access to high quantity, high purity and strong vitality of the uterine cells, rabbit uterus endometrial epithelial cells, stroma cells and smooth muscle cells were isolated and purified by digestion of trypsin, filtration, centrifugation and different tempo adherence, and amplified by RCCS and microcarrier, and cultured with different concentrations of estrogen (E2) and progesterone (P4). The cultured cells were observed by light microscope, characterized by immunocytochemical and immunofluorescence staining, and analyzed the purity by flow cytometry. The effect of E2 and P4 on cell proliferating was indirectly determined by MTT. Results:1 High purified endometrial epithelial cells, stroma cells and smooth muscle cells were obtained by digestion of trypsin, filtration, centrifugation and different tempo adherence. Immunocytochemical staining and flow cytometry analysis showed that the purities of the three cells were 96%, 95% and 98%.2 The cells were efficiently amplified using RCCS and microcarriers. The cell proliferation rate was fast and it was about 4 times about the proliferation rate of the general culture There were no difference beteween the cell characteristics of RCCS cultivation and general cultivation.3 Combination of estrogen (100nmol/mL) and a little progestogen (10nmol/L) can stimulate the proliferation of epithelial cell. Progestogen (100nmol/L) can stimulate the proliferation of stroma cell. And both of estrogen and progestogen also can stimulate the proliferation of smooth muscule cell.The method of isolating and culturing endometrial epithelial cells, stroma cells, and smooth muscle cells was improved. Large scale amplification of cells was implemented by using RCCS and microcarriers. It was proved that Estrogen and progestogen can stimulate the proliferation of above mentioned cells. All of above established the bases for research of uterine disease and embryo implantation...