OBJECTIVETo isolate, culture and identify of human endometrial stromal stem cells.

METHODSEndometrial tissue was obtained from 10 women undergoing hysterectomy. Purified stromal stem cell suspensions were then obtained by selecting cells with magnetic bead sorting and colony-forming. The surface markers of stromal cell were identified by flow cytometry. Proliferation of stromal stem cell was observed by MTT methods. The osteogenic potential was evaluated by alizarin red staining, the adipogenic potential by oil red O staining. Then the adipogenic and osteogenic specific markers of differentiated cells assayed by RT-PCR method. Expression of cell surface antigen OCT-4 was detected with immunocytochemical staining.

RESULTSEndometrial stem cells were successfully isolated from human endometrial tissue. They were stably proliferated and subcultured in vitro. Most of the passage cells expressed mesenchymal stem cell markers CD90, CD105 but not hemopoietic markers CD34, CD45. The analysis of cell cycle indicated that the percentage of G2-M phase and S phase cells increased. The growth curves of the third passage presented in "S" shape. After cultured in differentiation medium, the cells differentiated toward adipoblasts and osteoblasts as verified by positive staining with Oil Red O and alizarin red staining. Under induction,cells expressed osteogenic and adipogenic marker genes. The immunocytochemical staining of OCT-4 was positive.

CONCLUSIONHuman endometrium contains endometrial stromal stem cells, which present characteristics of mesenchymal stem cells and may be used as seed cells for tissue engineering.