CD133^+人脐血造血祖细胞的干性维持培养及鉴定

Long-term culture and identification of CD133^+ hematopoietic progenitor cells from human umbilical cord blood

目的从人脐带血中分选出CD133+造血祖细胞,并进行长时间干性维持培养。方法通过免疫磁珠法分选出人脐带血中的CD133+造血祖细胞,经流式细胞仪检测免疫磁珠分选后的CD133+造血祖细胞。采用五种方法扩增培养该细胞,8周后,再通过细胞形态学、流式细胞术、免疫细胞化学和免疫荧光对细胞进行干性鉴定,探索最佳干性维持培养方法。结果通过免疫磁珠法可以从人脐带血中分选出80%以上的CD133+造血祖细胞。采用优化的无血清培养基培养8周之后,CD133+造血祖细胞可得到有效扩增。而其他的培养基会使CD133+造血祖细胞由半悬浮细胞分化为梭形贴壁细胞,并且细胞状态欠佳。结论利用免疫磁珠法分选出的CD133+造血祖细胞,采用优化的无血清培养基能够有效扩增该细胞,并可长期有效的维持其干性。

Objective To isolate CD133＋ hematopoietic progenitor cells from human umbilical cord blood and optimize the culture condition for maintaining their stem cell characteristics.Methods CD133＋ hematopoietic progenitor cells were isolated from human umbilical cord blood using magnetic cell sorting system,and the cells were detected by flow cytometry.Four methods were used for culturing cells.After 8 weeks＇ culture,cytomorphology,flow cytometry,immunocytochemistry and immunofluorescence assay were used to identify the characteristics of the stem cells.Results Over 80% of CD133＋ hematopoietic progenitor cells were isolated from human umbilical cord blood using magnetic cell sorting system.The cells were effectively expanded using optimized serum-free medium after 8 weeks of cell culture,whereas the cells in other media differentiated into adherent cells in a poor state.Conclusion The optimized serum-free medium allows effective expansion of CD133＋ hematopoietic progenitor cells that maintain stem cell characteristics after a long-term culture.