Karyotyping, immunophenotyping, and apoptosis analyses on human hematopoietic precursor cells derived from umbilical cord blood following long-term ex vivo expansion

By means of flow cytometry, CD34+/CD38− hematopoietic stem cells (HSC) were collected from umbilical cord blood (UCB) of 10 healthy women at the time of delivery and cultivated in stem-cell culture media supplemented with cell growth stimulating factors (IL-3, IL-6, GM-CSF, EPO, IGF-1, and SCF) for long periods. Apoptotic status, cell surface marker expression, and karyotypes of the cultured UCB-derived CD34+/CD38− stem-cells were investigated by flow cytometry and GTG-banding methods. The UCB-derived CD34+/CD38− stem-cells were able to divide and proliferate in vitro for at least 6 months. They did not show significantly increased apoptosis following ex vivo expansion for 20 and 32 days, respectively, in 2 cases and retained the same cell surface marker expression pattern (i.e., CD34+ and CD38−) in the majority of the cells of 2 cases following 20 and 37 days of incubation, respectively. In another 2 cases, chromosome analysis showed no evidence of numerical and structural abnormalities in the CD34+/CD38− stem-cells obtained after 20 and 43 days in culture, respectively. Our findings indicated that UCB-derived CD34+/CD38− stemcells are able to maintain their basic biologic and genetic characteristics after dividing and proliferating in vitro for a long period of time. UCB-derived HSC following ex vivo expansion can serve as a reliable resource for hematopoietic precursor cells transplantation.