CD133-enriched CD34− (CD33/CD38/CD71)− cord blood cells acquire CD34 prior to cell division and hematopoietic activity is exclusively associated with CD34 expression

We compared the cell division behavior of CD34− and CD34+ (CD33/CD38/CD71)–negative (Lin− ) CD133+ cord blood cells stimulated with the cytokines Flt3-ligand, stem cell factor, and thrombopoietin. Within a 4-day time frame, Lin− CD34− CD133+ (CD34− ) cells underwent more cell divisions in serum-free culture than their Lin− CD34+ CD133+ (CD34+ ) counterparts. The majority of CD34− cells acquired expression of CD34 in vitro, including most undivided cells. Moreover, hematopoietic activity from both CD34− and CD34+ cells was exclusively retained within the cell fraction expressing CD34 after 4 days in culture. Most strikingly, in cultures from Lin− CD34− cells hematopoietic activity was associated with the fraction of divided cells, whereas in cultures of CD34+ cells, hematopoietic activity associated with the undivided cell fraction. Therefore, clonogenic CD34+ cells either do not divide or lose their clonogenic capacity upon cell division in vitro, while CD34− cells divide and retain this capacity under the same specific conditions. In conclusion, we demonstrate that CD133-enriched Lin− CD34− cord blood cells acquire CD34 prior to cell division and that long-term hematopoietic activity is associated exclusively with expression of CD34.