We have recently shown by in vitra cell tracking with the fluorescent membrane dye, PKH2, that apoptosis among dividing CD34+ cells from human BM, CB, and MPB Increases In association with the number of in vitro cellular divisions and correlates with a loss in hematopoietic potential in cultured CD34+ cells. To investigate mechanisms behind this increase in apoptosis, we examined the potential roles of FAS (CD95) and BCL2 in mediating apoptosis in dividing CD34+ cells. CD34+ cells from normal human BM, CB, and MPB were stained with PKH2 and cultured for 7 days with SCF, IL-3, and IL-6. and then analyzed for FAS or BCL2 expression In various fractions of expanded CD34+ cells. While CD34+ cells from all 3 tissues expressed low FAS on do, this percentage gradually increased among CD34+ cells during short-term culture such that by d7 40-80% of CD34+ cells were CD95+. Conversely, the percentage of CD34+ cells expressing BCL2 decreased from 90-100% on 00 to <50% by d7. When CD34+ cells were analyzed based on the number of in vitro cellular divisions, it was found that the percentage of CD95+ cells increased concomitantly with the number of In vitro cellular divisions, suggesting that expression of FAS may play a role in the proliferation-associated apoptosis observed in cultured CD34+ cells. This supposition was further substantiated when d7 CD34+/PKH2B" [cylokine non-responsive (CNR)] cells were fractionated based on their expression of CD95 and examined in long-term cultures (LTC). CNR cells lacking CD95 (CNR/CD95-) produced 3-fold more cells and 17-fold more progenitors by d25 in LTC than CNR/CD95+ celb. Furthermore, a greater percentage of CNR/CD95cells expressed BCL2 at d25 and with greater fluorescence intensity than CNR/CD95+ cells (68% @channel 281 vs. 56% @channel 185, respectively) Taken together, these results suggest that FAS-mediated apoptosis may play a role in the diminished hematopoietic potential observed among proliferating human BM, CB, and MPB CD34+ cells, and that this effect may be in part mediated by decreased expression of BCL2.
|Original language||English (US)|
|Number of pages||1|
|State||Published - Dec 1 1996|
ASJC Scopus subject areas
- Molecular Biology
- Cell Biology
- Cancer Research