Cell cycle analysis of fresh, ex vivo–cultured and BM- and spleen-homed MPB and BM CD34⁺ cells recovered from control and cytokine-treated NOD/SCID recipients.

CD34⁺ cells were isolated from fresh MPB or BM and stained with CFSE as described in “Materials and methods.” A sample was retained for cell cycle analysis at time zero and for initiating short-term ex vivo cultures maintained as described in the legend of Figure 2. Remaining cells were transplanted into control and cytokine-treated, conditioned NOD/SCID mice. Cytokine treatment of recipient mice consisted of 4 daily intraperitoneal injections on days −2, −1, 0 (day of transplantation), and +1 of a cytokine cocktail delivering per mouse 10 μg SCF, 5 μg Flt-3 ligand, 5 μg MGDF, 6 μg IL-3, 2 μg IL-6, 6 μg GM-CSF, and 10 U erythropoietin. At 24 and 48 hours AT, BM and spleen (Spl) cells were recovered from individual mice, and human CFSE⁺ cells were isolated by flow cytometric cell sorting. Cell cycle status of all groups of cells was determined by PI staining. Each bar represents the mean of 1 to 3 measurements of MPB or BM samples at the 24-hour time point and 2 to 3 measurements at the 48-hour time point from 3 independent experiments. Statistical analysis of measurements made at 48 hours between control and cytokine-treated recipients was not significant (P > .2).