Ex vivo expansion of human CD34⁺ PBSCs cultured in serum-free media in the presence of TPO, SCF, IL-3, IL-6, IL-11, and Fl3 and platelet production in NOD/SCID mice.

Human CD34⁺ PBSCs were transplanted before (day 0) and after 1, 3, 4, and 8 days of in vitro stimulation with TPO, SCF, IL-3, IL-6, IL-11, and Fl3. Mice were serially bled at weeks 2, 3, and 4 after transplantation. The transplant was performed without exogenous cytokines. BM and spleen cells were analyzed for human engraftment at week 4. (A) Uncultured (day 0) or cultured (days 3-8) human CD34⁺ PBSCs were analyzed using PE–anti-CD34 and FITC–anti-CD41. Isotype controls for each time point are shown in the upper panel. CD34/CD41 phenotypes are shown in the lower panel. The percentage of each subpopulation is indicated. (B) Human platelet production in 4 mice transplanted with human CD34⁺ PBSCs prior to expansion (day 0). Evidence of human engraftment at week 4 was only detected in mice 1 and 3. (C) Human platelet production in mice that received human CD34⁺ PBSCs that had been expanded for 1 (black), 3 (dotted), 4 (striped), or 8 (clear) days. Each column represents an individual mouse. No human platelets were detected at weeks 2 or 4.