In vivo cycling up-regulates Fas expression on Lin^-CD34⁺ CB cells. Freshly isolated CD34⁺ CB cells were transplanted into irradiated NOD-SCID mice that subsequently received 3 injections of PBS (-TNF) or 5 μg TNF (+TNF) 24, 64, and 96 hours after transplantation. (A) At 24 hours after the last injection (day 5), mice were killed, and the BM was analyzed for human (HuCD45/71) engraftment. Samples were also stained with murine CD45.1 (MuCD45.1) to distinguish murine and human cells. Cells positive for HuCD45/71 were analyzed for coexpression of HuCD34. Results shown are from 1 of 2 experiments. (B) Engrafted human cells coexpressing CD34 and lacking expression of lineage antigens (Lin^-CD34⁺) were gated and investigated for Fas expression. Broken lines represent engrafted Lin^-CD34⁺ cells stained with isotype control Ab; solid lines, Fas expression on Lin^-CD34⁺ cells from mice injected with PBS; and filled histograms, Fas expression on Lin^-CD34⁺ cells from mice treated with TNF. Results from 2 independent experiments are shown.