Fas inactivation can partially prevent the depletion of Tak1^−/−HSPCs in vitro but not in vivo. (A-B) c-kit⁺ HSPCs were isolated from Tak1^fx/fxTnfr1^−/−r2^−/− mice. (A) These cells were infected with MSCV-Cre-GFP virus to induce Tak1 deletion and cultured in HSPC medium with or without inflammatory cytokine treatment. MSCV-GFP infection was studied in parallel as a control. The ratios of GFP⁺ cells (normalized to MSCV-GFP infection) were compared among different treatment groups. **Significant reduction compared with vehicle-treated group (P < .01). (B) These infected cells were purified by FACS and cultured in HSPC medium with or without IFN-γ treatment. Cell death was examined by annexin V staining. Data shown in this figure are representative of 3 independent experiments. (C-D) c-kit⁺ HSPCs were isolated from Tak1^fx/fx and Tak1^fx/fxFas^−/− mice. These cells were infected with MSCV-Cre-GFP virus to induce Tak1 deletion and cultured in HSPC medium to examine dynamic changes in the ratios of infected (GFP⁺) cells (C) or seeded for CFU assay (D). MSCV-GFP infections were studied in parallel as a control. Normalized ratios of GFP⁺ cells were compared between Tak1^fx/fx and Tak1^fx/fxFas^−/− HSPCs. **Significant increase compared with Tak1^fx/fx group (P < .01). (E-J) Tak1^−/−Fas^−/− mice were injected with poly I:C every other day for a total 3 injections; BM and spleens were collected 8 days hence. WT and Tak1^−/− mice were treated and studied in parallel as controls. BM histology (20 × 0.7 air; bar represents 100 μm in length; E), MNCs in BM (F), MNCs in spleen (G), percentages (H), and absolute numbers (I) of HSCs and CPs in BM were compared among WT, Tak1^−/−, and Tak1^−/−Fas^−/− groups. J. As described in Figure 4, BM MNCs from Tak1^−/−Fas^−/− (CD45.2⁺) mice (before Tak1 deletion was induced) were mixed with WT (CD45.1⁺) competitor cells (10:1 ratio) and transplanted into lethally-irradiated recipient mice (CD45.1⁺). After examining the percentages of donor-derived cells in PB of recipient mice, these mice were injected with poly I:C every other day for a total of 3 injections to induce Tak1 deletion. BM MNCs from WT and Tak1^−/− mice were transplanted and studied in parallel as controls. The percentages of donor-derived cells in PB of the recipients were examined 10 and 20 days later. (F-J) **Significant reduction compared with WT control (P < .01). (K-L) c-kit⁺ HSPCs were collected from Tak1^fx/fx Tnfr1^−/−r2^−/−, Tak1^fx/fxTnfr1^−/−r2^−/−Fas^−/−, and Tak1^fx/fx mice and infected with MSCV-Cre-GFP to induce Tak1 deletion. Infected cells were incubated in either HSPC medium to examine the dynamic changes in ratios of infected cells (K), or sorted for CFU assay (L). MSCV-GFP–infected Tak1^fx/fx HSPCs were studied in parallel as controls. (K-L) **Significant increase compared with Tak1^fx/fx group (P < .01).