Proliferation and apoptosis of RhoH-transduced progenitor cells. LDBM cells were transduced with MIEG3-HA-RhoH and vector control. The EGFP⁺, c-Kit⁺ cells were sorted by FACS and used for the following assays. (A) Proliferation. Cells were starved in 1% serum-containing medium for 8 hours and stimulated with 100 ng/mL SCF for 48 hours. [³H]-thymidine was incorporated for 6 hours at 37°C. Data represent the mean ± SD; n = 6. (B) Apoptosis. Cells were stained with Annexin V-APC and 7-AAD after 100 ng/mL SCF stimulation for 48 hours. Data represent the mean ± SD; n = 3. *P < .01, RhoH-versus vector-transduced cells. These results shown in panels A and B are representatives of a minimum of 3 experiments. (C-D) In vivo apoptosis and proliferation analyses of RhoH-transduced BM cells from the representative recipient mice after 5-FU treatment. At 4 months after transplantation, the recipient mice of vector- or RhoH-transduced cells were injected intraperitoneally with 5-FU, and 48 hours later these mice were fed with 1 mg/mL BrdU for 36 hours. LDBM cells were harvested from each mouse and stained with APC-conjugated antibody to c-Kit receptor and PE-conjugated antibodies to BrdU or Annexin V. EGFP⁺, c-Kit⁺, Annexin V⁺ (C) or EGFP⁺, c-Kit⁺, BrdU⁺ (D) cells were analyzed by flow cytometry. Results shown in the top panels of panels C and D are gated on EGFP⁺ cells from a representative mouse. The lower panels show the summary data on apoptosis (C) and proliferation (D). Data represent the mean ± SD; n = 8 mice per group. *P < .01, RhoH-versus vector-transduced cells.