Effects of shN-cads on HSPC cell division in vitro. (A) Cell division assay experimental scheme. LSK cells were cotransduced with lentiviruses expressing shN-cad-1 or -scramble and retroviruses expressing control-mKO or N-cad with a silencing mutation in the shRNA targeting sequence (sil.mut.N-cad). Gene-transduced LSK cells were clone-sorted onto control- or N-cad-Fc–coated plates and cultured in SF-O3 medium with 1.0% BSA, SCF, and TPO. The number of cells in each colony on day 7 (shown in B) and the days required for a single cell to reach more than 30 cells (shown in panel C) were analyzed. (B) The number of cells in each colony on control- and N-cad-Fc–coated plates at day 7 of culture. *P < .05. (C) The percentage of colonies with more than 30 cells per colony on each day of culture are shown. Transduction of N-cad shRNA into LSK cells reduced the number of days required for a single cell to reach 30 cells or more, whereas sil.mut.N-cad rescued the accelerated cell division induced by shN-cad-1. (D) Hoechst staining of GFP⁺ LSK cells 2 weeks after BMT. (E) The percentage of GFP⁺LSK cells in G₀/G₁. Data are mean ± SD. **P < .05.