PKB inhibits migration of hematopoietic progenitors. CD34⁺ cells, cultured in presence of SCF and FLT-3L, were retrovirally transduced with myrPKB or eGFP alone. Three days after transduction, a transwell migration assay was performed. (A) The percentage of migrated cells was determined by flow cytometric analysis. Data were also depicted as fold induction compared with controls both in the (B) absence and (C) presence of SDF-1. (D) Hematopoietic progenitors were starved overnight in the absence of cytokines and in the presence of 0.5% FCS. Cells were left untreated (lanes 1 and 2) or incubated with 10μM PKB inhibitor VIII for 1 hour (lanes 3 and 4), before cells were stimulated with 0.5 μg/mL SCF for 15 minutes (lane 2 and 4). Protein lysates were made, and Western blot analysis was performed with an antibody against phosphorylated PKB and as a control for equal loading an antibody against tubulin. (E) Cells were cultured either in the absence or presence of the PKB inhibitor VIII for 48 hours, after which time a transwell migration assay was performed. Data were depicted as the percentage of migrated cells (n = 6). (F) Cells were cultured in the presence of the PKB inhibitor VIII for 2, 24, and 48 hours, after which time a transwell migration assay was performed. Data were depicted as the percentage of migrated cells. (G) Cells were cultured in the presence of the PKB inhibitor VIII for 24 and 48 hours, after which time a transwell migration assay was performed. The transwell inserts were preincubated for 1 hour with either fibronectin or fibrinogen. Data were depicted as the fold induction compared with untreated cells and uncoated inserts (n = 3). Error bars represent SEM. Samples significantly different (P < .05) are indicated with horizontal lines and asterisks.