Manipulation of SK-1 levels alters ECU formation. ECU numbers were determined from: (A) BM cells from SK-1-KO mice infected with adenovirus containing either control EV (SK-1-KO^EV) or SK-1 (SK-1-KO^SK); (B) WT and SK-1-Tg BM cells; (C) WT BM cells cultured with the SK inhibitor SKi (5 μM) administered at initial seeding and every 48 hours thereafter; and (D) WT and SK-1-KO BM cells cultured without (□ and with FTY720 (100 nM, ■) administered at initial seeding and every 48 hours thereafter. All experiments show the mean plus or minus SEM of 4-day cultured ECU from at least 3 separate experiments. SK-1 levels shown numerically leading the figure (activity for panels A-C and protein for panel D) identified successful SK-1 manipulation. Results are expressed as U/mg protein normalized to WT BM cells at day 4 of culture plus or minus SEM. *P < .05, compared with controls using a 2-tailed, paired t test. (E) Six-day cultured WT BM cells plus or minus FTY720 (100 nM) administered at initial seeding and every 48 hours thereafter, seeded into Matrigel. Images show tube formation; numbers reflect mean plus or minus SEM tubes per experiment; identify tubes formed at 30 hours. Results are representatives of 3 separate experiments.