SDF-1 protects PB CD34⁺ cells from apoptosis through the PI3-K pathway.

PB CD34⁺ cells purified immediately after density gradient separation (Inc⁻, ░) or after incubation on a plastic support (Inc⁺, ■) were incubated (1 × 10⁵cells/mL) for 24 hours under apoptosis-inducing culture conditions (serum- and cytokine-free medium [IMDM]) in presence or absence of SDF-1 (0.05 and 0.5 ng/mL) and processed for apoptosis assessment by using the annexin-V apoptosis detection kit as indicated in “Materials and methods.” The role of endogenous SDF-1 was evaluated by adding an anti–SDF-1 neutralizing antibody (10 ng/mL) to the culture. Involvement of PI3-K/AKT and MAP kinase pathways was evaluated by using LY294002- and PD98059-specific inhibitors at concentrations varying from 10 to 100 μM. Data from 3 to 5 independent experiments are presented as mean percentage of annexin-V⁺ cells ± SD. The antiapoptotic effect of exogenous SDF-1 is shown in conditions ➁/➀. The role of PI3-K in the autonomous survival of CD34⁺ cells is shown in conditions ③/➁. Requirement of PI3-K axis for the antiapoptotic effect of exogenous SDF-1 is shown in conditions ➀/④. The antiapoptotic effect of endogenous SDF-1 produced by Inc⁻cells is shown in conditions ➁/⑤. Requirement of PI3-K axis for the antiapoptotic effect of endogenous SDF-1 is shown in conditions ⑥/③. No requirement of MAP kinase pathway in the autonomous survival of CD34⁺ cells and in the antiapoptotic effect of SDF-1 is shown in conditions ⑦/➁, and ⑧/⑨, ➀/⑤, respectively.