Figure 4.

Interleukin-7 (IL-7) -mediated cell survival is abrogated by the Janus kinase/signal transducer and activator of transcription (JAK/STAT) inhibitor. (a) IL-7-mediated cell survival is reversed by the Pan-JAK inhibitor. Human effector/memory T cells were treated or not in the absence of cytokines with the Pan-JAK inhibitor (1 μm), the MEK-1 inhibitor U0126 (20 μm), or with the phosphatidylinositol 3-kinase (PI3K)/AKT inhibitor LY294002 (25 μm) for 1 hr. The cells were then stimulated or not with IL-7 (2 ng/ml) for 72 hr. Cells were collected and apoptosis was determined using Annexin V/7-aminoactinomycin D staining and flow cytometry analysis. The results are presented as mean percentages (± SE) of Annexin-V-positive cells from three independent experiments performed in triplicate with T cells isolated from different blood donors. *P < 0·05 between IL-7- or IL-7 + LY294002- or IL-7 + U0126-treated samples and non-treated samples (no stimulation) or IL-7 + Pan-JAK-treated samples. **P < 0·05 between LY294002-treated samples and non-treated or Pan-Jak- or U0126-treated samples. (b) IL-7-up-regulation of Bcl-2 proteins is abrogated by the Pan-JAK inhibitor. Cells were treated as described in (a) and the expression levels of Bcl-2 proteins were detected by immunoblot analysis. The results are representative of five independent experiments performed with T cells from different blood donors. (c) Densitometric quantification of Bcl-2 proteins in non-stimulated cells, versus cells stimulated with IL-7 and cells stimulated with IL-7 plus the different inhibitors as indicated. The results represent mean values (± SE) from three independent experiments and are expressed as the ratio between Mcl-1, Bcl-2 or Bcl-xL values and β-actin values. *P < 0·05 between IL-7- or IL-7 + LY294002- or IL-7 + U0126-treated samples and -non-treated samples (medium) or IL-7 + Pan-JAK-treated samples.