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. 2015 Mar 26;6:6692. doi: 10.1038/ncomms7692

Figure 6. Inhibition of PI3K/Akt and MEK/Erk pathways is involved in the metabolic reprogramming of activated T cells.

Figure 6

(a) CD4+ primary human T cells were cultured with tosyl-activated magnetic beads conjugated with aCD3/aCD28/IgG (TCC) in the presence of either LY294002 (LY, 10 μM), UO126 (UO, 10 μM) or their combination. Cell lysates were prepared at the indicated time points and expression of CPT1A and β-actin was assessed by SDS–PAGE and immunoblot. Results are representative of three experiments. (b–d) In parallel experiments, rate of fatty acid β-oxidation was examined. Values of TCC+inhibitor cultures were compared with TCC (*P<0.05, Student’s t-test; n=3) and values of TCC and TCC+inhibitor cultures were compared with unstimulated (UT) (♦P<0.05, analysis of variance (ANOVA); n=3). (e) At 72 h of culture under the indicated conditions, spare respiratory capacity (SRC) was determined. Values of TCC and TCC+inhibitor cells were compared with unstimulated (UT) cells (♦P<0.05, ANOVA; n=3) and values in TCC+inhibitor cells were compared with TCC (*P<0.05, Student’s t-test; n=3). (f) T cells were cultured with tosyl-activated magnetic beads conjugated with aCD3/aCD28/PD-L1-Ig or with tosylatcivated magnetic beads conjugated with aCD3/aCD28/IgG in the presence of either LY294002 (LY, 10 μM), UO126 (UO, 10 μM) or their combination. Cell lysates were prepared at the indicated time points and expression of ATGL and β-actin was assessed by SDS–PAGE and immunoblot.