Figure 8: PD-1 alters the metabolic programme of activated T cells from glycolysis to FAO.
CD4+ human T cells were pre-activated with anti-CD3-and-anti-CD28 mAbs for 4 h or for 24 h and subsequently were collected, were left to rest for 3 h and re-cultured with tosyl-activated magnetic beads conjugated with αCD3/αCD28/PD-L1-Ig (Tpr4hr→PD-1 and Tpr24hr→PD-1). In the same experiment CD4+ T cells without pre-activation were stimulated with tosyl-activated magnetic beads conjugated with αCD3/αCD28/IgG (TCC) and used as positive control for optimal stimulation without PD-1. (a) Expression of Glut1 after culture under the indicated conditions and time intervals was examined by flow cytometry. (b,c) mRNA for HK2 and CPT1A was analysed by real-time quantitative PCR and relative expression of mRNA of each time point and culture condition over the levels expressed in unstimulated cells (defined as 1) is shown. (d) Fatty acid β-oxidation rate after culture under the indicated conditions for various time intervals was examined. (e,f) Analysis of lactate (e) and 3-hydroxybutyrate (f), end metabolites of glycolysis and fatty acid β-oxidation, respectively, was performed in culture supernatants (for the studies shown in all panels: *P<0.05, Tpr4hr→PD-1 versus TCC or Tpr4hr→PD-1 versus TCC, Student’s t-test; ♦P<0.05, TCC versus UT, Tpr4hr→PD-1 versus UT and Tpr4hr→PD-1 versus UT, analysis of variance; n=3 experiments).
