Figure 6. PDHK is required for Th17, but not Treg, function in vitro.

CD4⁺CD25^– T cells were polarized in vitro for 3 days, split 1:2, and cultured with IL-2 alone for an additional 2 days to generate Th1, Th17, or Tregs. (A) Schematic showing the different fates of pyruvate and mechanism of DCA inhibition and measurement of ¹⁴C-pyruvate oxidation in Th17 cells and Tregs. (B) Real-time PCR and (C) immunoblots are shown, representative of 4 independent experiments. (D–F) Cells were treated with 10 mM DCA, and then cytokine production (D) and transcription factor staining (E) were determined by flow cytometry. (F) Treg function was assessed by an in vitro Treg-suppression assay. (G and H) T cells were polarized and infected with lentivirus expressing PDHK1 shRNA. (G) transcription factor staining for FoxP3 and RORγT or (H) intracellular cytokine staining for IL-17 was performed. Data are shown as mean ± SD of triplicate samples (A, B, and H), and all data are representative of at least 3 independent experiments. *P < 0.05.