Transcriptional profiling of human in vitro primed Th9 T cells (day 20) treated with GW9662 against untreated control. Transcriptional profiling of human in vitro primed Th9 T cells (day 20) treated with GW9662 against untreated control

To investigate the role of PPAR-γ in human TH cells, transcriptional response in vitro primed Th9 cells to treatment with GW9662, a potent PPAR-γ antagonist was assessed. In vitro primed Th9 cells (day 20) were incubated with GW9662 for 48h. Transcriptomic profiling was performed by bulk RNA-seq. To generate in vitro primed Th9 cells, human naive T cells were isolated from PBMCs using the EasySep™ Human naive CD4+ T Cell Isolation Kit (Stemcell Technologies) as per the manufacturer’s instructions. Naive T cells were stimulated with αCD3/CD2/CD28 beads (T cell/bead = 2:1, Miltenyi) and primed into effector CD4+ Th9 cells with IL-4 (50 ng/ml) and TGF-β (5 ng/ml) (R&D Systems). From cell culture initiation to analysis, the culture medium was supplemented with the indicated cytokines every other day. Cells were harvested for RNA sequencing (RNA-seq) after 20 days.