GSE162725
HLADR expression marks recently divided memory CD4 T cells in latent tuberculosis infected subjects upon in vitro stimulation with Mycobacterium tuberculosis antigens
Upon antigen encounter, T cells can rapidly divide and form short-lived effector cells which plays an important role fighting an active infection. However, little is known about the molecular markers that distinguish such effector cells from other T cell populations in humans. Due to previous exposure with Mycobacterium tuberculosis (Mtb), we expect latent tuberculosis infected (LTBI) subjects to have Mtb-specific memory CD4 T cells that are capable of rapid expansion upon antigen re-encounter. To investigate this further we designed an in vitro MTP-AE (Memory T cell Proliferation upon Antigen Exposure) assay to track and isolate proliferated CD4 T cells in response to antigen stimulation. PBMCs from LTBI subjects were cultured with the MTB300 (Mtb specific peptide pool, PMID: 27409590) for 8 days before sorting for HLADR+ vs HLADR- and divided vs undivided CD4 T cells. We found that HLADR is a marker of recently divided memory CD4 T cells and that differentially expessed genes in HLADR+/recently divided cells were enriched for cytotoxic markers and cytokines compared to HLADR-/undivided cells. Overall design: Bulk RNA-sequencing of sorted memory CD4 T cells based on HLADR expression (HLADR+/HLADR-) or based on Cell Trace Violet staining (undivided (Div0), cells that have undergone one division (Div1) and cells that have undergone at least two divisions (Div2)) after 8 days post in vitro stimulation of PBMCs from latent TB donors with DMSO, MTB300 (Mtb-specific peptide pool,PMID: 27409590)  or anti-CD3/CD28. Additionally, on D0 (ex vivo) and D1 (24 hrs post stimulation) memory CD4 T cells were directly sorted for bulk RNA-sequencing