GARP and CTLA4 identify a potently suppressive antigen specific Treg population in demyelinating autoimmunity 3183

Description In demyelinating disease, Tregs with the most effective suppressive capacity are self-antigen specific; however, it is unclear how the parameters of self-antigen stimulation (affinity, force, and bond lifetime) modulate suppressive potency. A critical unmet need limiting these studies is the ability to isolate antigen specific Tregs without prior knowledge of TCR reactivity. Here, we utilize the well-described myelin oligodendrocyte glycoprotein (MOG) 35-55 peptide driven experimental autoimmune encephalomyelitis (EAE) in B6 mice and have identified GARP and CTLA4 as markers that predict Treg antigen reactivity. In this system, we show that tetramer staining in the spleen is increased in enriched GARP+ CTL4+ Tregs (30%) compared to bulk Tregs (10%). We found that GARP+ CTLA4+ Tregs are more potent antigen specific suppressors by in vitro suppression assay using 2D2 responders stimulated with MOG. We compared bond lifetime and found that peak bond lifetime shifted to higher force in CTLA4+ GARP+ Tregs. We hypothesized that trogocytosis may be a force dependent mechanism of suppression by removal of self-antigen and we found that CTLA4+ GARP+ Tregs pulled more MOG loaded MHCII from the surface of antigen presenting cells than CTLA4- Tregs. This research uncovers an understanding of surface markers that will identify potently suppressive antigen specific Tregs, which will allow the design of therapeutic strategies for Tregs in autoimmune conditions.