Oxidized Phospholipid oxPAPC Alters Regulatory T-Cell Differentiation and Decreases Their Protective Function in Atherosclerosis in Mice

Regulatory T cells (T ) are protective in atherosclerosis but reduced during disease progression due to cell death and loss of stability. However, the mechanisms of T dysfunction remain unknown. Oxidized phospholipids are abundant in atherosclerosis and can activate innate immune cells, but little is known regarding their impact on T cells. Given T loss during atherosclerosis progression and oxidized phospholipid levels in the plaque microenvironment, we investigated whether oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (oxPAPC), an oxidized phospholipid associated with atherosclerotic plaques, alters T differentiation and function. CD4 T cells were polarized to T , T helper (Th) 1, and Th17 cells with or without oxPAPC and assessed by flow cytometry. Gene expression in oxPAPC-treated T was analyzed by bulk RNA sequencing. Functional studies of oxPAPC-induced T were performed by coculturing T with CellTrace Violet-labeled cells in vitro, and by adoptively transferring T to hyperlipidemic mice to measure atherosclerosis progression. Compared with controls, oxPAPC-treated T were less viable, but surviving cells expressed higher levels of the Th1-associated markers T-bet, CXCR3, and IFN (interferon)-γ. Th1 and Th17 skewing cultures were unaltered by oxPAPC. IFN-γ is linked to T instability, thus T polarization experiments were repeated using CD4 T cells. IFNγR1 (INF gamma receptor 1) deficiency did not improve cell viability in oxPAPC-treated T ; however, T-bet and IFN-γ expression was not increased in surviving cells suggesting a role for IFN-γsignaling. OxPAPC-treated T were less suppressive in vitro, and adoptive transfer studies in hyperlipidemic mice showed that oxPAPC-induced T possessed altered tissue homing and were insufficient to inhibit atherosclerosis progression. OxPAPC elicits T -specific changes altering T differentiation and inducing a Th1-like phenotype in surviving cells partially through IFN-γ signaling. This is biologically relevant as oxPAPC-treated T do not reduce atherosclerosis progression in mice. This study supports the role of oxidized phospholipids in negatively impacting T differentiation and atheroprotective function.