Oxidized phospholipid oxPAPC induces a Th1-like phenotype in regulatory T cells and inhibits their protective function in atherosclerosis

Background Regulatory T cells (T regs ) are protective in atherosclerosis but reduced during disease progression due to cell death and loss of stability. However, the mechanisms of T reg dysfunction remain unknown. Oxidized phospholipids (oxPLs) are abundant in atherosclerosis and can activate innate immune cells, but there is limited information regarding their impact on T cells. Given T reg loss during atherosclerosis progression and oxPL levels in the plaque microenvironment, we sought to determine whether oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (oxPAPC), an oxPL associated with atherosclerotic plaques, alters T reg differentiation and function. Methods Naïve CD4 + T cells were cultured under T reg, Th1, and Th17 polarizing conditions with or without oxPAPC and assessed by flow cytometry. Gene expression in oxPAPC-treated T regs was analyzed by bulk RNA sequencing. Functional studies of oxPAPC-induced T regs were performed by co-culturing T regs with CTV-labeled CD8 + cells in vitro. In vivo suppression of atherosclerosis was evaluated by adoptively transferring control or oxPAPC-treated T regs to hyperlipidemic Ldlr -/- mice. Results Compared to controls, oxPAPC-treated T regs were less viable but expressed higher levels of the Th1-associated markers T-bet, CXCR3, and IFN-γ. Th1 and Th17 skewing cultures were unaltered by oxPAPC. IFN-γ is linked to T reg instability, thus T reg polarization experiments were repeated using Ifngr1 -/- CD4 + T cells. IFNγR1 deficiency did not improve cell viability in oxPAPC-treated T regs , however, T-bet and IFN-γ expression was not increased suggesting a role for IFN-γ signaling. OxPAPC-treated T regs were less suppressive in vitro , and adoptive transfer studies in hyperlipidemic Ldlr -/- mice showed that oxPAPC-induced T regs possessed altered tissue homing and were insufficient to inhibit atherosclerosis progression. Conclusions OxPAPC elicits T reg -specific changes that induce a Th1-like phenotype dependent on IFN-γ signaling. This is biologically relevant as oxPAPC-treated T regs are unable to reduce atherosclerosis progression in Ldlr -/- mice. This study supports a role for oxPLs in negatively impacting T reg differentiation and atheroprotective function.