title: Atherosclerosis-Driven Treg Plasticity Results in Formation of a Dysfunctional Subset of Plastic IFNγ+ Th1/Tregs.
abstract: BACKGROUND
  Forkhead box P3+T regulatory cells (Tregs) are key players in maintaining immune homeostasis. Evidence suggests that Tregs respond to environmental cues to permit or suppress inflammation. In atherosclerosis, Th1-driven inflammation affects Treg homeostasis, but the mechanisms governing this phenomenon are unclear.
  
  
  OBJECTIVE
  Here, we address whether atherosclerosis impacts Treg plasticity and functionality in Apoe-/-mice, and what effect Treg plasticity might have on the pathology of atherosclerosis.
  
  
  RESULTS
  We demonstrate that atherosclerosis promotes Treg plasticity, resulting in the reduction of CXCR3+Tregs and the accumulation of an intermediate Th1-like interferon (IFN)-γ+CCR5+Treg subset (Th1/Tregs) within the aorta. Importantly, Th1/Tregs arise in atherosclerosis from bona fide Tregs, rather than from T-effector cells. We show that Th1/Tregs recovered from atherosclerotic mice are dysfunctional in suppression assays. Using an adoptive transfer system and plasticity-prone Mir146a-/-Tregs, we demonstrate that elevated IFNγ+Mir146a-/-Th1/Tregs are unable to adequately reduce atherosclerosis, arterial Th1, or macrophage content within Apoe-/-mice, in comparison to Mir146a+/+Tregs. Finally, via single-cell RNA-sequencing and real-time -polymerase chain reaction, we show that Th1/Tregs possess a unique transcriptional phenotype characterized by coexpression of Treg and Th1 lineage genes and a downregulation of Treg-related genes, including Ikzf2, Ikzf4, Tigit, Lilrb4, and Il10. In addition, an ingenuity pathway analysis further implicates IFNγ, IFNα, interleukin-2, interleukin-7, CTLA-4 (cytotoxic T-lymphocyte-associated protein 4), T-cell receptor, and Csnk2b-related pathways in regulating Treg plasticity.
  
  
  CONCLUSIONS
  Atherosclerosis drives Treg plasticity, resulting in the accumulation of dysfunctional IFNγ+Th1/Tregs that may permit further arterial inflammation and atherogenesis.
authors.affiliation: From the Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk (M.J.B., A.R.F., T.C.W., C.M.M., K.J.C., P.S.L., E.V.G.); and Department of Molecular and Cellular Biology, Beckman Research Institute, City of Hope, Duarte, CA (N.M., M.B.).
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Butcher
authors.affiliation: From the Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk (M.J.B., A.R.F., T.C.W., C.M.M., K.J.C., P.S.L., E.V.G.); and Department of Molecular and Cellular Biology, Beckman Research Institute, City of Hope, Duarte, CA (N.M., M.B.).
authors.forename: Adam R
authors.initials: AR
authors.lastname: Filipowicz
authors.affiliation: From the Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk (M.J.B., A.R.F., T.C.W., C.M.M., K.J.C., P.S.L., E.V.G.); and Department of Molecular and Cellular Biology, Beckman Research Institute, City of Hope, Duarte, CA (N.M., M.B.).
authors.forename: Tayab C
authors.initials: TC
authors.lastname: Waseem
authors.affiliation: From the Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk (M.J.B., A.R.F., T.C.W., C.M.M., K.J.C., P.S.L., E.V.G.); and Department of Molecular and Cellular Biology, Beckman Research Institute, City of Hope, Duarte, CA (N.M., M.B.).
authors.forename: Christopher M
authors.initials: CM
authors.lastname: McGary
authors.affiliation: From the Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk (M.J.B., A.R.F., T.C.W., C.M.M., K.J.C., P.S.L., E.V.G.); and Department of Molecular and Cellular Biology, Beckman Research Institute, City of Hope, Duarte, CA (N.M., M.B.).
authors.forename: Kevin J
authors.initials: KJ
authors.lastname: Crow
authors.affiliation: From the Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk (M.J.B., A.R.F., T.C.W., C.M.M., K.J.C., P.S.L., E.V.G.); and Department of Molecular and Cellular Biology, Beckman Research Institute, City of Hope, Duarte, CA (N.M., M.B.).
authors.forename: Nathaniel
authors.initials: N
authors.lastname: Magilnick
authors.affiliation: From the Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk (M.J.B., A.R.F., T.C.W., C.M.M., K.J.C., P.S.L., E.V.G.); and Department of Molecular and Cellular Biology, Beckman Research Institute, City of Hope, Duarte, CA (N.M., M.B.).
authors.forename: Mark
authors.initials: M
authors.lastname: Boldin
authors.affiliation: From the Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk (M.J.B., A.R.F., T.C.W., C.M.M., K.J.C., P.S.L., E.V.G.); and Department of Molecular and Cellular Biology, Beckman Research Institute, City of Hope, Duarte, CA (N.M., M.B.).
authors.forename: Patric S
authors.initials: PS
authors.lastname: Lundberg
authors.affiliation: From the Department of Microbiology and Molecular Cell Biology, Eastern Virginia Medical School, Norfolk (M.J.B., A.R.F., T.C.W., C.M.M., K.J.C., P.S.L., E.V.G.); and Department of Molecular and Cellular Biology, Beckman Research Institute, City of Hope, Duarte, CA (N.M., M.B.). galkinev@evms.edu.
authors.forename: Elena V
authors.initials: EV
authors.lastname: Galkina
mesh_terms: D000818:Animals; D050197:Atherosclerosis / Q000276:immunology / Q000378:metabolism*; D000066670:Cell Plasticity / Q000502:physiology*; D007371:Interferon-gamma / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D016176:T-Lymphocyte Subsets / Q000276:immunology / Q000378:metabolism; D050378:T-Lymphocytes, Regulatory / Q000276:immunology / Q000378:metabolism*; D018417:Th1 Cells / Q000276:immunology / Q000378:metabolism*
chemical_list: D007371:Interferon-gamma
keywords: Treg cells; atherosclerosis; cell plasticity; immune system; immunology; inflammation; lymphocytes