title: Characterization of protective human CD4CD25 FOXP3 regulatory T cells generated with IL-2, TGF-β and retinoic acid.
abstract: BACKGROUND
  Protective CD4+CD25+ regulatory T cells bearing the Forkhead Foxp3 transcription factor can now be divided into three subsets: Endogenous thymus-derived cells, those induced in the periphery, and another subset induced ex-vivo with pharmacological amounts of IL-2 and TGF-β. Unfortunately, endogenous CD4+CD25+ regulatory T cells are unstable and can be converted to effector cells by pro-inflammatory cytokines. Although protective Foxp3+CD4+CD25+ cells resistant to proinflammatory cytokines have been generated in mice, in humans this result has been elusive. Our objective, therefore, was to induce human naïve CD4+ cells to become stable, functional CD25+ Foxp3+ regulatory cells that were also resistant to the inhibitory effects of proinflammatory cytokines.
  
  
  RESULTS
  The addition of the vitamin A metabolite, all-trans retinoic acid (atRA) to human naïve CD4+ cells suboptimally activated with IL-2 and TGF-β enhanced and stabilized FOXP3 expression, and accelerated their maturation to protective regulatory T cells. AtRA, by itself, accelerated conversion of naïve to mature cells but did not induce FOXP3 or suppressive activity. The combination of atRA and TGF-β enabled CD4+CD45RA+ cells to express a phenotype and trafficking receptors similar to natural Tregs. AtRA/TGF-β-induced CD4+ regs were anergic and low producers of IL-2. They had potent in vitro suppressive activity and protected immunodeficient mice from a human-anti-mouse GVHD as well as expanded endogenous Tregs. However, treatment of endogenous Tregs with IL-1β and IL-6 decreased FOXP3 expression and diminished their protective effects in vivo while atRA-induced iTregs were resistant to these inhibitory effects.
  
  
  CONCLUSIONS
  We have developed a methodology that induces human CD4(+) cells to rapidly become stable, fully functional suppressor cells that are also resistant to proinflammatory cytokines. This methodology offers a practical novel strategy to treat human autoimmune diseases and prevent allograft rejection without the use of agents that kill cells or interfere with signaling pathways.
authors.affiliation: Division of Rheumatology, Department of Medicine, Keck School of Medicine at University of Southern California, Los Angeles, California, United States of America.
authors.forename: Ling
authors.initials: L
authors.lastname: Lu
authors.forename: Xiaohui
authors.initials: X
authors.lastname: Zhou
authors.forename: Julie
authors.initials: J
authors.lastname: Wang
authors.forename: Song Guo
authors.initials: SG
authors.lastname: Zheng
authors.forename: David A
authors.initials: DA
authors.lastname: Horwitz
mesh_terms: D000818:Animals; D015496:CD4-Positive T-Lymphocytes / Q000166:cytology*; D051858:Forkhead Transcription Factors / Q000096:biosynthesis* / Q000378:metabolism*; D006801:Humans; D007249:Inflammation; D007376:Interleukin-2 / Q000378:metabolism*; D053645:Interleukin-2 Receptor alpha Subunit / Q000096:biosynthesis*; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D010641:Phenotype; D050378:T-Lymphocytes, Regulatory / Q000166:cytology*; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D014212:Tretinoin / Q000378:metabolism*
chemical_list: C418974:FOXP3 protein, human; D051858:Forkhead Transcription Factors; D007376:Interleukin-2; D053645:Interleukin-2 Receptor alpha Subunit; D016212:Transforming Growth Factor beta; D014212:Tretinoin