title: Dichotomous response to transforming growth factor β after T cell receptor activation by naive CD4+ T cells from DBA/1 mice: enhanced retinoic acid receptor-related orphan nuclear receptor γt expression yet reduced FoxP3 expression.
abstract: OBJECTIVE
  To investigate the molecular mechanism for biased interleukin-17 (IL-17) production by DBA/1 CD4+ T cells upon T cell receptor (TCR) and transforming growth factor β (TGFβ) stimulation.
  
  
  METHODS
  Purified naive CD4+ T cells were stimulated with anti-CD3/CD28 under Th1, Th2, Th17, and induced T regulatory (iTreg) cell conditions. Cytokine production was assayed by intracellular staining and enzyme-linked immunosorbent assay. Expression of transcription factors was determined by reverse transcription-polymerase chain reaction, flow cytometry, and immunoblotting techniques.
  
  
  RESULTS
  Naive CD4+ T cells from DBA/1 mice produced more IL-17 under Th17 cell polarizing conditions as compared with those from C57BL/6 or BALB/c mice. Further investigation revealed no difference among the strains in terms of CD4+ T cell survival, upstream TCR signaling, or CD69 expression or in the phosphorylation of STAT-3 and expression of suppressor of cytokine signaling 3 that positively or negatively regulate IL-17 cell production. However, DBA/1 CD4+ T cells expressed increased levels of retinoic acid-related orphan receptor γt (RORγt). Furthermore, under iTreg cell polarizing conditions, DBA/1 CD4+ T cells showed a strikingly reduced level of FoxP3 expression. When interferon-γ and IL-4 were blocked, FoxP3 expression increased but remained lower in DBA/1 CD4+ T cells following exposure to TGFβ as compared with C57BL/6 CD4+ T cells. Moreover, DBA/1 CD4+ T cells showed reduced phosphorylation of Smad2 and Smad3 under both Th17 and iTreg cell polarizing conditions.
  
  
  CONCLUSIONS
  These results indicate that naive CD4+ T cells from DBA/1 mice have a dichotomous response to TGFβ: enhanced RORγt, yet reduced FoxP3, up-regulation. This observation may help to elucidate the branch point of TGFβ signaling that leads to skewed Th17, but reduced iTreg, cell differentiation.
authors.affiliation: University of Washington, Seattle, WA, USA.
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Morita
authors.forename: Doaa M
authors.initials: DM
authors.lastname: Ismail
authors.forename: Keith B
authors.initials: KB
authors.lastname: Elkon
authors.forename: Cong-Qiu
authors.initials: CQ
authors.lastname: Chu
mesh_terms: D000818:Animals; D015153:Blotting, Western; D015496:CD4-Positive T-Lymphocytes / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D002478:Cells, Cultured; D016207:Cytokines / Q000276:immunology / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D051858:Forkhead Transcription Factors / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D008811:Mice, Inbred DBA; D057094:Nuclear Receptor Subfamily 1, Group F, Member 1 / Q000276:immunology / Q000378:metabolism*; D011948:Receptors, Antigen, T-Cell / Q000276:immunology / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013045:Species Specificity; D053773:Transforming Growth Factor beta1 / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D016207:Cytokines; D051858:Forkhead Transcription Factors; C498833:Foxp3 protein, mouse; D057094:Nuclear Receptor Subfamily 1, Group F, Member 1; D011948:Receptors, Antigen, T-Cell; C539739:Rora protein, mouse; D053773:Transforming Growth Factor beta1