title: Mycobacterium tuberculosisInduces Expansion of Foxp3 Positive CD4 T-cells with a Regulatory Profile in Tuberculin Non-sensitized Healthy Subjects: Implications for Effective Immunization against TB.
abstract: OBJECTIVE
  Infection by MTB or exposure to MTB constituents is associated with intense microbial stimulation of the immune system, through both antigenic and TLR components, and induction of a milieu that is rich in pro-inflammatory/anti-inflammatory cytokines. Here, we addressed the basis of induced regulatory T-cell (iT-reg) expansion in response to MTB stimulation, in the absence of prior T cell antigen responsiveness.
  
  
  METHODS
  PBMC from HIV-1 un-infected TST negative and TST positive control subjects were stimulated by virulent MTB H37Rv lysate (L), a French press preparation of MTB that includes all bacterial components. Phenotype of MTB H37RvL induced iT-reg was assessed using immunostaining and flow cytometry. Functional capacity of iT-reg was assessed using3H-Thymidine incorporation and IFNγ production of non-adherent T cells (NAC) in the presence or absence of iT-reg in corresponding culture supernatants in response to TCR stimulation. Realtime PCR was used to assess IDO and FoxP3 mRNA expression.
  
  
  RESULTS
  The capacity of MTB H37RvL to induce CD4+CD25hi+Foxp3+T-cells in PBMC from TST negative subjects was robust (p<0.001), and in fact comparable to induction of iT-reg in PBMC from TST positive subjects. MTB-induced CD4+CD25hi+T-reg were TGFβ positive (p<0.05). Further, MTB H37RvL induced CD4+CD25hi+Foxp3+iT-reg suppressed3H-Thymidine incorporation and IFNγ production of non-adherent T cells (NAC) in response to TCR stimulation. MTB H37RvL induction of iT-reg was significantly stronger (p<0.01) than that by TLR-2, TLR-4, TLR-9 ligands, or combination of all TLR ligands. MTB H37RvL inducted indoleamine 2,3-dideoxygenase (IDO) mRNA expression in monocytes (p<0.001), and co-culture with the IDO inhibitor, D-1MT, decreased frequencies of T-reg (p<0.05). Inhibition of TGFβ by siRNA reduced Foxp3 mRNA expression in CD4 T cells (p<0.05).
  
  
  CONCLUSIONS
  Therefore, MTB and its components expand functional iT-reg in human mononuclear cells from MTB non-sensitized subjects. Also, MTB-induced iT-reg expansion depends on mononuclear phagocyte expression of both TGFβ and IDO.
authors.affiliation: Division of Infectious Diseases, Case Western Reserve University, Cleveland OH, USA.
authors.forename: Christina S
authors.initials: CS
authors.lastname: Hirsch
authors.affiliation: Division of Infectious Diseases, Case Western Reserve University, Cleveland OH, USA.
authors.forename: Roxana
authors.initials: R
authors.lastname: Rojas
authors.affiliation: Division of Infectious Diseases, Case Western Reserve University, Cleveland OH, USA.
authors.forename: Mianda
authors.initials: M
authors.lastname: Wu
authors.affiliation: Division of Infectious Diseases, Case Western Reserve University, Cleveland OH, USA; Veterans Affairs Medical Center, Cleveland, OH, USA.
authors.forename: Zahra
authors.initials: Z
authors.lastname: Toossi
keywords: IDO; IL-6; IL-8; T-reg; TGFβ; TLR2; Tuberculosis