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Bacteria-triggered CD4(+) T regulatory cells suppress Helicobacter hepaticus-induced colitis.

Kullberg MC, Jankovic D, Gorelick PL, Caspar P, Letterio JJ, Cheever AW, Sher A - J. Exp. Med. (2002)

Bottom Line: The disease-protective WT CD4(+) cells are contained within the CD45RB(low) fraction and unexpectedly were found in both the CD25(+) and the CD25(-) subpopulations of these cells, their frequency being higher in the latter.The mechanism by which CD25(+) and CD25(-) CD45RB(low) CD4(+) cells block colitis involves IL-10 and not transforming growth factor (TGF)-beta, as treatment with anti-IL-10R but not anti-TGF-beta monoclonal antibody abrogated their protective effect.The induction of such cells in response to gut flora may be a mechanism protecting normal individuals against inflammatory bowel disease.

View Article: PubMed Central - PubMed

Affiliation: Immunobiology Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases/NIH, Building 50, Room 6146, 50 South Drive, Bethesda, MD 20892, USA. MKullberg@niaid.nih.gov

ABSTRACT
We have previously demonstrated that interleukin (IL)-10-deficient (IL-10 knockout [KO]) but not wild-type (WT) mice develop colitis after infection with Helicobacter hepaticus. Here, we show that infected recombination activating gene (RAG) KO mice develop intestinal inflammation after reconstitution with CD4(+) T cells from IL-10 KO animals and that the cotransfer of CD4(+) T cells from H. hepaticus-infected but not uninfected WT mice prevents this colitis. The disease-protective WT CD4(+) cells are contained within the CD45RB(low) fraction and unexpectedly were found in both the CD25(+) and the CD25(-) subpopulations of these cells, their frequency being higher in the latter. The mechanism by which CD25(+) and CD25(-) CD45RB(low) CD4(+) cells block colitis involves IL-10 and not transforming growth factor (TGF)-beta, as treatment with anti-IL-10R but not anti-TGF-beta monoclonal antibody abrogated their protective effect. In vitro, CD45RB(low) CD4(+) cells from infected WT mice were shown to produce IL-10 and suppress interferon-gamma production by IL-10 KO CD4(+) cells in an H. hepaticus antigen-specific manner. Together, our data support the concept that H. hepaticus infection results in the induction in WT mice of regulatory T cells that prevent bacteria-induced colitis. The induction of such cells in response to gut flora may be a mechanism protecting normal individuals against inflammatory bowel disease.

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CD45RBlow CD4+ cells from infected WT mice inhibit IFN-γ production by IL-10 KO CD4+ cells and produce IL-10 after in vitro stimulation with SHelAg. (A) Responder CD4+ cells from infected IL-10 KO mice were cultured with APC, SHelAg, and indicated numbers of CD45RBlow CD4+ cells from naive (○) or infected (•) WT mice. IFN-γ was measured in 72-h supernatants. (B) CD45RBlow CD4+ cells from naive (□) or infected (▪) WT mice were cultured with IL-10–deficient APC and indicated Ag. IL-10 was measured in 72-h supernatants. No IFN-γ, IL-4, or IL-5 was detected in cultures stimulated with SHelAg plus IL-2. (C) CD45RBlow cells from naive or infected WT mice were cultured with IL-10–deficient APC and SHelAg plus IL-2. After removal of supernatants at 72 h, IL-2 was added for an additional 18 h before cells were stimulated with PMA plus ionomycin and analyzed for the expression of IL-10 and IFN-γ by intracellular staining. The FACS® dot plots shown are gated on CD4+ cells. In the same assay, the cells were negative for intracellular IL-4. The results shown in A–C are representative of two or three experiments performed.
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fig8: CD45RBlow CD4+ cells from infected WT mice inhibit IFN-γ production by IL-10 KO CD4+ cells and produce IL-10 after in vitro stimulation with SHelAg. (A) Responder CD4+ cells from infected IL-10 KO mice were cultured with APC, SHelAg, and indicated numbers of CD45RBlow CD4+ cells from naive (○) or infected (•) WT mice. IFN-γ was measured in 72-h supernatants. (B) CD45RBlow CD4+ cells from naive (□) or infected (▪) WT mice were cultured with IL-10–deficient APC and indicated Ag. IL-10 was measured in 72-h supernatants. No IFN-γ, IL-4, or IL-5 was detected in cultures stimulated with SHelAg plus IL-2. (C) CD45RBlow cells from naive or infected WT mice were cultured with IL-10–deficient APC and SHelAg plus IL-2. After removal of supernatants at 72 h, IL-2 was added for an additional 18 h before cells were stimulated with PMA plus ionomycin and analyzed for the expression of IL-10 and IFN-γ by intracellular staining. The FACS® dot plots shown are gated on CD4+ cells. In the same assay, the cells were negative for intracellular IL-4. The results shown in A–C are representative of two or three experiments performed.

Mentions: To further address whether H. hepaticus–reactive Treg lymphocytes exist within the CD45RBlow CD4 population, we used an in vitro model in which the suppressive effect of Treg cells on a responder IL-10 KO CD4+ cell population was examined. Using this system, infected but not naive WT CD45RBlow CD4+ cells inhibited IFN-γ production by IL-10 KO CD4+ cells in an H. hepaticus Ag–specific manner (Fig. 8 A). In addition, the infected WT CD45RBlow CD4+ cells were shown to produce IL-10 after stimulation with SHelAg plus IL-2 as measured by both ELISA and intracellular cytokine staining, a response not detected from comparable cell populations from naive mice (Fig. 8, B and C). As expected, CD45RBlow cells from both naive and infected WT mice produced IL-10 in response to anti-CD3 plus IL-2 (Fig. 8 B). Taken together, our results argue that the mechanism by which CD4+ cells from infected WT mice block colitis involves the production of IL-10 from CD45RBlow CD4+ Treg cells induced by exposure to H. hepaticus.


Bacteria-triggered CD4(+) T regulatory cells suppress Helicobacter hepaticus-induced colitis.

Kullberg MC, Jankovic D, Gorelick PL, Caspar P, Letterio JJ, Cheever AW, Sher A - J. Exp. Med. (2002)

CD45RBlow CD4+ cells from infected WT mice inhibit IFN-γ production by IL-10 KO CD4+ cells and produce IL-10 after in vitro stimulation with SHelAg. (A) Responder CD4+ cells from infected IL-10 KO mice were cultured with APC, SHelAg, and indicated numbers of CD45RBlow CD4+ cells from naive (○) or infected (•) WT mice. IFN-γ was measured in 72-h supernatants. (B) CD45RBlow CD4+ cells from naive (□) or infected (▪) WT mice were cultured with IL-10–deficient APC and indicated Ag. IL-10 was measured in 72-h supernatants. No IFN-γ, IL-4, or IL-5 was detected in cultures stimulated with SHelAg plus IL-2. (C) CD45RBlow cells from naive or infected WT mice were cultured with IL-10–deficient APC and SHelAg plus IL-2. After removal of supernatants at 72 h, IL-2 was added for an additional 18 h before cells were stimulated with PMA plus ionomycin and analyzed for the expression of IL-10 and IFN-γ by intracellular staining. The FACS® dot plots shown are gated on CD4+ cells. In the same assay, the cells were negative for intracellular IL-4. The results shown in A–C are representative of two or three experiments performed.
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Related In: Results  -  Collection

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fig8: CD45RBlow CD4+ cells from infected WT mice inhibit IFN-γ production by IL-10 KO CD4+ cells and produce IL-10 after in vitro stimulation with SHelAg. (A) Responder CD4+ cells from infected IL-10 KO mice were cultured with APC, SHelAg, and indicated numbers of CD45RBlow CD4+ cells from naive (○) or infected (•) WT mice. IFN-γ was measured in 72-h supernatants. (B) CD45RBlow CD4+ cells from naive (□) or infected (▪) WT mice were cultured with IL-10–deficient APC and indicated Ag. IL-10 was measured in 72-h supernatants. No IFN-γ, IL-4, or IL-5 was detected in cultures stimulated with SHelAg plus IL-2. (C) CD45RBlow cells from naive or infected WT mice were cultured with IL-10–deficient APC and SHelAg plus IL-2. After removal of supernatants at 72 h, IL-2 was added for an additional 18 h before cells were stimulated with PMA plus ionomycin and analyzed for the expression of IL-10 and IFN-γ by intracellular staining. The FACS® dot plots shown are gated on CD4+ cells. In the same assay, the cells were negative for intracellular IL-4. The results shown in A–C are representative of two or three experiments performed.
Mentions: To further address whether H. hepaticus–reactive Treg lymphocytes exist within the CD45RBlow CD4 population, we used an in vitro model in which the suppressive effect of Treg cells on a responder IL-10 KO CD4+ cell population was examined. Using this system, infected but not naive WT CD45RBlow CD4+ cells inhibited IFN-γ production by IL-10 KO CD4+ cells in an H. hepaticus Ag–specific manner (Fig. 8 A). In addition, the infected WT CD45RBlow CD4+ cells were shown to produce IL-10 after stimulation with SHelAg plus IL-2 as measured by both ELISA and intracellular cytokine staining, a response not detected from comparable cell populations from naive mice (Fig. 8, B and C). As expected, CD45RBlow cells from both naive and infected WT mice produced IL-10 in response to anti-CD3 plus IL-2 (Fig. 8 B). Taken together, our results argue that the mechanism by which CD4+ cells from infected WT mice block colitis involves the production of IL-10 from CD45RBlow CD4+ Treg cells induced by exposure to H. hepaticus.

Bottom Line: The disease-protective WT CD4(+) cells are contained within the CD45RB(low) fraction and unexpectedly were found in both the CD25(+) and the CD25(-) subpopulations of these cells, their frequency being higher in the latter.The mechanism by which CD25(+) and CD25(-) CD45RB(low) CD4(+) cells block colitis involves IL-10 and not transforming growth factor (TGF)-beta, as treatment with anti-IL-10R but not anti-TGF-beta monoclonal antibody abrogated their protective effect.The induction of such cells in response to gut flora may be a mechanism protecting normal individuals against inflammatory bowel disease.

View Article: PubMed Central - PubMed

Affiliation: Immunobiology Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases/NIH, Building 50, Room 6146, 50 South Drive, Bethesda, MD 20892, USA. MKullberg@niaid.nih.gov

ABSTRACT
We have previously demonstrated that interleukin (IL)-10-deficient (IL-10 knockout [KO]) but not wild-type (WT) mice develop colitis after infection with Helicobacter hepaticus. Here, we show that infected recombination activating gene (RAG) KO mice develop intestinal inflammation after reconstitution with CD4(+) T cells from IL-10 KO animals and that the cotransfer of CD4(+) T cells from H. hepaticus-infected but not uninfected WT mice prevents this colitis. The disease-protective WT CD4(+) cells are contained within the CD45RB(low) fraction and unexpectedly were found in both the CD25(+) and the CD25(-) subpopulations of these cells, their frequency being higher in the latter. The mechanism by which CD25(+) and CD25(-) CD45RB(low) CD4(+) cells block colitis involves IL-10 and not transforming growth factor (TGF)-beta, as treatment with anti-IL-10R but not anti-TGF-beta monoclonal antibody abrogated their protective effect. In vitro, CD45RB(low) CD4(+) cells from infected WT mice were shown to produce IL-10 and suppress interferon-gamma production by IL-10 KO CD4(+) cells in an H. hepaticus antigen-specific manner. Together, our data support the concept that H. hepaticus infection results in the induction in WT mice of regulatory T cells that prevent bacteria-induced colitis. The induction of such cells in response to gut flora may be a mechanism protecting normal individuals against inflammatory bowel disease.

Show MeSH
Related in: MedlinePlus