An essential role for interleukin 10 in the function of regulatory T cells that inhibit intestinal inflammation

Abstract

A T helper cell type 1-mediated colitis develops in severe combined immunodeficient mice after transfer of CD45RB(high) CD4(+) T cells and can be prevented by cotransfer of the CD45RB(low) subset. The immune-suppressive activities of the CD45RB(low) T cell population can be reversed in vivo by administration of an anti-transforming growth factor beta antibody. Here we show that interleukin (IL)-10 is an essential mediator of the regulatory functions of the CD45RB(low) population. This population isolated from IL-10-deficient (IL-10(-/-)) mice was unable to protect from colitis and when transferred alone to immune-deficient recipients induced colitis. Treatment with an anti-murine IL-10 receptor monoclonal antibody abrogated inhibition of colitis mediated by wild-type (WT) CD45RB(low) CD4(+) cells, suggesting that IL-10 was necessary for the effector function of the regulatory T cell population. Inhibition of colitis by WT regulatory T cells was not dependent on IL-10 production by progeny of the CD45RB(high) CD4(+) cells, as CD45RB(low) CD4(+) cells from WT mice were able to inhibit colitis induced by IL-10(-/-) CD45RB(high) CD4(+) cells. These findings provide the first clear evidence that IL-10 plays a nonredundant role in the functioning of regulatory T cells that control inflammatory responses towards intestinal antigens.

Figure 1

Representative photomicrographs of the descending colon of RAG-2^−/− mice after transfer of subpopulations of CD4⁺ T cells from WT or IL-10^−/− mice. (A) Severe colitis in a mouse injected with CD45RB^high CD4⁺ T cells from WT mice. (B) Normal appearance of the colon in a mouse restored with WT CD45RB^high and WT CD45RB^low CD4⁺ T cells, indicating that the WT CD45RB^low population is able to inhibit disease induced by WT CD45RB^high CD4⁺ T cells. (C) Severe colitis in a mouse cotransferred with WT CD45RB^high CD4⁺ T cells and IL-10^−/− CD45RB^low CD4⁺ cells, indicating that the IL-10^−/− CD45RB^low subpopulation is unable to protect from the disease. (D) Severe colitis in a mouse receiving only IL-10^−/− CD45RB^low cells, indicating that this population is able to induce disease. Hematoxylin and eosin; original magnifications: ×50.

Figure 2

IFN-γ and TNF-α production by LP CD4⁺ T cells from colons. RAG-2^−/− or C.B-17 SCID recipients were reconstituted with cell subsets as indicated. 8–12 wk after cell reconstitution, LP cells were isolated and stimulated for 12 h with anti-CD3∈ antibody. Levels of cytokine expression were determined by cytofluorography. (A) Absolute number of CD4⁺ cytokine-positive cells per colon. Numbers were determined by multiplying the frequency of cytokine-secreting CD4⁺ T cells by the total number of CD4⁺ cells. Data represent the mean ± SEM of two to five animals per group. (B) Frequency of cytokine-secreting CD4⁺ T cells. Data are gated on CD4⁺ T cells and are representative examples for each group. Horizontal lines represent staining with isotype control mAb.

Figure 2

IFN-γ and TNF-α production by LP CD4⁺ T cells from colons. RAG-2^−/− or C.B-17 SCID recipients were reconstituted with cell subsets as indicated. 8–12 wk after cell reconstitution, LP cells were isolated and stimulated for 12 h with anti-CD3∈ antibody. Levels of cytokine expression were determined by cytofluorography. (A) Absolute number of CD4⁺ cytokine-positive cells per colon. Numbers were determined by multiplying the frequency of cytokine-secreting CD4⁺ T cells by the total number of CD4⁺ cells. Data represent the mean ± SEM of two to five animals per group. (B) Frequency of cytokine-secreting CD4⁺ T cells. Data are gated on CD4⁺ T cells and are representative examples for each group. Horizontal lines represent staining with isotype control mAb.