Gαi2-deficient mice spontaneously develop colitis. Using xMAP technology and RT-PCR, we investigated cytokine/chemokine profiles during histologically defined phases of disease: (i) no/mild, (ii) moderate, (iii) severe colitis without dysplasia/cancer and (iv) severe colitis with dysplasia/cancer, compared with age-matched wild-type (WT) littermates. Colonic dysplasia was observed in 4/11 mice and cancer in 1/11 mice with severe colitis. The histology correlated with progressive increases in colon weight/cm and spleen weight, and decreased thymus weight, all more advanced in mice with dysplasia/cancer. IL-1β, IL-6, IL-12p40, IL-17, TNF-α, CCL2 and CXCL1 protein levels in colons, but not small intestines increased with colitis progression and were significantly increased in mice with moderate and severe colitis compared with WT mice, irrespective of the absence/presence of dysplasia/cancer. CCL5 did not change during colitis progression. Colonic IL-17 transcription increased 40- to 70-fold in all stages of colitis, whereas IFN-γ mRNA was gradually up-regulated 12- to 55-fold with colitis progression, and further to 62-fold in mice with dysplasia/cancer. IL-27 mRNA increased 4- to 15-fold during the course of colitis, and colonic IL-21 transcription increased 3-fold in mice with severe colitis, both irrespective of the absence/presence of dysplasia/cancer. FoxP3 transcription was significantly enhanced (3.5-fold) in mice with moderate and severe colitis, but not in mice with dysplasia/cancer, compared with WT mice. Constrained correspondence analysis demonstrated an association between increased protein levels of TNF-α, CCL2, IL-1β, IL-6 and CXCL1 and dysplasia/cancer. In conclusion, colonic responses are dominated by a mixed T(h)1/T(h)17 phenotype, with increasing T(h)1 cytokine transcription with progression of colitis in Gαi2(-/-) mice.

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BackgroundMice deficient in the inhibitory G protein subunit Gαi2 spontaneously develop a T helper 1 dominated colitis. We examined whether a defect in CD4+FoxP3+ regulatory T cells (Treg) underpins the pathogenesis of colitis in the Gαi2−/− (Gαi2-deficient) colitis model.Methodology/Principal FindingsUsing flow cytometry, we found that thymus and colonic lamina propria, but not spleen and mesenteric lymph nodes, of colitic Gαi2−/− mice contained increased frequencies of Treg, whereas FoxP3 expression intensity was similar in Gαi2−/− compared to Gαi2+/− or Gαi2+/+ wild type (WT) mice. The frequency of CD4+FoxP3+ T cells expressing CD103 was significantly increased in Gαi2−/− compared to WT mice. Treg in colons from WT mice clustered in the T cell areas of colonic lymphoid patches (CLP), with relatively few Treg in the lamina propria, as demonstrated by immunohistochemistry. In Gαi2−/− mice, CLP were not observed but lamina propria Treg were increased in number and frequency within the CD4+ infiltrate, compared to WT mice. Using an in vitro co-culture system and flow cytometric analysis of cell division we could demonstrate that the in vitro suppressive function of WT and Gαi2−/− CD4+FoxP3+ regulatory T cells (WT-Treg and KO-Treg) was indistinguishable, but that T effector cells (CD4+25− T cells) from Gαi2−/− mice were less readily suppressed than WT effectors (WT-Teff) by Treg from either source. However, neither WT nor Gαi2−/− Treg was able to suppress colitis induced by adoptive transfer of Gαi2−/− effector T cells (KO-Teff) to RAG2−/− recipients. The enhanced inflammatory activity of Gαi2−/− effectors was accompanied by increased expression of an effector/memory T cell phenotype and increased cytokine secretion, especially IL-4, IL-6 and IFN-γ.ConclusionsThere is an increased frequency of Gαi2−/− Treg in the colon, and they demonstrate no endogenous functional defect. However, Gαi2−/− T effector cells are dramatically less susceptible to suppression in vitro, and in vivo, despite increased effective numbers of Treg, they cannot prevent disease.