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There is now compelling evidence that TNFR2 is constitutively expressed on Tregs and TNF-TNFR2 interaction is critical for the activation, expansion and functional stability of CD4+Foxp3+ regulatory T cells (Tregs). However, we also showed that the expression of TNFR2 could be up-regulated on CD4+Foxp3- effector T cells (Teffs) upon TCR stimulation, and TNFR2-expressing Teff cells were more activated and more resistant to Treg-mediated inhibition. In order to further define the role of TNFR2 in the proliferative expansion of pathogenic CD4 T cells in a setting of autoimmune disorder, we transferred naïve CD4 cells from WT mice and TNFR2-/- mice into Rag1-/- recipients. The results showed that TNFR2-deficient Teff cells failed to induce full-fledged colitis unlike WT Teffs. This was due to defective proliferative expansion of TNFR2-deficient Teff cells in the lymphopenic mice, as well as their reduced capacity to produce proinflammatory Th1 cytokines on a per cell basis. In vitro, TNF blockade inhibited the up-regulation of TNFR2 expression, as well as proliferative responses and cytokine production, by TCR-stimulated naïve CD4 cells. This effect of TNF blockade can be replicated by anti-TNFR2 Ab, but not by anti-TNFR1 Ab. Therefore, TNFR2 is critical for the proliferative expansion of pathogenic CD4 T cells, and thus may be a therapeutic target in the treatment of CD4 T cell-mediated autoimmune inflammation, although at a cost of attenuation of Treg activity.

The Triggering Receptors Expressed on Myeloid cells (TREM) gene cluster contains receptors that regulate a variety of leukocytes. TREM1 synergizes with Toll Like Receptors to promote cytokine production in neutrophils, macrophages and dendritic cells (DC). Paradoxically, although TREM1 and TREM2 both signal via DAP12, TREM2 is a negative regulator of inflammation in macrophages and DC. TREM2 is expressed in the colon but its role in colitis and colitis-associated cancer (CAC) is poorly understood. We found TREM2 expressed in monocytic myeloid-derived suppressor cells (MDSC) and tumor-associated macrophages (TAM). Moreover, TAM expressed a apparent TREM2 ligand. To test the role of TREM2 in inflammation-induced cancer we studied the azoxymethane (AOM)-dextran sulfate sodium (DSS)-induced model of CAC. DSS administration led a pronounced influx of myeloid cells and upregulation of both TREM1 and TREM2. Surprisingly, we found that Trem2-/- mice had less severe colitis as indicated by reduced histological scores, colon shortening and inflammatory cytokine production in the colon. AOM-DSS carcinogenesis in Trem2-/- mice resulted in slightly reduced total polyp counts and mitotic epithelial cells but a substantial reduction in the numbers of advanced carcinomas compared to wild type mice. Taken together, our data suggest an important role for TREM2 in the regulation of colitis and CAC likely via control of epithelial proliferation during colonic injury and inflammation.

Our previous results showed that TNF-TNFR2 signaling promotes activation and expansion of Tregs. Thus we hypothesize that TNF-TNFR2 signaling may play a critical role in maintaining phenotypic and functional stability of Tregs during an inflammatory response. We were able to confirm that, in colitis model induced by transfer of naïve CD4 cells into Rag KO mice, the disease could be inhibited by co-transfer of WT Tregs, but not by co-transfer of TNFR2-deficient Tregs. Furthermore, in the colon lamina propria of the colitis model, the majority of WT Tregs maintained FoxP3 expression. In contrast, increased number of TNFR2-deficient Tregs lost FoxP3 expression. In vitro, FoxP3 expression by highly purified Tregs was reduced to almost undetectable levels after potent TCR-stimulation, which was partially abrogated by TNF. This effect of TNF was blocked by anti-TNFR2 Ab, but not by anti-TNFR1 Ab. In addition, TNF was not able to increase FoxP3 expression by TNFR2-deficient Tregs undergoing TCR-stimulation. A full mouse genome array study revealed that, as compared with TNFR2- Tregs, TNFR2+ Tregs from normal mice expressed 4-fold lower levels of a gene encoding SATB1, in line with a recent study show that the repression of this genome organizer play a crucial role in maintenance of Treg phenotype and function. Thus, our data clearly show that the TNF-TNFR2 signaling is critical for the phenotypic and functional stability of Treg in the inflammatory environment.

Our previous study showed that TNFR2 is preferentially expressed by CD4+FoxP3+ regulatory T (Treg) cells and expression of this molecule identified the maximally suppressive Treg cells. TNFR2 is also expressed by a small fraction of CD4+FoxP3- conventional T (Tconv) cells in normal mice and its expression is up-regulated by T cell activation. Our data indicate that TNFR2 also co-stimulates Tconv cells. In this study, by using FoxP3/gfp KI mice, we showed that TNFR2 signaling did not induce FoxP3- CD4 cells to become suppressive. TNFR2 is associated with greater suppressive functions when expressed by Treg cells and is associated with greater resistance to suppression when expressed by Tconv cells, because TNFR2+ Tconv cells were potently suppressed by FoxP3+TNFR2+ Treg cells, while resisting suppression by FoxP3+TNFR2- Treg cells. Furthermore, in mice with 4T1 breast tumor and LLC, intratumoral Tconv cells expressing elevated levels of TNFR2 acquired the capacity to resist the suppression mediated by LN-derived Treg cells. However, they remained susceptible to inhibition by more suppressive tumor infiltrating Treg cells which expressed higher levels of TNFR2. Intratumoral Treg cells express more TNFR2 and they are able to overcome the inhibition-resistance of intratumoral Tconv cells, resulting in a dominant immunosuppressive tumor environment. (Funded by NCI Contract N01-CO-12400)