Background Peripheral induced regulatory T cells (iTreg) play an important role for immune homeostasis. Dysregulation of iTreg differentiation is frequently associated with autoimmune diseases or tumors. Tumor necrosis factor (TNF) α is a pleiotropic cytokine with multiple biological function. Some studies suggest that mouse TNFα inhibits the differentiation of iTregs via TNFR2, providing TNFα-TNFR2 as an attractive target for drug development, for potential control of various diseases including malignant tumors and autoimmune diseases. However, whether human TNFα-TNFR2 signaling pathway plays similar role on iTreg differentiation remains unknown. Moreover, no optimal animal model is currently available for candidate drug evaluation targeting human TNFα-TNFR2. In this study, a humanized TNFR2 mouse model (TNFR2 HuGEMM) was developed to evaluate the effect of human TNFR2 agonist or antagonist on iTreg differentiation, which may be useful in the development of immunotherapies to treat tumors or autoimmune diseases, respectively.
Methods A humanized TNFR2 mouse model (HuGEMM) in C57BL/6 background was generated by replacing the extracellular domain of mouse TNFR2 with its human counterpart. Naïve CD4+ T cells were isolated from the splenocytes of homozygous TNFR2 HuGEMM. To validate the signal transduction function of this humanized chimeric TNFR2 upon human TNFα stimulation and its effect on iTreg differentiation, the isolated naïve mouse CD4+ cells were cultured under iTreg differentiation condition with mouse cytokines in the presence or absence of human TNFα. To test the effect of human TNFα-TNFR2 signaling on iTreg function, standard Treg suppression was performed.
Results The expression of humanized chimeric TNFR2 in mouse CD4+ T cells were determined by flow cytometry. Exogenous human TNFα significantly inhibited iTreg differentiation but had no effect on iTreg function in the suppression of naïve mouse CD3+ T cell proliferation.
Conclusions This study provided a novel and useful murine model to evaluate the drug candidate targeting hTNFa-hTNFR2 signaling pathway. In vitro iTreg differentiation assay can be used to evaluate the drug candidate’s effect on hTNFa-hTNFR2 signaling. These data also suggest that TNFa-TNFR2 signaling plays an important inhibitory role on iTreg differentiation, while it has no obvious effect on iTreg suppressive function on T cell proliferation.
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