Background/Purpose: In rheumatoid arthritis (RA), regulatory T cells (Tregs) are defective in their suppressive capacities and fail to control chronic inflammation. TNF-α is involved in inhibition of Treg differentiation and activation, likely via activation of TNF-α type 1 receptor (TNFR1)¹. Conversely, activation of TNFR2 on Tregs is critical for their phenotypic and functional stability in the inflammatory environment². Moreover, it has been shown that therapeutic TNF blockade with the anti-TNF monoclonal antibodies restores the potency of Treg cell suppression in RA by binding to membrane TNF- α on monocytes and promoting Treg cell expansion through enhanced TNFR2 signaling³.

In the present work, we studied the role of TNFR2 on Tregs in control of inflammation at multiple levels, by: 1) studying the action of TNF on Treg function in the presence and absence of TNFR2 in vitro, 2) testing the severity of a model of skin inflammation in TNFR2KO mice, 3) evaluating the evolution of TNFR2-expressing Tregs from RA patients during anti-TNF treatment.

Methods: Mice deficient in the TNFR2 gene (TNFR2 KO) and TNFR2 lox/lox mice to conditionally delete TNFR2 specifically in Tregs were used. CD4⁺CD25⁺Treg cells were purified by magnetic sorting. Cell phenotype was evaluated by flow cytometry. Tregs stability was evaluated by analyzing methylation status of 9 CpG motifs of the Foxp3 locus, assessed by bisulfite sequencing of CD4⁺ CD25⁺ purified cells. Skin inflammation was induced by cutaneous application of an imiquimod-containing ointment. Peripheral blood Tregs were characterized before and after 3 months of anti–TNF treatment in 12 RA patients and in 19 patients with axial spondylaorthritis (AxSpA) that were used as control.

Results: In vitro, TNF-α enhanced Foxp3 maintenance in cultured Tregs. With two models of TNFR2 inactivation (a TNFR2-blocking antibody and a TNFR2 knock-out mouse) this effect was mediated by TNF-TNFR2 -and not TNFR1- interaction. In vivo, TNFR2-negative Treg cells, from both TNFR2KO and TNFR2 lox/lox mice, had lower spontaneous suppressive capacities (lower inhibition of effector T cell proliferation and IFN-g production). FoxP3 methylation was higher in Tregs from TNFR2 KO mice than wt mice. This suggested that TNFR2 expression confers higher stability to Tregs.

Compared to wt mice, TNFR2KO mice had enhanced skin-inflammation and decreased Tregs and CD39⁺ Tregs frequencies in lymph nodes. In RA patients responding to anti-TNF treatment, an increase in the frequency of TNFR2-expressing Tregs was evident at 3 months of treatment vs. the baseline. Conversely, no variation in the frequency of these cells was observed in AxSpA patients

Conclusion: TNFR2 signaling on Tregs may play a major role in controlling inflammation and can be activated both by TNF-α and anti-TNF treatment. Further studies to dissect TNFR2 dependent pathways on Tregs are warranted.

References

1. Nie H, Zheng Y, Li R, et al. Nat Med 2013;19:322-8.

2. Chen X, Wu X, Zhou Q, et al. J Immunol. 2013;190:1076-84.

3. Nguyen DX, Ehrenstein MR. J Exp Med 2016;213:1241-53.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/tnfr2-regulatory-t-cellssubpopulations-are-highly-suppressive-and-are-increased-on-anti-tnf-treatment/