Relevance of Foxp3+ regulatory T cells for early and late phases of murine sepsis

The role of Foxp3⁺ regulatory T (Treg) cells in the course of the early hyper-inflammatory and subsequent hypo-inflammatory phases of sepsis is ambiguous. Whereas Nrp1 expression has been reported to discriminate natural Treg cells from induced Treg cells, the Treg cell stability depends on the methylation status of foxp3-TSDR. To specifically evaluate the role of Foxp3⁺ Treg cells in the early and late phases of sepsis, we induced sepsis by caecal ligation and puncture and subsequent Pseudomonas aeruginosa lung infection in a DEREG (DEpletion of REGulatory T cells) mouse model. We found an increase of Foxp3⁺ Treg cells to all CD4⁺ T cells during murine sepsis. Using a new methylation-sensitive quantitative RT-PCR method and deep amplicon sequencing, we demonstrated that natural (Nrp1⁺ Foxp3⁺) Treg cells and most induced (Nrp1⁻ Foxp3⁺) Treg cells are stable and exhibit unmethylated foxp3-TSDR, and that both Treg populations are functionally suppressive in healthy and septic mice. DEREG mice depleted of Foxp3⁺ Treg cells exhibit higher disease scores, mortality rates and interleukin-6 expression levels than do non-depleted DEREG mice in early-phase sepsis, a finding indicating that Foxp3⁺ Treg cells limit the hyper-inflammatory response and accelerate recovery. Treg cell depletion before secondary infection with P. aeruginosa 1 week after caecal ligation and puncture does not influence cytokine levels or the course of secondary infection. However, a moderate Treg cell recurrence, which we observed in DEREG mice during secondary infection, may interfere with these results. In summary, Treg cells contribute to a positive outcome after early-phase sepsis, but the data do not support a significant role of Treg cells in immune paralysis during late-phase sepsis.