# Treg cells retain stable lineage commitment during pregnancy in mice after late gestation inflammatory challenge

## TL;DR

This study shows that Treg cells in pregnant mice maintain their identity even when inflammation is induced, which could help understand preterm birth.

## Contribution

The study reveals that Treg cells retain lineage stability during pregnancy despite inflammatory challenges.

## Key findings

- Ex-Foxp3 cells were found in uterine-draining lymph nodes but did not increase with inflammation.
- Treg cells retained their lineage program and did not adopt proinflammatory phenotypes during late gestation.
- RNA sequencing showed loss of Treg lineage markers in ex-Foxp3 cells but no expansion due to inflammation.

## Abstract

Inflammation is a major driver of preterm birth, a common pregnancy disorder and the leading cause of childhood death. T regulatory (Treg) cells are essential mediators of maternal fetal tolerance and are critical for constraining uterine inflammation. In some tissue settings, loss of Foxp3 expression can cause instability in Treg cell lineage commitment, elevated production of proinflammatory cytokines and compromised suppressive function. Whether preterm birth susceptibility is associated with loss of lineage fidelity and adoption of proinflammatory phenotypes in Treg cells is unknown. In this study, we investigated the lineage stability of Treg cells in vivo in pregnant mice using a Foxp3 fate‐mapping system and models of preterm birth induced by late‐gestation inflammatory challenge with lipopolysaccharide (LPS) or interleukin‐1β (IL‐1β). Ex‐Foxp3‐expressing (ex‐Foxp3) cells were observed in the uterus‐draining lymph nodes (udLNs) in non‐pregnant mice and in similar abundance across normal gestation, and a proportion expressed proinflammatory cytokines IFNγ and/or IL‐17A. Bulk RNA‐sequencing of sorted Treg and ex‐Foxp3 cells from late‐gestation udLNs revealed substantial loss of the Treg cell lineage program in ex‐Foxp3 cells, characterized by reversal in expression of canonical Treg cell genes and pathways. Late gestation LPS or IL‐1β administration to induce preterm birth did not expand the ex‐Foxp3 cell population in the uDLNs or uterine decidua. We conclude that uterine Treg cells exhibit a high level of lineage stability in pregnancy regardless of proinflammatory challenge. Whether there is any biological or pathophysiological significance of ex‐Foxp3 cells in gestational tissues remains to be defined.

In this study, we used Foxp3‐fatemapping mice to examine the cell lineage stability of Treg cells in pregnancy. Ex‐Foxp3 cells were identified in gestational tissues. However, Treg cells retained lineage stability with no increased ex‐Foxp3 generation, regardless of inflammatory challenges that induce preterm birth.