Regulatory T cells are paramount effectors in progesterone regulation of embryo implantation and fetal growth.

Progesterone (P4) is essential for embryo implantation, but the extent to which the pro-gestational effects of P4 depend on the maternal immune compartment is unknown. Here, we investigate whether regulatory T cells (Treg cells) act to mediate luteal phase P4 effects on uterine receptivity in mice. P4 antagonist RU486 administered to mice on days 0.5 and 2.5 postcoitum to model luteal phase P4 deficiency caused fewer CD4+Foxp3+ Treg cells and impaired Treg functional competence, along with dysfunctional uterine vascular remodeling and perturbed placental development in midgestation. These effects were linked with fetal loss and fetal growth restriction, accompanied by a Th1/CD8-skewed T cell profile. Adoptive transfer at implantation of Treg cells - but not conventional T cells - alleviated fetal loss and fetal growth restriction by mitigating adverse effects of reduced P4 signaling on uterine blood vessel remodeling and placental structure and by restoring maternal T cell imbalance. These findings demonstrate an essential role for Treg cells in mediating P4 effects at implantation and indicate that Treg cells are a sensitive and critical effector mechanism through which P4 drives uterine receptivity to support robust placental development and fetal growth.

Hormonal regulation of the cytokine microenvironment in the mammary gland.

The mammary gland is a unique organ that undergoes hormone-driven developmental changes over the course of the ovarian cycle during adult life. Macrophages play a role in regulating cellular turnover in the mammary gland and may affect cancer susceptibility. However, the immune microenvironment that regulates macrophage function has not been described. Hormonal regulation of the cytokine microenvironment across the ovarian cycle was explored using microbead multiplex assay for 15 cytokines in mammary glands from C57Bl/6 mice at different stages of the oestrous cycle, and in ovariectomised mice administered oestradiol and progesterone. The cytokines that were found to fluctuate over the course of the oestrous cycle were colony-stimulating factor (CSF)1, CSF2, interferon gamma (IFNG) and tumour necrosis factor alpha (TNFA), all of which were significantly elevated at oestrus compared with other phases. The concentration of serum progesterone during the oestrus phase negatively correlated with the abundance of cytokines CSF3, IL12p40, IFNG and leukaemia inhibitory factor (LIF). In ovariectomised mice, exogenous oestradiol administration increased mammary gland CSF1, CSF2, IFNG and LIF, compared with ovariectomised control mice. Progesterone administration together with oestradiol resulted in reduced CSF1, CSF3 and IFNG compared with oestradiol administration alone. This study suggests that the cytokine microenvironment in the mammary gland at the oestrus phase of the ovarian cycle is relatively pro-inflammatory compared with other stages of the cycle, and that the oestradiol-induced cytokine microenvironment is significantly attenuated by progesterone. A continuously fluctuating cytokine microenvironment in the mammary gland presumably regulates the phenotypes of resident leukocytes and may affect mammary gland cancer susceptibility.

Stress response genes are suppressed in mouse preimplantation embryos by granulocyte-macrophage colony-stimulating factor (GM-CSF).

BACKGROUND: Granulocyte-macrophage colony-stimulating factor (GM-CSF) is known to promote the development and survival of human and mouse preimplantation embryos; however, the mechanism of action of GM-CSF in embryos is not defined. METHODS: Mouse blastocysts were cultured from zygote stage in vitro with and without recombinant mouse GM-CSF (rmGM-CSF), and in vivo developed blastocysts were flushed from Csf2 null mutant and wild-type mice. The effect of GM-CSF on blastocyst expression of stress response and apoptosis genes was evaluated by microarray, qPCR and immunochemistry. RESULTS: Microarray analysis of the gene transcription profile showed suppression of stress response and apoptosis gene pathways in blastocysts exposed to rmGM-CSF in vitro. qPCR analysis confirmed that rmGM-CSF inhibited expression of heat shock protein (HSP) and apoptosis pathway genes Cbl, Hspa5, Hsp90aa1, Hsp90ab1 and Gas5 in in vitro blastocysts. Immunocytochemical analysis of HSP 1 (HSPA1A/1B; HSP70), BAX, BCL2 and TRP53 (p53) in in vitro blastocysts showed that HSPA1A/1B and BCL2 proteins were less abundant when embryos were cultured with rmGM-CSF. BAX and TRP53 were unchanged at the protein level, but Bax mRNA expression was reduced after GM-CSF treatment. In in vivo developed blastocysts, Csf2 null mutation caused elevated expression of Hsph1 but not other stress response genes. CONCLUSIONS: We conclude that GM-CSF inhibits the cellular stress response and apoptosis pathways to facilitate embryo growth and survival, and the protective effects of GM-CSF are particularly evident in in vitro culture media, whereas in vivo other cytokines can partly compensate for absence of GM-CSF.