*2.4. Dendritic Cells*

DCs are the third cell group of innate immune cells present in the decidua. DCs have a key role in triggering the immune response by inducing the activation and differentiation of naïve T cells and simultaneously play a critical role for the development of tolerance [212]. The biological plasticity of DCs in promoting immunity or tolerance appears to be dependent on both their maturation state and the microenvironment in which they dwell. Thus, immature DCs are specifically efficient in antigen uptake and presentation, but the low expression levels of MHC gene products and T cell costimulatory molecules may contribute to peripheral tolerance under homeostatic conditions [213].

Concerning the uterine microenvironment, various stimuli such as soluble mediators (e.g., hormones, cytokines, and growth factors) or interactions with other immune cells modulate the tolerogenic activity of DCs, indicating that this behavior is context dependent. It has been reported that estrogen inhibited the ability of DCs to stimulate T cell proliferation and the production of both Th1⁄Th2 cytokines by upregulating IDO that supported the maturation of DCs with tolerogenic properties in rodents [214].

A recent review that focused on the pleiotropic nature of IFN-γ could help clarify the role played by DCs during early pregnancy, a phase characterized by inflammation [215]. Although IFN-γ exhibits extensive proinflammatory effects and has been associated with pathological pregnancies, it can also paradoxically exert an immunosuppressive role on both innate and adaptive immune cell types, promoting DC tolerance through the induction of IDO expression. IFN-γ induced IDO competence is not limited to immune cells but extends to other cell types such as epithelial cells, mesenchymal stem cells, and tumor cells. Therefore, the depletion of tryptophan leads to the inhibition of the effector T cells and promotes the differentiation of the FoxP3<sup>+</sup> Treg cells [215]. In addition, uMϕ may also be an additional source of IFN-γ [216], although uMϕ has been reported to suppress IFN-γ production by T cells costimulatory B7-H1/PD-1 signaling during early pregnancy. Therefore, B7-H1, expressed by uMϕ, functions as a key regulator of local IFN-γ production, and thus contributes to the development of appropriate maternal immune responses [217]. Several studies have shown that when DCs were stimulated by IFN-γ in the absence of danger or pathogen-related signals, its effects induced predominantly tolerogenic features [218]. In this scenario, the crosstalk between decidua and immune cells contributes to the adequate vascular remodeling of the spiral arterioles and to the protection of the fetus from infections, and also guarantees the development of the tolerogenic microenvironment.

Uterine DCs (uDCs) appear to also have a role in implantation and decidua formation because the presence of CD11c<sup>+</sup> DCs is critical during early placentation, as the number of uDCs increases at the implantation period and the depletion of these cells leads to implantation failure in mouse models [219,220].

The close contact between uDCs and uNK cells in decidua suggests that there are important reciprocal interactions between them in shaping the decidualizing microenvironment during the early stages of pregnancy [169]. Indeed, uDCs appear to promote the differentiation of uNK cells, because uNK cell functions are impaired in association with decreased levels of IL-15 and IL-12 in implantation sites depleted of uDCs [159,221]. Therefore, the depletion of uDCs, as well as the altered uNK cells maturation, impair tissue remodeling and angiogenesis [221–224]. In addition, studies have shown that uterine cell proliferation was dependent on a synergistic effect between uDCs and uNK cells, confirming that uDCs-uNK cells crosstalk may be important for the endometrial stroma differentiation during implantation [160,166,169].

Implant failure was also observed in combinations of allogeneic and syngeneic mating after DCs depletion, as DCs depletion altered decidua response [219]. Again, the expression of sFlt-1/sVEGFR-1 in mature human monocyte-derived dendritic cells counteracted their angiogenic properties, and thus prevented adequate implantation [223]. However, since uDCs are generally in an immature state, their role in implantation, on the one hand, is likely redundant and potentially compensated by other immune cells. On the other hand, they likely are more important in playing an early key role in promoting tolerance to paternal antigens. Indeed, altered levels of Treg cells and DCs have been demonstrated in the peripheral blood of women with PE [222,224].

Concerning the PlGF role in inducing tolerance, it has been reported that PlGF inhibited the activation and maturation of human DCs differentiated from CD14<sup>+</sup> monocytes, effectively and rapidly through the NF-κB signaling pathway [225], confirming the immunosuppressive effect shown by angiogenic growth factors [226,227]. PlGF-treated DCs resulted in the downregulation of the expression of maturation markers CD80, CD86, CD83, CD40, and MHC-II expression, as well as the inhibition of IL-12, IL-8, and TNF-α production in response to LPS stimulation, with respect to untreated DCs. PlGF inhibited DCs maturation through the VEGFR-1, and this PlGF-induced DCs dysfunction was rescued by anti-human Flt-1/VEGFR-1 mAb. Additionally, the treatment of DCs with PlGF resulted in the suppression of naïve CD4<sup>+</sup> T cell proliferation in an allogeneic mixed lymphocyte reaction. The results from this study indicated that PlGF could downregulate Th1 immune responses by modulating the function of DCs [225].

Last but not least, PlGF is able to recruit bone marrow-derived myelomonocytic cells through its receptor Flt-1/VEGFR-1 via NFAT-1 transcriptional activation [34,228]. These myeloid-derived cells have been shown to exert immunomodulatory effects on the immune cells, especially DCs. Common features of bone marrow-derived myelomonocytic cells are the immature state and a remarkable ability to suppress T cell responses [226,227]. In addition to their suppressive effects on adaptive immune responses, they have also been reported to regulate innate immune responses by modulating cytokines production by macrophages such as IL-6, IL-10, CCL-22, and TGF-β. These cytokines, in turn, promote the differentiation of monocytes to mature macrophages but block their differentiation to DC, decrease the expression of DCs maturation markers, attenuate the Th1 immune response, as well as impair the activity of cytotoxic T lymphocytes and NK cells.
