*3.1. Altered Hormonal Milieu Effect on Placental Development*

The interpretation of the effects of ARTs on pregnancy and the possible associated pathologies is complicated by the supra-physiological hormonal levels in the recipient, as a consequence of the ovarian stimulation treatment [37]. Bourgain and Devroey in 2003 suggested that increased hormone blood levels might alter the timing of endometrial receptivity, with possible suboptimal embryo implantation and development [60]. It was also suspected, by studies in animal models, that high estrogen levels exert a detrimental effect on spiral artery remodeling by the trophoblast [61]. Moreover, it has been observed that, in FET procedures, levels of progesterone higher than 32.5 ng/mL on the day of embryo transfer, were associated with no pregnancy [62]. This cut off level seems improbable in ovarian controlled stimulation conditions of fresh IVF cycles, where many corpora lutei are functioning and progesterone level, together with estradiol, are much higher. Conversely, the lack of hormones might also affect placental development by creating conditions leading to increased risks for HDP, as it has been suggested for FET following hormonal endometrial preparation [63,64]. Using these protocols, ovulation does not occur, and the corpus luteum, which secretes important protective vasoactive substances, is not formed [9,10,65]. Among these factors, relaxin is a potent vasodilator promoting vascular

compliance [66] and facilitating the adaptation of the maternal cardiovascular system to pregnancy [67]. Moreover, in programmed FET cycles, the use of estradiol and progesterone has been recently associated with obstetric adverse outcomes, i.e., PTB, LBW, and PE, which are small for gestational age (SGA) and large for gestational age (LGA) when compared to natural FET cycle [9]. Thyroid hormone (TH) has been also demonstrated as an important player in reproduction; both hypothyroidism and hyperthyroidism have been associated with subfertility, recurrent miscarriages, and adverse pregnancy outcomes [68,69]. It has been recently demonstrated that TH regulates protease expression and activation of Notch signaling in implantation and embryo development [70]. Moreover, levothyroxine (LT4) treatment has a positive effect on conception capacity and reduces miscarriages when administered to euthyroid women with autoimmune thyroid disease affected by recurrent miscarriages [71].

Several common infertility conditions, such as endometriosis and PCOS, often treated with ART, are characterized by a hormonal dysregulation that affects proper placentation. In endometriosis, a defective deep placentation may derive from functional abnormalities of the eutopic endometrium, as well as an imbalance in endocrine and inflammatory markers [72]. PTB, placenta previa, placental abruption, gestational hypertension, PE, LBW, SGA, cesarean delivery, postpartum hemorrhage, and stillbirth have been significantly associated with endometriosis in a systematic review and meta-analyses, including in 39 studies [73]. PCOS is another condition leading to infertility, in which the defective trophoblast invasion and placentation may be caused by mother's hyperandrogenism [74,75]. Testosterone can act directly on trophoblast invasion, with modifications of placenta morphology and function [76,77]. PCOS is a chronic low-grade inflammation associated with metabolic dysfunction that is enhanced in pregnancy by the induction of an endothelial dysfunction [78]. This condition might in turn prevent normal remodeling of spiral vessels and the physiologic decrease of uterine artery impedance, thus reducing the depth of endovascular trophoblast invasion. As a consequence of these patterns, in patients with PCOS the placental weight, thickness, density and volume was found significantly reduced [76]. In women with PCOS undergoing IVF, the risk of adverse pregnancy outcomes was found to be significantly elevated. In a recent meta-analysis including 29 observational studies, Sha et al. (2019) described an increased risk for GDM, pregnancy-induced hypertension (PIH), LGA, miscarriage and PTB in women with PCOS undergoing ART [79].
