*3.4. Mechanical Stress on Embryo and Placental Development (the Case of PGT-A)*

PGT-A is an ART technique that is increasingly performed, considering that it represents 40% of all IVF cycles in the USA [115]. Its aim is to discover if produced embryos are aneuploid and, in this case, they should not be transferred into the uterus, in order to prevent implantation failure and miscarriages. PGT-A preliminarily requires the in vitro development of embryos up to the blastocyst stage, in order to collect 5 to 10 trophoblast cells. Embryos are then frozen while waiting for the genetic analysis results. Embryos with no or few aneuploidies are thawed and transferred (FET) in the uterine cavity after endometrial hormonal preparation using estrogen and progesterone. In this condition, natural ovulation and formation of the corpus luteum are suppressed. Similar to PGT-A, preimplantation genetic test for monogenic diseases (PGT-M) allows to screen for embryos affected by monogenic diseases.

Some authors have expressed concerns on the possibility that trophectoderm biopsy might disturb embryo development, leading to potential adverse consequences on obstetric outcomes [65,116]. It has been hypothesized that removal of cells from the tissue that will become the placenta does not affect embryo implantation rate but may negatively influence later phases of placentation. Placenta-derived anomalies, such as placenta previa and accreta, HDP, and FGR might be consequences of this abnormal placentation [117–120].

In a comparative study investigating obstetric and neonatal outcome following PGT-A in blastocyst-stage biopsy with frozen embryo transfer and cleavage-stage biopsy with fresh embryo transfer, Jing et al. (2016) found that PGT-A-FET was associated with a higher incidence of gestational hypertension (9.0% vs. 2.3%; *p* = 0.017) [121]. These data might indicate a higher risk of placental injury as a consequence of PGT-A procedure on the trophectoderm. Similar indications came from the results of a study on 345 singleton and 76 twin deliveries after PGT-M. PGT-M showed an increased risk of obstetric complications, when compared with pregnancies conceived spontaneously or by IVF without PGT [122]. In particular, these data showed a higher rate of HDP (6.9%), compared with spontaneous conception patients (2.3%) and the IVF group (4.7%). It was reported, as well, a higher rate of SGA neonates (12.4%), in comparison with those from spontaneous conception group (3.9%) and the IVF patients (4.5%). Moreover, Makhijani et al. (2021) observed a significantly higher probability of developing HDP in patients who underwent PGT, compared with

those who did not (adjusted odds ratio (aOR) 1.943, 95% CI 1.072–3.521; *p* = 0.029) [123]. In this study, the potential influence of other factors causing HDP, such as the hormonal endometrial preparation was considered. In their binary logistic regression model, they found that, with the adjusted odds of HDP, the risk associated with trophectoderm biopsy remained significantly higher. A recent large single-center study found a two-fold higher risk of pregnancy-related hypertension following 241 blastocyst biopsy and frozen transfer than in 515 non-biopsied controls [123].

Conversely, is it worthwhile to look for embryonic aneuploidies. It was shown that, among embryos with aneuploid cells, 31% were of meiotic origin and 74% related to mitotic aneuploidies, with maximum aneuploidy originating by days 4–5 after fertilization, falling to 5–6% by day 7 [124]. Consequently, we should consider the environment influence on the mitotic origin of aneuploidies in developing embryos. From a strict clinical point of view, PGT-A does not improve IVF outcomes nor does it reduce miscarriage rates [125]. Moreover, healthy babies have been born after the transfer of aneuploid mosaic embryos [126–128]. This is not surprising since it is well known the self-correction ability of aneuploid embryos. It has been demonstrated that, in mice, a *p*-53-dependent autophagy-mediated apoptotic process is able to eliminate aneuploid cells [129]. In humans, this process is promoted by the bone morphogenetic protein-4 (BMP4) [130]. These mechanisms, together with the fact that the results obtained from a trophectoderm biopsy cannot be representative of the whole embryo [131], contradict the assumption that all aneuploid embryos should be eliminated and constitutes a strong criticism for the PGT-A clinical utility [125]. The Scientific and Clinical Advances Advisory Committee of HFEA recently changed the rating of PGT-A to "red", meaning that there is no evidence to show that the treatment is effective and safe [125].
