Francesca Evangelista

Polarized light microscopy-detectable structures of human oocytes and embryos are related to the likelihood of conception in IVF

PurposeTo clarify if birefringent structures of human oocytes and embryos, measurable by polarized light microscopy, have any value in predicting the chance of pregnancy in human in vitro fertilization and may halp to identify the most competent oocytes and embryos.MethodsThe inner layer of the zona pellucida (IL-ZP) and the meiotic spindle (MS) were analyzed by polarized light microscopy in 258 oocytes and in the 209 embryos deriving from them. Data obtained from 102 ICSI cycles with conception were compared with those obtained in 156 cycles without conception. The retardance and area of the IL-ZP, as well as the retardance, length of the major axis, and area of the MS were measured. Furthermore, polarized light microscopy parameters were related to the embryo morphological score by multiple regression analysis.Result(s)The mean area of the IL-ZP of both oocytes and embryos was significantly lower in conception than in non-conception cycles (p = 0.0001 for oocytes and p = 0.002 for embryos). The area of the IL-ZP in embryos was significantly, inversely related to the embryo morphological score (p = 0.011). The area, the major axis length and the retarcance of the MS, as well as the retardance of the IL-ZP in oocytes and embryos were comparable in conception and non-conception cycles.ConclusionThe area of the IL-ZP of the human oocytes may represent a marker of oocyte competence, as oocytes with a low IL-ZP area are more frequently obtained in conception cycles. When measured in embryos, a low IL-ZP area identifies embryos with a high chance of implantation.

Inducible Nitric Oxide Synthase and Heme Oxygenase 1 Are Expressed in Human Cumulus Cells and May Be Used as Biomarkers of Oocyte Competence

The interplay between oocyte and surrounding cumulus cells (CCs) during follicular growth influences oocyte competence to undergo fertilization and sustain embryo development. The expression of many genes and proteins in CCs has been suggested as potential biomarker of oocyte competence in human in vitro fertilization (IVF). In the present study, we analyzed 90 human cumulus–oocyte complexes obtained during IVF procedure: 30 CCs were analyzed using quantitative real-time polymerase chain reaction and 60 CCs using Western blotting analysis to detect gene and protein expression of some enzymes related to oxidative stress, that is, the 3 nitric oxide synthase (NOS) isoforms and heme oxygenase 1 (HO-1). In the group of 60 CCs, we also investigated the expression and phosphorylation of IkBα, a known inhibitor of the nuclear factor κB (NF-κB) pathway, which controls several redox-sensitive genes. The expression of the messenger RNAs (mRNAs) was related to the oocyte morphological analysis performed by polarized light microscopy and to the occurrence of normal fertilization after intracytoplasmic sperm injection. We observed that the amount of iNOS and HO-1 mRNAs and proteins is significantly higher, and that in the meanwhile the NF-κB pathway is activated, in CCs corresponding to oocytes that were not fertilized in comparison to CCs whose corresponding oocyte showed normal fertilization. Instead, no correlation between the fertilization and the oocytes’ morphological data was observed. These results suggest that the increase in iNOS and HO-1 mRNAs expression in CCs is a negative index of oocyte fertilizability and might be an useful tool for oocyte selection.

Large randomized trial comparing transabdominal ultrasound‐guided embryo transfer with a technique based on uterine length measurement before embryo transfer

To compare in a randomized, non‐inferiority trial the embryo transfer (ET) technique based on uterine length measurement before transfer (ULMbET) with transabdominal ultrasound‐guided embryo transfer (TA‐UGET) in a large population of patients undergoing in‐vitro fertilization (IVF).

A large prospective randomized trial comparing trans‐abdominal ultrasound‐guided embryo transfer (UGET) with an embryo transfer technique based on previous uterine length measurement (ULMb‐ET)

To compare in a randomized, non‐inferiority trial the embryo transfer (ET) technique based on uterine length measurement before transfer (ULMbET) with transabdominal ultrasound‐guided embryo transfer (TA‐UGET) in a large population of patients undergoing in‐vitro fertilization (IVF).

Evaluation of equine oocyte developmental competence using polarized light microscopy

The purpose of this study was to observe in vitro-matured equine oocytes with an objective computerized technique that involves the use of a polarized light microscope (PLM) in addition to the subjective morphological evaluation obtained using a classic light microscope (LM). Equine cumulus-oocyte complexes (COCs, n = 922) were subjected to different in vitro maturation times (24, 36 or 45 h), however, only 36-h matured oocytes were analyzed using CLM. The 36-h matured oocytes that reached maturity were parthenogenetically activated to evaluate the quality and meiotic competence. Average maturation percentages per session in groups 1, 2 and 3 (24-, 36- and 45-h matured oocytes respectively) were 29.31 ± 13.85, 47.01 ± 9.90 and 36.62 ± 5.28%, whereas the average percentages of immature oocytes per session were 28.78 ± 20.17, 7.83 ± 5.51 and 22.36 ± 8.39% respectively. The zona pellucida (ZP) birefringent properties were estimated and correlated with activation outcome. ZP thickness and retardance of the inner layer of the zona pellucida (IL-ZP) were significantly increased in immature oocytes compared with mature oocytes (P < 0.001 and P < 0.01 respectively). The comparison between parthenogenetically activated and non-activated oocytes showed a significant increase in the area and thickness of the IL-ZP in parthenogenetically activated oocytes (P < 0.01). These results show that the 36-h in vitro maturation (IVM) protocol allowed equine oocytes to reach maturity, and PLM observation of ZP can be used to distinguish mature and immature oocytes as well as activated and non-activated oocytes.

A slowly reabsorbed, echogenic surgical thread provides a long-lasting ultrasound- detectable marker of grafted ovarian tissue

This communication reports a novel technical solution for the orthotopic transplant of cryostored-thawed ovarian tissue. The described technique was applied to three young women with iatrogenic ovarian failure. An echogenic thread that is reabsorbed after 6 months was used to fasten the thawed ovarian small fragments before grafting them onto the atrophic ovary. This technical solution made it possible to avoid the loss of small tissue pieces during laparoscopic grafting as well as to precisely localize the grafted tissue by transvaginal ultrasound during the following months. The precise localization of the grafted tissue was particularly helpful when its revascularization and functional recovery were followed up using, respectively, colour Doppler and transvaginal follicle growth examination. In conclusion, the use of a slowly reabsorbed, ultrasound-detectable surgical thread as an ultrasound-detectable marker able to improve the localization of the exact site at which ovarian tissue was grafted is proposed.

La crioconservazione del tessuto gonadico femminile

RiassuntoLa crioconservazione del tessuto ovarico è una tecnica innovativa per preservare la fertilità delle giovani pazienti a rischio di insufficienza ovarica a causa di trattamenti oncostatici gonadotossici oppure per cause benigne quali patologie genetiche, autoimmuni od ovariche. Il tessuto ovarico crioconservato, dopo scongelamento, può essere reimpiantato orto elo eterotopicamente oppure posto in coltura per ottenere la maturazione in vitro dei follicoli primordiali. Ad oggi nel mondo sono nati 17 bambini da ritrapianto ortotopico di tessuto ovarico. Il primo e unico caso di bambino nato con questa tecnica in Italia è stato recentemente ottenuto dalla nostra équipe.