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Study Question: Can sperm donation increase live birth rates following ICSI in advanced maternal age (AMA) patients?Summary Answer: Sperm donation increases the live birth rate in AMA ICSI cycles.What Is Known Already: In ICSI practice, sperm donation has been predominantly applied to overcome male infertility. The involvement of paternal age and lower sperm quality in the severe reduction in fertility observed in AMA patients remains to be clarified.Study Design, Size, Duration: Retrospective multicenter cohort study including data generated between 2015 and 2019 from 755 ICSI cycles achieving a fresh embryo transfer, of which 337 were first homologous cycles (normozoospermic partner sperm and homologous oocytes) and 418 were first sperm donation cycles (donor sperm and homologous oocytes). The association of sperm origin (partner vs donor) with live birth was assessed by multivariate analysis in non-AMA (<37 years, n = 278) and AMA (≥37 years, n = 477) patients, separately, including in the model all variables previously found to be associated with live birth in a univariate analysis (number of MII oocytes recovered, number of embryos transferred, and maternal age). ICSI outcomes were compared between sperm donation and homologous cycles in overall, non-AMA and AMA patients.Participants/materials, Setting, Methods: The study was conducted in three fertility clinics and included 755 Caucasian patients aged 24-42 years undergoing their first homologous or sperm donation ICSI cycle achieving a fresh embryo transfer.Main Results and the Role Of Chance: The multivariate analysis revealed that sperm donation was positively associated with the likelihood of a live birth independently of all other variables tested in AMA (P = 0.02), but not in non-AMA patients. Live birth, delivery, and miscarriage rates differed substantially between sperm donation and homologous AMA cycles; live birth and delivery rates were 70-75% higher (25.4% vs 14.5% and 22.5% vs 13.5%, respectively; P < 0.01), while miscarriage occurrence was less than half (18.0% vs 39.5%; P < 0.01) in sperm donation compared to homologous AMA cycles.Limitations, Reasons For Caution: This study is limited by its retrospective nature, differences in patients profiles between sperm donation and homologous-control groups and varying proportion of donor cycles between fertility centers, although these variations have been controlled for in the statistical analysis.Wider Implications Of the Findings: The findings suggest that sperm donation increases live birth rates while reducing miscarriage occurrence in AMA patients, and thus may be a valid strategy to improve ICSI outcomes in this growing and challenging patient group.Study Funding/competing Interest(s): N/A.Trial Registration Number: N/A. [ABSTRACT FROM AUTHOR]

Background: Fertility loss during female ageing is associated with increasing basal FSH and decreasing anti-Müllerian hormone (AMH) concentrations, together with compromised oocyte quality, presumably due to increased oxidative stress (OS) and DNA damage, as well as reduced metabolic and meiotic competences. Basal FSH and AMH circulatory concentrations have been broadly utilized as IVF success predictors, regardless of fluctuations in prognostic accuracy; basal FSH and AMH perform better in pre-advanced maternal age (AMA: >35 years) and AMA patients, respectively. The relationships between FSH and AMH intrafollicular levels and IVF outcomes suggest, nevertheless, that both hormones regulate oocyte competence, supporting the hypothesis that changes in FSH/AMH levels cause, at least in part, oocyte quality degradation during ageing. To understand the reasons behind the fluctuations in FSH and AMH prognostic accuracies and to clarify their participation in mechanisms determining oocyte competence and age-related subfertility, a deeper knowledge of the regulation of FSH and AMH intrafollicular signalling during the female reproductive lifespan, and of their effects on the cumulus-oocyte complex, is required., Objective and Rationale: An extensive body of information on the regulation of FSH and AMH intrafollicular availability and signalling, as well as on the control of folliculogenesis and oocyte metabolism, has been accumulated. However, these datasets have been explored within the relatively narrow boundaries of their specific subjects. Given the aforementioned gaps in knowledge and their clinical relevance, herein we integrate clinical and basic data, within a wide biological perspective, aiming to shed light on (i) the reasons for the variability in the accuracy of serum FSH and AMH as fertility markers, and on (ii) the potential roles of these hormones in mechanisms regulating oocyte quality, particularly those associated with ageing., Search Methods: The PubMed database encompassing the period between 1960 and 2021 was searched. Principal search terms were FSH, FSH receptor, AMH, oocyte, maternal age, cumulus, transzonal projections (TZPs), actin, OS, redox, reactive oxygen species, mitochondria, DNA damage, DNA repair, aneuploidy, spindle, meiosis, gene expression, transcription, translation, oocyte secreted factors (OSFs), cAMP, cyclic guanosine monophosphate, natriuretic peptide C, growth differentiation factor 9, bone morphogenetic protein 15 and fibroblast growth factor., Outcomes: Our analysis suggests that variations in the accuracy of fertility prognosis reflect a modest association between circulatory AMH levels and oocyte quality as well as increasing basal FSH inter-cycle variability with age. In addition, the basic and clinical data articulated herein support the hypothesis that increased intrafollicular FSH levels, as maternal age advances, may override the physiological protective influences of AMH and OSFs against excessive FSH signalling in cumulus cells. This would result in the disruption of oocyte homeostasis via reduced TZP-mediated transfer of cumulus-derived molecules essential for meiotic competence, gene expression, redox activity and DNA repair., Wider Implications: In-depth data analysis, encompassing a wide biological perspective has revealed potential causative mechanisms of age-related subfertility triggered by alterations in FSH/AMH signalling during the female reproductive life. Insights from new mechanistic models arising from this analysis should contribute to advancing our comprehension of oocyte biology in humans and serve as a valuable reference for novel AMA subfertility treatments aimed at improving oocyte quality through the modulation of AMH/FSH action., (© The Author(s) 2021. Published by Oxford University Press on behalf of European Society of Human Reproduction and Embryology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Chronic endometritis (CE) is a subtle pathology. Despite being difficult to detect and probably underdiagnosed, it has great clinical relevance, representing as it does a reversible cause of infertility. Nowadays, histological examination with identification of endometrial stromal plasma cells is considered the gold standard for diagnosis. Diagnostic difficulties persist, however, as a result of the technical limitations of this method and the lack of standardized histological diagnostic criteria. Hysteroscopy has been proposed as an aid for CE diagnosis. The method works by detecting signs of inflammation (focal or diffuse hyperemia, stromal edema, presence of micropolyps and the typical strawberry aspect) on the endometrial surface. Yet, the jury is still out on how reliable this technique is. Hysteroscopy displays a high sensitivity (over 86% and up to 100%) and high negative predictive value (over 92% and up to 100%) in the diagnosis of CE, and it should probably be performed routinely in the assessment of patients with unexplained infertility, repeated implantation failure and repeated pregnancy loss; however, since values in the literature regarding specificity are conflicting, in cases of suspected CE, hysteroscopy may be combined with histological examination, which remains the gold standard to confirm CE. Considering that histopathological evaluation probably underdiagnoses CE, and that hysteroscopy tends to overdiagnose, further studies are needed to determine which technique (or combination of techniques) has greater value for patients.

Given the importance of embryo developmental competence assessment in reproductive medicine and biology, the aim of this study was to compare the performance of fertilization and cleavage morphokinetics with embryo morphology to predict post-ICSI live birth. Data from embryos cultured in a time-lapse microscopy (TLM) incubator and with known live birth outcomes (LB: embryos achieving live birth, n = 168; NLB: embryos not achieving live birth, n = 1633) were used to generate receiver operating characteristic (ROC) curves based on morphokinetic or morphological scores, and the respective areas under the curve (AUC) were compared. The association between live birth and 12 combinations of four morphokinetic quality degrees (A-D) with three morphological quality degrees (A-C) was assessed using multivariate analysis. Morphokinetic parameters from tPNa to t8 were reached earlier in LB compared with NLB embryos. The ROC curve analysis indicated that morphokinetic information is more accurate than conventional morphology to predict live birth [AUC = 0.64 (95% CI 0.58-0.70) versus AUC = 0.58 (95% CI 0.51-0.65)]. The multivariate analysis was in line with AUCs, revealing that embryos with poor morphokinetics, independently of their morphology, provide lower live birth rates (P < 0.001). A considerable percentage of embryos with top morphology presented poor morphokinetics (20.10%), accompanied by a severely reduced live birth rate in comparison with embryos with top morphology and morphokinetics (P < 0.001). In conclusion, TLM-derived early morphokinetic parameters were more predictive of live-birth achievement following ICSI than conventional morphology.

Research Question: Does the association of basal FSH and anti-Müllerian hormone (AMH) concentrations with post-IVF/intracytoplasmic sperm injection (ICSI) live birth change with maternal age?, Design: A total of 2003 IVF/ICSI patients were stratified according to basal FSH/AMH in concordant favourable (CF; AMH >1 ng/ml and FSH ≤10 IU/l), concordant unfavourable (CU; AMH ≤1 ng/ml and FSH >10 IU/l), discordant with favourable AMH (DFA) and discordant with favourable FSH (DFF) groups, as well as according to age in pre-advanced maternal age (pre-AMA; <35), AMA-1 (≥35, ≤37), AMA-2 (>37, ≤40) and AMA-3 (>40). IVF/ICSI outcomes were compared among CF, CU, DFA and DFF groups, and the association of basal FSH and AMH concentrations with live birth was tested by univariate and multivariate analysis in total, pre-AMA and AMA groups, separately., Results: Different outcome patterns were observed in discordant AMH/FSH groups from different age categories; favourable basal FSH concentrations were associated with higher delivery rates in pre-AMA patients, but with lower delivery rates in AMA groups. Within pre-AMA patients, DFF patients presented higher delivery rates but lower oocyte yield compared with DFA patients. In the univariate analysis, favourable AMH (P < 0.02) and oocyte yield (P < 0.002) were positively associated with live birth in all AMA groups. The multivariate analysis revealed that favourable basal FSH, but not AMH or oocyte yield, is associated with live birth in pre-AMA patients independently of other variables (P = 0.012)., Conclusions: The relationship of basal FSH and AMH with IVF/ICSI success changes with maternal age; basal FSH better reflects clinical outcomes probably determined by oocyte quality in pre-AMA patients, while AMH better suits AMA patients., (Copyright © 2020 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.)

Aim: Define how and when fixed starting gonadotropin doses can be used in current clinical ART practices in Italy., Methods: A Delphi conference consisting of three rounds was performed in order to define the ideal clinical conditions in which fixed-gonadotropin-dose during COS should be applied. During the conference, 19 statements about the current ART practice were provided to a panel of twenty-nine national experts. Median score was 5 (IQ:4-6) in all Delphi rounds., Results: Eleven statements (57.9%) were classified as shareable with high-degree of convergence, 2 (10.5%) as shareable with low convergence and 6 (31.6%) as un-shareable with high convergence. The panel reached high consensus regarding some statements: (i) fixed FSH-dose in normoresponders and poor-responder, (ii) importance of predicting ovarian response before COS, considering multiple markers to select the right stimulation protocol for each patient, (iii) importance of therapy simplification and standardization to improve efficiency during COS. Moreover, a low-convergence was reached about use of GnRH antagonist as first treatment line and drug storage at room temperature. However from these findings, the debate remains open regarding some other statements: (a) usefulness of Bologna-criteria to define poor-responders; (b) efficacy to change always stimulation protocol after a failure IVF; (c) utility of AMH-dosed with standardized automatic mode to define normo-responder patients; (d) usefulness to modify the dosage of 12.5 IU/die during COS to improve stimulation effectiveness., Conclusion: Controlled ovarian stimulation remains a challenging clinical step in Assisted Reproductive Technique, especially in some specific patient groups for which no clinical consensus is available. This study is the first attempt to describe the shared clinical opinion regarding the fixed versus variable gonadotropin dose in the real IVF practice.

Objective: To assess the relationship of early developmental kinetics with competence to provide a live birth and the impact of maternal age in this context., Design: Retrospective cohort study including 4,915 embryos, of which 1,390 were transferred and provided a clinical outcome paired with morphokinetic data; 168 of them resulted in a live birth (LB), and 1,222 did not (NLB). Early morphokinetic parameters were compared between LB and NLB embryos from patients stratified into two age groups (<37 and ≥37 years), and between embryos at the same competence group from patients aged <37 and ≥37 years. The association of morphokinetic parameters with live birth was tested by univariate and multivariate analyses., Setting: Fertility clinic., Patient(s): The study population included 1,066 patients undergoing autologous intracytoplasmic sperm injection cycles with fresh single (SET), double (DET) or triple (TET) embryo transfers on day 2 or 3. Of them, 669 patients produced NLB embryos and 134 produced LB embryos., Intervention(s): None., Main Outcome Measure(s): Fertilization and cleavage morphokinetic parameters and live birth., Result(s): In the total patient population, all morphokinetic parameters were achieved earlier in LB compared with NLB embryos. The same was observed in patients aged <37 years (P<.015), but not ≥37 years. Except for the t8 (time at which an 8-blastomere embryo was identified), all morphokinetic parameters were reached earlier in LB embryos from patients aged <37 years compared with LB embryos from patients aged ≥37 years. Univariate analysis revealed that earlier occurrence of all morphokinetic parameters was associated with live birth, although only earlier t2 (time at which two separate and distinct cells were identified) was associated with live birth independently from maternal age in the multivariate analysis., Conclusion(s): Despite its retrospective nature and performance in a single IVF center, this study presents novel data indicating that embryos competent to provide a live birth display overall faster early developmental kinetics compared with embryos that do not achieve a live birth after transfer, a difference that, however, narrows as maternal age advances. The findings suggest that fertilization and cleavage morphokinetic parameters may constitute valuable references for embryo selection strategies aiming to improve live birth rates, specifically before advanced maternal age while holding limited usefulness in advanced maternal age., (Copyright © 2020 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Research Question: How much the variability in patients' response during in vitro fertilization (IVF) may add to the initial predicted prognosis based only on patients' basal characteristics?, Design: Anonymous data were obtained from the Human Fertilization and Embryology Authority (HFEA). Data involving 114,882 stimulated fresh IVF cycles were retrospectively analyzed. Logistic regression was used to develop the models., Results: Prediction of live birth was feasible with moderate accuracy in all of the three models; discrimination of the model based only on basal patients' characteristics (AUROC 0.61) was markedly improved adding information of number of embryos (AUROC 0.65) and, mostly, number of oocytes (AUROC 0.66)., Conclusions: The addition to prediction models of parameters such as the number of embryos obtained and especially the number of oocytes retrieved can statistically significantly improve the overall prediction of live birth probabilities when based on only basal patients' characteristics. This seems to be particularly true for women after the first IVF cycle. Since ovarian response affects the probability of live birth in IVF, it is highly recommended to add markers of ovarian response to models based on basal characteristics to increase their predictive ability., Competing Interests: Declaration of Competing Interest A.L.M., M.C., V.D., M.M.R., C.D.G., R.D.A. and S.K.S. have nothing to declare., (Copyright © 2020 Elsevier Masson SAS. All rights reserved.)

Study Question: What is the clinical efficacy of an oocyte donation program based on the transportation of frozen semen and embryos between two countries?Summary Answer: The transnational oocyte donation program is efficient and reliable and it could provide a first-line strategy to overcome the lack of donors in some countries.What Is Known Already: While there is increasing need for donated oocytes, in many countries the availability of donors is still insufficient to cover the therapeutic demands, and patients are referred abroad for treatment. Since embryo cryopreservation is reliable and efficient, we propose a strategy based on frozen embryos instead of frozen oocytes to satisfy the increasing demand for cross border oocyte donation.Study Design, Size, Duration: This is a retrospective cohort study including 630 patients treated from December 2015 to July 2017.Participants/materials, Setting, Methods: Infertile women were treated with elective vitrified-thawed embryo shipping and embryo transfer (ET) between two IVF clinics, one in Spain and one in Italy.Main Results and the Role Of Chance: A total of 2617 embryos were created for the 630 patients and the survival rate after warming was 98.5%. After the first ET the live birth rate (LBR) was 30.6%. In 476 patients (75.5%), embryos were transferred at the cleavage stage (Day 2 or 3) and the LBR was 29.2%. Vitrified blastocysts were available for 154 patients (24.5%) and the LBR was 35%. Among patients who did not achieve a pregnancy after the first frozen ET (FET), 92.5% had at least one frozen embryo for successive procedures. 213 patients underwent a second FET. The LBR at the second FET was 30%. The cumulative LBR at the end of the observation period was 39.3%.Limitations, Reasons For Caution: The study design was retrospective. A direct comparison with vitrified oocyte donors cycle and subsequent fresh ET would have permitted to compare this strategy versus the current standard based on vitrified gametes.Wider Implications Of the Findings: The LBR found in our study is more than acceptable and seems to be higher than what reported with vitrified oocytes. The transnational fresh oocyte donation program may have several advantages over the shipment of vitrified oocytes: similarly to the fresh oocyte donation program it allows for personalized care in oocyte recipient, which is provided by assigning a flexible number of oocytes, and at the same time it maintains the benefit of a frozen ART program permitting scheduling flexibility. The TOD program is efficient and may be proposed as a first-line strategy for distance and inter-countries oocyte donation programs.Study Funding, Competing Interest(s): None.Trial Registration Number: NA. [ABSTRACT FROM AUTHOR]

As highlighted by European statistics, the employment of donor oocytes is a growing option for women who cannot make use of their own gametes. As the potential recipients are continuously increasing in number, a donor programme which satisfies this demand is mandatory. Improvements in cryopreservation techniques, like oocyte and embryo vitrification, have led to the overcoming of the sequence of stimulation-retrieval-transfer both from a spatial and a temporal point of view, with the development of cryobanks of oocytes permitting crossborder donation. However, while some studies report comparable success when using vitrified and fresh oocytes we still need to investigate whether the use of fresh oocytes give higher live birth rate than cryopreserved ones, when the same number of oocytes are given. The performance of embryo cryopreservation, conversely, seems to be more reliable. A novel approach based on the shipment of frozen sperm from the recipient's country to the oocyte donor's one, where fresh oocytes are inseminated and the resulting embryos frozen and transported back to the referring IVF centre to perform a frozen embryo transfer may be a good strategy. We believe that the use of frozen embryos from fresh oocytes could be associated with a higher cumulative live birth rate per cycle, while favouring personalised oocyte recipient care with a flexible number of oocytes assigned and limiting the burden of travelling abroad., (Copyright © 2020 Facts, Views & Vision.)

Background: The addition of a GnRH analogue to the luteal phase in in vitro fertilization programs has been seldom proposed due to the presence of GnRH receptors in the endometrium. The aim of the study was to evaluate the effect of triptorelin addition in short antagonist cycles, compared to cycles where the only supplementation was progesterone., Methods: The primary objective of this study was the study of the effect of Triptorelin addiction during the luteal phase on the live birth rate. Secondary objectives of efficacy were pregnancy rates and implantation rates, as well as safety in terms of OHSS risks. The study was a prospective, randomized, open study, performed in two independent Centers from July 2013 to October 2015. Patients were divided into three groups: a) Regular antagonist protocol, with only luteal progesterone; b) Antagonist protocol with luteal triptorelin as multiple injections, c) Antagonist protocol with luteal triptorelin as single bolus. Descriptive statistics were obtained for all the parameters. Mean and standard deviation were used for all quantitative parameters. Differences between percentages were studied using Chi-square test generalized to the comparison of several proportions., Results: A total number of 1344 patients completed the study, 786 under the age of 35 years, and 558 over 35 years. It was observed an increase of positive HCG results, Clinical pregnancy rates and Delivery rates when triptorelin was added in the luteal phase, irrespective whether as a single bolus or five injections. This increase was statistically significant both for pregnancy rates and delivery rates. The statistic difference between pregnancies and deliveries obtained with or without luteal triptorelin reached p < 0,01. No increase of OHSS risk was observed., Conclusions: From this large study it appears that the concept of luteal phase supplementation should be revisited. From our study it appears that triptorelin addition to the luteal phase of antagonist cycles, either as a single bolus or using multiple injections, is a good tool to optimize ART results., Trial Registration: The study was approved by the Ethics Committee of Provincia di Bergamo (n 1203/2013).

Ultrasound follicular count (antral follicle count, AFC) is a necessary tool for measuring ovarian reserve, whereby the estimated number of follicles responsive to FSH can predict the number of oocytes retrieved in IVF cycles and may be the basis for individualized ovarian stimulation therapy. Advances in the ultrasound technology have recently lead to the improvement in resolution and quality of the image. Moreover the automatic measurements of follicular diameter by using some specific 3D software seems associated to several advantages when compared to the 2D technique. Examination time is reduced because the ultrasound scan data are stored and can be analyzed in detail at a later time. These data can be reconstructed in any plane, regardless of the original scan plane facilitating the detailed analysis. Another advantage is that this new technique reduces the operator's influence on scan interpretation and objectivity; therefore, interobserver variability is reduced. Using follicular volume obtained with sono AVC as the measure of follicular growth combined with volume-based criteria for the hCG triggering may in the future improve the treatment outcome compared to that achieved with conventional monitoring with follicular diameter. Better knowledge in this area could be helpful to optimize IVF outcome, by refining ovarian stimulation protocols and obtain high quality oocytes.

Study Question: What is the clinical efficacy of an oocyte donation program based on the transportation of frozen semen and embryos between two countries?, Summary Answer: The transnational oocyte donation program is efficient and reliable and it could provide a first-line strategy to overcome the lack of donors in some countries., What Is Known Already: While there is increasing need for donated oocytes, in many countries the availability of donors is still insufficient to cover the therapeutic demands, and patients are referred abroad for treatment. Since embryo cryopreservation is reliable and efficient, we propose a strategy based on frozen embryos instead of frozen oocytes to satisfy the increasing demand for cross border oocyte donation., Study Design, Size, Duration: This is a retrospective cohort study including 630 patients treated from December 2015 to July 2017., Participants/materials, Setting, Methods: Infertile women were treated with elective vitrified-thawed embryo shipping and embryo transfer (ET) between two IVF clinics, one in Spain and one in Italy., Main Results and the Role of Chance: A total of 2617 embryos were created for the 630 patients and the survival rate after warming was 98.5%. After the first ET the live birth rate (LBR) was 30.6%. In 476 patients (75.5%), embryos were transferred at the cleavage stage (Day 2 or 3) and the LBR was 29.2%. Vitrified blastocysts were available for 154 patients (24.5%) and the LBR was 35%. Among patients who did not achieve a pregnancy after the first frozen ET (FET), 92.5% had at least one frozen embryo for successive procedures. 213 patients underwent a second FET. The LBR at the second FET was 30%. The cumulative LBR at the end of the observation period was 39.3%., Limitations, Reasons for Caution: The study design was retrospective. A direct comparison with vitrified oocyte donors cycle and subsequent fresh ET would have permitted to compare this strategy versus the current standard based on vitrified gametes., Wider Implications of the Findings: The LBR found in our study is more than acceptable and seems to be higher than what reported with vitrified oocytes. The transnational fresh oocyte donation program may have several advantages over the shipment of vitrified oocytes: similarly to the fresh oocyte donation program it allows for personalized care in oocyte recipient, which is provided by assigning a flexible number of oocytes, and at the same time it maintains the benefit of a frozen ART program permitting scheduling flexibility. The TOD program is efficient and may be proposed as a first-line strategy for distance and inter-countries oocyte donation programs., Study Funding, Competing Interest(s): None., Trial Registration Number: NA.

Purposes: The aim of this study is to determine whether a clinical advantage is gained with use of LH in combination with FSH or as a component of human menopausal gonadotropin (hMG) to achieve optimal ovarian stimulation., Methods: In this study, we compared retrospectively two regimens, r-FSH/r-LH and hMG, for the treatment of women with reduced ovarian reserve, identified as subjects with antral follicle count (AFC) < 11 and AMH ≤ 1.1 ng/ml., Results: Overall, the clinical pregnancy per started cycle was higher in the r-FSH/r-LH group (12.5 vs. 8.1%, P < 0.02), while implantation (11.1 vs. 9.5%) and miscarriage rates (29.9 vs. 35.9%) were comparable. Data were further analysed performing separate comparisons in subpopulations with different ranges of AFC, i.e. < 4, 4-6 and 7-10. Major differences between the two regimens were observed in women with AFC < 4. In this subpopulation, not only was the clinical pregnancies per started cycle higher in the r-FSH/r-LH group (10.2 vs. 1.5%, P < 0.01), but also implantation was significantly higher (13.0 vs. 2.8%, P < 0.02)., Conclusions: A r-FSH/r-LH regimen appears to be beneficial for the treatment of women with extremely poor ovarian reserve. It should be considered however that, being retrospective, this study is affected by obvious limitations, such as post-treatment patient selection criteria and absence of randomisation.

Backround: The goal of assisted reproduction is for a couple treated with IVF techniques to end the treatment by giving birth to a healthy baby. A neccessary presumption for success is the identification of the best embryo with high implantation and developmental potential. One option is to select an euploid embryo by invasive preimplantaion genetic testing for aneuploidy (PGT-A) or it is possible to select the best embryo by non-invasive time-lapse monitoring (TLM), specifically based on morphokinetic parameters and morphological markers that are able to identify an embryo with high developmental potential., Materials and Methods: The study involved a total of 1060 embryos (585 euploid and 475 aneuploid embryos after PGT-A) with good morphology from 329 patients in the period 01/2016-10/2021. All embryos were cultured in a time-lapse incubator, trophectoderm (TE) cells biopsies for PGT-A examination were performed on day 5 (D5) or day 6 (D6) of culture. During the study period, 225 frozen embryo transfers (FET) of one euploid embryo were performed. Based on the treatment outcome, the embryos were divided into 2 groups - euploid embryos, which led to the birth of a healthy child, and euploid embryos that did not show fetal heartbeat (FHB) after FET., Results: Based on the statistical analysis of the embryos without implantation and the embryos with live birth, it is clear that the morphokinetic parameters t5 (time of division into 5 cells) and tSB (time of start of blastulation) are significantly different., Conclusion: The results suggest that of the morphokinetic parameters tSB and t5 are predictive indicators for selecting an embryo with high developmental potential and with a high probability of achieving the birth of a healthy child., Competing Interests: The authors report no conflicts of interest in this work.

Study Question: Are specific morphological anomalies in human mature oocytes, as revealed by transmitted light microscopy, associated with intrinsic damage to the meiotic spindle and actin cytoskeleton?, Summary Answer: Aggregates of smooth endoplasmic reticulum (SER) and domains of centrally localized granular cytoplasm (GC) reflect intrinsic damage to the oocyte cytoskeleton, namely alterations in spindle size, chromosome misalignment and cortical actin disorganization., What Is Known Already: In preparation for ICSI, oocytes are often selected for use in treatment by morphological criteria, but the rationale and implications of this practice are controversial. Very little information is available on the relationship between oocyte morphology and intrinsic cellular characteristics, such as the actin cytoskeleton, meiotic spindle and chromosome alignment., Study Design, Size, Duration: A total of 170 metaphase II (MII) oocytes were donated by consenting IVF patients and analysed; 62 were classified as morphologically normal (control), 54 had SER clusters and 54 had centrally localized GC., Participants/materials, Setting, Methods: Supernumerary oocytes were fixed within 3 h from recovery and stained for tubulin, chromatin and actin. Spindles were analysed for 1D and 2D characteristics by high-performance confocal microscopy. Chromosomes were classified as scattered or aligned and the conformation and intensity of cortical actin was evaluated., Main Results and the Role of Chance: In comparison with control oocytes, both SER and GC oocytes showed greater spindle length (P = 0.033 and 0.003, respectively) and GC oocytes also showed greater spindle width (P= 0.049) and area (P= 0.036). Control and SER oocytes had statistically comparable rates of chromosome displacement from the metaphase plate, unlike GC oocytes where chromosome displacement occurred at higher rate (P = 0.013). In situations where a complete Z-stack was reconstructed from a polar angle, chromosome disposition was classified as being normal when two sets of concentric arrays were visible. Based on these parameters, the proportions of oocytes with normal chromosomal arrangement or partial/total disarrangement was not statistically different between control and SER oocytes. Conversely, in GC oocytes, chromosome disarrangement was higher (P = 0.002). All control oocytes displayed a continuous meshwork of suboolemmal actin, which appeared as an uninterrupted ring in thin optical sections. In contrast, in SER and  GC groups, integrity of suboolemmal actin was observed in only 66.7 and 42.9% of oocytes, respectively (P = 0.0001)., Large Scale Data: N/A., Limitations Reason for Caution: Only two of several known oocyte dysmorphisms were investigated, while oocyte quality was assessed only by cytoskeletal criteria., Wider Implications of the Findings: This study represents a significant step toward a more objective assessment of oocyte morphology, offering information that can assist embryologists to make a more aware and rationally founded decision on whether, and with what possible implications, oocytes with certain dysmorphic characters should be used for treatment or discarded. More generally, it also demonstrates that morphometric parameters of the cytoskeleton and chromosome organization can be used as biomarkers of oocyte quality., Study Funding and Competing Interest(s): This study was funded by Biogenesi Reproductive Medicine Centre (Monza, Italy). All authors declare no conflict of interests., (© The Author 2017. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com)

Differences in cumulus cell gene expression after oocyte maturation in vitro (IVM) or in vivo have been described in previous studies. However, the possible impact of follicle stage on gene expression deregulation during human oocyte IVM remains unknown. Expression of selected genes of interest was compared in cumulus cell of three classes of human cumulus cell-oocyte complexes (COCs): a) COCs derived from human chorionic gonadotropin (hCG)-triggered IVM cycles, collected at the germinal vesicle (GV) stage from mid-sized follicles (4-12 mm) and matured in vitro (IVM-GV); b) COCs derived from hCG-triggered IVM cycles, collected from mid-sized follicles (4-12 mm) and matured in vivo (IVM-MII); c) COCs derived from controlled ovarian stimulation in vitro fertilization (IVF) cycles, collected from large/preovulatory follicles and matured in vivo (IVF-MII). Overall, mRNA levels of the large majority of the 20 genes of different regulative and metabolic pathways subject to analysis were altered in IVM samples compared with in vivo matured COCs. In some cases, follicle size appeared to have a role in determining transcription deregulation. For example, in comparison to the IVF-MII control, the luteinizing hormone receptor was largely overexpressed in both IVM-GV and IVM-MII COCs, therefore irrespective of IVM. However, in other circumstances follicle size and IVM had distinct and opposite impacts on gene expression, as shown by transcription of amphiregulin, which was increased in IVM-MII COCs, but decreased in COCs matured in vitro (IVM-GV) compared with the IVF-MII control. This study confirms and extends previous data on gene expression dysregulation during IVM and indicates that the size of follicles from which immature oocytes are retrieved can be an independent factor of differential transcriptional regulation.

Study Hypothesis: How does the ultrastructure of human oocytes matured in vitro compare with oocytes collected from women after full hormonal stimulation?, Study Finding: The ultrastructure of human oocytes matured in vitro is largely, but not entirely, similar to those matured in vivo., What Is Known Already: Embryos derived from in vitro-matured oocytes often have limited developmental potential, possibly as an effect of inappropriate in vitro maturation (IVM) conditions. Transmission electron microscopy (TEM) is a valuable research tool to compare in vivo and in vitro matured oocytes. However, previous studies on the ultrastructure of human IVM oocytes were done with inadequate material or inappropriate IVM conditions, and have limited significance., Study Design, Samples/materials, Methods: Immature cumulus cell-enclosed oocytes, retrieved from mid-sized antral follicles of women requiring IVM treatment, were matured in vitro for 30 h. No leftover germinal vesicle-stage oocytes collected from fully stimulated cycles were used. Control in vivo matured oocytes were obtained from age-matched women undergoing full ovarian stimulation. In vitro and in vivo matured oocytes were analysed by TEM and compared according to previously established morphometric criteria of oocyte quality., Main Results and the Role of Chance: All oocytes had normal ooplasm showing uniform distribution of organelles. Mitochondrial morphology appeared similar between the maturation conditions. Cortical granules were found typically stratified in a single, mostly continuous row just beneath the ooplasm in all oocytes. Microvilli were well preserved after IVM. Vacuoles were only occasionally found in all oocytes and, if present, they were frequently associated with lysosomes. Mitochondria-smooth endoplasmic reticulum (M-SER) aggregates and mitochondria-vesicles (MV) complexes were commonly found in in vivo matured oocytes. However, large MV complexes partially replaced M-SER aggregates in IVM oocytes., Limitations, Reasons for Caution: As a note of caution it should be noticed that, being laborious and technically demanding, TEM cannot be applied to a large number of samples in a single investigation. Therefore, our data require further independent confirmation., Wider Implications of the Findings: Our data suggests the notion that TEM remains a valuable research tool that can also offer quantitative data if associated with morphometric criteria of evaluation. Therefore, it can be adopted to test pre-clinically the performance of novel in vitro systems that are demanded to make oocytes IVM more successful in the human., Large Scale Data: Not applicable., Study Funding and Competing Interests: This study was independently funded by Biogenesi Reproductive Medicine Centre, Monza, Italy. All authors declare that their participation in the study did not involve factual or potential conflicts of interests., (© The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Purpose: In in vitro maturation (IVM) cycles primed with human chorionic gonadotropin (hCG), both immature and mature oocytes are retrieved from antral follicles sized 8-12 mm. Using time-lapse microscopy, we compared the morphokinetic behavior of embryos developed from oocytes matured in vivo and in vitro, testing the hypothesis that IVM affects preimplantation development. Furthermore, we extended the morphokinetic analysis of these embryos by a comparison with embryos obtained in stimulated assisted reproduction technology (ART) cycles., Methods: In IVM cycles primed with follicle-stimulating hormone (FSH)/hCG, prior to sperm microinjection, oocytes surrounded by an expanded cumulus at retrieval and presumably mature (EC-MII) were incubated for 6 h, while immature oocytes enclosed in a compact cumulus (CC) were matured in vitro for 30 h. The morphokinetics of embryos selected for transfer or cryopreservation, derived from EC-MII and CC oocytes, were comparatively and retrospectively analyzed in terms of cleavage times (t2, t3, t4, t5, and t8) and intervals (cc2, cc3, s2, s3). For further comparison, the morphokinetics of embryos selected for transfer or cryopreservation (ICSI) or giving rise to ongoing pregnancies (model) in stimulated ART cycles was also assessed., Results: The morphokinetic behavior of EC-MII and CC embryos was entirely comparable, as suggested by the absence of statistical differences in the averages of all cleavage times and intervals. Almost all cleavage and interval times were also similar between EC-MII, CC, ICSI, and model groups, with the exception of t4 and s2, which were delayed and longer, respectively, in embryos generated in IVM cycles (EC-MII and CC)., Conclusions: These findings do not support the hypothesis that maturation in vitro affects embryo morphokinetics, while they suggest only marginal differences in the morphokinetics of embryos developed from oocytes matured in vivo and in vitro in IVM cycles and embryos developed from mature oocytes recovered in stimulated cycles.

Purpose: Only 50-60 % of immature human oocytes attain the mature stage in vitro. Such a deficiency may be a reflection of inadequate conditions of in vitro maturation (IVM) or a manifestation of intrinsic oocyte defects. In the present study, we explored the possibility that the DNA of immature oocytes may be damaged and that such a condition, or inability to trigger a repair action, is associated to germinal vesicle (GV) arrest., Methods: Immature oocytes (GV-stage oocytes) were obtained from women undergoing stimulated (Stim-C) or IVM (IVM-C) cycles. GV oocytes obtained from stimulated cycles were fixed for successive analysis either after recovery (T0) or following 30 h (T30) of culture if still arrested at the GV stage. Oocytes retrieved in IVM cycles were used only if they were found arrested at the GV stage after 30 h (T30) of culture. All oocytes were fixed and stained to detect chromatin and actin. They were also assessed for positivity to γH2AX and Rad51, markers revealing the presence of double-strand DNA breaks and the activation of a DNA repair response, respectively. Labelled oocytes were analysed using a Leica TCS SP2 laser scanning confocal microscope., Results: In Stim-C oocytes, γH2AX positivity was 47.5 and 81.5 % in the T0 and T30 groups, respectively (P = 0.003), while γH2AX-positive oocytes were 58.3 % in the IVM-C T30 group (Stim-C T0 vs. IVM-C T30, P = 0.178; Stim-C T30 vs. IVM-C T30, P = 0.035). Positivity for nuclear staining to Rad51 occurred in 42.1 and 74.1 % of Stim-C in the T0 and T30 subgroups, respectively (T = 0.006), while 66.7 % of IVM-C T30 oocytes resulted positive for a DNA repair response (Stim-C T0 vs. IVM-C T30, P = 0.010; Stim-C T30 vs. IVM-C T30, P = 0.345)., Conclusions: The present data document the existence of double-strand DNA breaks (DSBs) in human immature oocytes. Also, they are consistent with the hypothesis that insults to DNA integrity may be an important factor affecting meiotic resumption.

Purpose: To assess the effects of oocyte central granularity and its underlying endocrine environment on developmental competence of dysmorphic and morphologically normal oocytes., Methods: Retrospective cohort study including 1,082 patients undergoing autologous ICSI cycles. Of these, 211 patients provided 602 oocytes with central granularity (CG) and 427 morphologically normal cycle companion oocytes (NCG). The remaining 871 patients provided only morphologically normal oocytes in cycles not yielding dysmorphic oocytes (N). Patient profile associated with CG was characterized, and fertilization rates, early morphokinetics and live birth rates were compared between N, CG and NCG groups. Patient characteristics associated with implantation and delivery performance of CG-derived embryos were assessed., Results: CG was associated with higher maternal age, basal FSH concentrations and total FSH dose, but with lower circulating AMH (p ≤ 0.035). Fertilization rates were reduced and early morphokinetic parameters were delayed in CG (p < 0.025) and NCG (p < 0.05) groups as compared to the N group. Embryos derived from CG oocytes achieved a markedly lower live birth rate (14.9%) as compared to those derived from NCG (36.8%; p = 0.03) and N oocytes (29.8%; p = 0.002). The negative relationship between CG and live birth was confirmed by a multivariate analysis controlling for potential confounders (OR:2.59, IC:1.27-5.31; P = 0.009). Implantation and delivery rates following transfers of CG-derived embryos were inversely associated with maternal age., Conclusion: CG oocytes, but not their morphologically normal cycle companions, have severely compromised developmental competence. Maternal age should be a key parameter in deciding whether or not to utilize CG oocytes in ICSI cycles., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: In a growth phase occurring during most of folliculogenesis, the oocyte produces and accumulates molecules and organelles that are fundamental for the development of the preimplantation embryo. At ovulation, growth is followed by a phase of maturation that, although confined within a short temporal window, encompasses modifications of the oocyte chromosome complement and rearrangements of cytoplasmic components that are crucial for the achievement of developmental competence. Cumulus cells (CCs) are central to the process of maturation, providing the oocyte with metabolic support and regulatory cues., Methods: PubMed was used to search the MEDLINE database for peer-reviewed original articles and reviews concerning oocyte maturation in mammals. Searches were performed adopting 'oocyte' and 'maturation' as main terms, in association with other keywords expressing concepts relevant to the subject. The most relevant publications, i.e. those concerning major phenomena occurring during oocyte maturation in established experimental models and the human species, were assessed and discussed critically to offer a comprehensive description of the process of oocyte maturation., Results: By applying the above described search criteria, 6165 publications were identified, of which 543 were review articles. The number of publications increased steadily from 1974 (n = 7) to 2013 (n = 293). In 2014, from January to the time of submission of this manuscript, 140 original manuscripts and reviews were published. The studies selected for this review extend previous knowledge and shed new and astounding knowledge on oocyte maturation. It has long been known that resumption of meiosis and progression to the metaphase II stage is intrinsic to oocyte maturation, but novel findings have revealed that specific chromatin configurations are indicative of a propensity of the oocyte to resume the meiotic process and acquire developmental competence. Recently, genetic integrity has also been characterized as a factor with important implications for oocyte maturation and quality. Changes occurring in the cytoplasmic compartment are equally fundamental. Microtubules, actin filaments and chromatin not only interact to finalize chromosome segregation, but also crucially co-operate to establish cell asymmetry. This allows polar body extrusion to be accomplished with minimal loss of cytoplasm. The cytoskeleton also orchestrates the rearrangement of organelles in preparation for fertilization. For example, during maturation the distribution of the endoplasmic reticulum undergoes major modifications guided by microtubules and microfilaments to make the oocyte more competent in the generation of intracellular Ca(2+) oscillations that are pivotal for triggering egg activation. Cumulus cells are inherent to the process of oocyte maturation, emitting regulatory signals via direct cell-to-cell contacts and paracrine factors. In addition to nurturing the oocyte with key metabolites, CCs regulate meiotic resumption and modulate the function of the oocyte cytoskeleton., Conclusions: Although the importance of oocyte maturation for the achievement of female meiosis has long been recognized, until recently much less was known of the significance of this process in relation to other fundamental developmental events. Studies on chromatin dynamics and integrity have extended our understanding of female meiosis. Concomitantly, cytoskeletal and organelle changes and the ancillary role of CCs have been better appreciated. This is expected to inspire novel concepts and advances in assisted reproduction technologies, such as the development of novel in vitro maturation systems and the identification of biomarkers of oocyte quality., (© The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Purpose: The aim of this retrospective study was to compare the competence of oocytes obtained from preovulatory and antral follicles., Methods: Mature oocytes from preovulatory follicles were retrieved from women selected for standard IVF treatment (Group A). Mature oocytes from antral follicles were recovered from women undergoing hCG-primed in vitro maturation (IVM) treatment (Group B). Patients groups were matched for age, BMI, FSH, AMH and antral follicle count (AFC) values. In vivo matured oocytes from both groups were microinjected and resulting embryos were culture and selected on day 3 for embryo transfer., Results: Oocyte pick-ups (OPU) were 315 and 204 in Groups A and B, respectively. Fertilization rates were comparable (72.8 and 75.9 %, respectively; P = 0.137). In Group A, in which the average number of embryos transferred was higher, clinical pregnancy rates per OPU (37.5 %) and embryo transfer (38.4 %) were superior in comparison to Group B (27.0 %, P = 0.013; 29.4 %, P = 0.041; respectively). On the other hand, implantation rates (Group A, 23.7 %; Group B, 20.8 %) and proportions of babies born per transferred embryo (Group A, 19.5 %; Group B, 16.9 %) were similar (P = 0.528 and 0.332, respectively)., Conclusions: Overall, this suggests that oocyte competence is already achieved at the antral stage of follicle development.

Background: Sterigmatocystin (STE) is a mycotoxin widely found in contaminated food and foodstuffs, and excessive long-term exposure to STE is associated with several health issues, including infertility. However, there is little information available regarding the effects of STE toxin on the female reproductive system, particularly concerning oocyte maturation. Methods: In the present study, we investigated the toxic effects of STE on mouse oocyte maturation. We also used Western blot, immunofluorescence, and image quantification analyses to assess the impact of STE exposure on the oocyte maturation progression, mitochondrial distribution, oxidative stress, DNA damages, oocyte ferroptosis and asymmetric division defects. Results: Our results revealed that STE exposure disrupted mouse oocyte maturation progression. When we examined the cellular changes following 100 µM STE treatment, we found that STE adversely affected polar body extrusion and induced asymmetric division defects in oocytes. RNA-sequencing data showed that STE exposure affects the expression of several pathway-correlated genes during oocyte meiosis in mice, suggesting its toxicity to oocytes. Based on the RNA-seq data, we showed that STE exposure induced oxidative stress and caused DNA damage in oocytes. Besides, ferroptosis and α-tubulin acetylation were also found in STE-exposed oocytes. Moreover, we determined that STE exposure resulted in reduced RAF1 protein expression in mouse oocytes, and inhibition of RAF1 activity also causes defects in asymmetric division of mouse oocytes. Conclusions: Collectively, our research provides novel insights into the molecular mechanisms whereby STE contributes to abnormal meiosis. [ABSTRACT FROM AUTHOR]

Simple Summary: In the last years, many domesticated and wild animal species, as well as local and transboundary breeds, have been threatened with genetic depletion and extinction. The application of conservation strategies to sheep breeds is necessary to safeguard the productive characteristics but also the historical and cultural value that they exert on their territory and inhabitants. Despite many advances that have been made in gamete cryopreservation, in vitro embryo production (IVEP) and juvenile in vitro production (JIVET), the efficiency of these technologies is still low, and the application is restricted in the ovine species. In the present study, a high concentration-rapid exposure (HC-RE) and a low concentration-slow exposure (LC-SE) vitrification protocol, using dimethyl sulfoxide (DMSO) and ethylene glycol (EG) as permeating cryoprotectants (CPAs), were applied to pre-pubertal lamb immature cumulus–oocyte complexes (COCs) and evaluated on nuclear and cytoplasmic parameters after in vitro maturation (IVM). Slightly more encouraging results were observed with the LC-SE vitrification protocol, leading to the hypothesis that low CPA concentrations in association with prolonged exposure times could be more promising to pursue in order to improve pre-pubertal lamb immature COC vitrification. Oocyte vitrification allows for the storing of endangered breed female gametes. Cryoprotectant (CPA) concentration and exposure time should ensure cell protection with minimal toxicity. In the present study, a high concentration-rapid exposure (HC-RE) and a low concentration-slow exposure (LC-SE) vitrification protocol, using dimethyl sulfoxide (DMSO) and ethylene glycol (EG) as permeating CPAs, were evaluated on meiotic competence and bioenergetic-oxidative status of pre-pubertal lamb immature COCs after in vitro maturation (IVM). For each protocol, COCs vitrified through a traditional protocol and fresh ones were used as controls. Both protocols allowed COC morphology preservation after vitrification-warming (V-W) and cumulus expansion after IVM. The maturation rate (7% and 14%) was comparable to the vitrified control (13% and 21%) but not satisfactory compared to fresh ones (58% and 64%; p < 0.001). The rate of mature oocytes displaying a perinuclear/subcortical (P/S) mitochondrial distribution pattern, an index of cytoplasmic maturity, was comparable between vitrified and fresh oocytes. The LC-SE vitrification protocol did not affect quantitative bioenergetic-oxidative parameters compared to both controls whereas HC-RE protocol significantly reduced intracellular reactive oxygen species (ROS) levels, indicating cell viability loss. In conclusion, to improve pre-pubertal lamb immature COC vitrification, the combination of low CPA concentrations with prolonged exposure time could be more promising to investigate further. [ABSTRACT FROM AUTHOR]