A Pilot Analysis of Whole Transcriptome of Human Cryopreserved Sperm.

Sperm cryopreservation is a procedure widely used to store gametes for later use, to preserve fertility in patients prior to gonadotoxic treatments or surgery, and for sperm donation programs. The purpose of the study was to assess the impact of cryopreservation on human sperm transcriptome. Semen samples were collected from 13 normospermic men. Each sample was divided into two aliquots. The total RNA was immediately extracted from one aliquot. The second aliquot was frozen and total RNA was extracted after a week of storage in liquid nitrogen. The RNA samples were randomized in four pools, each of six donors, and analyzed by microarrays. The paired Significance Analysis of Microarray was performed. We found 219 lower abundant transcripts and 28 higher abundant transcripts in cryopreserved sperm than fresh sperm. The gene ontology analysis disclosed that cryopreservation alters transcripts of pathways important for fertility (i.e., spermatogenesis, sperm motility, mitochondria function, fertilization, calcium homeostasis, cell differentiation, and early embryo development), although the increase of some transcripts involved in immune response can compensate for the harmful effects of freezing.

Algorithm vs. clinical experience: controlled ovarian stimulations with follitropin-delta and individualised doses of follitropin-alpha/beta.

In the registrational trials, follitropin delta was compared with a fixed dose of 150 UI of follitropin alpha/beta, finding higher chances to reach a target response of 8-14 oocytes compared to controls. For this reason, follitropin delta is marketed as particularly useful in expected hyper-responder patients. The main outcome of this study is to report if comparable results are reached in a real-life scenario with follitropin alpha/beta personalized doses, based on patients' characteristics. This is a retrospective study performed in two public fertility centres. All first cycles from January 2020 to June 2022 with either follitropin delta (cases) or alpha/beta (controls) in patients with antiMüllerian hormone >2.5 ng/ml were compared by an inverse probability weighting approach based on propensity score. The follitropin total dose was higher in controls (1179.06 ± 344.93 vs. 1668.67 ± 555.22 IU, p<0.001). The target response of 8-14 oocytes was reached by 40.2% of cases and 40.7% of controls (odds ratio (OR) 0.99, 95% confidence interval (CI) 0.65-1.53, p=0.98). Fewer than 8 oocytes were collected in 24.1% of cases and 22% of controls (OR 1.10, 95% CI 0.71-1.69, p=0.67); more than 14 oocytes in 35.7% of cases and 37.3% of controls (OR 0.83, 95% CI 0.54-1.28, p=0.40). Our experience did not find worse results in term of proportion of patients who reached the target response with an algorithm-chosen dose of follitropin delta compared to a personalised starting dose of follitropin alpha/beta, with follitropin delta having the advantage of objectivity. Larger numbers are needed to confirm these results.

Oxidative Stress: The Role of Estrogen and Progesterone.

The effect of estrogen and progesterone on oxidative status is not yet very clear, improvements and detrimental effects having been reported with the use of menopausal hormone therapy or hormonal contraceptives, respectively. In this study, we evaluated the role played by estrogen and progesterone separately, on the oxidative status of 32 women, 18 to 43 years old, by inducing high levels of estrogen and then adding high levels of progesterone. During a cycle of in vitro fertilization, blood samples were collected prior to gonadotrophin stimulation (low estradiol levels), on the day of oocyte retrieval (high levels of estrogen), and on the day of embryo transfer (high levels of estrogen and progesterone). Total blood levels of oxidants (FORT), antioxidants (FORD), and their ratio FORT/FORD were measured using a colorimetric method based on the Fenton reaction. Seven women measured their early morning body temperature at the same time points. FORT significantly decreased from the low- to the high-estrogen phase (p = 0.023) and increased from the high-estrogen to the high-estrogen-progesterone phase (p = 0.006). FORD showed an opposite but non-significant trend. The FORT/FORD ratio decreased from the low- to the high-estrogen phase (p = 0.0104) and increased from the high-estrogen to the high-estrogen -progesterone phase (p = 0.004). Body temperature (n = 7) decreased in the high-estrogen phase (p = 0.001) and increased from the high-estrogen to the high-estrogen-progesterone phase (p = 0.001). In the seven women, FORT (p = 0.009) and FORT/FORD (p = 0.0056) were linearly related to body temperature values. Our data show opposite effects of estrogen and progesterone on oxidative status. These effects seem to be related to the effect exerted on body temperature regulation.

Optimizing the "Time to pregnancy" in women with multiple sclerosis: the OPTIMUS Delphi survey.

Background: The debate on how to manage women affected by multiple sclerosis (MS) during reproductive age is still open, as is the issue of fertility in such patients. Main issue regard the identification of the optimal window for pregnancy and how to deal with medical therapy before and during conception. The aim of this Delphi consensus was to collect the opinions of a multidisciplinary group, involving reproductive medicine specialists and neurologists with experience in the management of multiple sclerosis women with reproductive desire. Methods: Four experts plus scientific coordinators developed a questionnaire distributed online to 10 neurologists and later discussed the responses and amended a list of statements. The statements were then distributed via an online survey to 23 neurologists (comprising the first 10), who voted on their level of agreement/disagreement with each statement. Consensus was achieved if agreement or disagreement with a statement exceeded 66%. Results: Twenty-one statements reached consensus after two rounds of voting, leading to the following main recommendations: (1) Fertility evaluation should be suggested to wMS, in case of the need to shorten time to pregnancy and before treatment switch in women on DMTs contraindicated in pregnancy, particularly in case of highly active disease and age > 35 years. (2) ART should not be discouraged in wMS, but the use of DMTs until pregnancy confirmation should be suggested; ART may be considered in order to reduce time to pregnancy in MS women with a reduced ovarian reserve and/or age > 35 years, but in case of an expected poor ART prognosis and the need for more than one ART cycle, a switch to a high-efficacy DMD before ART should be offered. (3) Oocyte cryopreservation may be considered in women with reduced ovarian reserve, with unpredictable time to complete diagnostic workup and achieve disease control; a risk/cost-benefit analysis must be performed in women >35 years, considering the diminished ovarian reserve. Conclusion: This consensus will help MS neurologists to support family planning in wMS, respecting MS therapeutic needs while also taking into account the safety and impact of advancing age on fertility.

Does SARS-CoV-2 Affect the Prostate? A Molecular Analysis from a Case Series of COVID-19 Patients.

Severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2) causes COVID-19 that has been spreading worldwide since December 2019. Viral entry into cells requires expression of both angiotensin-converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) on the surface of the host cell. The male reproductive system, including the prostate, was supposed to be a potential target for SARS-CoV-2 since the presence of ACE and TMPRS2 receptors. This paper investigated for the first time the presence of SARS-CoV-2 mRNA in the prostatic tissue of a patient with active infection. In addition, we searched for the virus in the prostate of five patients after their recovery from COVID-19. The SARS-CoV-2 RNA was not detected in any of the prostate tissues tested even during the acute phase of infection. As case series have limitations, causality cannot be excluded and sporadic evidence of prostatic tissue invasion by SARS-CoV-2 may be detectable.

Collecting semen samples at home for fertility assessment: time for a new standard?

BACKGROUND: In fertility clinics the standard approach to semen collection involves a private room close to the laboratory to avoid fluctuations in temperature and to control the time between collection and processing. There are still no firm conclusions whether collecting semen at home has any influence on sperm quality and reproductive competence. The purpose of this study was to assess whether the site of semen collection affects semen parameters. METHODS: This retrospective cohort study performed at a tertiary level public fertility center included 8634 semen samples from 5880 men undergoing fertility assessment from 2015 to 2021. The impact of sample collection site was evaluated using a generalized linear mixed model. A subgroup analysis comparing clinic to home collection within the same patient was performed on 1260 samples from 428 men by paired t-test or Wilcoxon Signed Rank Test. RESULTS: Samples collected at home (N.=3240) had significantly higher semen volume, sperm concentration and total sperm count respect to samples collected at clinic (N.=5530) (median (range): 2.9 (0.0-13.9) mL versus 2.9 (0.0-11.5) mL, P=0.016; 24.0 (0.0-252.0) million/mL versus 18.0 (0.0-390.0), P<0.0001; 64.6 (0.0-946.0) million versus 49.3 (0.0-1045.0), P<0.0001, respectively). There was no difference in abstinence period and sperm motility. Paired comparisons of semen characteristics in 428 patients with home-collected (N.=583) and clinic-collected (N.=677) samples confirmed a no negative effect on volume and total sperm count. CONCLUSIONS: Our data provide evidence for a not disadvantage with collection at home.

Luteal phase support in assisted reproductive technology centers: Italian survey.

BACKGROUND: In assisted reproductive cycles (ART), the fine balance of controlling corpus luteum function is severely disrupted. To challenge this iatrogenic deficiency, clinicians aim to provide exogenous support. Several reviews have investigated progesterone route of administration, dosage and timing. METHODS: A survey about luteal phase support (LPS) after ovarian stimulation was conducted among doctors in charge in Italian II-III level ART centers. RESULTS: With regards to the general approach to LPS, 87.9% doctors declare to diversify the approach; the reasons for diversifying (69.7%) were based on the type of cycle. For all the most important administration routes (vaginal, intramuscular, subcutaneous) it appears that in frozen cycles there is a shift towards higher dosages. The 90.9% of the centers use vaginal progesterone, and when a combined approach is required, in 72.7% of cases vaginal administration is combined with injective route of administration. When Italian doctors were asked about the beginning and duration of LPS, 96% of the centers start the day of the pickup or the day after, while 80% of the centers continue LPS until week 8-12. The rate of participation of the centers confirms the low perceived importance of LPS among Italian ART centers, while may be considered quite surprising the relatively higher percentage of centers that measures P level. Tailorization to women's needs is the new objective of LPS: self-administration, good tolerability are the main aspects for Italian centers. CONCLUSIONS: In conclusion, results of Italian survey are consistent to results of main international surveys about LPS.

Telehealth for infertile patients during SARS-CoV-2 pandemic: far, and yet close.

BACKGROUND: In Italy during the first pandemic wave of SARS-CoV-2 the activity of fertility centers was stopped, with the exception of fertility preservation in oncological patients. We adopted telehealth and we evaluated whether it could help in the management of infertile couples at a fertility center. METHODS: A longitudinal study performed at a public fertility center. Telehealth was offered to 72 couples referred to our center for a first consultation from March 17th to May 31st, 2020. Percentage of patients who performed the first assisted reproduction technology (ART) cycle or intrauterine insemination (IUI) within 6 months from the first visit and drop-out rate were analyzed during COVID-19 pandemic and compared to historical controls (couples admitted to our center in 2017-2019). RESULTS: Eighty-five (61/72) percent of the couples accepted telehealth. Time to first treatment after online consultation in telehealth group (4.5±1.8 months) was significantly shorter (P=0.033) respect to time to first treatment after face-to-face visit of historical controls (7.5±6.9 months). After telehealth consultation, we observed a significant reduction (P=0.002) of drop-out rate from 39% in historical controls to 17% of telehealth group. Telehealth significantly diminished the drop-out rate also during the COVID-19 pandemic respect to 73% after traditional face-to-face visits (P=0.0005), with a time to first treatment of 3.7±2.1 months in couples who refused telehealth. CONCLUSIONS: Telehealth could be a useful tool to facilitate the path of patients in a fertility center.

mRNA and Viral Vector COVID-19 Vaccines Do Not Affect Male Fertility: A Prospective Study.

PURPOSE: To assess whether mRNA and viral vector coronavirus disease 2019 (COVID-19) vaccines detrimentally affected semen parameters. MATERIALS AND METHODS: In this prospective study, we enrolled 101 men vaccinated for COVID-19 (76% received mRNA vaccines, 20% viral vector vaccines, 2% a mixed formulation, and for 2 men no information about vaccine type was available) in 2021 and with a previous semen analysis. For each man we compared semen parameters before and after vaccination. RESULTS: Post-vaccine samples were obtained at a median of 2.3±1.5 months after the second dose. After vaccination, the median sample volume significantly decreased (from 3.0 to 2.6 mL, p=0.036), whereas the median sperm concentration, the progressive motility, and total motile sperm count increased (from 25.0 to 43.0 million/mL, p<0.0001; from 50% to 56%, p=0.022; from 34.8 to 54.6 million, p<0.0001, respectively). Thirty-four patients were oligospermic before the vaccine, and also in these patients we observed a significant increase of sperm parameters after vaccine. Finally, we confirmed the aforementioned results in men who received a mRNA or a viral vector vaccine. CONCLUSIONS: The semen parameters following COVID-19 vaccination did not reflect any causative detrimental effect from vaccination, and for the first time we demonstrated that this applies to both mRNA and viral-vector vaccines. The known individual variation in semen and the reduced abstinence time before the post-vaccine sample collection may explain the increases in sperm parameters.

Contraception and Hormone Replacement Therapy in Healthy Carriers of Germline BRCA1/2 Genes Pathogenic Variants: Results from an Italian Survey.

Several myths and misconceptions exist about hormones in women with familial predisposition to cancer, and there are few real-life data on their prescription and uptake. To better understand how they are prescribed and accepted in healthy carriers of a BRCA1/2 pathogenetic variant, an online survey was uploaded on Google Forms and shared through social media closed groups of patients' associations, aBRCAcadabra and ACTO Campania. A total of 241 questionnaires were collected. Sexual quality of life was considered of the utmost importance by most of the respondents (mean score of 7 ± 2.8/10), but they felt the counseling they received by healthcare professionals on the topic was insufficient (4.9 ± 3.2/10). Only 57 women out of 233 (24.5%) had used hormonal contraception after being diagnosed as carriers of a BRCA pathogenetic variant, and 42 out of 148 (28.4%) underwent menopause hormonal therapy. The majority of women (53.6% for contraception and 61.5% for menopause) reported being dissatisfied with the counseling received, and 58.2% were not aware of the protective effect of hormonal contraception on the risk of ovarian cancer. An educational effort is desirable to guarantee healthy BRCA carriers reliable contraception and evidence-based menopause counseling.

Hyaluronic acid-sperm selection significantly improves the clinical outcome of couples with previous ICSI cycles failure.

BACKGROUND: Beneficial effects of hyaluronic acid (HA)-based selection of spermatozoa for intracytoplasmic sperm injection (ICSI) are still controversial, and further studies are needed to categorize patients that might benefit from such a method. OBJECTIVE: We investigated whether HA sperm selection improved ICSI outcome of couples with previous ICSI cycle failure. MATERIALS AND METHODS: In this retrospective study, we prospectively collected data of (i) Group 1: 96 couples who performed one failed ICSI cycle ("1st procedure," n = 96) followed by another ICSI cycle ("2nd procedure," n = 101); ii) Group 2: 87 couples who performed one failed ICSI cycle (n = 87) followed by an HA-ICSI cycle (n = 104). Differences between procedures and groups were measured by paired and independent statistical tests, respectively. A generalized linear mixed model analyzed the effect of procedure on the outcomes and the interaction between procedures and groups. RESULTS: Injection of HA-bound sperm significantly improved cleavage rate with respect to standard ICSI (p = 0.026). No evolutive pregnancies were obtained in the 1st ICSI attempts. The 2nd ICSI cycles resulted in successfully seven pregnancies. In HA-ICSI cycles, the better quality of embryos with respect to ICSI (p = 0.034) increased the choice of day 5 embryo transfer (p = 0.030), which resulted in successfully 28 pregnancies. No differences were observed in clinical outcomes of the two ICSI procedures in Group 1, while pregnancy and implantation rates were significantly higher in HA-ICSI with respect to ICSI cycles (p = 0.001, p < 0.0001, respectively). No negative perinatal outcomes were recorded. DISCUSSION: In couples where previous 1st ICSI failed, selection of HA-bound spermatozoa significantly improved clinical outcomes with respect to further standard ICSI. CONCLUSION: This study identified couples with previous ICSI cycles failure as a category of infertile patients that really may benefit from HA sperm selection before ICSI.

Semen parameters and male reproductive potential are not adversely affected after three or more months of recovery from COVID-19 disease.

Background: The male reproductive system may be a potential target for SARS-CoV-2 since the presence of ACE and TMPRS2 receptors. After a first report of the presence of SARS-CoV-2 in semen of COVID-19 patients, several papers reported that SARS-CoV-2 was not detected in the semen. However, some evidences indicated that COVID-19 disease could impair semen parameters. During the infection, or in a short period after, a reduction in sperm concentration and motility and an increase in DNA fragmentation were observed, even in asymptomatic patients. There is no conclusive data exploring whether this damage changes with time. We investigated whether COVID-19 disease has a negative impact on semen parameters and male reproductive potential after recovery. Methods: In this longitudinal retrospective study, we enrolled 20 men who had COVID-19 disease. We compared sperm parameters in samples collected before COVID-19 and after infection (8.3 ± 4.8 months). We also evaluated the reproductive potential in pre- and post-COVID-19 infertility treatments of 8 self-controlled couples as well as in 40 cycles after COVID-19 infection of the male partner. Results: For most patients, we obtained results of more than one semen analysis before and after COVID-19. After adjusting for age, days of sexual abstinence, frequency of ejaculations and presence of fever, we found no significant difference over time in any semen parameter. The interval between COVID-19 infection and subsequent infertility treatments was 10.7 ± 7.5 months. There were no differences in the embryological and clinical outcomes of infertility treatments performed before and after male infection. One couple obtained a single pregnancy in the post COVID-19 IUI. Normal fertilization (65%), cleavage (99%) and blastocyst development (40%) rates in treatments performed after male infection were within the expected range of competencies. A total of 5 singleton and 1 twin clinical pregnancies were obtained, and 6 healthy children were born. A total of 10 blastocysts have been cryopreserved. Conclusion: Our data are reassuring that COVID-19 disease has no negative effect on semen quality and male reproductive potential when semen samples are collected three months or more after infection.

Impact of the COVID-19 pandemic on voluntary terminations of pregnancy in an Italian metropolitan area.

PURPOSE: The COVID-19 pandemic and national lockdown from 9 March to 4 May 2020 changed social, familial, and sexual relationships, as well as how citizens interact with the health services. How these profound changes have affected sexuality, contraception and voluntary terminations of pregnancy is still largely undescribed. The main objective of this study was therefore to find out whether the COVID-19 pandemic and ensuing lockdown affected the demand for legal abortion. MATERIAL AND METHODS: The study period was divided into three phases: the pre-pandemic (January and February 2020); lockdown (March and April); and post-lockdown (May and June). The number and characteristics of women requesting pregnancy termination each month during that time were compared with the stats for the same months in the preceding three years (2017-2019). RESULTS: Immediately after national lockdown, the number of voluntary abortions markedly declined (-40.45%). The effect was more evident in women below 20 years of age (-66.67%), employed versus unemployed women (-42.71% vs. -21.05), and non-Italian versus Italian citizens (-53.01 vs. -32.85). No difference was found in the mean time from request to execution of the procedure, or in the type of the procedure used. CONCLUSION(S): National lockdown reduced the number of unwanted pregnancies, especially in younger women. The Italian health service's response to the demand appears to have been unaffected by the pandemic. However, as the demand for abortion is still high, probably due to unplanned pregnancies among cohabitants within a stable relationship, contraception guidance should be improved among women traditionally deemed low-risk in terms of sexual behaviour.

Effect of Multiple Sclerosis and Its Treatments on Male Fertility: Cues for Future Research.

Multiple sclerosis is a chronic disease that may lead to different types of symptoms and disabilities. with the better quality of life and decreased disability due to early diagnosis and the availability of disease-modifying therapies (DMTs), the treating physician is increasingly asked to counsel patients on its effects on fertility and reproduction. In particular, reproductive issues are still scarcely studied and discussed in men. Among the still open questions are the following: (a) Does multiple sclerosis cause infertility per sè? (b) Is multiple sclerosis correlated with conditions that increase the risk of infertility? (c) Do DMTs or other therapies for multiple sclerosis impact gonadal function in men? The aim of this review is to provide an overview on the available literature data about the reproductive issues unique to men with multiple sclerosis, underlining the numerous areas where evidence is lacking and, therefore, the priorities for future research.

Reproductive issues in carriers of germline pathogenic variants in the BRCA1/2 genes: an expert meeting.

BACKGROUND: Healthy individuals and patients with cancer who are carriers of germline pathogenic variants in the BRCA1/2 genes face multiple reproductive challenges that require appropriate counseling and specific expertise. MAIN BODY: On December 5th-7th, 2019, patient advocates and physicians with expertise in the field of reproductive medicine, fertility preservation, and oncology were invited to "San Giuseppe Moscati" Hospital in Avellino (Italy) for a workshop on reproductive management of women with germline pathogenic variants in the BRCA1/2 genes. From the discussion regarding the current evidence and future prospective in the field, eight main research questions were formulated and eight recommendations were developed regarding fertility, fertility preservation, preimplantation genetic testing, and pregnancy in healthy carriers and patients with cancer. CONCLUSION: Several misconceptions about the topic persist among health care providers and patients often resulting in a discontinuous and suboptimal management. With the aim to offer patient-tailored counseling about reproductive issues, both awareness of current evidences and research should be promoted.

Oral Antioxidant Treatment of Men Significantly Improves the Reproductive Outcome of IVF Cycles.

Some 30% to 80% of male sub-fertility may be associated with oxidative stress that damages spermatozoa and can decrease success of in vitro fertilization (IVF) techniques. This multicenter, longitudinal, prospective study aimed to investigate whether oral antioxidant supplementation improved the reproductive competence of men who had had low fertilization rates in their previous intracytoplasmic sperm injection (ICSI) cycles without azoospermia or severe oligozoospermia or any identifiable andrological disease. Seventy-seven men from couples who had an ICSI attempt with unexplained <60% fertilization rate took an antioxidant mix of myo-inositol, alpha-lipoic acid, folic acid, coenzyme Q10, zinc, selenium, and vitamins B2, B6, and B12. Semen parameters were analyzed before (T0) and after 90 days (T90) of treatment, and outcomes of the paired T0 and T90 cycles were compared. After the treatment there was an increase in sperm concentration (p = 0.027), total motile sperm count (p = 0.003), progressive motility (p < 0.0001), and a decreasing trend of DNA-fragmented spermatozoa. Embryological outcomes (fertilization, embryo quality, blastocyst development) were significantly higher in T90 than T0 cycles. No T0 cycle resulted in an evolutive pregnancy. Conversely, in T90 cycles 29 singleton clinical pregnancies were obtained. No negative neonatal outcomes were recorded in newborns after antioxidant treatment. Diet supplementation of men who have had low fertilization rates in their previous ICSI cycles with a combination of myo-inositol, alpha-lipoic acid, folic acid, coenzyme Q10, zinc, selenium, betaine, and vitamins may improve semen reproductive potential and ICSI clinical outcome.

The PREgnancy and FERtility (PREFER) Study Investigating the Need for Ovarian Function and/or Fertility Preservation Strategies in Premenopausal Women With Early Breast Cancer.

BACKGROUND: Offering ovarian function and/or fertility preservation strategies in premenopausal women with newly diagnosed breast cancer candidates to undergo chemotherapy is standard of care. However, few data are available on uptake and main reasons for refusing these options. METHODS: The PREFER study (NCT02895165) is an observational, prospective study enrolling premenopausal women with early breast cancer, aged between 18 and 45 years, candidates to receive (neo)adjuvant chemotherapy. Primary objective is to collect information on acceptance rates and reasons for refusal of the proposed strategies for ovarian function and/or fertility preservation available in Italy. RESULTS: At the study coordinating center, 223 patients were recruited between November 2012 and December 2020. Median age was 38 years (range 24 - 45 years) with 159 patients (71.3%) diagnosed at ≤40 years. Temporary ovarian suppression with gonadotropin-releasing hormone agonists (GnRHa) was accepted by 58 out of 64 (90.6%) patients aged 41-45 years and by 151 out of 159 (95.0%) of those aged ≤40 years. Among patients aged ≤40 years, 57 (35.8%) accepted to access the fertility unit to receive a complete oncofertility counseling and 29 (18.2%) accepted to undergo a cryopreservation technique. Main reasons for refusal were fear of delaying the initiation of antineoplastic treatments and contraindications to the procedure or lack of interest in future childbearing. Patients with hormone-receptor positive breast cancer had a tendency for a higher acceptance rates of ovarian function and/or fertility preservation strategies than those with hormone-receptor negative disease. CONCLUSIONS: More than 90% of premenopausal women with early breast cancer, and particularly those with hormone receptor-positive disease, were concerned about the potential risk of chemotherapy-induced premature ovarian insufficiency and/or infertility and accepted GnRHa administration. Less than 1 out of 5 women aged ≤40 years accepted to undergo cryopreservation strategies.

Pronuclear score improves prediction of embryo implantation success in ICSI cycles.

BACKGROUND: In assisted reproduction technology embryo competence is routinely evaluated on morphological criteria but efficacy remains relatively low. Additional information could be obtained by evaluating pronuclear (PN) morphology. Up to now controversial results have been reported about the prognostic value of PN score. One of the main limitations of literature data is the use of different PN classification methods. In this regard, in 2011 the ESHRE and Alpha Scientists in Reproductive Medicine defined three PN categories to standardize zygote assessment. In this study we evaluated whether the consensus ESHRE-Alpha system for the pronuclear scoring could be an useful additional criterion to improve prediction of embryo implantation potential. METHODS: This is a retrospective, longitudinal, observational, cohort study. We included 3004 zygotes from 555 women who underwent ICSI treatment at our Center between January 2014 and June 2019. The PN were categorized as score 1: symmetrical, 2: non-symmetrical, 3: abnormal. A subset of 110 zygotes did not cleaved. On day 2-3 1163 embryos were transferred, 232 arrested, and 9 were cryopreserved. Among the 1490 embryos cultured up to day 5-7, 516 became blastocysts: 123 were transferred on day 5 and 393 were cryopreserved. Comparisons of age, cleavage and blastocyst rate, quality of embryos, implantation success among PN score groups were evaluated by chi-square test or Kruskal-Wallis test as appropriate. Potential predictors of embryo implantation were first tested in univariable analysis using generalized estimating equations taking into account correlation between embryos originated from the same patient. Then, variables potentially associated with implantation success (P<0.05) were included in a multivariable analysis for calculating the adjusted odds ratio (OR) and 95% confidence interval (CI). RESULTS: There was no significant difference in patients'age, cleavage and blastulation rates, and embryo morphology among the three PNscore groups. The PN score 1-embryos had a greater implantation success respect to score 2-3-ones (OR 1.83; 95% CI 1.34-2.50, P=0.0001). Consistently, the pronuclear score remained predictive of implantation in top quality embryos (OR 1.68; 95%CI 1.17-2.42, P= 0.005). CONCLUSIONS: The consensus pronuclear score may be routinely included among criteria for embryo evaluation to increase patients' chance of becoming pregnant.

Menstrual cycle resumption and female fertility after autologous hematopoietic stem cell transplantation for multiple sclerosis.

Data on fertility after autologous hematopoietic stem cell transplantation (aHSCT) in women with multiple sclerosis (MS) are inconclusive. This study aims to report on post-aHSCT menstrual resumption in a multi-center MS-women cohort. Out of 43 women, 30 (70%) recovered menses after a mean time of 6.8 months. Older age (odds ratio (OR) = 0.5, p < 0.0001) and previous pulsed cyclophosphamide (OR = 0.44, p = 0.005) were independently associated with a reduced menstrual recovery probability. Conditioning regimens' intensity resulted not associated with post-procedure amenorrhea. Our results highlight younger age as significantly associated with menses recovery; proper fertility counseling for MS women candidated to aHSCT both prior- and post-transplantation is therefore warranted.

Occurrence of smooth endoplasmic reticulum aggregates in metaphase II oocytes: relationship with stimulation protocols and outcome of ICSI and IVF cycles.

STUDY QUESTION: Is there any association between the appearance of smooth endoplasmic reticulum aggregates (SERa) in oocytes and ovarian stimulation, embryological, clinical and neonatal outcomes of ICSI and IVF cycles? SUMMARY ANSWER: A suboptimal prolonged ovarian stimulation is detrimental to oocytes by inducing the occurrence of SERa, which reduces the reproductive potential of oocytes. WHAT IS KNOWN ALREADY: Controlled ovarian stimulation recruits oocytes of different qualities. Based on current evidence, it was agreed that non-homogeneous cytoplasm may represent the normal variability among oocytes rather than a dysmorphism with developmental significance. The only exception is the appearance of SERa within the ooplasm. Owing to the lack of univocal evidence in this literature about the safety of injecting oocytes with SERa and the mechanism responsible for the occurrence of SERa, this topic is still a matter of debate. STUDY DESIGN, SIZE, DURATION: A retrospective, longitudinal cohort study performed at a tertiary level public infertility center. We included 1662 cycles (180 SERa+ and 1482 SERa-) from 1129 women (age: 20-44 years) who underwent IVF/ICSI treatments in 2012-2019. The SERa+ cycles had at least one SERa+ oocyte in the oocyte cohort. The SERa- cycles had morphologically unaffected oocytes. PARTICIPANTS/MATERIALS, SETTING, METHODS: We collected stimulation data and embryological, clinical, neonatal outcomes of SERa- and SERa+ cycles and oocytes. MAIN RESULTS AND THE ROLE OF CHANCE: Overall, 347 out of 12 436 metaphase II oocytes (2.8%) were affected by SER. We performed only 12 transfers involving at least one SERa+ embryo. Stimulation length (P = 0.002), serum progesterone (P = 0.004) and follicle size (P = 0.046) at trigger, number of retrieved (P = 0.004) and metaphase II (P = 0.0001) oocytes were significantly higher in SERa+ than SERa- cycles. Fertilization rate was significantly (P < 0.0001) reduced in SERa+ cycles and oocytes compared to SERa- counterparts. Embryos of SERa+ cycles had a lower blastocyst formation rate compared to embryos of SERa- cycles (P = 0.059). Statistical analysis according to a generalized estimating equation model performed at patient level demonstrated that the duration of ovarian stimulation was predictive of SERa+ oocytes appearance. The clinical success of SERa+ cycles was lower than SERa- cycles, although no differences in neonatal birthweights or malformations were recorded in sibling unaffected oocytes of SERa+ cycles. LIMITATIONS, REASONS FOR CAUTION: Given that SERa+ oocytes were discarded in our center for years and transfers of embryos originating from affected oocytes were generally avoided, clinical outcomes of SERa+ cycles are largely attributable to the transfer of embryos derived from unaffected oocytes of SERa+ cycles and we did not have data about newborns from affected oocytes, since none of the transfers involving SERa+ embryos resulted in a progressive clinical pregnancy. WIDER IMPLICATIONS OF THE FINDINGS: For the first time, we speculate that the late-follicular phase elevated serum progesterone caused by a suboptimal prolonged ovarian stimulation may be detrimental to the oocytes by inducing the occurrence of SERa, resulting in negative effects on their reproductive potential. This raises the question of whether some stimulation regimens could be worse than others and a change in stimulation protocol would reduce the possibility of producing oocytes with suboptimal maturation. In particular, our data highlight the importance of correct timing of the trigger in order to maximize oocyte collection, not only in terms of numerosity but also their reproductive potential. STUDY FUNDING/COMPETING INTEREST(S): None. TRIAL REGISTRATION NUMBER: N/A.