Successful Pregnancy Following Platelet-Rich Plasma Intraovarian Injection and In Vitro Maturation of Oocytes in a 47-Year-Old Woman: A Case Report.

In vitro maturation (IVM) of oocytes represents an assisted reproductive technique that involves the minimal or absence of ovarian stimulation and is beneficial to specific groups of patients. It is based on the collection of immature cumulus-oocyte complexes (COCs) from antral follicles, which are cultured in vitro until they reach the metaphase II (MII) stage. Once maturation is completed, IVM oocytes are normally fertilized, as during a conventional IVF protocol. On the other hand, ovarian rejuvenation through the intraovarian platelet-rich plasma (PRP) injection represents an innovative procedure intended to restore ovarian fertility and development, and it is used to enhance ovarian stimulation outcomes. Here, we report a case of a 47-year-old woman who underwent an assisted reproductive technology cycle (ART) with PRP injection and IVM, which resulted in a successful pregnancy.

In vitro maturation of oocytes in light of ovarian mitochondrial improvement: effectiveness and safety.

In vitro maturation of oocytes (IVM) represents an assisted reproductive technique that involves the minimal or absence of ovarian stimulation and is beneficial to specific groups of patients. These may include women with polycystic ovarian syndrome and/or patients who need a fertility preservation option before undergoing gonadotoxic treatment. However, when IVM is applied in cases where it is not recommended, it can be considered as an add-on technique, as described by the ESHRE Guideline Group on Female Fertility Preservation. Interestingly, IVM has not been proven yet to be as effective as conventional IVF in the laboratory, in terms of clinical pregnancy and live birth rates, while concerns have been raised for its long-term safety. As a result, both safety and efficacy of IVM remain still questionable and additional data are needed to draw conclusions.

Poland Syndrome in a Pregnancy From an Assisted Reproductive Technology (ART) Cycle With In Vitro Maturation (IVM) and Rescue Intracytoplasmic Sperm Injection (ICSI).

Poland syndrome is a congenital anatomical anomaly, characterised by partial or total aplasia of one side of the body causing abnormalities affecting the chest, shoulder, and upper limb. The exact mechanism that leads to this syndrome is unknown, but an abnormality in the vasculature formation or interruption of the blood supply of the subscapular artery and its branches early in development may be the main cause. Depending on the underlying mechanism, the syndrome has several expressions with some hardly being detectable and others not even being compatible with life. Here, we present a case of pregnancy from an assisted reproductive technology (ART) cycle with in vitro maturation (IVM) and rescue intra cytoplasmic sperm injection (ICSI), which resulted in the in-utero death of the foetus. The subsequent necropsy revealed a variation of Poland syndrome.

Evaluation of Sperm DNA Fragmentation Using Two Different Methods: TUNEL via Fluorescence Microscopy, and Flow Cytometry.

Background and Objectives: Sperm DNA fragmentation refers to any break in one or both of the strands of DNA in the head of a sperm. The most widely used methodologies for assessing sperm DNA fragmentation are the sperm chromatin structure assay (SCSA), the sperm chromatin dispersion assay (SCD), the single-cell gel electrophoresis assay (SCGE-comet), and the terminal-deoxynucleotidyl-transferase (TdT)-mediated dUTP nick end labelling (TUNEL) assay. The aim of this study was to compare the efficiency and sensitivity of the analysis of sperm DNA fragmentation using TUNEL via fluorescence microscopy, and flow cytometry. Materials and Methods: Semen samples were collected and analyzed for standard characteristics using light microscopy, and for sperm DNA fragmentation using both TUNEL via fluorescence microscopy, and flow cytometry. Results: There were no significant differences in the values of the sperm DNA fragmentation index (DFI) obtained when the analysis was performed using TUNEL or flow cytometry (p = 0.543). Spearman's correlation analysis revealed a significant negative correlation between sperm motility (%) and sperm DNA fragmentation (p < 0.01), as well as between sperm concentration and sperm DNA fragmentation (p < 0.05). The Mann-Whitney U test showed no significant difference in the DFI among couples with repeated implantation failure (RIF) and miscarriages (p = 0.352). Conclusions: Both methods (TUNEL via fluorescence microscopy, and flow cytometry) have a high efficiency and sensitivity in accurately detecting sperm DNA fragmentation, and can be effectively used to assess male fertility.

The impact of cryopreservation on both sperm HPV-negative and positive subtypes.

It is well known that various human papillomavirus (HPV) genotypes are present in semen specimens. Also, it has been demonstrated that sperm parameters are negatively affected when HPV infection is present in the sperm sample. Besides all these, the effect of cryopreservation on HPV sensitivity and resistance is not known. The aim of the present study is to evaluate first the prevalence of HPV and secondly to elucidate whether cryopreservation of sperm HPV-positive samples has any effect on the viability of HPV. For this purpose, a cohort of 78 sperm specimens was used from a respective number of patients. After giving informed consent, semen analysis was performed. Each sperm sample was divided into four equal aliquots. The first one (fresh) was evaluated for the prevalence of HPV, while the other three aliquots were cryopreserved by adding an equal quantity of cryoprotectant and plunged into the LN. Each of the three aliquots was thawed 3, 6, and 12 months later, respectively, so as to evaluate whether there is a time-resistance period of HPV prevalence. HPV infection was found to be in eleven sperm samples, demonstrating a 14.1% (11/78) HPV prevalence. Among the HPV-positive samples, six of them were high-risk and the remaining were low-risk genotypes. Moreover, the high-risk fresh samples demonstrated higher motility values than the low-risk samples (60% ± 2.7 vs 45.6% ± 3.7, p < .05), while semen volume in the high-risk samples was significantly lower than the respective volume in the low-risk samples (2.26 ± 0.2ml vs 3.5 ± 0.6ml, p < .05). Interestingly, cryopreservation of the HPV-positive samples resulted in the sustainability and time-resistance of HPV in all high-risk HPV-positive samples, something that was not the case with the low-risk HPV-positive samples. Conclusively, sperm samples infected with high-risk HPV, demonstrate lower sperm parameters and time-resistance activity during cryopreservation.

Oxytocin preprotein and oxytocin receptor mRNA expression is altered in semen samples with abnormal semen parameters.

RESEARCH QUESTION: Are oxytocin preprotein and the oxytocin receptor expressed in human spermatozoa and is their mRNA expression different between normal semen samples and samples with at least one abnormal parameter? DESIGN: An in-vitro prospective study of 175 semen samples from Greek men, according to World Health Organization criteria, 2010. mRNA expression levels were compared between different categories of semen samples, classified according to their concentration, total number, motility and morphology. Immunohistochemistry was used to detect oxytocin preprotein and its receptor on spermatozoa smears. RESULTS: Compared with normal samples (normal motility and normal concentration), samples with at least one abnormal sperm parameter had statistically significantly lower oxytocin preprotein mRNA expression (P = 0.019) and higher oxytocin receptor mRNA expression levels (P < 0.001). Oligozoospermic samples had statistically significantly higher oxytocin preprotein mRNA expression levels (P = 0.002) and lower oxytocin receptor mRNA expression levels (P = 0.047). Asthenozoospermic samples had statistically significantly lower oxytocin preprotein mRNA expression levels (P < 0.001). Teratozoospermic samples had statistically significantly lower oxytocin preprotein mRNA expression levels (P = 0.049) and higher oxytocin receptor mRNA expression levels (P < 0.001). Oxytocin preprotein mRNA expression was positively associated with total progressive motility (P < 0.001) and negatively associated with the percentage of immotile spermatozoa (P = 0.001). Oxytocin receptor mRNA expression was negatively associated with the percentage of normal forms (P < 0.001). CONCLUSION: Oxytocin preprotein and oxytocin receptor mRNA expression in spermatozoa could be used as a novel and unbiased diagnostic tool for male infertility.

Clinical Outcome, Socioeconomic Status and Psychological Constrains of Patients Undergoing Preimplantation Genetic Testing (PGT) in Northern Greece.

Background and objectives: Preimplantation genetic testing (PGT) offers patients the possibility of having a healthy baby free of chromosomal or genetic disorders. The present study focuses on the application of PGT for patients located in Northern Greece, investigating their clinical outcomes, their motives, and their overall physical and emotional experience during the treatment, in association with their socioeconomic background. Materials and Methods: Couples who underwent PGT for a monogenic condition (PGT-M, n = 19 cycles) or aneuploidy (PGT-A, n = 22 cycles) participated in the study. Fertilization, implantation, and pregnancy rates were recorded for all cycles. The couples were asked to fill in a questionnaire about the consultation they had received prior to treatment, their sociodemographic information, and the psychological impact PGT had on both the female and male partner. Results: The fertilization, implantation, and ongoing pregnancy rates for the PGT-M and PGT-A cycles were 81.3%, 70.6%, and 52.9%, and 78.2%, 64.3%, and 57.1%, respectively. Females experienced more intense physical pain than their male partners while psychological pain was encountered by both partners and occasionally in higher instances in males. No typical socioeconomic background of the patients referred for PGT in Northern Greece was noticed. Conclusion: PGT is an attractive alternative to prenatal diagnosis (PND), aiming to establisha healthy pregnancy by identifying and avoiding the transfer of chromosomally or genetically abnormal embryos to the uterus. Although the benefits of PGT were well-received by all patients undergoing the procedure, psychological pain was evident and especially prominent in patients with a previous affected child or no normal embryos for transfer. Holistic counseling is of utmost importance in order to make patients' experience during their journey to have a healthy baby less emotionally demanding and help them make the right choices for the future.

The human embryo following biopsy on day 5 versus day 3: viability, ultrastructure and spindle/chromosome configurations.

RESEARCH QUESTION: Are there any differences in viability, spindle abnormalities and mitochondrial and other organelle structures amongst embryos biopsied on day 3 versus day 5 before and after vitrification? DESIGN: A total of 240 day 3 biopsied embryos that developed to blastocysts but were rejected for transfer following preimplantation genetic testing for monogenic/single gene defects (PGT-M) (n = 115) or for aneuploidies (PGT-A) (n = 125) were divided into two groups: (i) 120 blastocysts treated for viability, spindle/chromosome configuration (SCC) analysis and transmission electron microscopy (TEM) analysis (fresh n = 20, n = 20, n = 20 and following vitrification/warming n = 20, n = 20, n = 20); (ii) 120 embryos were re-biopsied at the blastocyst stage and treated for viability, SCC and TEM analysis (fresh n = 20, n = 20, n = 20 and following vitrification/warming n = 20, n = 20, n = 20). Also, 60 vitrified blastocysts biopsied only on day 5 that were rejected for transfer following PGT-M (n = 6) or PGT-A (n = 54) were treated following warming for viability (n = 20), SCC (n = 20) and TEM analysis (n = 20). RESULTS: No differences were observed in SCC and ultrastructure between embryos biopsied on day 5 and day 3 but following vitrification higher numbers of abnormal spindles, distension of mitochondria, multivesicular bodies, lipofuscin droplets, altered cell junctions and occasionally excessive accumulation of glycogen granules were evident. The fresh day 3 biopsied group also had a lower incidence of damaged (propidium iodide-stained) cells compared with the fresh day 3+5 (P = 0.02) and the vitrified day 5 (P = 0.001) biopsied groups. CONCLUSIONS: Biopsies on day 5 and day 3 do not adversely affect embryo viability, SCC or ultrastructure, although following vitrification minimal embryo quality-dependent increases in spindle abnormalities and cell damage are observed.

Effects of Different Drug Therapies and COVID-19 mRNA Vaccination on Semen Quality in a Man with Ankylosing Spondylitis: A Case Report.

Background and Objectives: Ankylosing spondylitis (AS) is a condition that affects 0.1% to 0.5% of the adult population. The aim of this case report was to investigate the possible effects of the drugs taken for treatment of AS as well as mRNA vaccination for COVID-19 on semen quality by performing a highly detailed analysis. Materials and Methods: Sperm characteristics were examined by light microscopy, DNA fragmentation (DFI) was analysed by flow cytometry and morphology was evaluated by transmission electron microscopy (TEM). Results: Semen analysis under therapy with (1) celecoxib and sulphasalazine showed: concentration 47 million/mL, 53% progressive motility, 7% normal morphology and 9.6% DFI, (2) Golimumab and before mRNA Vaccination showed: concentration 108 million/mL, 82% progressive motility, 1% normal morphology and 7.6% DFI, and (3) Golimumab and after 3 doses of mRNA Vaccination showed: concentration 142 million/mL, 85% progressive motility, 1% normal morphology and 6.8% DFI. TEM revealed head, neck and tail abnormalities, as well as the presence of cells with incomplete spermiogenesis white cells and phagocytes in the sample under therapy with celecoxib and sulphasalazine. Golimumab treatment lead to an increased incidence of elongated heads but in general reduced inflammation as no white cells were evident in TEM. Conclusion: The anti-inflamatory drugs celecoxib and sulphasalazine had no adverse effect on sperm quality as all parameters were within normal limits and the patient achieved under that treatment 2 pregnancies following natural conception that lead to the birth of a healthy boy and girl respectively. Anti-TNFa treatment with Golimumab exerted a negative effect on morphology but not on concentration, motility and DFI. After 3 doses of mRNA Vaccination, sperm concentration increased while motility, morphology and DFI remained similar to the values before vaccination suggesting no negative effect of the mRNA vaccine for COVID-19 on sperm quality.

Insights into the Role of Telomeres in Human Embryological Parameters. Opinions Regarding IVF.

Telomeres promote genome integrity by protecting chromosome ends from the activation of the DNA damage response and protecting chromosomes from the loss of coding sequences due to the end replication problem. Telomere length (TL) is progressively shortened as age progresses, thus resulting in cellular senescence. Therefore, TL is in strong adverse linear correlation with aging. Mounting evidence supports the notion that telomeres and male/female infertility are in a close relationship, posing the biology of telomeres as a hot topic in the era of human-assisted reproduction. Specifically, the length of sperm telomeres is gradually increasing as men get older, while the telomere length of the oocytes seems not to follow similar patterns with that of sperm. Nonetheless, the telomere length of the embryos during the cleavage stages seems to have a paternal origin, but the telomere length can be further extended by telomerase activity during the blastocyst stage. The latter has been proposed as a new molecular biomarker with strong predictive value regarding male infertility. As far as the role of telomeres in assisted reproduction, the data is limited but the length of telomeres in both gametes seems to be affected mainly by the cause of infertility rather than the assisted reproductive therapy (ART) procedure itself. The present review aims to shed more light into the role of telomeres in human embryological parameters, including gametes and embryos and also presents opinions regarding the association between telomeres and in vitro fertilization (IVF).

Viability assessment using fluorescent markers and ultrastructure of human biopsied embryos vitrified in open and closed systems.

RESEARCH QUESTION: Are there any differences in viability and ultrastructure amongst embryos biopsied on Day 5 versus Day 3 following vitrification in open and closed systems and compared to fresh embryos? DESIGN: One hundred human embryos (40 blastocysts biopsied on Day 5 and subsequently vitrified in open or closed systems and 60 Day 3 biopsied embryos that developed to blastocysts but were rejected for transfer following preimplantation genetic testing for monogenic/single gene defects and for aneuploidies were either treated fresh [n = 20] or vitrified [n = 40] in open or closed systems) and following warming and culture for 4 h were subjected to viability staining with carboxyfluorescein-diacetate succinimidylester/propidium iodide or processed for transmission electron microscopy. RESULTS: No statistically significant differences were observed in the viability of human biopsied embryos following vitrification in open and closed systems. Compared to fresh embryos, vitrified ones had a higher incidence of damage (propidium iodide-stained cells) irrespective of the vitrification method (P = 0.005). These damaged cells were more prominent in Day 5 biopsied blastocysts and mainly located at the position of cutting. Characteristic lipofuscin droplets (representative of apoptosis) and a higher number of vacuoles and distension of mitochondria were also more evident in vitrified embryos, although this was not statistically assessed. CONCLUSIONS: Vitrification in open and closed systems does not adversely affect the viability and ultrastructure of Day 5 and Day 3 biopsied embryos as revealed by the minimal yet statistically significant cell damage observed. This damage may be compensated by the embryos, which in their attempt to fully recover following vitrification, potentially enable 'rescue' processes to eliminate it.

The Impact of SARS-CoV-2 on Sperm Cryostorage, Theoretical or Real Risk?

Cryopreservation of human gametes and embryos as well as human reproductive tissues has been characterized as an essential process and aspect of assisted reproductive technology (ART). Notably, sperm cryopreservation is a fundamental aspect of cryopreservation in oncological patients or patients undergoing gonadotoxic treatment. Given that there is a risk of contamination or cross-contamination, either theoretical or real, during the procedures of cryopreservation and cryostorage, both the European Society for Human Reproduction and Embryology (ESHRE) and the American Society for Reproductive Medicine (ASRM) have provided updated guidelines for preventing or reducing the contamination risk of sexually transmitted viruses. Given the ongoing and worldwide COVID-19 pandemic, there is considerable interest in what measures should be taken to mitigate SARS-CoV-2 contamination during cryopreservation and cryostorage of semen samples. The SARS-CoV-2 virus is the virus that causes COVID-19, and whose transmission and infection is mainly aerosol-mediated. Several ART professional societies, including ESHRE and ASRM have proposed measures to mitigate the spread of the SARS-CoV-2 virus. Whether the proposed safety directives are enough to mitigate the possible SARS-CoV-2-contamination of sperm samples during cryopreservation or whether the policies should be re-evaluated will be discussed in this review. Additionally, insights regarding the possible impact of COVID-19 vaccination on the safety of sperm cryopreservation will be discussed.

Influence of conception and delivery mode on stress response marker Oct4B1 and imprinted gene expression related to embryo development: A cohort study.

BACKGROUND: Recent scientific data support that the mode of conception and delivery may influence epigenetic regulation and therefore embryo development. Octamer-binding transcription factor 4-B1 (OCT4B1), a novel variant of OCT4 with yet unknown biological function, is suggested to have a potential role in mediating cellular stress response. Furthermore, Insulinlike Growth Factor 2 (IGF2), Mesoderm-specific Transcript (MEST) and paternally expressed gene 10 (PEG10) are genes known as imprinted and are regulated via means of epigenetic regulation. The influence of delivery mode and conception on epigenetic regulation is an active research field. OBJECTIVE: Our aim was to correlate the expression level of Oct4B1 and the expression and methylation level of IGF2, MEST, and PEG10 imprinted genes with the mode of delivery and conception in the umbilical cord blood of newborns. MATERIALS AND METHODS: Samples of umbilical cord blood from infants born after vaginal delivery, caesarean section (CS) with the infant in cephalic position and CS due to breech position were examined. Furthermore, the investigation included infants conceived through means of assisted reproductive technology. RESULTS: No statistically significant differences were found in mRNA expression levels between different modes of conception and delivery (p = 0.96). Oct4B1, IGF2, MEST, and PEG10 expression levels do not seem to be significantly affected by different modes of conception and delivery. CONCLUSION: These results indicate that the expression and methylation patterns of Oct4B1, IGF2, MEST and PEG10 in umbilical cord blood are not affected by the conception and delivery mode.

Expression of genes that regulate follicle development and maturation during ovarian stimulation in poor responders.

RESEARCH QUESTION: Sex hormone-binding globulin (SHBG), androgen receptor (AR), LH beta polypeptide (LHB), progesterone receptor membrane component 1 (PGRMC1) and progesterone receptor membrane component 2 (PGRMC2) regulate follicle development and maturation. Their mRNA expression was assessed in peripheral blood mononuclear cells (PBMC) of normal and poor responders, during ovarian stimulation. DESIGN: Fifty-two normal responders and 15 poor responders according to the Bologna criteria were enrolled for IVF and intracytoplasmic sperm injection and stimulated with 200 IU of follitrophin alpha and gonadotrophin-releasing hormone antagonist. HCG was administered for final oocyte maturation. On days 1, 6 and 10 of stimulation, blood samples were obtained, serum hormone levels were measured, RNA was extracted from PBMC and real-time polymerase chain reaction was carried out to identify the mRNA levels. Relative mRNA expression of each gene was calculated by the comparative 2-DDCt method. RESULTS: Differences between mRNA levels of each gene on the same time point between the two groups were not significant. PGRMC1 and PGRMC2 mRNA levels were downregulated, adjusted for ovarian response and age. Positive correlations between PGRMC1 and AR (standardized beta = 0.890, P < 0.001) from day 1 to 6 and PGRMC1 and LHB (standardized beta = 0.806, P < 0.001) from day 1 to 10 were found in poor responders. PGRMC1 and PGRMC2 were positively correlated on days 6 and 10 in normal responders. CONCLUSIONS: PGRMC1 and PGRMC2 mRNA are significantly decreased during ovarian stimulation, with some potential differences between normal and poor responders.

The effects of postnatal exposure of endocrine disruptors on testicular function: a systematic review and a meta-analysis.

BACKGROUND: Despite many epidemiological studies having been conducted, the impact of postnatal exposure of endocrine disruptors (EDs) on testicular function remains a controversial issue. AIM: To systematically review the literature and perform a quantitative synthesis to evaluate the effect of EDs on testicular function. MATERIALS AND METHODS: A comprehensive search was conducted in the MEDLINE, Scopus, and CENTRAL databases. Eligible for the systematic review were observational (cross-sectional and cohort) studies with (i) adult men who had a high probability of postnatal exposure to EDs ("exposed"), (ii) adult men who had a low probability of postnatal exposure to EDs ("non-exposed"), and (iii) an outcome of interest [seminal parameters and reproductive hormone concentrations]. The continuous outcomes in each of the studies were synthesized by the random effects model and expressed as standardized mean difference (SMD) with 95% confidence interval (CI). RESULTS: Thirteen studies, including 959 exposed and 907 non-exposed men, fulfilled the inclusion criteria. Exposure to EDs was associated with decreased LH [SMD - 0.17, 95% CI - 0.33 to - 0.02, 10 studies (616 exposed, 563 non-exposed), I2 40%, p = 0.09], progressive motility [SMD - 0.45, 95% CI - 0.77 to - 0.13, three studies (133 cases, 153 controls), I2 38%, p = 0.20], and normal morphology [SMD - 0.50, 95% CI - 0.85 to - 0.14, eight studies (562 cases, 540 controls), I2 87%, p < 0.01] compared with non-exposure. No difference was observed between the other study groups. CONCLUSIONS: Postnatal exposure to EDs is associated with decreased semen quality. Nevertheless, there is no evidence that a disruption of testicular function mediates the deterioration in semen quality.

In-utero stress and mode of conception: impact on regulation of imprinted genes, fetal development and future health.

BACKGROUND: Genomic imprinting is an epigenetic gene regulatory mechanism; disruption of this process during early embryonic development can have major consequences on both fetal and placental development. The periconceptional period and intrauterine life are crucial for determining long-term susceptibility to diseases. Treatments and procedures in assisted reproductive technologies (ART) and adverse in-utero environments may modify the methylation levels of genomic imprinting regions, including insulin-like growth factor 2 (IGF2)/H19, mesoderm-specific transcript (MEST), and paternally expressed gene 10 (PEG10), affecting the development of the fetus. ART, maternal psychological stress, and gestational exposures to chemicals are common stressors suspected to alter global epigenetic patterns including imprinted genes. OBJECTIVE AND RATIONALE: Our objective is to highlight the effect of conception mode and maternal psychological stress on fetal development. Specifically, we monitor fetal programming, regulation of imprinted genes, fetal growth, and long-term disease risk, using the imprinted genes IGF2/H19, MEST, and PEG10 as examples. The possible role of environmental chemicals in genomic imprinting is also discussed. SEARCH METHODS: A PubMed search of articles published mostly from 2005 to 2019 was conducted using search terms IGF2/H19, MEST, PEG10, imprinted genes, DNA methylation, gene expression, and imprinting disorders (IDs). Studies focusing on maternal prenatal stress, psychological well-being, environmental chemicals, ART, and placental/fetal development were evaluated and included in this review. OUTCOMES: IGF2/H19, MEST, and PEG10 imprinted genes have a broad developmental effect on fetal growth and birth weight variation. Their disruption is linked to pregnancy complications, metabolic disorders, cognitive impairment, and cancer. Adverse early environment has a major impact on the developing fetus, affecting mostly growth, the structure, and subsequent function of the hypothalamic-pituitary-adrenal axis and neurodevelopment. Extensive evidence suggests that the gestational environment has an impact on epigenetic patterns including imprinting, which can lead to adverse long-term outcomes in the offspring. Environmental stressors such as maternal prenatal psychological stress have been found to associate with altered DNA methylation patterns in placenta and to affect fetal development. Studies conducted during the past decades have suggested that ART pregnancies are at a higher risk for a number of complications such as birth defects and IDs. ART procedures involve multiple steps that are conducted during critical windows for imprinting establishment and maintenance, necessitating long-term evaluation of children conceived through ART. Exposure to environmental chemicals can affect placental imprinting and fetal growth both in humans and in experimental animals. Therefore, their role in imprinting should be better elucidated, considering the ubiquitous exposure to these chemicals. WIDER IMPLICATIONS: Dysregulation of imprinted genes is a plausible mechanism linking stressors such as maternal psychological stress, conception using ART, and chemical exposures with fetal growth. It is expected that a greater understanding of the role of imprinted genes and their regulation in fetal development will provide insights for clinical prevention and management of growth and IDs. In a broader context, evidence connecting impaired imprinted gene function to common diseases such as cancer is increasing. This implies early regulation of imprinting may enable control of long-term human health, reducing the burden of disease in the population in years to come.

Addition of procyanidine to semen preserves progressive sperm motility up to three hours of incubation.

Several studies on semen physiology and sperm fertilizing capacity have shown a beneficial effect of antioxidants. Procyanidine is a natural antioxidant, more efficient compared with vitamin C and E, with many applications in the food, agriculture, pharmaceutical and cosmetic industry. Thus, we tested whether the addition of procyanidine to the semen of infertile men has a beneficial effect on spermatozoa during their in vitro incubation and during the cryopreservation process. Semen samples of 25 infertile men were divided in to two aliquots, in which procyanidine was added or not. Semen analysis, measurement of sperm DNA fragmentation index (DFI) and measurement of reactive oxygen species (ROS) were performed 3 h after incubation at 37 °C and after sperm cryopreservation and thawing. In-vitro addition of procyanidine to semen of infertile men resulted in a lesser decrease in progressive motility [-4 (-31:+6) vs. -6 (-31:+5), p < 0.001] and total motility [-5 (-29:+3) vs. -9 (-32:+2), p < 0.001] after 3 h of incubation compared with no addition of procyanidine. Sperm morphology was decreased only in the control group after 3 h of incubation [2 (0:+6) vs. 1 (0:+4), p = 0.009]. Furthermore, a larger increase in sperm DFI was observed in the control compared with the procyanidine group [9 (-7:+27) vs. 3 (-3:+18), p = 0.005] after thawing of cryopreserved semen samples. In conclusion, in-vitro addition of procyanidine to the semen of infertile men exerts a protective effect on progressive motility during handling and after 3 h of incubation as well as on sperm DFI during the process of cryopreservation.

Spindle and chromosome configuration analysis of human biopsied versus non-biopsied embryos by confocal laser scanning microscopy following vitrification.

SummaryThe aim of this study was to investigate the effects of zona drilling and biopsy on day 3 followed by vitrification on day 5 on the cytoskeleton and development of human embryos, by analysing survival rates and spindle and chromosome configurations by fluorescence and confocal laser scanning microscopy in human biopsied and non-biopsied embryos. In total, 98 human blastocysts (50 non-biopsied and 48 following biopsy on day 3) were vitrified on day 5 using either a commercial dimethyl sulphoxide (DMSO)-free vitrification kit or increasing concentrations of DMSO/EG (5%/5-10%/10-20%/20%). Following warming, the blastocysts were allowed to recover in culture for 24 h and were immunostained with α-tubulin, acetylated tubulin, and/or γ-tubulin antibodies in combination with 4',6-diamidino-2-phenylindole (DAPI). Labelled embryos were examined by both fluorescence and confocal laser scanning microscopy. The survival rates following warming (92% non-biopsied vs 83.3% biopsied) and the incidence of normal spindle chromosome configurations was not statistically different between the two groups (65.2% non-biopsied vs 59.2% biopsied, P>0.05). The incidence of spindle abnormalities including multipolarity, chromosome lagging, congression failure and chromosome bridging were also similar between the two groups (P>0.05). This study is the first to compare the incidence of cytoskeletal abnormalities in biopsied and non-biopsied human embryos following vitrification. We conclude that there was no significant difference in the survival rates and the incidence of spindle abnormalities between the two groups.

Artificial oocyte activation: physiological, pathophysiological and ethical aspects.

Infertile couples with low oocyte yield in combination with abnormal semen parameters may experience intra-cytoplasmic sperm injection (ICSI) failure. An established factor associated with ICSI failure is oocyte activation deficiency (AOD). The latter originates from seminal contributors, such as phospholipase C-zeta (PLCζ) that is not adequate to produce calcium (Ca2+) oscillations for oocyte activation. Apart from this natural activator, other stimulants, such as A23187, ionomycin, strontium chloride or even electric pulses, have been used in embryological laboratories to overcome AOD and ICSI failure. The aim of the present narrative review is to discuss the role of Ca+2 oscillations in oocyte activation and summarize the evidence concerning the use of oocyte activators as agents for artificial oocyte activation (AOA). Studies in humans and animals have emerged many physiological, pathophysiological and ethical aspects of AOA. In conclusion, in mammalian eggs, the cytosolic Ca+2 oscillations derive from a periodic release of Ca+2 from intracellular pools. PLCζ, as well as artificial stimulants, have been used to produce Ca+2 oscillations for AOA. As the latter may increase the risk of epigenetic induced malformations, further studies are required to clarify whether AOA constitutes an effective and safe method to overcome ICSI failure. Abbreviations: AOA: artificial oocyte activation; AOD: oocyte activation deficiency; Ca+2: Calcium; CAMKII: Ca+2/calmodulin-dependent protein kinase II; CICR: calcium-induced calcium-release; DAG: diacylglycerol; GM-CSF: granulocyte-macrophage colony-stimulating factor; ICSI: intra-cytoplasmic sperm injection; InsP3R: inositol-trisphosphate receptor; IP3: inositol 1,4,5-trisphosphate; IVF: in vitro fertilization; MAP: mitogen-activated protein; MII: metaphase II; NADP: nicotinic acid adenine dinucleotide phosphate; NO: nitric oxide; PAWP: post-acrosomal WW-binding domain protein; PIP2: phosphatidylinositol 4,5-bisphosphate; PLC: phospholipase C; PLCζ: phospholipase C-zeta; SOAFs: spermatozoon-released oocyte-activating factors; Sr+2: strontium; TFF: total fertilization failure.

Testicular versus ejaculated spermatozoa for ICSI in patients without azoospermia: A systematic review.

The use of testicular spermatozoa in men without azoospermia has been proposed as a means to increase the chances of pregnancy following assisted reproductive treatment. The purpose of this systematic review is to assess whether clinical outcomes are better when testicular rather than ejaculated spermatozoa are used for intracytoplasmic sperm injection in patients with abnormal semen parameters without azoospermia. A literature search identified four eligible studies out of 757 initially found. In a prospective study in men with high DNA fragmentation index (DFI) and oligozoospermia, the probability of live birth was significantly higher with testicular compared to ejaculated spermatozoa (risk ratio [RR]: 1.75, 95% confidence interval [CI]: 1.14-2.70). This was not the case in a retrospective study in men with high DFI only (RR: 2.36, 95% CI: 0.98-5.68). Clinical pregnancy rates were similar in a randomized controlled trial in men with asthenozoospermia with or without teratozoospermia (RR: 2.85, 95% CI: 0.76-10.69) and in a retrospective study in men with isolated asthenozoospermia (RR: 1.09, 95% CI: 0.56-2.14). Currently, there is limited, low-quality evidence suggesting that a higher probability of pregnancy might be expected using testicular rather than ejaculated spermatozoa, only in men with high DFI and oligozoospermia.