Antioxidants and Polyphenols: Concentrations and Relation to Male Infertility and Treatment Success

Oxidative stress is induced by reactive oxygen substances (ROS) that are known to affect male fertility. The aims of this study were to prospectively investigate and characterize total antioxidant and specifically polyphenols concentrations and their relations to sperm quality and fertility treatment success. During their infertility treatment, sixty-seven males were prospectively recruited to this study. After separation of the sperm from the semen sample, the semen fluid samples antioxidants and polyphenols concentrations were determined. Antioxidant concentration was significantly associated with sperm concentration and total motile count. Antioxidants concentration in the group of male with sperm concentration ≥ 15 × 106 was significantly higher than in the group of male with antioxidants concentration < 15 × 106 (830.3 ± 350 μM and 268.3 ± 220 μM, resp., p<0.001). Polyphenols concentration did not differ between the groups of sperm concentration above and below 15 × 106 (178.7 ± 121 μM and 161.7 ± 61 μM, resp., p-NS). No difference was found between fertilization rates and antioxidants or polyphenols concentrations. This is the first study that reports on polyphenols concentration within semen fluid. More studies are needed in order to investigate polyphenols role in male fertility

Absence of SCAPER causes male infertility in humans and Drosophila by modulating microtubule dynamics during meiosis

Mutation in S-phase cyclin A-associated protein rin the endoplasmic reticulum (SCAPER) have been found across ethnicities and have been shown to cause variable penetrance of an array of pathological traits, including intellectual disability, retinitis pigmentosa and ciliopathies. Human clinical phenotyping, surgical testicular sperm extraction and testicular tissue staining. Generation and analysis of short spindle 3 (ssp3) (SCAPER orthologue) Drosophila CAS9-knockout lines. In vitro microtubule (MT) binding assayed by total internal reflection fluorescence microscopy. We show that patients homozygous for a SCAPER mutation lack SCAPER expression in spermatogonia (SPG) and are azoospermic due to early defects in spermatogenesis, leading to the complete absence of meiotic cells. Interestingly, Drosophila null mutants for the ubiquitously expressed ssp3 gene are viable and female fertile but male sterile. We further show that male sterility in ssp3 null mutants is due to failure in both chromosome segregation and cytokinesis. In cells undergoing male meiosis, the MTs emanating from the centrosomes do not appear to interact properly with the chromosomes, which remain dispersed within dividing spermatocytes (SPCs). In addition, mutant SPCs are unable to assemble a normal central spindle and undergo cytokinesis. Consistent with these results, an in vitro assay demonstrated that both SCAPER and Ssp3 directly bind MTs. Our results show that SCAPER null mutations block the entry into meiosis of SPG, causing azoospermia. Null mutations in ssp3 specifically disrupt MT dynamics during male meiosis, leading to sterility. Moreover, both SCAPER and Ssp3 bind MTs in vitro. These results raise the intriguing possibility of a common feature between human and Drosophila meiosis

EGF increases expression and activity of PAs in preimplantation rat embryos and their implantation rate

BACKGROUND: Embryo implantation plays a major role in embryogenesis and the outcome of pregnancy. Plasminogen activators (PAs) have been implicated in mammalian fertilization, early stages of development and embryo implantation. As in-vitro developing embryos resulted in lower implantation rate than those developed in-vivo we assume that a reduced PAs activity may be involved. In the present work we studied the effect of EGF on PAs activity, quantity and embryo implantation. METHODS: Zygotes were flushed from rat oviducts on day one of pregnancy and grown in-vitro in R1ECM supplemented with EGF (10 ng/ml) and were grown up to the blastocyst stage. The control groups were grown in the same medium without EGF. The distribution and quantity of the PAs were examined using fluorescence immunohistochemistry followed by measurement of PAs activity using the chromogenic assay. Implantation rate was studied using the embryo donation model. RESULTS: PAs distribution in the embryos was the same in EGF treated and untreated embryos. Both PAs were localized in the blastocysts' trophectoderm, supporting the assumption that PAs play a role in the implantation process in rats. EGF increased the quantity of uPA at all stages studied but the 8-cell stage as compared with controls. The tissue type PA (tPA) content was unaffected except the 8-cell stage, which was increased. The activity of uPA increased gradually towards the blastocyst stage and more so due to the presence of EGF. The activity of tPA did not vary with the advancing developmental stages although it was also increased by EGF. The presence of EGF during the preimplantation development doubled the rate of implantation of the treated group as compared with controls

A Live Birth Subsequent to IVF following Egg Retrieval Only 12 Hours after hCG Priming

Introduction. To report a live birth following egg retrieval after only 12 hours from hCG priming. Patients. A childless couple with five-years-lasting secondary infertility. Methods. IVF was performed according to the long protocol. Two immature oocytes were retrieved following only 12 hours after hCG priming due to the patient misunderstanding. The eggs were cultured in vitro and ICSI was performed following polar body extruded after 24 hours in culture. After additional 24 hours a 4-cell embryo was developed and ET was performed. Results. A viable pregnancy was achieved and a healthy baby girl was delivered at 38 weeks of gestation. Conclusion. In a rare and unexpected situation when immature oocytes are retrieved following a short hCG priming, the eggs should be cultured in vitro, late ICSI should be performed, and a pregnancy may be expected

Expression of plasminogen activators in preimplantation rat embryos developed <it>in vivo </it>and <it>in vitro</it>

Abstract Background Embryo implantation plays a major role in embryogenesis and the outcome of pregnancy. Plasminogen activators (PAs) have been implicated in mammalian fertilization, early stages of development and embryo implantation. The invasion of trophoblast cells into the endometrium during the implantation process can be blocked by inhibitors of serine proteases, illustrating the role of these enzymes in the invasion process. As in vitro developing embryos resulted in lower implantation rate than those developed in vivo we assume that a reduced PAs activity may lead to it. There is hardly any information regarding qualitative or quantitative differences in expression of PAs in preimplantation embryos, or comparisons between in vivo and in vitro developed embryos. The purpose of this study was to assess the expression of urokinase type (uPA) and tissue type (tPA) plasminogen activators in in vivo and in vitro preimplantation development in rat embryos using immunofluorescence confocal microscopy and computerized image analysis. Methods Zygotes, 2-cell, 4-cell, 8-cell, morula and blastocyst stages of development were flushed from the reproductive tract (control groups) of Wistar rats. Zygotes were flushed and grown in vitro to the above mentioned developmental stages and comprised the experimental groups. Immunofluorescence microscopy and computerized image analysis were used to evaluate both qualitative (localization) and quantitative expression of plasminogen activators. Results uPA and tPA were found to be expressed in rat embryos throughout their preimplantation development, both in vivo and in vitro. While uPA was localized mainly in the cell cytoplasm, the tPA was detected mainly on cell surface and in the perivitelline space. In blastocysts, both in vivo and in vitro, uPA and tPA were localized in the trophectoderm cells. Total uPA content per embryo was higher in the in vivo as compared with the in vitro developed embryos at all stages measured. Blastocyst uPA content was significantly low as compared with the four-cell, eight-cell, and morula stages. Total tPA content was higher in embryos developed in vivo than those developed in vitro except for the 4-cell and 8-cell stages. Conclusion In vitro embryo development leads to lower PAs expression in a stage dependent manner as compared with in vivo developing controls. The enzymes studied vary probably in the ratio of their active and inactive forms as there is no correlation between their content and the activity observed in our previous study. The localization of both PAs in the blastocysts' trophectoderm supports the assumption that PAs plays a role in the implantation process in rats.</p

An artificial intelligence algorithm for automated blastocyst morphometric parameters demonstrates a positive association with implantation potential

Abstract Blastocyst selection is primarily based on morphological scoring systems and morphokinetic data. These methods involve subjective grading and time-consuming techniques. Artificial intelligence allows for objective and quick blastocyst selection. In this study, 608 blastocysts were selected for transfer using morphokinetics and Gardner criteria. Retrospectively, morphometric parameters of blastocyst size, inner cell mass (ICM) size, ICM-to-blastocyst size ratio, and ICM shape were automatically measured by a semantic segmentation neural network model. The model was trained on 1506 videos with 102 videos for validation with no overlap between the ICM and trophectoderm models. Univariable logistic analysis found blastocyst size and ICM-to-blastocyst size ratio to be significantly associated with implantation potential. Multivariable regression analysis, adjusted for woman age, found blastocyst size to be significantly associated with implantation potential. The odds of implantation increased by 1.74 for embryos with a blastocyst size greater than the mean (147 ± 19.1 μm). The performance of the algorithm was represented by an area under the curve of 0.70 (p < 0.01). In conclusion, this study supports the association of a large blastocyst size with higher implantation potential and suggests that automatically measured blastocyst morphometrics can be used as a precise, consistent, and time-saving tool for improving blastocyst selection

Automated Evaluation of Human Embryo Blastulation and Implantation Potential using Deep‐Learning

In in vitro fertilization (IVF) treatments, early identification of embryos with high implantation potential is required for shortening time to pregnancy while avoiding clinical complications to the newborn and the mother caused by multiple pregnancies. Current classification tools are based on morphological and morphokinetic parameters that are manually annotated using time‐lapse video files. However, manual annotation introduces interobserver and intraobserver variability and provides a discrete representation of preimplantation development while ignoring dynamic features that are associated with embryo quality. A fully automated and standardized classifiers are developed by training deep neural networks directly on the raw video files of >6200 blastulation‐labeled and >5500 implantation‐labeled embryos. Prediction of embryo implantation is more accurate than the current state‐of‐the‐art morphokientic classifier. Embryo classification improves with video length where the most predictive images show only partial association with morphological features. Deep learning substitute to human evaluation of embryo developmental competence thus contributes to implementing single embryo transfer methodology

Absence of SCAPER causes male infertility in humans and Drosophila by modulating microtubule dynamics during meiosis.

BACKGROUND: Mutation in S-phase cyclin A-associated protein rin the endoplasmic reticulum (SCAPER) have been found across ethnicities and have been shown to cause variable penetrance of an array of pathological traits, including intellectual disability, retinitis pigmentosa and ciliopathies. METHODS: Human clinical phenotyping, surgical testicular sperm extraction and testicular tissue staining. Generation and analysis of short spindle 3 (ssp3) (SCAPER orthologue) Drosophila CAS9-knockout lines. In vitro microtubule (MT) binding assayed by total internal reflection fluorescence microscopy. RESULTS: We show that patients homozygous for a SCAPER mutation lack SCAPER expression in spermatogonia (SPG) and are azoospermic due to early defects in spermatogenesis, leading to the complete absence of meiotic cells. Interestingly, Drosophila null mutants for the ubiquitously expressed ssp3 gene are viable and female fertile but male sterile. We further show that male sterility in ssp3 null mutants is due to failure in both chromosome segregation and cytokinesis. In cells undergoing male meiosis, the MTs emanating from the centrosomes do not appear to interact properly with the chromosomes, which remain dispersed within dividing spermatocytes (SPCs). In addition, mutant SPCs are unable to assemble a normal central spindle and undergo cytokinesis. Consistent with these results, an in vitro assay demonstrated that both SCAPER and Ssp3 directly bind MTs. CONCLUSIONS: Our results show that SCAPER null mutations block the entry into meiosis of SPG, causing azoospermia. Null mutations in ssp3 specifically disrupt MT dynamics during male meiosis, leading to sterility. Moreover, both SCAPER and Ssp3 bind MTs in vitro. These results raise the intriguing possibility of a common feature between human and Drosophila meiosis