The effect of cult-active medium on pregnancy outcomes after intracytoplasmic sperm injection in azoospermic men: A case-control study

Background: Failed oocyte activation following intracytoplasmic sperm injection (ICSI) as a result of calcium deficiency is a major challenge. Objective: We compared the effect of cult-active medium (CAM) on ICSI outcomes in obstructive azoospermia cases. Materials and Methods: The present study was conducted with 152 ICSI cases, classified into CAM and control groups. The injected oocytes in the control group were cultured in the cleavage medium, while in the artificial oocyte activation group, oocytes were chemically activated through exposure to 200 µL of CAM for 15 min. Fertilization and cleavage rates, quality of embryos, and biochemical pregnancy and live birth rates were assessed in both groups. Results: There were significant differences between the groups in terms of fertilization and cleavage rates after using the CAM in the percutaneous epididymal sperm aspiration (PESA) subgroup (p = 0.05, p ≤ 0.001) and in the testicular sperm extraction subgroup (p = 0.02, p = 0.04), compared to their control groups. Also, the pregnancy rate was significantly higher in the PESA-CAM subgroup (p = 0.03). The PESA-CAM subgroup demonstrated a significant difference in embryo quality after ICSI (p = 0.04). Unsuccessful embryo transfer and abortion were lower in both subgroups compared to the control groups, but this difference was not significant. Surprisingly, live birth rate was higher in the PESA-CAM subgroup (p = 0.03). Conclusion: CAM treatment could improve fertilization and cleavage rates in obstructive azoospermia participants. It had a significant effect on embryo quality, and pregnancy and live birth rates in PESA cases. Key words: Calcium ionophore, Obstructive azoospermia, Fertilization, ICSI.

Effects of Extra-Long-Acting Recombinant Bovine FSH (bscrFSH) on Cattle Superovulation

Over the last few years, several commercial FSH products have been developed for cattle superovulation (SOV) purposes in Multiple Ovulation and Embryo Transfer (MOET) programs. The SOV response is highly variable among individuals and remains one of the main limiting factors in obtaining a profitable number of transferable embryos. In this study, follicle stimulating hormone (FSH) from different origins was included in two SOV protocols, (a) FSH from purified pig pituitary extract (NIH-FSH-p; two doses/day, 12 h apart, four consecutive days); and (b) extra-long-acting bovine recombinant FSH (bscrFSH; a single dose/day, four consecutive days), to test the effects of bscrFSH on the ovarian response, hormone profile levels, in vivo embryo production and the pluripotency gene expression of the obtained embryos. A total of 68 healthy primiparous red Angus cows (Bos taurus) were randomly distributed into two experimental groups (n = 34 each). Blood sample collection for progesterone (P4) and cortisol (C) level determination was performed together with ultrasonographic assessment for ovarian size, follicles (FL) and corpora lutea (CL) quantification in each SOV protocol (Day 0, 4, 8, and 15). Moreover, FSH profiles were monitorised throughout both protocols (Day 0, 4, 5, 6, 7, 8, 9, 10, and 15). In vivo embryo quantity and quality (total structures, morulae, blastocysts, viable, degenerated and blocked embryos) were recorded in each SOV protocol. Finally, embryo quality in both protocols was assessed by the analysis of the expression level of crucial genes for early embryo development (OCT4, IFNt, CDX2, BCL2, and BAX). P4 and cortisol concentration peaks in both SOV protocols were obtained on Day 15 and Day 8, respectively, which were statistically different compared to the other time-points (p < 0.05). Ovarian dimensions increased from Day 0 to Day 15 irrespective of the SOV protocol considered (p < 0.05). Significant changes in CL number were observed over time till Day 15 irrespective of the SOV protocol applied (p < 0.05), being non- significantly different between SOV protocols within each time-point (p > 0.05). The number of CL was higher on Day 15 in the bscrFSH group compared to the NIH-FSH-p group (p < 0.05). The number of embryonic structures recovered was higher in the bscrFSH group (p = 0.025), probably as a result of a tendency towards a greater number of follicles developed compared to the NIH-FSH-p group. IFNt and BAX were overexpressed in embryos from the bscrFSH group (p < 0.05), with a fold change of 16 and 1.3, respectively. However, no statistical differences were detected regarding the OCT4, CDX2, BCL2, and BCL2/BAX expression ratio (p > 0.05). In conclusion, including bscrFSH in SOV protocols could be an important alternative by reducing the number of applications and offering an improved ovarian response together with better embryo quality and superior performance in embryo production compared to NIH-FSH-p SOV protocols.

Hormonal Management for the Induction of Luteolysis and Ovulation in Andalusian Jennies: Effect on Reproductive Performance, Embryo Quality and Recovery Rate

Two prostanglandins (luprostiol, LUP, and dinoprost, DIN) and two ovulation-inducing agents (human Chorionic Gonadotropin, hCG, and deslorelin, DES) were evaluated for luteolysis and estrus induction, and for ovulation induction, respectively, in embryo donor jennies. Twenty-six fertile Andalusian jennies were used. In Experiment 1, jennies (n = 112 cycles) were randomly treated with either LUP or DIN after embryo flushing. In Experiment 2, donors (n = 84 cycles) were randomly treated with either hCG or DES to induce ovulation. No differences were found between prostaglandins for all variables studied (prostaglandin–ovulation interval (POI), interovulatory interval (IOI), embryo recovery rate (ERR), positive flushing rate (PFR) and embryo grade (EG)). The ovulation rate was similar for hCG and DES (60.9% vs. 78.7%). However, the interval to ovulation (ITO) was affected (62.61 ± 7.20 vs. 48.79 ± 2.69 h). None of the other variables studied (ERR, PFR and EG) were affected (p > 0.05), except for embryo quality (p = 0.009). In short, both prostaglandins evaluated are adequate to induce luteolysis and estrus. Both ovulation-inducing agents hastened ovulation, but DES seems to be more effective than hCG. Follicular diameter affected the interval from treatment to ovulation, and high uterine edema was related to low embryo quality.

Impact of Living in Different Altitudes on Intracytoplasmic Sperm Injection (ICSI) Outcomes for Infertile Couples in Southwestern Region of Saudi Arabia, a Retrospective Study

Objective: The aim of this study was to determine the effect of different altitudes in the Southwestern region of Saudi Arabia on ICSI outcomes, fertilization rate, embryo quality, pregnancy rate, and miscarriage rates for infertile couples. Materials and Methods: This is a retrospective study on 551 infertile couples carried out in the Assisted Reproductive Technology unit at the Maternity and Childern Hospital in Abha, Saudi Arabia between 2018 and 2019 to compare ICSI outcomes in different altitudes. Low altitude (205 patients), mild altitude (86 patients) and high altitude (260 patients). Main result measurements: fertilization rates, embryo quality, clinical pregnancy and miscarriage rates at different altitudes. Results: The data showed that there were no significant differences (P>0.67) in fertilization rates and the number of good-quality embryos between different altitudes. Importantly, clinical pregnancy rates were similar between groups and there was no significant difference in the miscarriage rates between high, mild and low altitudes. Conclusions: This study demonstrates that there is now increased risk of miscarriage or low pregnancy rates with different altitudes in Southwestern region of Saudi Arabia. This suggests that altitude changes has no obvious risk on pregnancy rate and pregnancy outcome.

Autophagy as a Therapeutic Target of Natural Products Enhancing Embryo Implantation

Infertility is an emerging health issue worldwide, and female infertility is intimately associated with embryo implantation failure. Embryo implantation is an essential process during the initiation of prenatal development. Recent studies have strongly suggested that autophagy in the endometrium is the most important factor for successful embryo implantation. In addition, several studies have reported the effects of various natural products on infertility improvement via the regulation of embryo implantation, embryo quality, and endometrial receptivity. However, it is unclear whether natural products can improve embryo implantation ability by regulating endometrial autophagy. Therefore, we performed a literature review of studies on endometrial autophagy, embryo implantation, natural products, and female infertility. Based on the information from these studies, this review suggests a new treatment strategy for female infertility by proposing natural products that have been proven to be safe and effective as endometrial autophagy regulators; additionally, we provide a comprehensive understanding of the relationship between the regulation of endometrial autophagy by natural products and female infertility, with an emphasis on embryo implantation.

Randomized allocation of oocytes to IVF or ICSI for IVF-naïve cases with unexplained infertility in an IVF-ICSI Split protocol favors ICSI to optimize live birth outcomes

In assisted reproduction treatments (ART), applying the ICSI method for fertilization of oocytes rather than traditional IVF method, is regarded as controversial for two reasons, namely utility and safety. Our study examines an IVF-ICSI Split model for couples with unexplained infertility, where male factor is meticulously excluded and ART is conducted by a strict algorithm, a commitment to blastocyst culture, along with single embryo transfers and a high commitment to cryopreservation. From 242 treatment cycles, 3346 oocytes recovered (13.8 per OPU) were randomly allocated to IVF or ICSI and the fertilization rates standardized to the number of 2PNS arising from each group applying the metaphase II oocyte number identified for the ICSI group, as the denominator for both groups. The fertilization rates were significantly higher overall for ICSI (83.2% vs 65.4%; p<0.0001), being most pronounced for women under 40 years. The resultant embryos had equivalent implantation rates in both fresh ET and frozen (FET) cycles with no significant differences in pregnancy rates, miscarriage rates or live birth outcomes indicating equivalent embryo quality. However, there were significantly higher numbers of ICSI-generated embryos cryopreserved and subsequent FET procedures showed higher live birth rates (21 births vs 6 births; p<0.005) and potential livebirths (214 births vs 104 births; p<0.0001). No congenital fetal abnormalities were detected in any of the 199 babies delivered during the study period to December 2020, neither IVF-generated nor ICSI-generated. Whilst the data strongly favors ICSI, there were 2 women (from 26 with fertilization in one arm only) who demonstrated fertilization only in the IVF arm of the study. We conclude that the IVF-ICSI Split model should be undertaken on all IVF-naïve women with unexplained infertility to determine the appropriate fertilization mode, albeit ICSI will be safely preferred for >90% of cases.

The Significance of Planned Fertility Preservation for Women With Endometrioma Before an Expected Ovarian Cystectomy

Endometrioma is known to reduce the ovarian reserve and the extent of the decrease is more severe when ovarian surgery is performed. Therefore, to prevent this decline in fertility, patients with endometrioma are considered candidates for preoperative fertility preservation (FP). In this study, we evaluate the efficacy of FP in women with endometrioma before planned ovarian surgery. A total of 95 cycles in 62 patients with endometrioma, undergoing controlled ovarian stimulation (COS) for FP using a gonadotropin-releasing hormone (GnRH) antagonist protocol before an expected ovarian surgery, were enrolled retrospectively. COS outcomes were compared according to endometrioma laterality. Additionally, first COS cycle outcomes in patients with endometrioma were compared with those in infertile patients, or in patients with a benign ovarian cyst using propensity score matching. When multiple COS cycles were performed, the results of cumulative cycles were analyzed. Embryo quality was worse in the bilateral endometrioma group. Compared with the infertile patient group, the patients with endometrioma had significantly lower Anti-Müllerian Hormone (AMH) and fewer numbers of oocytes retrieved (median, 3.3 vs. 1.2, p&lt;0.001; 7.0 vs. 4.0, p=0.009, respectively). Compared with mature oocytes in infertile patients or patients with a benign cyst, mature oocytes were fewer in patients with endometrioma, but this was not statistically significant (median, 4.0 vs. 3.0, p=0.085; 5.5 vs. 3.0, p=0.052, respectively). The median value of the cumulative number of cryopreserved oocytes or embryos was 14.5 up to the fourth cycle compared to 3 up to the first cycle, with cumulative effect. Women with endometrioma should be counseled for FP before planned ovarian cystectomy. The number of cryopreserved oocytes or embryos can be increased by repeated cycles.

Small Noncoding RNAs in Reproduction and Infertility

Infertility has been reported as one of the most common reproductive impairments, affecting nearly one in six couples worldwide. A large proportion of infertility cases are diagnosed as idiopathic, signifying a deficit in information surrounding the pathology of infertility and necessity of medical intervention such as assisted reproductive therapy. Small noncoding RNAs (sncRNAs) are well-established regulators of mammalian reproduction. Advanced technologies have revealed the dynamic expression and diverse functions of sncRNAs during mammalian germ cell development. Mounting evidence indicates sncRNAs in sperm, especially microRNAs (miRNAs) and transfer RNA (tRNA)-derived small RNAs (tsRNAs), are sensitive to environmental changes and mediate the inheritance of paternally acquired metabolic and mental traits. Here, we review the critical roles of sncRNAs in mammalian germ cell development. Furthermore, we highlight the functions of sperm-borne sncRNAs in epigenetic inheritance. We also discuss evidence supporting sncRNAs as promising biomarkers for fertility and embryo quality in addition to the present limitations of using sncRNAs for infertility diagnosis and treatment.

Maternal parity affects Day 8 embryo gene expression in old mares

As sport career is a priority in most of equine breeds, mares are frequently bred for the first time at an advanced age. Both age and first gestation were shown to have a deleterious effect on reproduction outcomes, respectively on fertility and offspring weight but the effect mare's parity in older mares on embryo quality has never been considered. The aim of this project was to determine the effect of old mare's nulliparity on gene expression in embryos. Day 8 post ovulation embryos were collected from old (10-16 years old) nulliparous (ON, N=5) or multiparous (OM, N=6) non-nursing Saddlebred mares, inseminated with the semen of one stallion. Pure (TE_part) or inner cell mass enriched (ICMandTE) trophoblast were obtained by embryo bisection and paired end, non-oriented RNA sequencing (Illumina, NextSeq500) was performed on each hemi-embryo. To discriminate gene expression in the ICM from that in the TE, deconvolution (DeMixT R package) was used on the ICMandTE dataset. Differential expression was analyzed (DESeq2) with embryo sex and diameter as cofactors using a false discovery rate <0.05 cutoff. Although the expression of only a few genes was altered by mare's nulliparity (33 in ICM and 23 in TE), those genes were related to nutrient exchanges and responses to environment signaling, both in ICM and TE, suggesting that the developing environment from these mares are not optimal for embryo growth. In conclusion, being nulliparous and old does not seem to be the perfect match for embryonic development in mares.