Search Results (15)

Cryopreserved pig semen tends to produce excessive reactive oxygen species (ROS) during the thawing process, which leads to a decline in semen quality during in vitro storage. Mogroside V (MV) has been proven to be an effective antioxidant, and previous research has shown that MV can delay oocyte aging and improve the in vitro maturation efficiency of pig oocytes. However, the role of MV in the cryopreservation capacity of animal sperm remains unclear. To evaluate the effect of MV on sperm motility after thawing, different concentrations of MV (0, 25, 50, 75, 100 μmol/L) were added to the thawing medium. By comparing the sperm motility and kinematic parameters in the thawing medium with different MV concentrations and incubation times (0, 1, 2, and 4 h), we ultimately selected sperm thawed immediately in the medium supplemented with 75 μmol/L MV for subsequent experiments. Compared with the control group, the sperm thawing medium containing MV improved sperm quality during the freeze–thaw process. Immediate evaluation after thawing at 37 °C showed that supplementation with 75 μmol/L MV produced an optimal effect on the maintenance of motility, plasma membrane integrity, the acrosome integrity, the ROS levels, and the T-AOC activity. In conclusion, MV supplementation improves the quality of frozen–thawed sperm by enhancing sperm function and preventing oxidative stress. Full article

This study aimed to analyze the effect of curcumin on the antioxidant properties and fertility of freeze–thawed bovine spermatozoa and bovine oocytes. In this study, curcumin concentrations of 0, 5, 10, 25, and 50 µM were added bovine sperm cryopreservation solution and oocyte IVM medium to assess sperm quality, antioxidant properties, oocyte maturation, IVF rate, and embryonic development. The results demonstrated that adding curcumin to the cryopreservation solution significantly improved the viability, motility, and acrosome integrity of bull sperm after freezing and thawing (p < 0.05). The addition of 25 µM curcumin resulted in the best sperm quality. Analysis of antioxidant capacity showed that 25 µM curcumin significantly increased the activities of MMP and antioxidant enzymes, such as CAT, SOD, and GSH-PX, and lowered the levels of MDA and ROS (p < 0.05). Adding curcumin to the in vitro maturation medium notably enhanced the maturation rate and decreased DNA fragmented nuclei of bovine oocytes (p < 0.05), with optimal outcomes observed at 25 and 50 µM curcumin. Totals of 25 and 50 µM curcumin markedly elevated GSH and MMP (p < 0.05), reduced ROS and malondialdehyde concentrations (p < 0.05), and significantly enhanced fertilization rates and blastocyst formation (p < 0.05). In conclusion, incorporating curcumin into both the bovine semen cryopreservation solution and the oocyte IVM medium significantly improved the quality of frozen–thawed sperm, antioxidant activity, oocyte maturation, IVF rate, and embryonic development. Full article

Antioxidants protect cellular function and structure by neutralizing the oxidative stress caused by increased reactive oxygen species (ROS) during sperm freezing. Studies on cryopreservation using various antioxidants have demonstrated encouraging results. Many studies have used antioxidants to increase the efficiency of sperm freezing and to improve the success rate of artificial insemination and pregnancy. Manganese (III) tetrakis (4-benzoic acid) porphyrin chloride (MnTBAP) is a newly synthesized antioxidant with positive effects on sperm morphology and capacitation in humans, rams, and stallions. In this study, porcine semen was treated with 0, 50, 100, and 150 μM of MnTBAP based on a Tris–egg-yolk extender and frozen to determine whether MnTBAP can assist the status of sperm during cryopreservation. First, motility was assessed using the computer-assisted sperm analysis (CASA) system, with the 100 μM treatment group showing the highest motile rate (66.8%) compared with that of the other groups (control, 51.1%; 50 μM and 150 μM, 59.6%); therefore, the remaining analyses were conducted comparing the two groups (control vs. 100 μM group; p < 0.01). Second, fluorescence staining was applied to examine the control and 100 μM groups using fluorescence microscopy. The viability (41.7% vs. 62.4%) and the acrosome integrity (77.9% vs. 86.4%) differed significantly (p < 0.05). In addition, the mitochondrial membrane potential (MMP) was 46.5% vs. 51.9%; the fragmentation rate, estimated using the Sperm-sus-Halomax kit, was 63.4% vs. 57.4%; and the detected caspase activity was 30.1% vs. 22.9%. These tended to be higher in the treated group but did not differ significantly. Third, measurements using FACSLyric revealed that the 100 μM treatment group exhibited a state of elevated normal lipid arrangement within the plasma membrane and diminished levels of apoptosis and ROS (p < 0.01). We assessed the expression of genes relevant to antioxidant effectiveness using real-time RT-qPCR. Our findings indicated significant alterations in the expression levels of various mRNA species, with the exception of NOX5 (p < 0.05). Finally, the straws were dissolved and used to treat matured denuded oocytes to investigate the effect on fertilization and embryo development in vitro. The cleavage rate was (77.6% vs. 84.1%), and the blastocyst rate was 9.7% vs. 11.4% (p < 0.05). In conclusion, these results suggest that MnTBAP positively affected sperm freeze–thawing, improving the fertilization capacity, and leading to increased embryo development. Full article

This paper uses a SWOT (strengths, weaknesses, opportunities, and threats) analysis to overview the option of fertility preservation in women with genetic diseases, who would later use preimplantation genetic testing for monogenic disorders, in order to not transmit their condition. Strengths associated with elective oocyte freezing are ethical considerations, overall maternal and fetal safety, and effectiveness, if performed at <35 years of age. Weaknesses are related to costs and rare but present (<1–3%) risks of maternal complications. Counselling on fertility management aimed at preventing infertility offers a valuable opportunity, the same as it has been in oncological patients’ care. The potentially high percentage of women with genetic conditions who would return to use their frozen oocytes also represents an opportunity together with the minimization of the need for egg donation, which has higher obstetrical risks compared to the use of autologous oocytes. Finally, a threat is represented by the potential psychological distress to young women who could never attempt to become pregnant through preimplantation genetic testing, or do it before any decline in their fertility. Potential unknown future long-term health risks for children conceived after egg vitrification/thawing are also a threat, but current knowledge is reassuring. Altogether, early counselling on the option of fertility preservation should thus be incorporated into standard care of all patients with any genetic condition. Full article

The purpose of this study is to evaluate the live birth outcome following oocyte thaw in women who underwent social egg freezing at Guy’s Hospital, alongside a detailed published literature review to compare published results with the current study. A retrospective cohort study was conducted between January 2016 and March 2022 for all women who underwent egg freezing during this period. Overall, 167 women had 184 social egg freezing cycles. The mean age at freeze was 37.1 years and an average of 9.5 eggs were frozen per retrieval. In total, 16% of the women returned to use their frozen eggs. The mean egg thaw survival rate post egg thaw was 74%. The mean egg fertilisation rate was 67%. The pregnancy rate achieved per embryo transfer was 48% and the live birth rate per embryo transfer was 35%. We also noted that irrespective of age at freezing, a significantly high live birth rate was achieved when the number of eggs frozen per patient was 15 or more. Despite the rapid increase in social egg freezing cycles, the utilisation rate remains low. Pregnancy and live birth rate post thaw are encouraging if eggs are frozen at a younger age and if 15 eggs or more were frozen per patient. Full article

This study aims to analyze the cycle characteristics, pregnancy, and neonatal outcomes in early rescue intracytoplasmic sperm injection (r-ICSI) cycles in normal and hyper-ovarian response women in their first IVF/ICSI attempts. Data from short-term in vitro fertilization (IVF, N = 7148), early r-ICSI (N = 618), and ICSI (N = 1744) cycles were retrospectively analyzed from normal and hyper-ovarian women who underwent their first IVF/ICSI cycles at our center from October 2015 to October 2021. The r-ICSI group was subdivided into partial r-ICSI (N = 451) and total r-ICSI (N = 167) based on the number of fertilized oocytes in the IVF part. Cyclic characteristics, pregnancy, delivery and neonatal outcomes in the fresh cycle were compared among the four groups; pregnancy, delivery and neonatal outcomes in frozen-thawed cycles were compared regarding cleavage and blastocyst transfers derived from r-ICSI cycles. Partial r-ICSI cycles showed different cyclic characteristics compared to total r-ICSI cycles, presenting as elevated AMH and estradiol levels on trigger day and an increased number of oocytes retrieved. Early r-ICSI delayed blastocyst development as seen by the increase in the number of day 6 blastocysts. There was no significant difference among the groups in clinical pregnancy, pregnancy loss, and live birth in fresh cleavage-stage embryo transfer cycles. However, early r-ICSI groups showed a reduction in clinical pregnancy and live birth rates in fresh blastocyst transfer cycles but not in the frozen-thawed cycles. For pregnant women, early r-ICSI did not show a negative effect on the risk of preterm birth, Cesarean section, neonatal birth weight, and sex ratio. In conclusion, early r-ICSI had comparable pregnancy, delivery, and neonatal outcomes when compared with short-term IVF and ICSI groups in fresh cleavage-stage embryo transfer cycles, but early r-ICSI did result in reduced pregnancy outcomes in fresh blastocyst embryo cycles, possibly due to delayed blastocyst development and asynchronization with the endometrium. Full article

Magnetized water is defined as the amount of water that has passed through a magnet. The magnetic field weakens the hydrogen bonds between the water molecules, leading to the magnetized liquid acquiring special characteristics such as easy supercooling and forming smaller ice crystals. We researched the influences of a magnetized freezing extender on cell membrane damage and in vitro fertilization of boar sperm during cryopreservation. The freezing extenders were passed through 0, 2000, 4000, and 6000 gausses (G) of magnetic devices using a liquid cycling pump system and then used for the sperm freezing process. The damage to plasma, acrosomal, and mitochondrial membranes in frozen-thawed spermatozoa was investigated by flow cytometry, and motility was assessed using the CASA system. The fertility of frozen-thawed sperm was estimated using in vitro fertilization. The damage to the membranes was significantly decreased in the magnetized freezing extender by the 6000 G magnetic field compared to that of the control in frozen-thawed sperm, and motility was increased in the 6000 G group. Although there were no significant differences in the cleavage rates of in vitro fertilized oocytes among the treatment groups, the ratio of blastocyst formation increased in the magnetized freezing extender groups compared with that in the control group. The number of blastocysts was significantly higher in the 4000 G group than in the 0 G group. In conclusion, these results suggest that a magnetized freezing extender could improve the freezability of sperm and the development of oocytes fertilized in vitro with frozen-thawed sperm. Full article

We aimed to assess the efficacy of accumulated embryo transfer (ACC-ET) through several controlled ovarian hyperstimulation (COS) cycles to increase the rates of pregnancy in patients with poor ovarian response (POR). We retrospectively reviewed the medical records of 588 patients with POR under 43-years old who underwent embryo transfer from January 2010 to December 2015. We compared the pregnancy rate (PR), clinical pregnancy rate (CPR), and live birth rate (LBR) between ACC-ET (frozen-thawed: 47; fresh + frozen-thawed: 24) group (n = 71) and fresh ET groups (n = 517). Characteristics of ACC-ET patients were similar to those of fresh ET groups (Age: 38.1 ± 3.5 vs. 38.2 ± 3.7, p = 0.88; Anti Müllerian Hormone (AMH; ng/mL): 0.5 ± 0.4 vs. 0.6 ± 0.6, p = 0.38; follicle stimulating hormone (FSH: mIU/mL): 11.9 ± 8.0 vs. 10.8 ± 9.0, p = 0.35). The total number of transferred embryos (3.1 ± 0.9 vs. 1.5 ± 0.7, p = 0.00), PR (29.6% (21/71) vs. 18.8% (97/517), p = 0.040), and CPR (23.5% (16/68) vs. 14.0% (71/508) p = 0.047) were significantly higher in the ACC-ET group than in the fresh ET group. In addition, PR, CPR, and LBR increased with the number of ET in the fresh ET group. However, there were no significant differences observed in LBR between ACC-ET and fresh ET groups (14.9% (10/67) vs. 9.8% (50/508), p = 0.203). From our knowledge, there is no clinical evidence reported to prove that transfer of multiple embryos of adequate quality obtained through multiple cycles can compensate for the limited number of retrieved oocytes from POR patients. We concluded that ACC-ET from several COS cycles could be an alternative method to increase PR and CPR in <43-year-old patients with POR. Full article

The objectives of this study were to evaluate the properties of sperm motility and morphology under induced oxidative stress, compare the antioxidant capacity of dithiothreitol (DTT) and glutathione (GSH) following the cryopreservation of Large White boar semen, investigate the ability of cryopreserved Large White boar semen to fertilize the matured gilts oocytes and compare the efficacy of DTT and GSH antioxidants in improving the oocyte fertilization by cryopreserved Large White boar semen. The semen was collected from three Large White boars (ten ejaculates per boar) and transported (37 °C) to the laboratory. Semen freezing extenders were supplemented with 5 mM DTT, 5 mM GSH and a combination of 2.5 mM DTT + 2.5 mM GSH. A liquid nitrogen vapor method was used to freeze boar semen. Gilts’ ovaries were collected from the local abattoir and transported (37 °C) to the laboratory. The slicing method was used to retrieve the oocytes from the ovaries. Fresh semen and frozen-thawed semen were used for in vitro fertilization (IVF). For frozen-thawed semen, four treatments (control, 5 mM DTT, 5 mM GSH, and a combination of 2.5 mM DTT + 2.5 mM GSH) were used during IVF in order to evaluate the fertilizing ability of the antioxidants. The supplementation of 5 µM DTT to H₂O₂-treated semen significantly improved progressive motility (PM) by 14.82%. A combination of 2.5 mM DTT + 2.5 mM GSH treatment reduced percentage of sperm total motility (TM) and rapid motility (RAP) following thawing (p < 0.05). Fresh semen and a combination of 2.5 mM DTT + 2.5 mM GSH treatment recorded a higher percentage of zygotes with polyspermy (p < 0.05). The control treatment numerically recorded a high percentage of zygotes with 1 PN, while the 5 mM DTT treatment recorded a high percentage of zygotes with 2 PN. Full article

In vitro produced (IVP) embryos show large metabolic variability induced by breed, culture conditions, embryonic stage and sex and gamete donors. We hypothesized that the birth potential could be accurately predicted by UHPLC-MS/MS in culture medium (CM) with the discrimination of factors inducing metabolic variation. Day-6 embryos were developed in single CM (modified synthetic oviduct fluid) for 24 h and transferred to recipients as fresh (28 ETs) or frozen/thawed (58 ETs) Day-7 blastocysts. Variability was induced with seven bulls, slaughterhouse oocyte donors, culture conditions (serum + Bovine Serum Albumin [BSA] or BSA alone) prior to single culture embryonic stage records (Day-6: morula, early blastocyst, blastocyst; Day-7: expanding blastocyst; fully expanded blastocysts) and cryopreservation. Retained metabolite signals (6111) were analyzed as a function of pregnancy at Day-40, Day-62 and birth in a combinatorial block study with all fixed factors. We identified 34 accumulated metabolites through 511 blocks, 198 for birth, 166 for Day-62 and 147 for Day-40. The relative abundance of metabolites was higher within blocks from non-pregnant (460) than from pregnant (51) embryos. Taxonomy classified lipids (12 fatty acids and derivatives; 224 blocks), amino acids (12) and derivatives (3) (186 blocks), benzenoids (4; 58 blocks), tri-carboxylic acids (2; 41 blocks) and 5-Hydroxy-l-tryptophan (2 blocks). Some metabolites were effective as single biomarkers in 95 blocks (Receiver Operating Characteristic – Area Under the Curve [ROC-AUC]: 0.700–1.000). In contrast, more accurate predictions within the largest data sets were obtained with combinations of 2, 3 and 4 single metabolites in 206 blocks (ROC-AUC = 0.800–1.000). Pregnancy-prone embryos consumed more amino acids and citric acid, and depleted less lipids and cis-aconitic acid. Big metabolic differences between embryos support efficient pregnancy and birth prediction when analyzed in discriminant conditions. Full article

This study aimed to investigate the effect of L-Carnitine (LC) supplementation during in vitro maturation (IVM) of canine oocytes on nuclear maturation, fertilization status, and preimplantation development. Cumulus–oocyte complexes (COCs) collected from the ovaries of ovariohysterectomized female dogs were matured in vitro for 72 h in a TCM-199 medium supplemented with (0.1, 0.3, 0.6, 1.0, or 2.0 mg/mL) or without (0.0 mg/mL) LC. Matured oocytes were fertilized in vitro with frozen–thawed spermatozoa, and zygotes were cultured in a SOF medium for 7 days. IVM rates were higher (p ≤ 0.05) in 0.3 and 0.6 mg/mL LC supplemented groups than in the control (0.0 mg/mL LC) and other LC groups. Fertilization (18 h postinsemination (pi)) and cleavage (2–16-cell stage at day 3 pi) rates were higher (p ≤ 0.05) in the 0.6 mg/mL LC group than in the control and 0.1, 1.0, and 2 mg/mL LC supplemented groups. Interestingly, 4.5% of fertilized oocytes developed to morula (day 5 pi) in the 0.6 mg/mL LC group, which was higher (p ≤ 0.05) than those developed in the 0.3 mg/mL group (1.0%). No cleaved embryos developed to morula in other groups. In conclusion, LC supplementation at 0.6 mg/mL during IVM of canine oocytes improved their maturation, fertilization, and preimplantation embryo development rates following IVF and in vitro culture (IVC). Full article

The objective of the present study was to establish conditions for using technology that can potentially enhance the efficiency of bovine embryos derived from in vitro fertilization (IVF) with frozen semen. Frozen semen from selected bulls can be stored indefinitely in liquid nitrogen as genetic resources; however, these resources are considered consumable because they cannot be regenerated. Therefore, to optimize the utilization of frozen semen, as many oocytes as possible should be fertilized with one straw. However, a sufficient number of prepared oocytes might not be available for one experiment, which can limit the use of the total spermatozoa population. Thus, an economical method for producing embryos needs to be established by optimizing technology for transplantable embryos. In this study, the utilization of frozen semen was increased by dividing the straw with an ultrasonic cutter. The post-thaw survival rate of uncut straws from Korean Proven Bulls did not differ from that of half cuttings. When ultrasonic cutting was applied to frozen semen, spermatozoa could be prepared for IVF trials at least four times, and blastocysts were produced. Therefore, cutting frozen semen with an ultrasonic cutter represents a potentially useful tool to expand genetic resources from excellent breeding stocks. This approach could also be valuable in the field of IVF of endangered species or rare breeds for their preservation, as well as in ovum pick-up (OPU) techniques. Full article

Background and objectives: Cancer incidence is growing with younger patients diagnosed with this disease every year. Improved cancer diagnostics and treatment lead to better survival of cancer patients. However, after aggressive chemo- or radiotherapy, cancer survivors suffer from various degrees of subfertility or infertility. Several fertility preservation technologies have been developed for young cancer patients: cryopreservation of germ cells, embryos, or reproductive tissues. The best results have been shown by cryopreservation of sperm and embryos. Yet the success of using cryopreserved oocytes or reproductive tissues (ovarian and testicular) is still insufficient. Therefore, this study was designed to assess the vitality, viability, general quality, and safety of frozen–thawed human ovarian tissue for retransplantation using modern molecular tests. Materials and Methods: The new miRNA array test was used to evaluate miRNA expression in thawed ovarian tissue in combination with standard xenotransplantation and pathological examination of microslides. Results: Our results demonstrated that slow freezing is an efficient way (80%) to cryopreserve ovarian tissue with no structural damage afterwards. We have shown that xenotransplantation into immunodeficient mice, histology, and immunohistochemistry could be potentially replaced by more recent molecular methods. Conclusions: The latter method has shown that altered expression of miRNAs might be used as identifiers of normal/damaged tissue after further analysis. Newer, safer, and more specific approaches need to be developed in order to eliminate the risk of disease reoccurrence. Full article

The aim was to compare the blastocyst stages of red deer embryos in respect of in vitro fertilization (IVF) efficiency, morphology, apoptotic and proliferative abilities, and antioxidative potential according to the reproductive status of hinds. We used three experimental groups, including the ovaries collected post mortem on the 4th and 13th days of the estrous cycle and during pregnancy (n = 18). After oocyte maturation, frozen-thawed epididymal semen was used for IVF. Blastocyst quality, apoptotic potential by determining the mRNA expression of BAX, BCL-2, OCT4, SOX2, and placenta-specific 8 gene (PLAC8), and antioxidative potential of blastocysts were evaluated by determining the mRNA expression of CuSOD, MnSOD, and GPX as well as the enzymatic activity of superoxide dismutase and reduced glutathione. The highest development rate of expanded blastocyst, mRNA expression of BCL-2, OCT4, SOX2, and PLAC8 and mRNA expression and enzymatic activity of the antioxidative factors increased (p < 0.05) in blastocysts developed from the oocytes collected on the 4th day, compared to those developed from the oocytes collected on the 13th day of the cycle and during pregnancy. Our study indicates that the 4th day of the estrous cycle is the most effective period for oocyte collection for IVF and embryo development in hinds, considering quality parameters and antioxidative potential of the blastocysts. Full article

Background and Objective: During the last few years, a trend has been noted towards embryos being transferred at the blastocyst stage, which has been associated with improved rates regarding implantation and clinical pregnancy in comparison to cleavage stage embryo transfers. There is a limited number of studies investigating this notion in oocyte donation cycles employing cryopreserved embryos. The aim of this study is to evaluate the implantation potential and clinical pregnancy rates between the day 3 cleavage stage and blastocyst stage embryo transfers in oocyte donation cycles employing vitrified embryos. Methods: This is a retrospective evaluation of oocyte donation frozen–thawed transfers completed in our clinic from January 2017 to December 2017. Intracytoplasmic sperm injection was conducted for all oocytes. Following fertilization, all embryos were cryopreserved either at the cleavage or blastocyst stage. Embryo transfer of two embryos was performed under direct sonographic guidance in all cases. Results: Our results confirmed a 55.6% clinical pregnancy (CP) resulting from day 3 embryo transfers, a 68.8% CP from day 5, and 71.4% CP from day 6. Significantly improved pregnancy rates were related to embryo transfers at the blastocyst stage when compared to cleavage stage transfers (68.9% and 55.6% respectively, p = 0.016). The risk with regards to multiple pregnancies was similar. Conclusion: Our findings indicate that in oocyte donation cycles employing vitrified embryos, embryo transfer at the blastocyst stage is accompanied with a significant improvement in pregnancy rates and merits further investigation. Full article