The ever-increasing number of genetically engineered mouse models highlights the need for efficient archiving and distribution of these lines. Sperm cryopreservation has become the preferred technique for the majority of these models due to its low requirement of costs, time, and experimental animals. Yet, current in vitro fertilization (IVF) protocols either exhibit decreased fertilization efficiency for the most popular C57BL/6 strain, as recently demonstrated by us, or require costly and difficult-to-prepare media, respectively. As a result, we previously developed SEcuRe, a modified IVF protocol with low costs and high fertilization efficiency. The popular basal fertilization medium, Cook's® proprietary "Research vitro fert" (RVF), used in this protocol has recently been discontinued. As a result, the application of the SEcuRe approach and other IVF protocols employing this medium has been severely limited.

We recently reported evidence suggesting associations between urine cadmium concentrations, reflecting long-term exposure, measured in 25 female patients (relative risk = 1.41, P = 0.412) and 15 of their male partners (relative risk = 0.19, P = 0.097) and oocyte fertilization in vitro. Blood cadmium concentrations reflect more recent exposure.

This study aimed to establish a protocol for in vitro embryo production using epididymal sperm (EP). Samples were obtained from ejaculated sperm (EJ) and the epididymis of 7 Gir bulls. First, the effect of heparin (+) on the viability, longevity (Experiment 1) and fertilization rates (Experiment 2) of the EP was evaluated. In experiment 2, a pool of EP and EJ sperm (n = 7) was coincubated with cumulus-oocyte complexes (COCs) for 0, 3, 6, 12 and 18 h, and the fertilization rate (FR) was evaluated. A third experiment was performed to test sperm treatments for IVP using the Percoll (P) or PureSperm (PS) gradients or a spTALP wash for sperm selection. Cleavage, blastocyst rate (BR) and embryo sex were evaluated. In experiment 4, embryos were produced using 6, 12, and 18 h of sperm-oocyte coincubation. The cleavage, BR, and total number and percentage of apoptotic cells were determined. Heparin affected EP viability, longevity and FR. After 6 h, 82% of the oocytes were fertilized in the EP+ group, a higher value (P<0.05) than that in the EJ (19%) and EP- (42%) groups. At 12 and 18 h, FR remained higher in the EP+ group, and a gradual increase in polyspermy was observed. The use of a P or PS gradient yielded a similar BR on D7 (54% and 52%), which was higher than the rate obtained using the washing method (37%). The embryos produced by EP and selected in a P or PS gradient resulted in a sex deviation in favor of male embryos (P>0.05). No differences (P>0.05) were observed among the groups that were coincubated for 6, 12 and 18 h with respect to embryo production, kinetics of development, total cell number and percentage of apoptotic cells. In conclusion, IVF time can be reduced to 6 h without affecting embryo production and quality. In addition, EP sperm selection can be performed by either a PS or P gradient.

No abstract available

We measured toxic metals in seminal plasma collected from 30 men using vitro fertilization (IVF), to evaluate associations with semen quality and IVF outcomes. A doubling in Hg-adjusted Pb concentration was associated with 47% lower total motile sperm. Positive associations were suggested for Hg with pregnancy and live birth, adjusted for Cd or Pb. A negative association was suggested for Hg-adjusted Cd with pregnancy. These data add to evidence indicating that toxic metals impact IVF.

In this study, we used more reliable experimental materials and methods to detect the effects of osteopontin (OPN) on boar sperm in vitro capacitation, acrosome reaction, and fertilization efficiency. We reorganized and obtained the OPN protein of the porcine source. Immunofluorescence and Western blot show the localization and expression of the OPN protein before and after sperm capacitation. To determine whether OPN can affect sperm during sperm capacitation, we examined cyclic adenosine monophosphate (cAMP) concentrations after sperm capacitation, and the results showed that OPN significantly increased the cAMP concentration in sperm (p < 0.05). Flow cytometry showed that 0.1 μg/mL OPN-treated sperm had better acrosome reaction ability. In vitro fertilization (IVF) showed that 0.1 μg/mL OPN significantly increased the rate of embryo division. In conclusion, this study found that 0.1 μg/mL porcine OPN protein can significantly improve porcine capacitated sperm motility, cAMP concentration after capacitation sperm, acrosome reaction ability, and embryo division during IVF and provides new clues to explore the mechanism of OPN's function on sperm.

Assisted reproductive technologies in the pig are critical for the production of genetically modified pigs as models of human disease and to improve production agriculture. Methods of oocyte maturation, fertilization, and culture all play an extremely important role in how the embryo, fetus, and offspring will develop. In this chapter, we discuss the historical methods and recent advances that have been essential in promoting efficient and competent embryo development. Here we describe the procedures that can be used to mature, fertilize, and culture pig embryos to the blastocyst stage.

Trace exposures to the toxic metals mercury (Hg), cadmium (Cd) and lead (Pb) may threaten human reproductive health. The aim of this study is to generate biologically-plausible hypotheses concerning associations between Hg, Cd, and Pb and in vitro fertilization (IVF) endpoints. For 15 female IVF patients, a multivariable log-binomial model suggests a 75% reduction in the probability for a retrieved oocyte to be in metaphase-II arrest for each microg/dL increase in blood Pb concentration (relative risk (RR)=0.25, 95% confidence interval (CI) 0.03-2.50, P=0.240). For 15 male IVF partners, each microg/L increase in urine Cd concentration is associated with an 81% decrease in the probability for oocyte fertilization (RR=0.19, 95% CI 0.03-1.35, P=0.097). Because of the magnitude of the effects, these results warrant a comprehensive study with sufficient statistical power to further evaluate these hypotheses.

In-vitro fertilization (IVF) is a popular method of resolving complications such as endometriosis, poor egg quality, a genetic disease of mother or father, problems with ovulation, antibody problems that harm sperm or eggs, the inability of sperm to penetrate or survive in the cervical mucus and low sperm counts, resulting human infertility. Nevertheless, IVF does not guarantee success in the fertilization. Choosing IVF is burdensome for the reason of high cost and uncertainty in the result. As the complications and fertilization factors are numerous in the IVF process, it is a cumbersome task for fertility doctors to give an accurate prediction of a successful birth. Artificial Intelligence (AI) has been employed in this study for predicting the live-birth occurrence. This work mainly focuses on making predictions of live-birth occurrence when an embryo forms from a couple and not a donor. Here, we compare various AI algorithms, including both classical Machine Learning, deep learning architecture, and an ensemble of algorithms on the publicly available dataset provided by Human Fertilisation and Embryology Authority (HFEA). Insights on data and metrics such as confusion matrices, F1-score, precision, recall, receiver operating characteristic (ROC) curves are demonstrated in the subsequent sections. The training process has two settings Without feature selection and With feature selection to train classifier models. Machine Learning, Deep learning, ensemble models classification paradigms have been trained in both settings. The Random Forest model achieves the highest F1-score of 76.49% in without feature selection setting. For the same model, the precision, recall, and area under the ROC Curve (ROC AUC) scores are 77%, 76%, and 84.60%, respectively. The success of the pregnancy depends on both male and female traits and living conditions. This study predicts a successful pregnancy through the clinically relevant parameters in In-vitro fertilization. Thus artificial intelligence plays a promising role in decision making process to support the diagnosis, prognosis, treatment etc.

Although conventional cryopreservation is a proven method for long-term, safe storage of genetic material, protocols used by the zebrafish community are not standardized and yield inconsistent results, thereby putting the security of many genotypes in individual laboratories and stock centers at risk. An important challenge for a successful zebrafish sperm cryopreservation program is the large variability in the post-thaw in vitro fertilization success (0 to 80%). But how much of this variability was due to the reproductive traits of the in vitro fertilization process, and not due to the cryopreservation process? These experiments only assessed the in vitro process with fresh sperm, but yielded the basic metrics needed for successful in vitro fertilization using cryopreserved sperm, as well. We analyzed the reproductive traits for zebrafish males with a strict body condition range. It did not correlate with sperm volume, or motility (P>0.05), but it did correlate with sperm concentration. Younger males produced more concentrated sperm (P<0.05). To minimize the wastage of sperm during the in vitro fertilization process, 10(6) cells/ml was the minimum sperm concentration needed to achieve an in vitro fertilization success of ≥ 70%. During the in vitro process, pooling sperm did not reduce fertilization success (P>0.05), but pooling eggs reduced it by approximately 30 to 50% (P<0.05). This reduction in fertilization success was due not to the pooling of the females' eggs, but to the type of tools used to handle the eggs. Recommendations to enhance the in vitro process for zebrafish include: 1) using males of a body condition closer to 1.5 for maximal sperm concentration; 2) minimizing sperm wastage by using a working sperm concentration of 10(6) motile cells/ml for in vitro fertilization; and 3) never using metal or sharp-edged tools to handle eggs prior to fertilization.

Autophagy contributes to the clearance and recycling of macromolecules and organelles in response to stress. We previously reported that vitrified mouse oocytes show acute increases in autophagy during warming. Herein, we investigate the potential role of Atg7 in oocyte vitrification by using an oocyte-specific deletion model of the Atg7 gene, a crucial upstream gene in the autophagic pathway.

Mouse embryos forming blastocoele early vs those forming late are predominantly male. We examined whether the male advantage could be recognized at an earlier stage of development.

Difructose dianhydride IV (DFA-IV) is produced from levan, which is a natural polysaccharide that belongs to the fructan family, through the activity of levan fructotransferase (LF) derived from microorganisms. Recently, DFA-IV has been expected to have diverse applications in the food and medical industry. Here, we examined the potential application of DFA-IV forin vitro fertilization (IVF) in pigs. In the assessment of acrosomal integrity during incubation, intact acrosomal or viable spermatozoa were highly sustained in 0.1% or 0.25% DFA-IV (69.8%-70.8%,P<0.05). Reactive oxygen species (ROS) levels during sperm incubation decreased following the addition of DFA-IV, and 0.1%-0.5% DFA-IV in particular significantly decreased ROS production relative to that seen with no addition or 0.75% DFA-IV. Total fertilization (mono+ polyspermic oocyte) rate was significantly higher in the addition of 0.1% DFA-IV (94.2%) than with other concentrations (71.8%-86.7%,P<0.05). When using reduced IVF times and lower sperm numbers, we found that addition of 0.1%-0.5% DFA-IV significantly increased the fertilization rate (P<0.05). Fertilized oocytes treated with 0.1% DFA-IV exhibited higher embryonic development and blastocyst formation than those treated with other concentrations (P<0.05). Consequently, the addition of DFA-IV during IVF improved fertilization and embryonic development, suggesting the possible use of novel sugars for enhancement of assisted reproductive technology (ART) in mammals.

Empiric therapy for patients with unexplained recurrent pregnancy loss (URPL) is not precise. Some patients will ask for assisted reproductive technology due to secondary infertility or advanced maternal age. The clinical outcomes of URPL patients who have undergone in vitro fertilization-embryo transfer (IVF-ET) require elucidation. The IVF outcome and influencing factors of URPL patients need further study.

Human embryos produced by in vitro fertilization (IVF) are usually cultured to day 6 after insemination, and good quality of embryos should develop to blastocyst stage at days 5 and 6. However, some embryos develop slowly, thus they may form blastocysts on day 7. Most IVF laboratories culture embryos to day 6 and then discard retarded embryos that do not develop to blastocyst stage. It has been reported that transfer of day 7 blastocysts can yield pregnancy although the rates were very low. In the present study, we evaluated the prevalence of aneuploidy in day 7 human blastocysts and also assessed embryo implantation after transfer of normal euploid blastocysts developed on day 7.

Infertility affects one in six of the population and increasingly couples require treatment with assisted reproductive techniques. In vitro fertilization (IVF) treatment is most commonly conducted using exogenous FSH to induce follicular growth and human chorionic gonadotropin (hCG) to induce final oocyte maturation. However, hCG may cause the potentially life-threatening iatrogenic complication "ovarian hyperstimulation syndrome" (OHSS), which can cause considerable morbidity and, rarely, even mortality in otherwise healthy women. The use of GnRH agonists (GnRHas) has been pioneered during the last two decades to provide a safer option to induce final oocyte maturation. More recently, the neuropeptide kisspeptin, a hypothalamic regulator of GnRH release, has been investigated as a novel inductor of oocyte maturation. The hormonal stimulus used to induce oocyte maturation has a major impact on the success (retrieval of oocytes and chance of implantation) and safety (risk of OHSS) of IVF treatment. This review aims to appraise experimental and clinical data of hormonal approaches used to induce final oocyte maturation by hCG, GnRHa, both GnRHa and hCG administered in combination, recombinant LH, or kisspeptin. We also examine evidence for the timing of administration of the inductor of final oocyte maturation in relationship to parameters of follicular growth and the subsequent interval to oocyte retrieval. In summary, we review data on the efficacy and safety of the major hormonal approaches used to induce final oocyte maturation in clinical practice, as well as some novel approaches that may offer fresh alternatives in future.

Sperm cryopreservation has been widely adopted for maintenance of the genetically engineered mouse (GEM). The cryopreserved sperm are being exchanged among many institutes worldwide. However, the recipients are not always able to obtain high fertilization rates with the frozen sperm shipped from senders. In this study, we cryopreserved mouse sperm via various methods and performed in vitro fertilization (IVF) in which the combination of methyl-beta-cyclodextrin for sperm preincubation and reduced glutathione for insemination was used (the MBCD-GSH IVF). In addition, frozen sperm sent from the Jackson Laboratory (USA) were thawed and used for IVF in the same manner. The fertilization rates of both the sperm cryopreserved via the methods applied in some countries and the cryopreserved GEM sperm improved when used with the MBCD-GSH IVF method. Therefore, we strongly believe that the MBCD-GSH IVF method brings about relatively high fertilization rates with any strain of frozen mouse sperm.

Limited research suggests ambient air pollution impairs fecundity but groups most susceptible have not been identified. We studied whether long-term ambient air pollution exposure prior to an in vitro fertilization (IVF) cycle was associated with successful livebirth, and whether associations were modified by underlying infertility diagnosis.

This study was conducted to assess the fertilization and development of porcine oocytes matured in a solo follicular fluid (pFF) using different in vitro culture systems and insemination periods. Cumulus-oocyte complexes (COCs), follicular cells (FCs), and pFF were collected from the follicles of ovaries. The pFF was used as a maturation medium (MpFF) after supplementation with follicle stimulating hormone (FSH) and antibiotics. The COCs were matured in a 15 ml test tube containing 3.5 ml of MpFF with FCs (5.2 × 10(6) cells/ml; rotating culture system) or 2 ml of MpFF without FCs in a 35-mm petri dish (static culture system) for 44 to 48 h. After maturation culture, oocytes were co-incubated with frozen-thawed spermatozoa for 5 h and then cultured for 7 days. The total mean rates of sperm penetration, normal fertilization, male pronucleus (MPN) formation, cleavage, and development to the blastocyst stage of oocytes after insemination were significantly higher (P<0.01) in the rotating culture system than in the static culture system. In conclusion, compared with the static culture system, the rotating culture system is adequate for the production of developmentally competent porcine oocytes when MpFF is used as a maturation medium.

Human follicular fluid (HFF) provides a key environment for follicle development and oocyte maturation, and contributes to oocyte quality and in vitro fertilization (IVF) outcome.