Sperm Inspection for In Vitro Fertilization via Self-Assembled Microdroplet Formation and Quantitative Phase Microscopy.
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ABSTRACT:
SUBMITTER: Atzitz Y
PROVIDER: S-EPMC8699486 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Magnetotactic bacteria synthesize highly uniform intracellular magnetite nanoparticles through the action of several key biomineralization proteins. These proteins are present in a unique lipid-bound organelle (the magnetosome) that functions as a nanosized reactor in which the particle is formed. A master regulator protein of nanoparticle formation, magnetosome membrane specific F (MmsF), was recently discovered. This predicted integral membrane protein is essential for controlling the monodispersity of the nanoparticles in Magnetospirillum magneticum strain AMB-1. Two MmsF homologs sharing over 60% sequence identity, but showing no apparent impact on particle formation, were also identified in the same organism. We have cloned, expressed, and used these three purified proteins as additives in synthetic magnetite precipitation reactions. Remarkably, these predominantly ?-helical membrane spanning proteins are unusually highly stable and water-soluble because they self-assemble into spherical aggregates with an average diameter of 36 nm. The MmsF assembly appears to be responsible for a profound level of control over particle size and iron oxide (magnetite) homogeneity in chemical precipitation reactions, consistent with its indicated role in vivo. The assemblies of its two homologous proteins produce imprecise various iron oxide materials, which is a striking difference for proteins that are so similar to MmsF both in sequence and hierarchical structure. These findings show MmsF is a significant, previously undiscovered, protein additive for precision magnetite nanoparticle production. Furthermore, the self-assembly of these proteins into discrete, soluble, and functional "proteinosome" structures could lead to advances in fields ranging from membrane protein production to drug delivery applications.
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Project description:ObjectiveTo report a unique case of total fertilization failure (TFF) after in vitro fertilization with intracytoplasmic sperm injection related to homozygous WEE2 gene mutation and summarize the current literature and management of TFF.DesignCase report.SettingAcademic fertility center.PatientsA 25-year-old woman and her 35-year-old partner with a history of near-complete fertilization failure after 2 cycles of in vitro fertilization/intracytoplasmic sperm injection.InterventionsConsultation with medical and commercial genetic testing for WEE2, PLCZ1, and TLE6.Main outcome measuresOocyte fertilization.ResultsThe patient was homozygous for WEE2 pathogenic variant impacting oocyte activation and resulting in infertility.ConclusionsIn the setting of TFF, early consideration should be given to genetic testing to assist couples in clinical decision-making and help limit the financial and emotional burden associated with unsuccessful fertility intervention.
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Project description:This is a method for zebrafish sperm cryopreservation that is an adaptation of the Harvey method (Harvey et al., 1982). We have introduced two changes to the original protocol that both streamline the procedure and increase sample uniformity. First, we normalize all sperm volumes using freezing media that does not contain the cryoprotectant. Second, cryopreserved sperm are stored in cryovials instead of capillary tubes. The rates of sperm freezing and thawing (delta degrees C/time) are probably the two most critical variables to control in this procedure. For this reason, do not substitute different tubes for those specified. Working in teams of 2 it is possible to freeze the sperm of 100 males per team in approximately 2 hrs. Sperm cryopreserved using this protocol has an average of 25% fertility (measured as the number of viable embryos generated in an in vitro fertilization divided by the total number of eggs fertilized) and this percent fertility is stable over many years.
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Project description:PurposeTo investigate whether in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI), influence the embryo's development and its quality using the mouse as a model.MethodsAssisted fertilization was performed using ICSI and IVF. Fluorescent beads were adhered to the fertilization cone or place of previous sperm injection in the natural mated (NM), IVF and ICSI embryos, respectively. Embryo examination was carried out at the two-cell and blastocyst stage to determine the position of fluorescent bead. Protein expression was detected by fluorescence immunocytochemical staining and confocal microscopic imaging of blastocysts.ResultsIVF and ICSI embryos developed at rates comparable to NM group. Embryos show similar expression patterns of two transcription factors, Oct4 and Cdx2. The most preferred place for spermatozoa attachment was the equatorial site of the egg, whether fertilization occurred in vitro or under natural conditions. We also link the sperm entry position (SEP) to embryo morphology and the number of cells at the blastocyst stage, with no influence of the method of fertilization.ConclusionsIVF and ICSI, do not compromise in vitro pre-implantation development. Additional data, related to sperm entry, could offer further criteria to predict embryos that will implant successfully. Based on embryo morphology, developmental rate and protein expression level of key transcription factors, our results support the view that ART techniques, such as IVF and ICSI, do not perturb embryonic development or quality.
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Project description:To determine the cost-effectiveness of split IVF-intracytoplasmic sperm injection (ICSI) for the treatment of couples with unexplained infertility.Adaptive decision model.Academic infertility clinic.A total of 154 couples undergoing a split IVF-ICSI cycle and a computer-simulated cohort of women <35 years old with unexplained infertility undergoing IVF.Modeling insemination method in the first IVF cycle as all IVF, split IVF-ICSI, or all ICSI, and adapting treatment based on fertilization outcomes.Live birth rate, incremental cost-effectiveness ratio (ICER).In a single cycle, all IVF is preferred as the ICER of split IVF-ICSI or all ICSI ($58,766) does not justify the increased live birth rate (3%). If two cycles are needed, split IVF/ICSI is preferred as the increased cumulative live birth rate (3.3%) is gained at an ICER of $29,666.In a single cycle, all IVF was preferred as the increased live birth rate with split IVF-ICSI and all ICSI was not justified by the increased cost per live birth. If two IVF cycles are needed, however, split IVF/ICSI becomes the preferred approach, as a result of the higher cumulative live birth rate compared with all IVF and the lesser cost per live birth compared with all ICSI.
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Project description:BACKGROUND: The sperm penetration assay (SPA) is used to predict the fertilizing capacity of sperm. Thus, some programs rely on SPA scores to formulate insemination plans in conjunction with in-vitro fertilization (IVF) cycles. The purpose of this study was to evaluate if a relationship exists between SPA scores and polyspermy rates during conventional IVF cycles. METHODS: A total of 1350 consecutive IVF patients using conventional IVF insemination were evaluated in the study. Oocytes were inseminated three hours post-retrieval by the addition of 150,000 to 300,000 progressively motile sperm. Approximately 18 hours after insemination, the oocytes were evaluated for fertilization by the visualization of pronuclei. The presence of three or more pronuclei was indicative of polyspermy. Polyspermy rates, fertilization success, embryo quality, and pregnancy rates were analyzed retrospectively to evaluate their relationship with SPA score, count, motility, number of progressively motile sperm inseminated, oocyte pre-insemination incubation time, patient age, and diagnosis. RESULTS: A significant positive relationship was observed between SPA score and polyspermy rate (rs = 0.10, p < 0.05). Patients with a normal SPA score had significantly higher polyspermy rates than those with abnormal SPA scores (6.3% +/- 1.5% vs. 2.0% +/- 0.7%, p < 0.05). Fertilization percentage was significantly lower in the group with severely abnormal SPA scores versus all other SPA groups (57.5% +/- 2.1% vs. 70.2% +/- 1.3%, p < 0.005). Although embryo quality was not affected, both clinical pregnancy and implantation rates improved slightly as SPA score increased. In addition, there was a decrease in the rate of spontaneous abortion as SPA score increased. CONCLUSIONS: These data indicate SPA score is positively correlated with polyspermy rates and IVF fertilization percentage. Additionally, there is a slight increase in clinical pregnancy rates, and embryo implantation rates with increased SPA. Furthermore, there is a slight decrease in spontaneous abortions rates related to increased SPA.
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Project description:ObjectiveTo determine the impact of neurotensin (NTS), a naturally occurring peptide, on the function of human and nonhuman primate sperm.DesignExperimental study.SettingUniversity-based research laboratory.Patient(s)/animal(s)Consenting normozoospermic human donors and cynomolgus macaques.Intervention(s)None.Main outcome measure(s)Sperm acrosome status was assessed. Computer-assisted semen analysis assessed sperm motility, progression, and velocity. Immunocytochemistry and receptor selective agonists were used to identify specific NTS receptors on sperm. Monkey oocytes were obtained after ovarian stimulation, and NTS-treated monkey sperm were used for in vitro fertilization.ResultsNeurotensin treatment of human sperm stimulated the acrosome reaction in both a dose-dependent (0.1-10 μmol/L) and time-dependent (5-30 minutes) manner. Neurotensin treatment did not alter sperm motility or progression. Both a general NTS receptor antagonist (SR142948) and a NTSR1 selective antagonist (SR48692) reduced the ability of NTS to stimulate the acrosome reaction. The neurotensin receptor NTSR1, but not NTSR2 or SORT1, was detected in monkey sperm using immunostaining. Neurotensin treatment also compromised the ability of sperm to fertilize an oocyte. Percentage of fertilization with untreated monkey sperm and monkey oocytes was 72%. Sperm pre-treated with NTS yielded a significantly lower fertilization rate of 18%.Conclusion(s)Neurotensin effectively stimulates the acrosome reaction in human and monkey sperm. Neurotensin produced by the oviduct or cumulus cells may promote natural fertilization. Pretreatment of sperm with NTS significantly reduces fertilization. Exposure of sperm to NTS prior to reaching the oviduct has the potential for contraceptive development. Identification of NTSR1 as the mediator of NTS action provides a specific target for future studies.
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