Human embryos derived from first polar body nuclear transfer exhibit comparatively abnormal morphokinetics during development / 대한생식의학회지

Objective@#Reconstructed oocytes after polar body genome transfer constitute a potential therapeutic option for patients with a history of embryo fragmentation and advanced maternal age. However, the rescue of genetic material from the first polar body (PB1) through introduction into the donor cytoplasm is not yet ready for clinical application. @*Methods@#Eighty-five oocytes were obtained following in vitro maturation (IVM) and divided into two groups: PB1 nuclear transfer (PB1NT; n=54) and control (n=31). Following enucleation and PB1 genomic transfer, PB1 fusion was assessed. Subsequently, all fused oocytes underwent intracytoplasmic sperm injection (ICSI) and were cultured in an incubator under a time-lapse monitoring system to evaluate fertilization, embryonic morphokinetic parameters, and cleavage patterns. @*Results@#Following enucleation and fusion, 77.14% of oocytes survived, and 92.59% of polar bodies (PBs) fused. However, the normal fertilization rate was lower in the PB1NT group than in the control group (56.41% vs. 92%, p=0.002). No significant differences were observed in embryo kinetics between the groups, but a significant difference was detected in embryo developmental arrest after the four-cell stage, along with abnormal cleavage division in the PB1NT group. This was followed by significant between-group differences in the implantation potential rate and euploidy status. Most embryos in the PB1NT group had at least one abnormal cleavage division (93.3%, p=0.001). @*Conclusion@#Fresh PB1NT oocytes successfully produced normal zygotes following PB fusion and ICSI in IVM oocytes. However, this was accompanied by low efficiency in developing into cleavage embryos, along with an increase in abnormal cleavage patterns.

Does Overnight Culture of Cleaved Embryos Improve Pregnancy Rate in Vitrified-Warmed Embryo Transfer Programme?

@#Background: Vitrification is a routine procedure in assisted reproductive technique (ART) lab. However, there is widespread variability between protocols of different centres. The aim of this study was to compare the chemical pregnancy, clinical pregnancy and live birth rates between one-day embryo culture and immediate transfer for frozen-thawed embryo transfer (FET) cycles. Methods: In this cohort retrospective study, 366 FET cycles were divided into two groups: Group A, the embryos were warmed one day before transfer, and were cultured overnight; Group B, the embryos were warmed on the same day of transfer, at least were cultured 1 h before embryo transfer (ET). Chemical and clinical pregnancy and live birth rates were compared between two groups. Results: The chemical pregnancy was higher in group A than B (37.9% versus 28.9%), but this difference was not significant (P = 0.07). Clinical pregnancy (30.8% versus 24.1%) and live birth (19.8% versus 22.05%) were similar in group A and B, (P = 0.15), and (P = 0.8). Conclusion: In conclusion, overnight culture and confirmation of mitosis resumption was not essential for FET cycles in vitrification method.

Vitrification increased vacuolization of human spermatozoa: application of MSOME technology

Background: sperm vitrification is a technique of ice and cryoprotectant free cryopreservation by direct plunging of sperm suspension into liquid nitrogen [LN2]. The aim of this study was to investigate the influence of cryoprotectant free-vitrification on human sperm fine structure by MSOME technology and the fertility potential by zona binding assay [ZBA]

Methods: 20 normo-ejaculates were prepared by swim up technique, and supernatants were divided into two parts of fresh and vitrified groups. For vitrification, sperm was dropped into LN2. Sperm motility, morphology, viability and MSOME were evaluated for each sample. In MSOM morphologically normal sperm [class 1], 2 small vacuoles [class 3] were evaluated. Also, fertility potential was evaluated by zona binding assay. Data was analyzed using paired t-test or Willcoxon's test and p-value <0.05 was considered significant

Results: vitrification significantly reduced progressive motility, viability and morphology. Also, normal morphology of spermatozoa decreased significantly after vitrification. In MSOME evaluation, normal motile spermatozoa [Class 1] decreased from 23.00+/-12.44 to 16.00.56+/-10.79 after vitrification [p=0.008]. Although spermatozoa classes 2 and 3 were increased, the difference was not significant. Moreover, fertility potential of motile spermatozoa was reduced after vitrification [9.0+/-13.87 vs. 13.40+/-22.73; p=0.07]

Conclusion: Vitrification increased the rate of vacuolization in motile sperm head. Therefore, MSOME technology is recommended for assessment of sperm fine morphology in ICSI program used cryopreserved spermatozoa

Revealing the secret life of pre-implantation embryos by time-lapse monitoring: a review

High implantation success following in vitro fertilization cycles are achieved via the transfer of embryos with the highest developmental competence. Multiple pregnancies as a result of the transfer of several embryos per cycle accompany with various complication. Thus, single-embryo transfer [SET] is the preferred practice in assisted reproductive technique [ART] treatment. In order to improve the pregnancy rate for SET, embryologists need reliable biomarkers to aid their selection of embryos with the highest developmental potential. Time-lapse technology is a noninvasive alternative conventional microscopic assessment. It provides uninterrupted and continues the survey of embryo development to transfer day. Today, there are four time-lapse systems that are commercially available for ART centers. In world and Iran, the first time lapse babies were born in 2010 and 2015, respectively, conceived by SET. Here, we review the use of time-lapse monitoring in the observation of embryogenesis as well as its role in SET. Although, the findings from our review support common use of time-lapse monitoring in ART centers; but, future large studies assessing this system in well-designed trials are necessary

Live birth in a woman with recurrent implantation failure and adenomyosis following transfer of refrozen-warmed embryos / 대한생식의학회지

The aim was to report a healthy live birth using re-vitrified-warmed cleavage-stage embryos derived from supernumerary warmed embryos after frozen embryo transfer (ET) in a patient with recurrent implantation failure (RIF). The case was a 39-year-old female with a history of polycystic ovarian syndrome and adenomyosis, along with RIF. After ovarian hyperstimulation, 33 cumulus-oocyte complexes were retrieved and fertilized with conventional in vitro fertilization and intracytoplasmic sperm injection. Because of the risk of ovarian hyperstimulation syndrome, 16 grade B and C embryos were vitrified. After 3 and 6 months, 3 and 4 B–C warmed embryos were transferred to the uterus, respectively. However, implantation did not take place. Ten months later, four embryos were warmed, two grade B 8-cell embryos were transferred, and two embryos were re-vitrified. One year later, the two re-vitrified cleavage-stage embryos were warmed, which resulted in a successful live birth. This finding showed that following first warming, it is feasible to refreeze supernumerary warmed embryos for subsequent ET in patients with a history of RIF.

Assessment of ovarian tissues autografted to various body sites followed by IVM in mouse

Ovarian tissue transplantation is emerging technologies for fertility preservation. In addition, in vitro maturation [IVM] of oocytes retrieved from ovarian tissues may overcome the fertility defects in certain cases. The aim was to evaluate the best site for ovarian tissue transplantation in mice. Also, feasibility of IVM of oocytes retrieved from auto grafted ovarian tissues was freshly assessed. Hemi-ovaries from 6 weeks old mice were auto grafted into kidney capsule [K] versus the back muscle [B] and leg muscle [L] in a mouse auto graft model which was stimulated with gonadotrophins. Then ovarian grafts were recovered and processed histologically for follicle assessment compared with control, also the ability of oocytes to mature with IVM was studied 14 days after transplantation. Total follicle count was significantly higher in K-graft [3.5 +/- 3.17] and the antral follicles were only observed in K-site model. The number of retrieved immature oocytes as well as successful IVM in K-grafts was significantly higher than other groups [p=0.008, p=0.016]. The kidney capsule is a promising site for ovarian tissue auto graft in mice. This resulted in better follicular survival and IVM outcomes

Comparing the roles of sperm chromatin integrity and apoptosis in intrauterine insemination outcomes of couples with mild male and female factor infertility

Intrauterine insemination [IUI] is one of the therapeutic approaches for infertility. The objective of this study was to evaluate DNA integrity and apoptosis role in success of IUI in both mild male and female factor infertility. Patients were divided into two groups: M [mild male factor; n=29] and F [female factor; n=31] undergoing single IUI. Ejaculates were analyzed and chromatin quality was assessed using chromomycin A3 [CMA3] staining. In addition, spermatozoal apoptosis was recognized using TUNEL assay. Statistical analyses were done using t-test and Mann Whitney test for sperm apoptosis and sperm chromatin by SPSS. Data were expressed in mean +/- SD for variables. P<0.05 was considered statistically significant. Sperm concentration and progressive motility were higher in F than M group. Sperm with normal morphology were statistically similar in M and F infertile patients [32.7 +/- 15.6% vs. 35.5 +/- 9.07%, p=0.39]. Sperm chromatin immaturity was higher in patients with mild male infertility, when compared with the other group [p<0.01]. Also, 32.0 +/- 5.6% and 30.8 +/- 6.1% of the spermatozoa showed signs of apoptosis in groups M and F, respectively [p=0.49]. Very low [3.4%] clinical pregnancy rates were noticed in patients with mild male factor infertility. Defect in sperm motility as well as high rates of DNA damage and apoptosis may be involved with very low rate of pregnancy outcomes in patients with mild male factor infertility. Therefore, it seems the application of IUI may have better outcomes in patients with female infertility compared to mild male factor infertility

[Quality control of disposable objects in ART laboratories performing human sperm motility assays]

ART laboratories are quality controlled to make sure that disposable objects used for the culture of gametes and embryos are toxin-free. To maintain a high standard, all disposable objects in our ART laboratory were tested by human sperm motility assay [HuSMA]. HuSMA was used as a measure for QC at the intended ART laboratory. Eighteen objects that are commonly used in IVF laboratories were tested by HuSMA. The objects included gloves, syringes, culture dishes, pipettes, tips and semen collection dishes. HuSMA was conducted at 10 and 30 minutes and also at 1, 2, 4, and 24 hours of incubation at room temperature. Sperm motility index [SMI] was calculated by dividing the percentage of progressive motile sperms of the test by that of the control at the specified intervals. An SMI value < 0.85 was defined to indicate sperm toxicity. The tests were repeated for three times. QC by HuSMA confirmed the toxicity of three objects, including embryo transfer [ET] gloves A and B, and puncture gloves A. ET gloves A [SMI=0.0] and puncture gloves A [SMI=0.0] were toxic after 10 minutes, but ET gloves B [SMI=0.63] were shown to be toxic after 24 hours [46% progressive motile sperm compared with 68% in the control group]. Moreover, two other objects including culture dish [SMI=0.42] and semen collection dish [SMI=0.67] had borderline values after 24 hours; different results in four repeats after 24 hours [twice toxic and twice nontoxic]. Some objects which are routinely used in ART laboratories may be toxic and their use should be discontinued as part of QC programs. To increase the efficiency of HuSMA, it seems necessary to do this test more than once for each object