BACKGROUND: Sugammadex is a new neuromuscular blockade reversal agent. Recently, it has been used in patients under general anesthesia. However, sugammadex could be toxic to fetuses and pediatric patients under 3 years of age. In this study, we demonstrated the safety of sugammadex in fetuses, using zebrafish larvae. Furthermore, its neurotoxicity was evaluated using neuronal cell lines.METHODS: We used SH-SY5Y cells to determine the viability of neuronal cells treated with sugammadex. Zebrafish larvae were used to determine the teratogenic effects of sugammadex.RESULTS: Sugammadex showed no adverse effects on neuronal cells and zebrafish larvae. The survival rates of neuronal cells were not different in all concentrations. In addition, the heart formation of zebrafish embryos, which were exposed to various concentrations of sugammadex, were not different.CONCLUSION: This study demonstrated the feasibility of using sugammadex during pregnancy. However, further clinical studies will be required to extrapolate these results to humans.
Anesthesia, General ; Cell Line ; Embryonic Structures ; Fetus ; Heart ; Humans ; Larva ; Neuromuscular Blockade ; Neurons ; Pregnancy ; Survival Rate ; Zebrafish

BACKGROUND AND OBJECTIVES: Genomic imprinting modulates growth and development in mammals and is associated with genetic disorders. Although uniparental embryonic stem cells have been used to study genomic imprinting, there is an ethical issue associated with the destruction of human embryos. In this study, to investigate the genomic imprinting status in human neurodevelopment, we used human uniparental induced pluripotent stem cells (iPSCs) that possessed only maternal alleles and differentiated into neural cell lineages. METHODS: Human somatic iPSCs (hSiPSCs) and human parthenogenetic iPSCs (hPgiPSCs) were differentiated into neural stem cells (NSCs) and named hSi-NSCs and hPgi-NSCs respectively. DNA methylation and gene expression of imprinted genes related neurodevelopment was analyzed during reprogramming and neural lineage differentiation. RESULTS: The DNA methylation and expression of imprinted genes were altered or maintained after differentiation into NSCs. The imprinting status in NSCs were maintained after terminal differentiation into neurons and astrocytes. In contrast, gene expression was differentially presented in a cell type-specific manner. CONCLUSIONS: This study suggests that genomic imprinting should be determined in each neural cell type because the genomic imprinting status can differ in a cell type-specific manner. In addition, the in vitro model established in this study would be useful for verifying the epigenetic alteration of imprinted genes which can be differentially changed during neurodevelopment in human and for screening novel imprinted genes related to neurodevelopment. Moreover, the confirmed genomic imprinting status could be used to find out an abnormal genomic imprinting status of imprinted genes related with neurogenetic disorders according to uniparental genotypes.
Alleles ; Astrocytes ; Cell Lineage ; DNA Methylation ; Embryonic Stem Cells ; Embryonic Structures ; Epigenomics ; Ethics ; Gene Expression ; Genomic Imprinting ; Genotype ; Growth and Development ; Humans ; In Vitro Techniques ; Induced Pluripotent Stem Cells ; Mammals ; Mass Screening ; Neural Stem Cells ; Neurons

Mucopolysaccharidosis type II (MPS II) is a rare X-linked recessive lysosomal storage disease caused by mutation of the iduronate-2-sulfatase gene. The mutation results in iduronate-2-sulfatase deficiency, which causes the progressive accumulation of heparan sulfate and dermatan sulfate in cellular lysosomes. The phenotype, age of onset, and symptoms of MPS II vary; accordingly, the disease can be classified into either the early-onset type or the late-onset type, depending on the age of onset and the severity of the symptoms. In patients with severe MPS II, symptoms typically first appear between 2 and 5 years of age. Patients with severe MPS II usually die in the second decade of life although some patients with less severe disease have survived into their fifth or sixth decade. Here, we report the establishment of a preimplantation genetic diagnosis (PGD) strategy using multiplex nested polymerase chain reaction, direct sequencing, and linkage analysis. Unaffected embryos were selected via the diagnosis of a single blastomere, and a healthy boy was delivered by a female carrier of MPS II. This is the first successful application of PGD in a patient with MPS II in Korea
Age of Onset ; Blastomeres ; Dermatan Sulfate ; Diagnosis ; Embryonic Structures ; Female ; Heparitin Sulfate ; Humans ; Korea ; Lysosomal Storage Diseases ; Lysosomes ; Male ; Mucopolysaccharidoses ; Mucopolysaccharidosis II ; Multiplex Polymerase Chain Reaction ; Parturition ; Phenotype ; Polymerase Chain Reaction ; Preimplantation Diagnosis ; Prostaglandins D

OBJECTIVE: We aimed to evaluate the effects of different oxygen conditions (20% [high O₂], 5% [low O₂] and 5% decreased to 2% [dynamic O₂]) on mouse pre- and peri-implantation development using a novel double-channel gas supply (DCGS) incubator (CNC Biotech Inc.) to alter the oxygen concentration during in vitro culture.METHODS: The high-O₂ and low-O₂ groups were cultured from the one-cell to the blastocyst stage under 20% and 5% oxygen concentrations, respectively. In the dynamic-O₂ group, mouse embryos were cultured from the one-cell to the morula stage under 5% O₂ for 3 days, followed by culture under 2% O₂ to the blastocyst stage. To evaluate peri-implantation development, the blastocysts from the three groups were individually transferred to a fibronectin-coated dish and cultured to the outgrowth stage in droplets.RESULTS: The blastocyst formation rate was significantly higher in the low-O₂ and dynamic-O₂ groups than in the high-O₂ group. The total cell number was significantly higher in the dynamic-O₂ group than in the low-O₂ and high-O₂ groups. Additionally, the apoptotic index was significantly lower in the low-O₂ and dynamic-O₂ groups than in the high-O₂ group. The trophoblast outgrowth rate and spread area were significantly higher in the low-O₂ and dynamic-O₂ groups than in the high-O₂ group.CONCLUSION: Our results showed that a dynamic oxygen concentration (decreasing from 5% to 2%) had beneficial effects on mouse pre- and peri-implantation development. Optimized, dynamic changing of oxygen concentrations using the novel DCGS incubator could improve the developmental competence of in vitro cultured embryos in a human in vitro fertilization and embryo transfer program.
Animals ; Apoptosis ; Blastocyst ; Cell Count ; Embryo Transfer ; Embryonic Structures ; Fertilization in Vitro ; Humans ; In Vitro Techniques ; Incubators ; Mental Competency ; Mice ; Morula ; Oxygen ; Trophoblasts

OBJECTIVE: To determine the clinical pregnancy (CP) and live birth (LB) rates arising from frozen embryo transfers (FETs) that had been generated under the influence of in vitro fertilization (IVF) adjuvants given to women categorized as poor-prognosis.METHODS: A registered, single-center, retrospective study. A total of 1,119 patients with first FETs cycle include 310 patients with poor prognosis (109 treated with growth hormone [GH], (+)GH group vs. 201 treated with dehydroepiandrosterone, (–)GH group) and 809 patients with good prognosis (as control, (–)Adj (Good) group).RESULTS: The poor-prognosis women were significantly older, with a lower ovarian reserve than the (–)Adj (Good) group, and demonstrated lower chances of CP (p<0.005) and LB (p<0.005). After adjusting for confounders, the chances of both CP and LB in the (+)GH group were not significantly different from those in the (–)Adj (Good) group, indicating that the poor-prognosis patients given GH had similar outcomes to those with a good prognosis. Furthermore, the likelihood of LB was significantly higher for poor-prognosis women given GH than for those who did not receive GH (p<0.028). This was further confirmed in age-matched analyses.CONCLUSION: The embryos cryopreserved from fresh IVF cycles in which adjuvant GH had been administered to women classified as poor-prognosis showed a significant 2.7-fold higher LB rate in subsequent FET cycles than a matched poor-prognosis group. The women with a poor prognosis who were treated with GH had LB outcomes equivalent to those with a good prognosis. We therefore postulate that GH improves some aspect of oocyte quality that confers improved competency for implantation.
Dehydroepiandrosterone ; Embryo Transfer ; Embryonic Structures ; Female ; Fertilization in Vitro ; Growth Hormone ; Humans ; Live Birth ; Melatonin ; Oocytes ; Ovarian Reserve ; Pregnancy ; Prognosis ; Retrospective Studies ; Single Embryo Transfer

OBJECTIVE: In vitro maturation (IVM) of immature oocytes can be useful for some infertile patients. In IVM programs, the rates of embryo formation and pregnancy are low. Therefore, it is essential to recognize the main factors involved in regulating oocyte maturation in vitro. The purpose of this study was to investigate the effects of growth differentiation factor 9 (GDF9) and cumulus cell (CC) supplementation in IVM medium on the rates of embryo formation and viability of human blastocysts.METHODS: A total of 80 germinal vesicle oocytes from stimulated cycles underwent an IVM program. The oocytes were divided into four groups, where group I consisted of IVM media only and served as the control, group II consisted of IVM+CCs, group III consisted of IVM+GDF9 (200 ng/mL), and group IV consisted of IVM+CCs+GDF9 (200 ng/mL). Intracytoplasmic sperm injection was performed on the IVM oocytes, and the cleavage embryos that were generated were vitrified. Following thawing, the embryos were cultured for 3 additional days, and the viability rates of the developed blastocysts were determined.RESULTS: The maturation rate of the oocytes did not differ significantly across the four groups. The fertilization rate in group II was significantly higher than that in the control group (76.5% vs. 46.2%). Embryo formation was significantly more frequent in all experimental groups than in the control group, while blastocyst formation did not show significant differences in the three experimental groups compared to the control. The mean viability rates in groups II, III, and IV were 58.16%, 55.91%, and 55.95%, respectively, versus 37.78% in the control group (p<0.05).CONCLUSION: Supplementation of IVM culture media with GDF9 and CCs enhanced the fertilization, embryo formation, and viability rates of blastocysts generated from vitrified cleavage embryos.
Blastocyst ; Culture Media ; Cumulus Cells ; Embryonic Structures ; Fertilization ; Growth Differentiation Factor 9 ; Humans ; In Vitro Techniques ; Oocytes ; Pregnancy ; Sperm Injections, Intracytoplasmic

BACKGROUND: The standard morphological evaluation has been widely used for embryo selection, but it has limitations. This study aimed to investigate the correlation between morphologic grading and euploidy rate of in vitro fertilization (IVF) preimplantation genetic screening (PGS) and compare the pregnancy rates in young and old ages. METHODS: This is a retrospective study using the medical records of patients who underwent IVF procedures with PGS between January 2016 and February 2017 in a single center. The embryo grades were categorized into 4 groups: excellent, good, fair, and poor. Basic characteristics, euploidy rates, clinical pregnancy (CP) rates and ongoing pregnancy rates were analyzed. RESULTS: The excellent group had significantly higher rate of euploid embryos than fair group (47.82% vs. 29.33%; P = 0.023) and poor group (47.82% vs. 29.60%; P = 0.005). When the four groups were recategorized into two groups (excellent and good vs. fair and poor), they also showed significant difference in euploidy rates (44.52% vs. 29.53%; P = 0.002). When the patients were divided into two groups by age 35, the CP rates for those under and over 35 years old were 44.74% and 47.83%, respectively, which showed no significant difference. CONCLUSION: The significant differences among the euploidy rates of different morphologic embryo grades demonstrated the positive correlations between the morphologic grading of the embryo and the euploidy rate of PGS. Additionally, there was no significant difference between the younger and older patients' CP rates. These findings emphasize the fact that old age patients might benefit from PGS whatever the indication of PGS is.
Blastocyst* ; Embryonic Structures ; Fertilization in Vitro* ; Genetic Testing* ; Humans ; In Vitro Techniques* ; Medical Records ; Pregnancy ; Pregnancy Rate ; Retrospective Studies

PURPOSE: To elucidate the correlation between ovarian reserve and the incidence of ectopic pregnancy (EP) following in vitro fertilization and embryo transfer (IVF/ET) cycles. MATERIALS AND METHODS: In this observational study, 430 fresh IVF/ET cycles were examined from patient data of two university hospital infertility clinics. All included patients were positive for β-human chorionic gonadotropin (hCG) at 2 weeks after oocyte retrieval via controlled ovarian stimulation. For each cycle, information on age, duration of infertility, basal follicle stimulating hormone (FSH), anti-Müllerian hormone (AMH), days of ovarian stimulation, numbers of retrieved oocytes and transferred embryos, and pregnancy outcomes was collected. Patients with AMH lower than 1.0 ng/dL or basal FSH higher than 10 mIU/mL were classified into the decreased ovarian reserve (DOR) group, and the remaining patients were classified into the normal ovarian reserve (NOR) group. RESULTS: In total, 355 cycles showed NOR, and 75 cycles DOR. There were no significant differences between the DOR and NOR groups regarding intrauterine (74.7% vs. 83.4%, respectively) or chemical (14.7% vs. 14.1%, respectively) pregnancies. The DOR group had a higher EP than that of NOR group [10.7% (8/75) vs. 2.5% (9/355), p=0.004]. In both univariate [odds ratio (OR) 5.6, 95% confidence interval (CI) 1.4–9.6, p=0.011] and multivariate (adjusted OR 5.1, 95 % CI 1.1–18.7, p=0.012) analysis, DOR was associated with a higher risk of EP. CONCLUSION: DOR may be associated with a higher risk of EP in IVF/ET cycles with controlled ovarian stimulation. More careful monitoring may be necessary for pregnant women with DOR.
Chorionic Gonadotropin ; Embryo Transfer ; Embryonic Structures ; Female ; Fertilization in Vitro ; Follicle Stimulating Hormone ; Humans ; In Vitro Techniques ; Incidence ; Infertility ; Observational Study ; Oocyte Retrieval ; Oocytes ; Ovarian Reserve ; Ovulation Induction ; Pregnancy ; Pregnancy Outcome ; Pregnancy, Ectopic ; Pregnant Women

PURPOSE: To investigate the associations between sperm DNA fragmentation (SDF) and embryo formation rate in normal responder women to in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI). MATERIALS AND METHODS: Fifty-three consecutive, fresh IVF/ICSI cycles performed from 2014 to 2017 were selected. All women were normal responders (4 to 14 mature oocytes were retrieved) and at least one normally fertilized oocyte with two pronuclei was obtained in all cycles. Semen was collected on the day of oocyte retrieval, and SDF levels were measured by sperm chromatin dispersion test (Halosperm assay). At day 3 after insemination, embryo quality was evaluated by morphologic criteria and categorized as A/B/C/D. Top quality embryo were defined as grade A embryos with seven cells or more. RESULTS: SDF levels showed a positive linear correlation with the male's age (r=0.307, p=0.025) and a negative linear correlation with sperm motility (r=−0.491, p<0.0001). To achieve top-quality or a grade A embryo formation rate >70%, the cut-off value SDF was <30.7% for each. Among individuals with SDF <30.7%, the median top-quality or grade A embryo formation rate was significantly higher than that among individuals with SDF ≥30.7% (38.1% vs. 20.0%, p=0.038; 50% vs. 25.0%, p=0.017). CONCLUSION: In normal responder women, high SDF level resulted in low day 3 embryo formation rates. Our results suggest a paternal effect on embryo quality in IVF/ICSI cycles.
Chromatin ; DNA Fragmentation ; DNA ; Embryonic Structures ; Female ; Fertilization in Vitro ; Humans ; In Vitro Techniques ; Insemination ; Oocyte Retrieval ; Oocytes ; Semen ; Sperm Injections, Intracytoplasmic ; Sperm Motility ; Spermatozoa

Somatic cell nuclear transfer (SCNT) has various applications in research, as well as in the medical field and animal husbandry. However, the efficiency of SCNT is low and the accurate mechanism of SCNT in murine embryo development is unreported. In general, the developmental rate of SCNT murine embryos is lower than in vivo counterparts. In previous studies, polo-like kinase 1 (Plk1) was reported to be a crucial element in cell division including centrosome maturation, cytokinesis, and spindle formation. In an initial series of experiments in this study, BI2536, a Plk1 inhibitor, was treated to in vivo-fertilized embryos and the embryos failed to develop beyond the 2-cell stage. This confirmed previous findings that Plk1 is crucial for the first mitotic division of murine embryos. Next, we investigated Plk1's localization and intensity by immunofluorescence analysis. In contrast to normally developed embryos, SCNT murine embryos that failed to develop exhibited two types of Plk1 expressions; a low Plk1 expression pattern and ectopic expression of Plk1. The results show that Plk1 has a critical role in SCNT murine embryos. In conclusion, this study demonstrated that the SCNT murine embryos fail to develop beyond the 2-cell stage, and the embryos show abnormal Plk1 expression patterns, which may one of the main causes of developmental failure of early SCNT murine embryos.
Animal Husbandry ; Cell Division ; Centrosome ; Cytokinesis ; Ectopic Gene Expression ; Embryonic Development ; Embryonic Structures ; Female ; Fluorescent Antibody Technique ; Nuclear Transfer Techniques ; Phosphotransferases ; Pregnancy