Retrospective study of single vitrified-warmed blastocyst transfer cycles according to the presence of morphokinetic variables / 대한생식의학회지

This study retrospectively assessed whether time-lapse data relating to developmental timing and morphology were associated with clinical outcomes, with the eventual goal of using morphokinetic variables to select embryos prospectively for cryopreservation. In this study, we examined the clinical outcomes of single vitrified-warmed blastocyst transfer cycles that were cultured in a time-lapse incubation system. The morphokinetic variables included uneven pronuclei, an uneven blastomere, multinucleation, and direct, rapid, and irregular division. A total of 164 single vitrified-warmed blastocyst transfer cycles were analyzed (102 cycles of regularly developed blastocysts and 62 cycles of blastocysts with morphokinetic variables). No significant differences in the age of females or the standard blastocyst morphology were found between these two groups. The regularly developed blastocysts showed significantly higher implantation and clinical pregnancy rates than the blastocysts exhibiting morphokinetic variables (30.4% vs. 9.7% and 37.3% vs. 14.5%, respectively; p < 0.01). The blastocysts that exhibited morphokinetic variables showed different mean development times compared with the regularly developed blastocysts. Although morphokinetic variables are known to have fatal impacts on embryonic development, a considerable number of embryos developed to the blastocyst stage. Morphokinetic variables had negative effects on the implantation and clinical pregnancy rates in vitrified-warmed blastocyst transfer cycles. These findings suggest that blastocysts cultured in a time-lapse incubation system should be considered for selective cryopreservation according to morphokinetic variables.

Impact of presence of antiphospholipid antibodies on in vitro fertilization outcome

OBJECTIVE: To investigate prevalence of antiphospholipid antibody (APA) in Korean infertile women undergoing the first in vitro fertilization (IVF) treatment and to evaluate the influence of APA on the subsequent IVF outcomes. METHOD: Two hundred nineteen infertile women who destined the first IVF were prospectively enrolled in 2 infertility centers. Male factor or uterine factor infertility and women with past or current endocrine or immunologic disorders were completely excluded. Plasma concentration of lupus anticoagulant was measured by clot-based method, and anticardiolipin antibody (IgG/IgM), and anti-β2-glycoprotein 1 antibody (IgG/IgM) was measured by enzyme-linked immunosorbent assay method before starting ovarian stimulation for IVF. RESULTS: APA was positive in 13 women (5.9%). Lupus anticoagulant was positive in 2 women (0.9%), anticardiolipin antibody was positive in 7 women (3.2%), and anti-β2-glycoprotein 1 antibody was positive in 4 women (1.8%). In 193 women entering embryo transfer, clinical characteristics and stimulation outcomes were comparable between APA-positive (n=12) and APA-negative group (n=181). The clinical pregnancy rate (66.7% vs. 45.9%), ongoing pregnancy rate (58.3% vs. 37.0%), and miscarriage rate (12.5% vs. 19.3%) were all similar between APA-positive and APA-negative group. CONCLUSION: The prevalence of APA is low in Korean infertile women undergoing the first IVF cycle, and the presence of APA appears to neither decrease their first IVF success nor increase abortion rate.

Evaluation of human embryo development in in vitro fertilization- and intracytoplasmic sperm injection-fertilized oocytes: A time-lapse study / 대한생식의학회지

OBJECTIVE: We investigated whether the insemination method (in vitro fertilization [IVF] or intracytoplasmic sperm injection [ICSI]) affected morphokinetic events and abnormal cleavage events in embryonic development. METHODS: A total of 1,830 normal fertilized embryos were obtained from 272 IVF and ICSI cycles that underwent ovum retrieval culture using a time-lapse system (Embryoscope) from June 2013 to March 2015. All embryos were investigated by a detailed time-lapse analysis that measured the developmental events in the hours after IVF or ICSI insemination. RESULTS: No significant differences were observed between the two groups regarding clinical outcomes (p>0.05). ICSI-derived embryos showed significantly faster morphokinetics than those derived from conventional IVF, from the time to pronuclear fading to the time to 6 cells (p0.05). There were no differences in abnormal cleavage events between the two groups (p>0.05); they showed the same rates of direct cleavage from 1 to 3 cells, 2 multinucleated cells, 2 uneven cells, and reverse cleavage. CONCLUSION: The morphokinetics of embryo development was found to vary between IVF- and ICSI-fertilized oocytes, at least until the 6-cell stage. However, these differences did not affect the clinical outcomes of the embryo. Additionally, no significant differences in abnormal cleavage events were found according to the fertilization method.

A retrospective study of single frozen-thawed blastocyst transfer / 대한생식의학회지

OBJECTIVE: To study the clinical outcomes of single frozen-thawed blastocyst transfer cycles according to the hatching status of frozen-thawed blastocysts. METHODS: Frozen-thawed blastocysts were divided into three groups according to their hatching status as follows: less-than-expanded blastocyst (≤EdB), hatching blastocyst (HgB), and hatched blastocyst (HdB). The female age and infertility factors of each group were evaluated. The quality of the single frozen-thawed blastocyst was also graded as grade A, tightly packed inner cell mass (ICM) and many cells organized in the trophectoderm epithelium (TE); grade B, several and loose ICM and TE; and grade C, very few ICM and a few cells in the TE. The clinical pregnancy and implantation rate were compared between each group. The data were analyzed by either t-test or chi-square analysis. RESULTS: There were no statistically significant differences in average female ages, infertility factors, or the distribution of blastocyst grades A, B, and C in each group. There was no significant difference in the clinical pregnancy and implantation rate of each group according to their blastocyst grade. However, there was a significant difference in the clinical pregnancy and implantation rate between each group. In the HdB group, the clinical pregnancy and implantation rate were similar regardless of the blastocyst quality. CONCLUSION: There was an effect on the clinical outcomes depending on whether the blastocyst hatched during single frozen-thawed blastocyst transfer. When performing single frozen-thawed blastocyst transfer, the hatching status of the frozen-thawed blastocyst may be a more important parameter for clinical outcomes than the quality of the frozen-thawed blastocyst.

Nomogram to predict the number of oocytes retrieved in controlled ovarian stimulation / 대한생식의학회지

OBJECTIVE: Ovarian reserve tests are commonly used to predict ovarian response in infertile patients undergoing ovarian stimulation. Although serum markers such as basal follicle-stimulating hormone (FSH) or random anti-Müllerian hormone (AMH) level and ultrasonographic markers (antral follicle count, AFC) are good predictors, no single test has proven to be the best predictor. In this study, we developed appropriate equations and novel nomograms to predict the number of oocytes that will be retrieved using patients' age, serum levels of basal FSH and AMH, and AFC. METHODS: We analyzed a database containing clinical and laboratory information of 141 stimulated in vitro fertilization (IVF) cycles performed at a university-based hospital between September 2009 and December 2013. We used generalized linear models for prediction of the number of oocytes. RESULTS: Age, basal serum FSH level, serum AMH level, and AFC were significantly related to the number of oocytes retrieved according to the univariate and multivariate analyses. The equations that predicted the number of oocytes retrieved (log scale) were as follows: model (1) 3.21-0.036×(age)+0.089×(AMH), model (2) 3.422-0.03×(age)-0.049×(FSH)+0.08×(AMH), model (3) 2.32-0.017×(age)+0.039×(AMH)+0. 03×(AFC), model (4) 2.584-0.015×(age)-0.035×(FSH)+0.038×(AMH)+0.026×(AFC). model 4 showed the best performance. On the basis of these variables, we developed nomograms to predict the number of oocytes that can be retrieved. CONCLUSION: Our nomograms helped predict the number of oocytes retrieved in stimulated IVF cycles.

Comparison of static culture, micro-vibration culture, and micro-vibration culture with co-culture in poor ovarian responders / 대한생식의학회지

OBJECTIVE: This study was conducted to compare the effects of static culture, dynamic culture, and the combination of dynamic culture with specialized surfaces involving co-culture on human embryonic development. Embryos cultured using conventional static culture (SC) techniques served as a control group. We compared dynamic culture using micro-vibration culture (MVC) and micro-vibration with co-culture (MCoC), in which autologous cumulus cells were used as a specialized surface. METHODS: We conducted a chart review of patients who were treated between January 2011 and November 2014 in order to compare embryonic development rates and pregnancy rates among the groups. Zygotes were cultured in micro-droplets, and embryos were subsequently selected for transfer. Some surplus embryos were cryopreserved, and the others were cultured for blastocyst development. A micro-vibrator was set at the frequency of 42 Hz for duration of 5 seconds per 60 minutes to facilitate embryo development. RESULTS: No significant differences among the groups were present in patient's characteristics. However, the clinical pregnancy rates were significantly higher in the MVC group and the MCoC group than in the SC group. No significant differences were found in the blastocyst development rate between the SC group and the MVC group, but the blastocyst development rate in the MCoC group was significantly higher than in the SC and MVC groups. CONCLUSION: The clinical pregnancy rate was significantly increased by the application of micro-vibration to the embryonic cultures of poor responders. The blastocyst development rate was significantly increased by the application of MCoC to surplus embryos.

Comparison between intracytoplasmic sperm injection and intracytoplasmic morphologically selected sperm injection in oligo-astheno-teratozoospermia patients / 대한생식의학회지

OBJECTIVE: The aim of this study was to evaluate the efficiency of the intracytoplasmic morphologically selected sperm injection (IMSI) technique compared with conventional ICSI and previous ICSI attempts in oligo-astheno-teratozoospermia (OAT) patients. METHODS: The sperms were selected under high magnification (6,600x) and used to induce fertilization in previous ICSI patients by IMSI. These results were compared with previous conventional ICSI cycles in patients with OAT infertility. RESULTS: These results demonstrated no significant difference in the fertilization rate between IMSI and previous ICSI cycles (67.7% vs. 65.0%). However, the pregnancy and implantation rates with IMSI were significantly higher than those of the ICSI cycles (33.3% vs. 12.5% and 14.6% vs. 5.4%, respectively; p<0.05). The miscarriage rate among pregnant patients (18.2% vs. 37.5%) showed no statistically significant difference between groups. CONCLUSION: Compared to conventional ICSI, this study found that IMSI increased the IVF-ET success rates in patients with OAT.

An effective method for improving outcomes in patients with a fertilization defect / 대한생식의학회지

The effect of artificial oocyte activation (AOA) with a calcium ionophore on intracytoplasmic morphologically selected sperm injection (IMSI) was examined in patients with histories of repeated failed implantation attempts. Four singleton pregnancies and one twin pregnancy were obtained after embryos transfer (5/14, 35.7%). Therefore, AOA combined with IMSI can be considered an option for cycles with a fertilization defect and recurrent implantation failures.

In vitro maturation: Clinical applications / 대한생식의학회지

Oocyte in vitro maturation (IVM) is an assisted reproductive technology in which oocytes are retrieved from the antral follicles of unstimulated or minimally stimulated ovaries. IVM of human oocytes has emerged as a promising procedure. This new technology has advantages over controlled ovarian stimulation such as reduction of costs, simplicity, and elimination of ovarian hyperstimulation syndrome. By elimination or reduction of gonadotropin stimulation, IVM offers eligible infertile couples a safe and convenient form of treatment, and IVM outcomes are currently comparable in safety and efficacy to those of conventional in vitro fertilization. IVM has been applied mainly in patients with polycystic ovary syndrome or ultrasound-only polycystic ovaries, but with time, the indications for IVM have expanded to other uncommon situations such as fertility preservation, as well as to normal responders. In this review, the current clinical experiences with IVM will be described.

Current status of assisted reproductive technology in Korea, 2009

Great advances have been made in the field of assisted reproductive technology (ART) since the first in vitro fertilization (IVF) baby was born in Korea in the year of 1985. However, it deserve to say that the invaluable data from fertility centers may serve as a useful source to find out which factors affect successful IVF outcome and to offer applicable information to infertile patients and fertility clinics. This article intended to report the status of ART in 2009 Korean Society of Obstetrics and Gynecology surveyed. The current survey was performed to assess the status and success rate of ART performed in Korea, between January 1 and December 31, 2009. Reporting forms had been sent out to IVF centers via e-mail, and collected by e-mail as well in 2012. With International Committee Monitoring Assisted Reproductive Technologies recommendation, intracytoplasmic sperm injection (ICSI) and non-ICSI cases have been categorized and also IVF-ET cases involving frozen embryo replacement have been surveyed separately. Seventy-four centers have reported the treatment cycles initiated in the year of 2009, and had performed a total of 27,947 cycles of ART treatments. Among a total of 27,947 treatment cycles, IVF and ICSI cases added up to 22,049 (78.9%), with 45.3% IVF without ICSI and 54.7% IVF with ICSI, respectively. Among the IVF and ICSI patients, patients confirmed to have achieved clinical pregnancy was 28.8% per cycle with oocyte retrieval, and 30.9% per cycle with embryo transfer. The most common number of embryos transferred in 2009 is three embryos (40.4%), followed by 2 embryos (28.4%) and a single embryo transferred (13.6%). Among IVF and ICSI cycles that resulted in multiple live births, twin pregnancy rate was 45.3% and triple pregnancy rate was 1.1%. A total of 191 cases of oocyte donation had been performed to result in 25.0% of live birth rate. Meanwhile, a total of 5,619 cases of frozen embryo replacement had been performed with 33.7% of clinical pregnancy rate per cycle with embryo transfer. When comparing with international registry data, clinical pregnancy rate per transfer from fresh IVF cycles including ICSI (34.1%,) was comparable to clinical pregnancy rate per transfer in European Society for Human Reproduction and Embryology report was 32.5% though lower than 45.0% for USA data. There was no remarkable difference in status of assisted reproductive technology in Korea between the current report and the data reported in 2008. The age of women trying to get pregnant was reconfirmed to be the most important factor that may have impact on success of ART treatment.

Cessation of gonadotropin-releasing hormone antagonist on triggering day in flexible multiple-dose protocol: A randomized controlled study / 대한생식의학회지

OBJECTIVE: To investigate outcomes of stimulated IVF cycles in which GnRH antagonist was omitted on the ovulation triggering day. METHODS: A total of 86 women who underwent controlled ovarian hyperstimulation with recombinant FSH and GnRH antagonist flexible multiple-dose protocols were recruited and prospectively randomized into the conventional group (group A) or cessation group (group B). The GnRH antagonist, 0.25 mg/day of cetrorelix, was started when the leading follicle reached 14 mm in diameter and was continuously administered until the hCG triggering day (group A, 43 cycles) or until the day before hCG administration (group B, 43 cycles). The maturity of oocytes, fertilization rate, embryo quality, and implantation and clinical pregnancy rates were evaluated. RESULTS: The duration of ovarian stimulation, total dose of gonadotropins, serum estradiol levels on hCG administration day, and number of oocytes retrieved were not significantly different between the two groups. The total dose of GnRH antagonist was significantly lower in group B than group A (2.5+/-0.9 vs. 3.2+/-0.8 ampoules, p<0.05). There was no premature luteinization in any of the subjects. The proportion of mature oocytes and fertilization rate were not significantly different in group B than group A (70.7% vs. 66.7%; 71.1% vs. 66.4%, respectively). There were no significant differences in the implantation or clinical pregnancy rates. CONCLUSION: Our prospective randomized study suggested that cessation of GnRH antagonist on the hCG administration day during a flexible multiple-dose protocol could reduce the total dose of GnRH antagonist without compromising its effects on pregnancy rates.

Vitrification of mouse embryos using the thin plastic strip method / 대한생식의학회지

OBJECTIVE: The aim of this study was to compare vitrification optimization of mouse embryos using electron microscopy (EM) grid, cryotop, and thin plastic strip (TPS) containers by evaluating developmental competence and apoptosis rates. METHODS: Mouse embryos were obtained from superovulated mice. Mouse cleavage-stage, expanded, hatching-stage, and hatched-stage embryos were cryopreserved in EM grid, cryotop, and TPS containers by vitrification in 15% ethylene glycol, 15% dimethylsulfoxide, 10 microg/mL Ficoll, and 0.65 M sucrose, and 20% serum substitute supplement (SSS) with basal medium, respectively. For the three groups in which the embryos were thawed in the EM grid, cryotop, and TPS containers, the thawing solution consisted of 0.25 M sucrose, 0.125 M sucrose, and 20% SSS with basal medium, respectively. Rates of survival, re-expansion, reaching the hatched stage, and apoptosis after thawing were compared among the three groups. RESULTS: Developmental competence after thawing of vitrified expanded and hatching-stage blastocysts using cryotop and TPS methods were significantly higher than survival using the EM grid (p<0.05). Also, apoptosis positive nuclei rates after thawing of vitrified expanded blastocysts using cryotop and TPS were significantly lower than when using the EM grid (p<0.05). CONCLUSION: The TPS vitrification method has the advantages of achieving a high developmental ability and effective preservation.

Clinical application of anti-Mullerian hormone as a predictor of controlled ovarian hyperstimulation outcome / 대한생식의학회지

OBJECTIVE: In 2009 anti-Mullerian hormone (AMH) assay was approved for clinical use in Korea. This study was performed to determine the reference values of AMH for predicting ovarian response to controlled ovarian hyperstimulation (COH) using the clinical assay data. METHODS: One hundred sixty-two women who underwent COH cycles were included in this study. We collected data on age, basal AMH and FSH levels, total dose of gonadotropins, stimulation duration, and numbers of oocytes retrieved and fertilized. Blood samples were obtained on cycle day 3 before gonadotropin administration started. Serum AMH levels were measured at a centralized clinical laboratory center. The correlation between the AMH level and COH outcomes and cut-off values for poor and high response after COH was analyzed. RESULTS: Concentration of AMH was significantly correlated with the number of oocytes retrieved (OPU; r=0.700, p or =20) response were 0.94+/-0.15 ng/mL, 2.79+/-0.21 ng/mL, and 6.94+/-0.90 ng/mL, respectively. The cut-off level, sensitivity and specificity for poor and high response were 1.08 ng/mL, 85.8%, and 78.6%; and 3.57 ng/mL, 94.4%, and 83.3%, respectively. CONCLUSION: Our data present clinical reference values of the serum AMH level for ovarian response in Korean women. The serum AMH level could be a clinically useful predictor of ovarian response to COH.

Effect of artificial shrinkage on clinical outcome in fresh blastocyst transfer cycles / 대한생식의학회지

OBJECTIVE: This study aimed to determine the safety and clinical effect of artificial shrinkage (AS) in terms of assisted hatching of fresh blastocysts. Also, we evaluated the correlation between patient age and the effect of AS on clinical outcome. METHODS: Two AS methods, using a 29-gauge needle and laser pulse, were compared. Seventy-three blastocysts were shrunk using a 29-gauge needle and the same number of other blastocysts were shrunk by a laser pulse. We evaluated the shrunken blastocysts hourly and considered them viable if they re-expanded >70%. Blastocyst transfer cycles (n=134) were divided into two groups: a control group consisted of the cycles whose intact embryos were transferred (n=100), while the AS group consisted of the cycles whose embryos were replaced following AS (n=34). The implantation and pregnancy rates of the control group and AS group were compared (p<0.05). RESULTS: The re-expansion rates of the 29-gauge needle and laser pulse AS groups were similar (56 [76.7%] vs. 62 [84.9%], respectively). All of the remaining shrunken blastocysts were re-expanded within 2 hours. There was no degeneration of shrunken blastocysts. The total and clinical pregnancy rate of the AS group (23 [67.6%]; 20 [58.8%], respectively) was significantly higher than that of the control group (47 [47.0%]; 39 [39.0%], respectively). In the older patient group, there was no difference in the clinical outcomes between the AS and control groups. CONCLUSION: These results suggest that AS of blastocoele cavity, followed by the transfer, would be a useful approach to improve the clinical outcome in cycles in which fresh blastocyst stage embryos are transferred.

Serum Anti-Mullerian Hormone and Inhibin B Levels at Ovulation Triggering Day Can Predict the Number of Immature Oocytes Retrieved in In Vitro Fertilization Cycles

The aim of this study was to investigate whether serum levels of anti-Mullerian hormone (AMH) and inhibin B at ovulation triggering day correlate with the number of immature oocytes obtained from stimulated in vitro fertilization (IVF) cycles. Fiftynine consecutive cycles of ovarian hyperstimulation and IVF were selected from 45 women who had tubal (n=18) or unexplained infertility (n=27) and obtained at least one oocyte. Serum levels of AMH and inhibin B at ovulation triggering day were measured by enzyme-linked immunosorbent assay (ELISA). Univariate analysis and multiple regressions revealed that serum AMH or inhibin B levels were significantly correlated with immature oocyte count and the correlation coefficients were higher compared to the mature oocyte count. Serum AMH and inhibin B levels on triggering day seems to be more closely related with the immature oocyte count and thus could be good predictors to determine the immature oocyte count in IVF cycle.

Comparative study between natural cycles and programmed cycles using exogenous steroid hormones for endometrial preparation in cryopreserved-thawed blastocyst transfer / 대한산부인과학회지

OBJECTIVE: The objective of this study was to compare the outcomes of cryopreserved-thawed blastocyst transfer (CT-BT) in natural or programmed cycles using exogenous steroid hormones. METHODS: A total of 221 CT-BT cycles were included and divided into two groups according to endometrial preparation protocols. In natural cycle group (n=116), monitoring was performed by transvaginal ultrasonography to detect ovulation. In programmed cycle group (n=105), oral estradiol valerate, 6 mg/day, was started on the third day of the menstrual cycle and administered continuously, and progesterone in oil 100 mg i.m. daily injection was started on cycle day 15. CT-BTs were performed on five days after ovulation in natural cycles and five days after the initiation of progesterone administration in programmed cycles. Pregnancy rates, implantation rates, and other clinical characteristics of the two groups were compared. RESULTS: Clinical characteristics of study subjects did not differ between the two groups. Post-thaw survival rates, number of transferred blastocysts, and number of good-quality blastocysts were not different. There were no statistically significant differences in implantation rates (21.1% vs. 19.4%), clinical pregnancy rates (36.2% vs. 36.2%), and ongoing pregnancy rates (28.4% vs. 27.6%) between the two groups. CONCLUSIONS: No statistically significant differences were found in pregnancy rates and implantation rates between the two protocols. Our results suggest that both protocols are equally effective for endometrial preparation in CT-BT cycles.

Effect of Fertilization Promoting Peptide (FPP) on the Acrosome Status of Cryopreserved Human Sperm / 대한불임학회지

No abstract available.

Estrogen Receptor Gene Polymorphism (PvuII and XbaI) and Outcomes of Controlled Ovarian Hyperstimulation for In-vitro Fertilization and Embryo Transfer / 대한산부인과학회잡지

OBJECTIVE: The purpose of this study was to investigate the association of the estrogen receptor alpha gene polymorphism (PvuII and XbaI) with outcomes of controlled ovarian hyperstimulation for in- vitro fertilization and embryo transfer (IVF-ET). METHODS: The PvuII and XbaI in the estrogen receptor gene was analyzed by PCR-RFLP in 189 infertile women undergoing controlled ovarian hyperstimulation for in-vitro fertilization and embryo transfer. Comparisons were done between the three PvuII or XbaI genotypes, concerning the numbers of follicles and oocytes, the ratio of oocytes to follicles, serum estradiol concentrations, total dose of gonadotropins, number of transferred embryos, implantation rate and pregnancy rate. RESULTS: In this study, there was no statistically significant difference in outcomes of controlled ovarian hyperstimulation, except total dose of gonadotropins used among three genotypes of PvuII or XbaI polymorphism. CONCLUSION: These results suggest that the PvuII and XbaI polymorphisms of the estrogen receptor gene are not associated with the outcomes of controlled ovarian hyperstimulation for IVF-ET in Korean infertile women.

Effects of Neurotrophic Factors on the Generation of Functional Dopamine Secretory Neurons Derived from in vitro Differentiated Human Embryonic Stem Cells / 대한불임학회지

OBJECTIVE: This study was to examine the in vitro neural cell differentiation patterns of human embryonic stem (hES) cells following treatment of various neurotrophic factors [basic fibroblast growth factor (bFGF), retinoic acid (RA), brain derived neurotrophic factor (BDNF) and transforming growth factor (TGF)-alpha], particulary in dopaminergic neuron formation. METHODS: The hES cells were induced to differentiate by bFGF and RA. Group I) In bFGF induction method, embryoid bodies (EBs, for 4 days) derived from hES were plated onto gelatin dish, selected for 8 days in ITSFn medium and expanded at the presence of bFGF (10 ng/ml) for another 6 days followed by a final differentiation in N2 medium for 7, 14 and 21 days. Group II) For RA induction, EBs were exposed of RA (10-6 M) for 4 days and allowed to differentiate in N2 medium for 7, 14 and 21 days. Group III) To examine the effects of additional neurotrophic factors, bFGF or RA induced cells were exposed to either BDNF (10 ng/ml) or TGF-alpha (10 ng/ml) during the 21 days of final differentiation. Neuron differentiation and dopamine secretion were examined by indirect immunocytochemistry and HPLC, respectively. RESULTS: The bFGF or RA treated hES cells were resulted in similar neural cell differentiation patterns at the terminal differentiation stage, specifically, 75% neurons and 11% glial cells. Additionally, treatment of hES cells with BDNF or TGF-alpha during the terminal differentiation stage led to significantly increased tyrosine hydroxylase (TH) expression of a dopaminergic neuron marker, compared to control (p<0.05). In contrast, no effect was observed on the rate of mature neuron (NF-200) or glutamic acid decarboxylase-positive neurons. Immunocytochemistry and HPLC analyses revealed the higher levels of TH expression (20.3%) and dopamine secretion (265.5+/-62.8 pmol/mg) in bFGF and TGF-alpha sequentially treated hES cells than those in RA or BDNF treated hES cells. CONCLUSION: These results indicate that the generation of dopamine secretory neurons from in vitro differentiated hES cells can be improved by TGF-alpha addition in the bFGF induction protocol.

A New Efficient Cryopreservation of Human Embryonic Stem Cells by a Minimum Volume Cooling Method / 대한불임학회지

No abstract available.