• Clinical and Experimental Reproductive Medicine
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• v.30 no.3
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• pp.193-202
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• 2003

Objective: To evaluate the difference of implantation rate (IR) and clinical pregnancy rate (CPR) between two protocols of endometrial preperation in women undergoing frozen-thawed embryo transfer (FET) cycles. Methods: This study was performed during the different time periods: A retrospective study from January 2000 to June 2001 (phase I) and a prospective study from July 2001 to March 2002 (phase II). All the patients received estradiol valerate (6 mg p.o. daily) starting from day 1 or 2 of the menstrual cycle without pituitary down regulation. Progesterone was administered around day 14 after sonographic confirmation of endometrial thickness $\geq$7 mm and no growing follicle. In Group A (n=88, 99 cycles) of phase I, progesterone was administered i.m. at a dose of 50 mg daily from one day prior to thawing of pronuclear (PN) stage frozen embryo or three days prior to thawing of 6-8 cell stage frozen embryo and then each stage embryos were trasnsferred 2 days or 1 day later after thawing. In Group B (n=246, 299 cycles) of phase I, patients recieved progesterone 100 mg i.m. from one day earlier than group A; two days prior to PN embryo thawing, four days prior to of 6-8 cell embryo thawing. During the phase II, to exclude any differences in embryo transfer procedures, in Group 1 (n=23, 28 cycles) of phase II embryo was transfered by one who have used the progesterone protocol since the phase I. In Group 2 (n=122, 139 cycles) of phase II embryo was transfered by one who use the progesterone protocol from the phase II. Results: When compared across the phase and group, there were no significant differences in the characteristics. During the phase I, there were significant increase in IR (14.4% vs 5.9%, p=0.001) and CPR (28.3% vs 14.5%, p=0.000) in group A. During the phases II, IR (11.8% vs 10.6%) and CPR (27.6% vs 27.3%) show no differences between two groups. Conclusions: In FET cycles, IR and CPR are increased significantly by the change of dosage and timing of progesterone administraton. And the timing is considered to be more important factor because the dosage of progesterone did not affect implantation window in previous studies. Therefore, we suggest that progesterone administration in FET cycle should begin from one day prior to PN stage embryo thawing and three days prior to 6-8 cell stage embryo thawing.

• Journal of Embryo Transfer
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• v.11 no.1
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• pp.7-14
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• 1996

The object of this study was to evaluate the effect of uterine epithelial cells on development of Korean native cattle(KNC) oocytes fertilized in vitro. Qocytes were collected from ovaries of slaughtered Korean Native Cows and matured in TCM199 with granulosa cells supplemented with 10% FBS, 5$\mu$g/ml FSH, 10 JU/ml hCG, and 1$\mu$g/ml estradiol-17$\beta$ for 24 hrs. For co-culture of in vitro development of fertilized ova, oviductal epithelial cells (l$\times$l0˚cells /ml) obtained from slaughtered cow and uterine epithelial cells(1$\times$10˚cells /ml) flushed from the superovulated holstein on Day 7 were incubated in 39$^{\circ}C$, 5% $CO_2$, 95% air. Frozen-thawed KNC sperm was capacitated with BO(Brackett & Oliphant, 1975) medium supplemented with 10mM, 5mM-caffein. Matured oocytes were inseminated for 20 hrs. And then fertilized oocytes were washed with culture medium and transferred to oviductal epithelial cells for in vitro development and three days later a portion of embryos were transferred to uterine epithelial cells. Stastical methods of developmental rates on KNC-IVF oocytes was ANOVA-test. Developmental rates of KNC-IVF oocytes was significant higher(P<0.01) when co-cul-tured with uterine epithelial cells(25.2%) than oviductal epithelial cells. Blatocyst cul-tured for 7 to 9 days were frozen by automatic freezer with 1.4M glycerol-PBS. Survival rates of blastocyst was 40.0%. Fourteen frozen-thawed blastocysts were transferred to five holstein heifers on day 7 after natural estrus. Three recipients were observed twin and one recipient was single by ultra-sound systems on days 45 after embryo transfer.

• Journal of Embryo Transfer
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• v.26 no.2
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• pp.97-104
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• 2011

This study was carried out to confirm the effects of luteotrophin, human chorionic gonadotrophin (hCG), and an anti-luteolytic agent, flunixin meglumin (FM), on pregnancy rates in Hanwoo with in vitro produced (IVP) embryo transfers (ET), and to research the effects on the estrus cycle. Treatments included hCG and FM administration 3~10 minutes prior to ET. Also, pregnancy rates were compared with lidocane treatment and FM treatment prior to ET. The results are shown below. 30-day pregnancy rate was 76.7% in the hCG-treated group and 75.7% in the FM-treated group. Both rates were higher than the 70% rate for the control group. 42-day pregnancy rate was 76.7% in the FM-treated group. This was higher than 66.7% recorded for both the hCG-treated and control groups. The pregnancy rate of the hCG-treated group was high at Day 30 (76.7%) but low at Day 40 (66.7%), and there were no differences from the FM-treated and control groups. The recurrent estrus rate of infertile individuals at 2 weeks after ET was 36.4% in the hCG-treated group, under 71.4% in the FM-treated group and 80.0% in the control group. The non-pregnancy rate of individuals without recurrent estrus was 18.2% in the hCG-treated group, which was higher than the 0% rate in both the FM-treated and control groups. The pregnancy rates were higher in the FM-treated group than the Lidocane-treated group with 72.3% versus 67.5% in the heifers and 48.9% versus 43.6% in the cows. From the above results, the FM treatment proved more effective than the hCG treatment and no treatment whatsoever in increasing pregnancy rates after ET. In addition, hCG treatment was shown to be undesirable due to the deviations it caused in the reproductive physiology of the hCG-treated recipients. Therefore, in our study, the FM treatment resulted in a higher pregnancy rate than either lidocaine treatment or no-treatment in the trials of ET.

• Journal of Embryo Transfer
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• v.23 no.1
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• pp.51-57
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• 2008

The present study investigated the efficient methods to produce in vitro Hanwoo embryos, and to improve the pregnancy rate. The developmental rate, total cell number and ICM ratio of in vitro embryos were compared amongst different culture media. Comparisons were also made on the status of recipients, pregnancy rate along with day of transfer after the estrus. Development of embryos into blastocyst stage in IVMD101 supplemented with 5% fetal bovine serum (FBS) group was significantly higher (34.2%) than that of TCM-199 supplemented with 5% FBS (26.8%) and IVMD101 without FBS (25.9%) (p<0.05). The development rate to blastocyst stage was significantly faster in IVMD101(5% FBS) than that of other groups ($0.2{\sim}2.3%$) (p<0.05). The average number of inner cell mass and trophectoderm were similar among treatment groups, which were $36.0{\sim}44.7$ and $83.3{\sim}106.7$. However, total cell number in IVMD101(5% FBS and 0% FBS) was significantly higher than that of TCM199(5% FBS). There were no differences in the pregnancy rate among treatment groups (32.0%, 33.9% and 28.6%, respectively). However, the pregnancy rate of Day 6 embryos cultured in IVMD101(5% FBS) was significantly (p<0.01) higher than IVMD101 without FBS and TCM-199 + 5% FBS (38.0% vs. 17.2% and 32.4%, respectively). No significant difference was observed for the pregnancy rate between heifer and cow transferred with Day 6 embryos cultured in IVMD 101(5% FBS) (42.7% and 39.3%, respectively). However, there was a significant difference of pregnancy rate (p<0.05) in heifer between one and two embryos transferred (31.4% and 41.9%). There was no difference of pregnancy rate among transfer days after estrus between heifer and cow, but the pregnancy rate of transfer to heifer with day 6 after estrus was significantly higher (p<0.05) than that of day 7 and 8 (22.2% vs. 49.0% and 38.7% respectively). Based on the above findings, there is a possibility to produce in vitro produced embryos cultured in IVMD101(5% FBS) showed higher blastocyst rate and the increased cell number. In terms of the pregnancy rate of in vitro produced embryos, the highest pregnancy rate was observed when two embryos were cultured in IVMD101(5% FBS) and transferred.

• Journal of Embryo Transfer
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• v.15 no.3
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• pp.255-261
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• 2000

The objective of this study was to improve the efficiency of bovine embryo transfer by transferring of Hanwoo embryos into Hanwoo or Holstein recipients. The cryopreserved or fresh in vitro produced(IVP) embryos were transferred into uterine horn contralaterally or ipsilaterally to the corpus luteum. The recipients were inseminated by artificially on the next day of estrus. The pregnancy was diagnosed by rectal palpation at 60∼90 days after transfer of the embryos. The pregnancy rate by transfer of one or two embryos was 78%(7/9) and 74%(31/42), respectively. The pregnancy rates according to the grade of corpus lutea of recipients was 75% (20/27) and 82.0%(18/22) at the grade of A and B, respectively. Ten(67.0%) of 15 Holstein recipients transferred with IVP Hanwoo embryos and 5(42.0%) of 12 Holstein recipients transferred with frozen IVP Hanwoo embryos were pregnant. The single and twin calving ratio in Hanwoos was 77.0%(10/13) and 23.0%(3.13) in the recipients transferred with IVP embryos and 64.0%(7/10) and 27.0%(3/10) in the recipients transferred with frozen IVP embryos, respectively. Twenty-four pregnant cows following transfer of IVP embryos, 21(88.0%) calved the normal calves, and 2(8.3%) aborted. When the frozen IVP embryos were transferred, 16 pregnant cows calved 14(88.0%) normal calves and 2(13.0%) aborted. In conclusion, when one or two IVP bovine embryos were transferred into recipients, the A and B grade of corpus luteum resulted in high pregnancy rates. For the production of twin calves, transfer of the IVP or frozen IVP embryos could be suitable.

• Journal of Embryo Transfer
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• v.29 no.4
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• pp.369-373
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• 2014

This study was performed to establish a new parameter for estimating gestational age and predicting parturition day by ultrasonographic measurement of deep portion of telencephalic vesicle (DPTV) diameter in small dogs. Fetal head diameter (HD) and DPTV diameter were measured in 15 pregnant Pekingese bitches, from Day 15 to the parturition day, and evaluated the correlation between gestational age. HD was measured from day 29 of pregnancy to parturition day and increased from $4.9{\pm}2mm$ to $25.5{\pm}0.7mm$. Especially, from day 38 of pregnancy to parturition day, HD uniformly increased about 0.6 mm per day and was significantly and linearly relative to gestational age during this period ($r^2$ >0.99). DPTV diameter was measured from day 35 to day 60 of pregnancy and increased from $3.2{\pm}0.9mm$ to $11.5{\pm}0.7mm$. Especially from day 38 to day 60 of pregnancy, DPTV diameter uniformly increased about 1 mm per 3 days and was significantly and linearly relative to gestational age during this period ($r^2$ >0.99). In conclusion, DPTV diameter could to be a useful parameter for the estimation of gestational age and the prediction of parturition day when used alone or in combination with HD during the second half of pregnancy.

• Clinical and Experimental Reproductive Medicine
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• v.35 no.3
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• pp.213-221
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• 2008

Objective: To evaluate predictor of IVF outcomes following single embryo transfer in patients with decreased ovarian reserve. Methods: A retrospective review was performed in 919 IVF cycles with elevated basal serum FSH (${\geq}12\;mIU/mL$), the number of retrieved oocytes ${\geq}4$ and serum $E_2$ concentration on hCG day <500 pg/ml between Jan. 1996 and Dec. 2006. Two hundred thirty five IVF cycles following single embryo transfer were included. Pregnancy rates and live birth rates was evaluated according to maternal age, serum $E_2$ on hCG day, basal FSH level, the number of blastomere on day 3 ET, stimulation protocol, the number of cycles of ET. Statistical analysis was used SPSS 12.0 program. Results: OPU cancellation rates were 25.6% (235 cycles), OPU failure rates were 18.5% (170 cycles), embryo transfer cancellation rates were 14.0% (129 cycles). Pregnancy rates following single embryo transfer was 8.1% (19 cycles) and live birth rates was 4.7% (11 cycles). Pregnancy rates and live birth rates of women under 35 years old was statistically higher than those of women above 35 years old (20% vs. 3.5% (p<0.0001), 12.3% vs. 1.8%, (p=0.002)). There was no difference in basal FSH, serum $E_2$ on hCG day, and the number of blastomere on ET, and stimulation protocol. Cumulative pregnancy rates according to the number of cycles of ET were $1^{st}$ 8.1%, $2^{nd}$ 9.2%, $3^{rd}$ 9.7%, $4^{th}$ 9.0%, and $5^{th}$ 9.5%. Conclusion: Pregnancy rates and live birth rates of IVF-ET cycles following single embryo transfer in patients with decreased ovarian reserve are statistically increased in women under 35 yrs old. There is no difference in cumulative pregnancy rates. These data may be helpful for counseling women with decreased ovarian reserve in attempting IVF with their own eggs or when choosing donor oocytes.

• Journal of Embryo Transfer
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• v.25 no.4
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• pp.221-227
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• 2010

Early cleavage is a reliable prognostic tool for successful embryo transfer in assisted reproduction because early cleaved embryo show better pregnancy rate after transfer. There for, preparation of good embryo recipient is important factor to optimize efficiency of pig cloning. The present study was performed to evaluate the effect of early cleavage on the in vivo development of cloned embryos and to analyze breed, parity and estrous synchrony to optimize recipient for pig cloning. In vitro matured porcine oocytes derived from local slaughterhouse and fibroblasts derived from miniature pig fetuses were used for somatic cell nuclear transfer (SCNT). Reconstructed embryos were transferred to recipient pigs on the same day of SCNT or after 1~2 days of in vitro culture for selecting early cleaved embryos. Breed, parity and date of standing estrous of recipients were recorded for analysis. After 25~35 days after embryo transfer pregnancy was diagnosed using ultrasonography, and pregnant recipients were monitored till delivery. Between purebred and crossbred, no significant difference was founded in both pregnancy and delivery rates. However, early cleaved embryos showed significantly higher pregnancy (46.2%) and delivery (12.8%) rates compared to non-selectively transferred group (24.8% and 4.5%, respectively). The results also showed that the recipients showing standing estrous on the same day of SCNT and less than 4 parities were most suitable for pig cloning.

• Journal of Embryo Transfer
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• v.21 no.3
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• pp.225-231
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• 2006

The objective of this study was to determine changes in serum hormone concentrations, blood chemical values and recovery rate of in vivo embryos during the estrous cycle following super-ovulation treatments in Jeju black cows. Superovulation was induced by subcutaneous administration of FSH twice a day for 4 days. Serum hormones were assayed by radioimmunoassay (RIA) and blood chemical values were analyzed by blood analyser system. Embryos were collected from all treated black cows using nonsurgical technique on day 7 after artificial insemination (AI). The results of this study were summarized as follows: 1. The progesterone concentrations were $7.2{\pm}3.8ng/ml$ at day -11 and $0.3{\pm}0.1ng/mL$ at day 0 (Day 0 is the first day of AI). The estradiol concentrations were $10.6{\pm}4.48pg/ml$ at day -11 and $15.0{\pm}2.2pg/ml$ at day 0. The lowest level of progesterone was measured at day 0. The highest levels of estradiol was measured at day 0. 2. The blood chemical values of treated black cows were no significant differences in normal cow values. 3. Sixty two embryos were collected in 12 black cows. Among the collected embryos, 37 embryos (59.7%) could be transferred into recipients. These results would be used as the basic informations for changing patterns of hormonal level and blood biochemistry in Jeju black cow with superovulation.

• Journal of Embryo Transfer
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• v.12 no.1
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• pp.67-74
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• 1997

This study was carried out to increase the viability of bovine frozen4hawed in vitro produced (IVP) embryos and pregnancy rate by direct transfer method. Cumulus-oocyte complexes were aspirated from excised Hanwoo ovaries and matured in TGM 199 for 20~22 hours at 38.5$^{\circ}C$ in 2% $CO_2$ in air. Matured oocytes were fertilized with capacitated sperm for 6 hours and then co-cultured with cumulus cells for 9 days. 63% of the oocytes cultured was deaved and 29% out of them developed into blastocysts. Good or excellent grade of blastocysts on D 7 or 8 were frozen with 1.8M ethylene glycol as a cryoprotectant for direct transfer. Frozen embryos were thawed at 2$0^{\circ}C$ water for 10 sec following 4~5 second in air. For the survival assay of frozen4hawed lVP blastocysts, they were cultured in TCM 199 supplemented with 100$\mu$M $\beta$-mercaptoethanol and 20% FCS for 72 hours. The percentage of embryos developed to re-expanded or hatched after 72 hours culture was 95. 5 and 77.3%, respectively. When frozen-thawed Ivp embryos were transferred to 43 synchronized recipients by direct transfer method, eighteen recipients (41.8%) was pregnant. The highest pregnant was in naturafly synchronized recipients (71.4%), but induced estrus by using PRID(29.2%) and PGF$_2$$\alpha$(20.0%) was showed lower pregnancy rate. The pregnancy rate was higher in day 7 blastocysts(56.0%) than day 8 blastocysts(22.2%). (Key words: in vitro produced, blastocyst, frozen-thawed, direct transfer)