• Clinical and Experimental Reproductive Medicine
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• 제38권1호
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• pp.31-36
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• 2011

Objective: To determine whether the serum ${\beta}$-human chorionic gonadotropin (hCG) profile following preimplantation genetic diagnosis (PGD) is lower than that of intracytoplasmic sperm injection (ICSI) cycles. Methods: A total of 129 PGD cycles and 1,161 age-matched ICSI cycles, which resulted in pregnancy (serum ${\beta}-hCG{\geq}5$ mIU/mL) on post-ovulation day (POD) 12 were included. We compared the mean serum ${\beta}$-hCG levels on POD 12, 14, 21, and 28, doubling time of serum hCG, and created a cut-off value for predicting a singleton pregnancy in each group. Results: The mean serum ${\beta}$-hCG concentration of the PGD group was significantly lower than that of the control group on POD 12, 14, and 21. The doubling time of serum ${\beta}$-hCG at each time interval showed no significant difference. The cut-off-value of serum ${\beta}$-hCG for predicting a single viable pregnancy was 32.5 mIU/mL on POD 12 and 113.5 mIU/mL on POD 14 for the PGD group, which was lower than that for the control group. Conclusion: Blastomere biopsy may decrease the ${\beta}$-hCG-producing activity of the trophoblasts, especially in early pregnancy. Setting a lower cut-off value of serum ${\beta}$-hCG for predicting pregnancy outcomes in PGD may be needed.

• 한국가축번식학회지
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• 제16권4호
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• pp.341-346
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• 1993

The development of isolated blastomeres from mammalian preimplantation embryos has been basically studied for the multiplication of embryos from superior animals. Therefore, this study was investigated the effect of co-culture with mouse fetal fibroblast cells(MFFC) on in vitro development of blastomeres from mouse preimplantation embryos. Mature female ICR mice were treated with hormone to induce superovulation and embryos were collected at each 2, 4, and 8-cell stage. Then, after removing zona pellucida with protease, blastomeres were isolated by micropipetting, or reconstituted with different stage blastomere, and incubated for 72 hrs either in T6 or TCM199 or on the monolayer of MFFC, which was prepared with fibroblast cells from 14∼14 day mouse fetus. After incubation, we examined their development rates every day and the nuclei numbers of each blastocyst by Hoechst-33342 staining. In the development rates of blastomeres, there were no significant differences between media but the higher rateswere found in the monolayer of MFFC, regardless of reconsititution. In addition, blastomeres cultured with MFFC had slightly greater number of nuclei than those cultured in single media. Generally, the higher development rates of blastomeres were found from earlier stage embryos than the later ones, regardless of culture conditions. Reconsitituted blastomeres had more nuclei but did not show the higher development rates, compared to the single blastomeres. Taken together, our results suggest that co-culture with MFFC have a beneficial effect on the in vitro development of blastomeres from mouse embryos.

• 한국가축번식학회지
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• 제16권4호
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• pp.347-352
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• 1993

This study was designed to develop the in vitro culture systemof mammalian preimplantation embryos. We proposed mouse fetal fibroblast cells (MFFC) from 14∼15 day mouse fetus. Zygotes from superovulated female ICR mice were cultured 96 hrs in simple defined media (T6) or on the monolayer of MFFC. In addition, to evaluate the action of the co-culture of MFFC, various diluted superoxide dismutase antibody (SOD-Ab) was supplemented into the monolayer of MFFC and zygotes were cultrued in presence or absence of SOD-Ab. The developmental rates of zygotes were significantly increased in co-culture with MFFC compared to the control. The rates of zygotes to the 4-cell stage in media treated with EDTA were higher than those cultured in MFFC but the proportions of morula and blastocyst were not differ between EDTA and MFFC. Interestingly blastocysts in co-culture with MFFC possessed as many as blastomere as those developing in vivo, but blastocysts cultured with EDTA had significantly fewer blastomeres. In addition, the treatment of SOD-Ab suppressed the beneficial effect of MFFC. Therefore, our findings suggest that co-cultrue system using MFFC may have an advantage in the development of mouse zygotes as well as embryonic differentiation.

• 한국가축번식학회지
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• 제18권2호
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• pp.121-126
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• 1994

This study was conducted to examine the condition of activation of the nuclear transplant bovine embryos. In vitro fertilized(IVF) and nuclear transplant embryos(NTs) were co-cultured with bovine oviduct epithelial tissue(BOET). NTs were treated with cycloheximide(CHXM) for 0 to 6 h after electrofusion to investigate the activation conditin of recipient ooplast. Then, the infljence of the CHXM treatment timing on the cleavage and development of NTs were investigated in relation to the nuclear transplant time. The cleavage rates of NTs were increased with the increasing time of the CHXM treatment from 0 to 6 h (54.7 to 91.3%, P<0.01). Similar trend was shown in the development into the morula or blastocyst stage, but very limitted. Activation of enucleated oocytes prior to fusion enhanced development of NTs compared with that post fustion. This result suggests that the frequency of activation of NTs can be greatly enhanced by treating with CHXM for 6 h. The result also suggests that if blastomeres of unknown cell cycle stage are used, activation of enucleated oocytes prior to fusion enhances development of NTs.

• 한국가축번식학회지
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• 제20권2호
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• pp.155-161
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• 1996

For a large sclase production of genetically identical or cloned animals, the effect of cryopreservation by vitrification on the post-thaw viability of nuclear transplant rabbit embryos were investigated. The embryos of 16-cell stage were collected from the mated does at 48 hours post-hCG injection, and they were synchronized to G1 phase of 32-cell stage were injected into enucleated recipient cytoplasms by micromanipulation. After culture until 20h post-hCG injection, the nuclear transplant oocytes were electrofused and activated by electrical stimulation. After in vitro culture for 48h, the nuclear transplant embryos developed to morula stage were cryoperserved with EFS solution by vitrification method. The forzen nuclear transplant embryos were thawed and cultured for 72h and the nuclear transplant of blastomeres under a fluorescence microscopy. The in vitro development to blastocyst of intact-fresh and intact-frozen 16-cell embryos was found to be 96.9 and 63.9%, respectively. The in vitro development to blastocyst of nuclear transplant and frozen-thawed nuclear transplant embryos was found to be 74.5 and 42.9%, respectively. Also, their mean blastomere numbers and mean cell cycles/day was 153 and 105, 145 and 1.34, respectively. From the above results it was concluded that the present cryopreservation by vitrification of nuclear transplant rabbit embryos might be useful though was decreased significantly.

• Clinical and Experimental Reproductive Medicine
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• 제46권4호
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• pp.206-210
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• 2019

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.

• 한국수정란이식학회지
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• 제8권1호
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• pp.13-19
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• 1993

These experiments were carried out to determine the effect of pregnancy in bisected embryo. The embryos of ICR mouse were microsurgically bisected at morula and blastocyst stage using microsurgical blade attached a micromanipulator. These bisected embryos without zona pellucida were cultured up to blastocyst stage and cell count and diameter of stained blastomere, and transferred pseudopregnant mice. And the development of these bisected embryos was compared with the results of production of young of the corresponding intact embryos or cell stage. When the bisected mouse embryos were cultured in vitro for 20 to 24 hours in morula stage(77.2%) or 3 to 6 hours in blastocyst stage(84.1%), them were developed to the expanded blastocyst stage. There were no significant(P<0.05) differences in the development rate of bisected embryos between in morula and blastocyst stages. The embryo size of blastocyst developed in vitro from bisected embryo was small(P<0.05)than intact embryo. However, the number of blastomeres with bisected embryo (24.7+1.3and 21.5+1.2 respectively) were significantly(P<0.05) reduced, compared with that of intacted embryos(36.3+1.1 and 41.4+1.2 respectively). When compared with the result of pregnancy rate(63.6%) after surgical transfer of bisected morulae, a similar result(65.4%) was obtained with bisected blastocyst stage(P< 0.05). However, production of youngs (38.8%) after transfer of bisected morula, a similar result (38.1%) was obtained with bisected blastocyst stage (P<0.05).

• 한국가축번식학회지
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• 제14권2호
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• pp.133-140
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• 1990

This study was conducted to develop a simple and efficient technique for fusing 2-cell mouse embryos to obtain tertraploid embryos. Various concentration of PEG and exposure times were compared in order to determine the best condition for fusion and subsequent of fused embryos. The results obtained were follows ; 1. The incidence of fusion induction treated with 40% PEG(70.8%) and 45%(62.7%) for 60 sec. exposure were higher than those of 40% and 45% PEG for 30 sec., 90 sec., or 120 sec. exposure group. Also, the highest incidence of fusion induction(76.9%) was achieved with 120 sec. exposure at 50% PEG concentration. 2. Fused embryos after PEG treatment were cleavaged 2-to 4-cell, 8-cell, morula and blastocyst at 20-24 hr., 30-34., 44-52 hr., respectively, and were not different from those obtained fleshly. 3. The high proportions of the embryos developed to blastocysts after blastomere fusion with 40% PEG for 60 sec., 45% PEG for 60 sec. and 50% for 120 sec. were 66.7%(42/63), 69.0%(29/42) and 32.0%(16/50), respectively, this trend indicated that the fusion rate was similar to the incidence of fused embryos forming blastocysts. 4. The cell number of blastocyst developed from fused embryos(18.7 2.6) was samller than that of untreated embryos(48.9 1.69)

• 한국가축번식학회지
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• 제17권4호
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• pp.287-294
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• 1994

This study was to investigate the effects of electrofusion, activation and developmental stage of donor embryos on in vitro development of nuclear transplant bovine embryos. A single blastomere nucleus from 8-cell to morula stage embryos produced by in vitro fertilization(IVF) was transferred into a recipient oocyte enucleated at 23∼25 h after in vitro maturation(IVM) or into a recipient oocyte enucleated and cultured for 14∼15 h. In one experiment the nuclear transplant embryos were subjected to additional activation treatments. Fusion rate of nuclear transplant eggs was high at direct current(D.C) voltages of 1.0 and 1.5 kV/cm 991.5 and 93.3%, respectively), but decreased at 2.0kV/cm (81.8%). Additional activation treatments by electric pulases or 7% ethanol did not affect the cleavage and development of nuclear transplant embryos. Development of nuclear transplant embryos slightly increased by delayed nuclear transfer and fusion (42∼43 h after IVM). With this system, blastocysts were obtained from transfer of 8-cell to morula stage donor nuclei (9.6%∼2.4%). The result of this study suggests that nucleo-cytoplasmic interactins, expecially activation of ooplast are very important for the development of nuclear transplant embryos, and donor cell stage does not affect the development of nuclear transplant embryos.

• Clinical and Experimental Reproductive Medicine
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• 제27권3호
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• pp.307-311
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• 2000

Objective: To report the successful delivery in a patient of recurrent spontaneous abortion caused by chromosomal abnormality. Material and Method: Case report. Results: Twelve oocytes were obtained by in vitro fertilization. Of eleven oocytes fertilized, two embryos turned out to be normal by using fluorescent in situ hybridization on blastomere biopsy. The patient succeeded in pregnancy and the result of amniocentesis was found to be normal. She delivered the healthy female baby by cesarean section. Conclusions: The successful delivery is possible in recurrent spontaneous abortion related with reciprocal translocation by using preimplantation genetic diagnosis.