• Molecular & Cellular Toxicology
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• v.4 no.3
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• pp.253-259
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• 2008

Human follicular fluid (HFF) is the in vivo microenvironment for oocyte maturation and includes a variety of proteins that could be involved in oocyte development and fertilization. We therefore used a proteomic approach to identify new HFF proteins. HFF from mature human follicles was obtained from five women following oocyte collection for in vitro fertilization (IVF). Ethanol-precipitated HFF run on two-dimensional gel electrophoresis (2DE) produced approximately 250 Coomassie brilliant blue-stained spots, 64 of which were identified using matrix-assisted laser desorption/ionization-mass spectrometry (MALDIMS). In this study, several proteins including complement factor H, inter-${\alpha}$ (globulin) inhibitor H4, inter-${\alpha}$-trypsin inhibitor heavy chain H4 precursor, human zinc-${\alpha}$-2-glycoprotein chain B, PRO2619, PRO02044, and complex-forming glycoprotein HC were new proteins that have not been previously reported in HFF using proteomic methods. Additionally, we identified alloalbumin venezia for the first time from trichloroacetic acid (TCA)-precipitated HFF. These HFF proteins could serve as new biomarkers for important human reproductive processes.

• Journal of Embryo Transfer
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• v.19 no.2
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• pp.113-119
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• 2004

The purpose of the present study was to examine whether collection time affects results of oocyte recovery from superovulated goats. Fiftyty-one mature Korean native goats, maintained in a pen under natural day length and fed hay ad libitum, were pretreated with progestagen impregnated CIDR for 10 days and then the goats were divided into two groups. One group of the goats received a single intramuscular injection of 1,000 IU PMSG on Day 8 of CIDR insertion. The other group of the goats received twice daily intramuscular injections of a total of 70 mg FSH for 3 days from Day 8 of CIDR. All the gonadotropin treated goats were injected with 10 mg $PGF_{2\alpha}$ on Day 8 and 400 IU hCG in the afternoon on Day 10. For oocyte recovery, donor goats were fasted 24 h before operation. Anesthesia was induced by intravenous injection of 2% xylazine(0.2 mg/kg body weight) and ketamin(11 mg/kg body weight). In vivo oocytes were recovered by follicle aspiration or oviduct flushing at 29 to 34, 35 to 40 and 41 to 50 h after hCG injection through mid-ventral incision. There was no significant difference in the mean number of CL and oocytes recovered. Oocyte collection at 29 to 40 h after hCG increased(P<0.05) the recovery rate of ovulated oocytes in oviducts compared to 41 to 50 h. The same results were also observed in the recovery of follicular oocytes. Oocyte grade was not affected by collection time. When oocytes were collected from follicular oocytes at 41 to 50 h after hCG, the recovery rate of Grade II oocytes was the lowest(P<0.05). From these results, it is suggested that oocyte recovery at 35 to 40 h after hCG will be successful for further use.

• Journal of Embryo Transfer
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• v.20 no.1
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• pp.63-69
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• 2005

This study were carried out to evaluate the possibility of nuclear progression of canine immature oocytes, of which was cultured in a reproductive tract, such as oviduct, ovarian bursa and uterus of estrus bitch for 4, 5 and 6 days following immediately collection. Cumulus intact oocytes(COC) fore collected from domestic dog following ovariohysterectomy at local veterinary clinics. In Exp. 1, COCs $of>110\;{\mu}m$ diameter were selected and cultured in vitro at $39^{\circ}C$, $5\%\;CO\_{2} $ in air atmosphere. The nuclear progression of canine oocytes checked at 24, 48 or 72 h after in vitro maturation. There was not increased the nuclear progression to the M II stage depending on culture periods at 24, 48 and 72h $(1.3\%,\;3.7\%\;and\;4.7\%)$. In Exp. 2, to evaluate of nuclear progression of immature oocytes, collected or in vitro cultured oocytes were transfer into a canine reproductive tract (oviduct, ovarian bursa and uterus). The recovery rates of canine oocytes from a reproductive tract after 4 days $(33.7\%)$ in vivo culture were significantly higher than those 5 $(17.7\%)$ 6 day $(3.4\%)$ (P<0.05). The survival rates of collected oocytes after 4 days $(60.0\%)$ were also significantly higher than those of 5 days $(30.2\%)$ and 6 days $(38.9\%)$ (P<0.05). The meiotic resumption rates of canine oocytes were not significantly difference among the culture periods at 4 days $(5.9\%)$, 5 days $(0.0\%)$ and 6 days $(0.0\%)$. These results show that the nuclear progression of canine immature oocytes from in in vivo culture was not affect the nuclear resumption of oocytes.

• Proceedings of the Korean Society of Developmental Biology Conference
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• 2003.10a
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• pp.128-128
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• 2003

The aims of this study are 1) to test oocytes and embryos collected from in-vivo and in-vitro to achieving the valuable protocol by culturing, vitrifying and thawing of oocytes/embryos, and 2) to transfer them to recipient, and finally have resulted in pregnancies from recipient females after surgical or nonsurgical transfer. In vitro maturation and fertilization were performed according to the procedures of Funahashi et al. Fertilized oocytes were cultured in glucose-free NCSU 23 supplemented with 5 mM sodium pyruvate, 0.5 mM sodium lactate and 4 mg/ml bovine serum albumin for 2 days at 39$^{\circ}C$, and 10% fetal bovine serum was added to the culture medium thereafter. Embryos were treated with 7.5$\square$g/ml cytochalasin-B for 30 min, centrifuged at 13,000 ${\times}$ g for 13 min and then exposed sequentially to an ethylene glycol (EG) vitrification solution, aspirated into OPSs, and plunged/thawed into/from liquid nitrogen. In vivo embryos were surgically collected from three donors after Al. Forty-six embryos (18, 9 and 19 embryos, respectively) were washed 3 times in mPBS+10%FBS, followed treatments : cultured, centrifuged, vitrified, recovered and transferred to recipients as in vitro prepared embryos. Three recipients received surgically 34(control), 188 and 184 embryos (derived from abattoir), respectively. Another three recipients were received nonsurgically 150, 100 and 150 embryos, respectively. All recipient sows exhibited delayed returns to estrus. To our knowledge, these results suggest that required an improved techniques, more vigorous embryos preparation and cleaner uterous condition(use gilt).

• Journal of Embryo Transfer
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• v.19 no.2
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• pp.147-153
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• 2004

본 실험에서는 수술적 방법을 이용하여 난관 절제술을 실시한 성숙 난자의 회수율과 카데터를 이용한 난자 회수율 그리고 새롭게 고안된 개 난자 회수용 니들을 이용한 회수율을 비교하였으며, 각각의 회수 방법이 성숙난자의 외형에 미치는 영향을 조사한 결과는 12두에서 난관절제술로 채취한 성숙난자의 회수율은 89.7% 이었다. 수술적 회수방법에서는 본 연구실에서 개발한 난자 회수용 니들을 난관내에 삽입-결찰한 후 난관-자궁 접합부에서 난자 회수용 배지를 관류하는 방법으로 평균 83.0%의 회수율을 얻었다. 이 같은 결과는 TomCat 카테터를 이용한 회수율 (68.9%)과 난자 회수용 니들을 결찰하지 않고 관류한 방법 (73.5%) 보다 유의적으로 높은 회수율을 나타내었다 (p<0.05). 또한 난관 절제술과 각각의 수술적 방법으로 회수한 난자의 형태학적 차이는 관찰할 수 없었으나 난관 절제술과 난자회수용 니들을 결찰하여 회수한 난자의 형태와 난질이 Tom Cat 카테터나 결찰하지 않은 니들을 이용하였을 때보다 영향을 덜 받는 경향을 나타내었다 (각각 72.0%, 73.8%와 62.8%, 69.6%).