• Reproductive and Developmental Biology
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• 제36권4호
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• pp.255-260
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• 2012

The purpose of this study was to assess follicular viability and competence through in vitro culture of preantral follicles isolated from vitrified mouse whole ovaries. Mouse preantral follicles were enzymatically isolated from vitrified- warmed and fresh ovaries and cultured for 10 days followed by in vitro oocyte maturation. In vitro matured oocytes were fertilized and cultured to the blastocyst stage. Five minutes pre-exposure to vitrification solution of whole ovaries had significantly higher (p<0.05) oocyte survival and maturation rates than between 10 min exposure groups. Oocyte diameter was significantly smaller (p<0.05) in the 5 and 10 min exposure groups ($69.4{\pm}2.8$ and $67.8{\pm}3.1$) when compared to that of control group ($71.7{\pm}2.1$). There was no statistical significant difference in blastocyst development rates between vitrification group (8.6%) and the fresh control group (12.0%). The mean number of cells per blastocyst was significantly lower (p<0.05) in the vitrification group ($41.9{\pm}20.2$) than in the fresh control group ($55.1{\pm}22.5$). The results show that mouse oocytes within preantral follicles isolated from the vitrified whole ovaries can achieve full maturation, normal fertilization and embryo development.

• 한국발생생물학회지:발생과생식
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• 제21권1호
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• pp.71-78
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• 2017

Nesfatin-1/NUCB2 is known to take part in the control of the appetite and energy metabolism. Recently, many reports have shown nesfatin-1/NUCB2 expression and function in various organs. We previously demonstrated that nesfatin-1/NUCB2 expression level is higher in the pituitary gland compared to other organs and its expression is regulated by $17{\beta}-estradiol$ and progesterone secreted from the ovary. However, currently no data exist on the expression of nesfatin-1/NUCB2 and its regulation mechanism in the pituitary of male mouse. Therefore, we examined whether nesfatin-1/NUCB2 is expressed in the male mouse pituitary and if its expression is regulated by testosterone. As a result of PCR and western blotting, we found that a large amount of nesfatin-1/NUCB2 was expressed in the pituitary and hypothalamus. The NUCB2 mRNA expression level in the pituitary was decreased after castration, but not in the hypothalamus. In addition, its mRNA expression level in the pituitary was increased after testosterone treatment in the castrated mice, whereas, the expression level in the hypothalamus was significantly decreased after the treatment with testosterone. The in vitro experiment to elucidate the direct effect of testosterone on NUCB2 mRNA expression showed that NUCB2 mRNA expression was significantly decreased with testosterone in cultured hypothalamus tissue, but increased with testosterone in cultured pituitary gland. The present study demonstrated that nesfatin-1/NUCB2 was highly expressed in the male mouse pituitary and was regulated by testosterone. This data suggests that reproductive-endocrine regulation through hypothalamus-pituitary-testis axis may contribute to NUCB2 mRNA expression in the mouse hypothalamus and pituitary gland.

• 한국발생생물학회지:발생과생식
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• 제18권4호
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• pp.301-309
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• 2014

Nesfatin-1, an anorexic nucleobindin-2 (NUCB2)-derived hypothalamic peptide, controls appetite and energy metabolism. Recent studies show that nesfatin-1/NUCB2 is expressed not only in the brain but also in gastric and adipose tissues. Thus, we investigated the distributions of nesfatin-1/NUCB2 in various tissues of male and female mice by real-time PCR, western blotting, and immunohistochemical staining. Real-time PCR analyses showed that NUCB2 mRNA was predominantly expressed in the pituitary and at lower levels in the hypothalamus, spleen, thymus, heart, liver, and muscle of both male and female mice. Expression was much higher in reproductive organs, such as the testis, epididymis, ovary, and uterus, than in the hypothalamus. Western blot analysis of the nesfatin-1 protein level showed similar results to the real-time PCR analyses in both male and female mice. These results suggest that nesfatin-1/NUCB2 have widespread physiological effects in endocrine and non-endocrine organs. In addition, immunohistochemical staining revealed that nesfatin-1 was localized in interstitial cells, including Leydig cells and in the columnar epithelium of the epididymis. Nesfatin-1 was also expressed in theca cells and interstitial cells in the ovary and in epithelial cells of the endometrium and uterine glands in the uterus. These results suggest that nesfatin-1 is a novel potent regulator of steroidogenesis and gonadal function in male and female reproductive organs. Further studies are required to elucidate the functions of nesfatin-1 in various organs of male and female mice.

• 생명과학회지
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• 제22권5호
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• pp.687-691
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• 2012

본 연구는 성숙마우스의 난소에서 황체형성호르몬수용체(luteinizing hormone receptor, LHR)와 beta-galctoside-binding animal lectin인 galetin-3 (Gal-3)의 발현분포를 면역조직화학방법으로 평가하였다. LHR은 활동(성숙) 황체에서 강하게 발현되는 반면에, Gal-3의 발현은 퇴행중인 황체와 폐쇄난포에서 높게 나타났다. 또한, 임신중인 마우스의 황체에서는 LHR의 면역반응이 강하게 나타났으나, Gal-3의 면역반응은 현저히 낮았다. 위의 결과를 종합해 볼 때, 성숙마우스의 난소에서 LHR과 Gal-3는 난소의 특정한 시기에 따라 다른 분포양상을 나타냄을 알 수 있었으며, 이들 각각의 단백질의 발현으로 보아 황체의 형성과 퇴행에서 상반된 역할을 할 것으로 여겨진다.

• 한국수산과학회지
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• 제42권1호
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• pp.1-7
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• 2009

The toxicity of two species of puffer fish, Takifugu poecilonotus (Heuinjeombok) and T. vermicularis (Gukmaeribok) collected from the coastal regions of Korea was determined using a mouse bioassay. In the T. poecilonotus collected in Jeju and Tongyeong, the proportion of toxic specimens containing ${\ge}10$ mouse units (MU) per gram exceeded 95% for the skin, liver, ovary, and fin, and approximately 30% for the testis and muscles. In each of the organs, the highest toxin levels were 79 MU/g in the muscle, hundreds (158-365) of MU per gram in the fin, intestine, testis, and gallbladder, but thousands (1,147-2,406) of MU per gram in the skin, liver, and ovary. In T. vermicularis collected from Incheon and Gunsan, the proportions of toxic specimens were 100% for the gallbladder, and 56-68% for the skin, fin, liver, and intestine however, no toxic muscle specimens were noted. The highest toxin scores were below 10 mouse units (MU) per gram in the muscle, 20-94 MU/g in the skin and fin, 319 MU/g in the intestine, and thousands (1,548-4,624) of MU per gram in the liver, gonad, and gallbladder. The toxicity in the muscle of T. vermicularis was deemed acceptable for human consumption, whereas the toxicities in the muscle of T. poecilonotus and the skin of both species of puffer fish were significantly high, such that special attention may be required when the fish is intended for human consumption.

• 한국발생생물학회지:발생과생식
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• 제2권1호
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• pp.1-8
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• 1998

본 연구에서는 자연세포사 (apoptosis)를 유발시키는 것으로 알려진 ceramide를 배양중인 생쥐 과립세포에 처리한 뒤 형광염색, in-situ 3'-end labeling(ISEL), 그리고 flow cytometry 기법을 이용하여, 자연세포사 및 세포주기에 미치는 ceramide의 영향을 조사하였다. Ceramide를 처리하지 않은 대조군에 비하여, ceramide를 처리한 실험군에서 세로의 생존율은 농도에 비레하여 유의하게 감소하였다. 또한 acridine orange에 의한 형광염색 결과, 자연세포사의 양상을 보이는 핵을 갖는 세포의 수가 ceramide의 농도가 증가함에 따라 현저하게 증가되었다. 또한 ISEL을 실시해 본 결과, 자연세포사가 ceramide의 처리농도가 증가됨에 따라 점차적으로 증가되었다. 한편, ceramide를 처리한 과립세포의 세포주기 분석을 위한 flow cytometry 결과도 자연세포사가 일어난 $A_{0}$기에 있는 세포들의 비율이 대조군에 비하여 농도 의존적으로 증가하였으며, $G_{0}$/$G_{1}$ 기에 있는 세포들이 비율은 현저하게 감소됨을 관찰할 수 있었다. 위의 결과로 보아 ceramide는 생쥐 과립세포의 $G_{0}$/$G_{1}$ 기에 특이저긍로 자\ulcorner하여 자연세포사를 유발하며, 난포의 폐쇄시 과립세포의 자연세포사를 유발할 것으로 사료된다.

• Reproductive and Developmental Biology
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• 제39권2호
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• pp.43-47
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• 2015

Sirtuin proteins are evolutionary conserved Sir2-related $NAD^+$-dependent deacetylases and regulate many of cellular processes such as metabolism, inflammation, transcription, and aging. Sirtuin contains activity of either ADP-ribosyltransferase or deacetyltranfease and their activity is dependent on the localization in cells. However, the expression pattern of Sirtuins has not been well studied. To examine the expression levels of Sirtuins, RT-PCR was performed using total RNAs from various tissues including liver, small intestine, heart, brain, kidney, lung, spleen, stomach, uterus, ovary, and testis. Sirtuins were highly expressed in most of tissues including the testis. Immunostaining assay showed that Sirt1 and Sirt6 were mainly located in the nucleus of germ cells, spermatocytes, and spermatids in the seminiferous tubules, whereas Sirt2 and Sirt5 were exclusively present in the cytoplasm of germ cells and spermatocytes. Our results indicate that Sirtuins may function as regulators of spermatogenesis and their activities might be dependent on their location in the seminiferous tubules.

• Nuclear Engineering and Technology
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• 제31권4호
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• pp.385-390
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• 1999

Ovarian follicles are faced with one of two fates, atresia or development. Up to 99% of follicles become degenerated rather than ovulated in female life span. Thus, atresia occurs at all stages of follicle development in mammalian ovaries. In the present experiment, the effect of ${\gamma}$-radiation on primordial follicles was morphologically analyzed in a mouse ovary. Thirty-seven percent of the primordial follicles in the non-irradiated control mice ovaries were abnormal. At day 8 post irradiation, most of primordial follicles became atretic. They lost their integrity of architecture in the follicular shape. Then, all the oocytes disappeared from the follicles. And only 3 to 4 granulosa cells lay down onto the basement membrane. Disappearance of granulosa cells or oocytes resulted from the radiation-induced apoptotic process. It is definitely clear that ${\gamma}$-radiation induces rapid apoptotic degeneration of the primordial follicles. The morphological degeneration induced by radiation in the primordial follicles can be used as an experimental model to draw out a deeper insight for radioprotectant researches.

• Asian-Australasian Journal of Animal Sciences
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• 제29권7호
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• pp.944-951
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• 2016

Tdrd12 is one of tudor domain containing (Tdrd) family members. However, the expression pattern of Tdrd12 has not been well studied. To compare the expression levels of Tdrd12 in various tissues, real time-polymerase chain reaction was performed using total RNAs from liver, small intestine, heart, brain, kidney, lung, spleen, stomach, uterus, ovary, and testis. Tdrd12 mRNA was highly expressed in testis. Antibody against mouse TDRD12 were generated using amino acid residues SQRPNEKPLRLTEKKDC of TDRD12 to investigate TDRD12 localization in testis. Immunostaining assay shows that TDRD12 is mainly localized at the spermatid in the seminiferous tubules of adult testes. During postnatal development, TDRD12 is differentially expressed. TDRD12 was detected in early spermatocytes at 2 weeks and TDRD12 was localized at acrosome of the round spermatids. TDRD12 expression was not co-localized with TDRD1 which is an important component of piRNA pathway in germ cells. Our results indicate that TDRD12 may play an important role in spermatids and function as a regulator of spermatogenesis in dependent of TDRD1.

• Reproductive and Developmental Biology
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• 제34권4호
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• pp.283-288
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• 2010

During normal early embryonic development in mammals, the global pattern of genomic DNA methylation undergoes marked. changes. The level of methylation is high in male and female gametes. Thus, we cloned the cDNA of the porcine DNA methyltransferase 1 (Dnmt1) gene to promote the efficiency of the generation of porcine clones. In this study, porcine Dnmt1 cDNA was sequenced, and Dnmt1 mRNA expression was detected by reverse transcription-polymerase reaction (RT-PCR) in porcine tissues during embryonic development. The porcine Dnmt1 cDNA sequence showed more homology with that of bovine than human, mouse, and rat. The complete sequence of porcine Dnmt1 cDNA was 4,774-bp long and consisted of an open reading frame encoding a protein of 1611 amino acids. The amino acid sequence of porcine DNMT1 showed significant homology with those of bovine (91%), human (88%), rat (76%), and mouse (75%) Dnmt1. The expression of porcine Dnmt1 mRNA was detected during porcine embryogenesis. The mRNA was detected at stages of porcine preimplantation development (1-cell, 2-cell, 4-cell, 8-cell, morula, and blastocyst stages). It was also abundantly expressed in tissues (lung, ovary, kidney and somatic cells). Further investigations are necessary to understand the complex links between methyltransferase 1 and the transcriptional activity in cloned porcine tissues.