• Archives of Pharmacal Research
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• v.26 no.11
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• pp.917-924
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• 2003

This research aims to test a new drug candidate based on a traditional medicinal herb, F1, an herbal extract obtained from Astragalus membranaceus and its main ingredient, 1-monolinolein that may have fewer side effects and less uterine hypertrophy. In vitro experiments, human osteoblast-like cell lines, MG-63 and Saos-2, were analyzed by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and an alkaline phosphatase (ALP) assays. Mouse osteoclasts were induced through a calcium-deficient diet and inhibition effects were measured. In vivo experiments were done using ovariectomized (OVX) rats for 9 weeks. At necropsy, uterus weights were measured, trabecular bone area (TBA) of tibia and lumbar vertebra were measured bone histomorphology. In results, cell proliferation and ALP activity in Saos-2 by ether F1 or 1-monolinolein did not increased significantly compared to the control. The F1 inhibited osteoclast development ($IC_{25}=3.37{\times}10^{-5}$mg/mL) less than 17$\beta$-estradiol. The OVX rats administered F1 (2 mg/kg/day and 10 mg/kg/day) showed an increase in TBA of the tibia significantly (136.3${\pm}4.2% and 138.5{\pm}$10.3% of control). In conclusions, the herbal extract, F1 inhibited tibia and lumbar bone loss and did not cause uterine hypertrophy. However, 1-monolinolein, the main ingredient of the herbal extract, did not inhibit bone loss.

• Development and Reproduction
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• v.21 no.1
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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.

• Asian-Australasian Journal of Animal Sciences
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• v.23 no.5
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• pp.649-658
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• 2010

Myogenic satellite cells (MSCs) are adult stem cells that activate and differentiate into myotubes. These stem cells are multipotent as they transdifferentiate into adipocyte-like cells, nerve cells and osteocytes. The effects of steroid hormones ($E_2$ and testosterone) were studied as a further step toward understanding the mechanism of MSCs proliferation and differentiation. In this study, MSCs were grown continuously for 87 days, implying that there may be a group of MSCs that continue to proliferate rather than undergoing differentiation. Isolated MSCs were cultured in Dulbecco's Modified Eagle's Medium supplemented with adult male, female or castrated bovine serum to observe the effect of steroid hormones on MSC proliferation. Cell proliferation was the highest in cultures supplemented with male serum followed by female and castrated serum. The positive effect of male hormone on MSC proliferation was confirmed by the observation of testosterone-mediated increased proliferation of cells cultured in medium supplemented with castrated serum. Furthermore, steroid hormone treatment of MSCs increased lipid accumulation in myotubes. Oil-Red-O staining showed that 17${\beta}$-estradiol ($E_2$) treatment avidly increased lipid accumulation, followed by $E_2$+testosterone and testosterone alone. To our knowledge, this is the first report of lipid accumulation in myotubes due to steroids in the absence of an adipogenic environment, and the effect of steroid hormones on cell proliferation using different types of adult bovine serum, a natural hormonal system. In conclusion, we found that sex steroids affect MSCs proliferation and differentiation, and lipid accumulation in myotubes.

• Biomedical Science Letters
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• v.8 no.4
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• pp.235-244
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• 2002

This study was carried out to determine the blood and milk progesterone by enzyme-linked immunosorbent assay (ELISA), and milk urea nitrogen (MUN) in cows. MUN and protein concentration were determined using automated infared procedures. The optimum conditions of ELISA system was investigated including the first and second antibody titres, bound percent, and enzyme conjugate and also the factors on MUN and protein concentration by sampling procedures and addition of preservatives. Progesterone antibodies did not react to pregnenlone, testosterone, estrone, estradiol-l7$\beta$, aldosterone, cortisol, corticosterone and 11$\alpha$-dehydroxycortisone (DOC), but reacted with only progesterone. The intra and inter-assay coefficient of variation 4.5%, 6.1~9.4% when used of bovine serum. The morning, MUN concentration (17.6$\pm$2.8 mg/100 ml) in the 13 herds was similar to that of evening MUN concentration of the lactating cows from the same herd. A significant relationship between morning and evening milk samples of upper parameters was found r=0.93. Difference in MUN concentration with sampling procedures and using of preservatives were investigated.

• Development and Reproduction
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• v.20 no.4
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• pp.305-314
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• 2016

The marine medaka, Oryzias dancena is a suitable sample as a laboratory animal because it has a small size and clearly distinguishes between female and male. Data on the growth and maturity of the diploid and triploid sea cucurbit species suitable for laboratory animals are very useful for studying other species. Triploidy was induced in the marine medaka by cold shock treatment ($0^{\circ}C$) of fertilized eggs for 45 min, applied two minutes after fertilization. The diploid and triploid male fish were larger than their female counterparts (P<0.05), and the concentrations of thyroid stimulating hormone (TSH) and thyroxine (T4) were higher in the induced triploids over 1 year (P<0.05). In both the diploid and tri-ploid groups the concentrations of TSH and T4 were higher in the male fish than in the females (P<0.05), while the testo-sterone and estradiol-$17{\beta}$ concentrations in the induced triploids were lower than in the diploids (P<0.05). The gonadosomatic index (GSI) of the triploid fish was lower than that for the diploids, and the GSI for females in each ploidy group were higher than that for the males. For both groups the GSI was highest at 4 months of age, and decreased thereafter to 12 months. Analysis of the gonads of one-year-old triploid fish suggested that the induction of triploidy probably causes sterility in this species; this effect was more apparent in females than in males.

• Korean Journal of Animal Reproduction
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• v.15 no.3
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• pp.161-172
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• 1991

This study was conducted to research the endocrine mechanisms of postpartum anestrus and determine if the morphology of the CL could be related to function in Korean native goats. At parturition 48 goats were assigned to a nonsuckled group and a suckled group. Serum concentrations of luteinizing hormone(LH), follicle stimulating hormone(FSH), prolactin(PRL), estradiol-17$\beta$(E2) and cortisol were measured at various times after parturitionin the goats. The corpora lutea of pregnancy were examined by a light microscope on the 6th hour and the first, 3rd, 10th, 11th, and 21st days after parturition. The results were summarized as follows : Mean serum LH concentrations were lower after parturition in all treatments and increased gradually with the intervals after parturition(P<0.01). These values did not differ between groups. The levels of serum FSH were lower after parturition and tended to increase gradually between 2 and 21 days. The levels of serum FSH are not significantly different between the groups of goats. Two days after kidding mean levels of serum PRL began to fall in nonsuckling goats but increased in suckling goats. During 3 weeks serum PRL concentrations were different between nonsuckling and suckling goats(P<0.01). Three days after parturition the levels of serum E2 decreased in all treatments. From parturition to day 21 serum E2 concentrations were greater in nonsuckling than in suckling goats(P<0.01). At the sixth hour after parturition the structure of the CL was well preserved. At days 1 and 3 the blood vessels were sparcely distributed, whereas, at days 1 and 3 the blood vessels were sparcely distributed, whereas, at days 10, 11 and 21 tortuous larger vessels with thick walls were observed on the luteal tissue. At days 1, 3, 10, 11 and 21 after parturition the CL of pregnancy showed degeneration and the proportion of tissue occupied by intercelluar substances increased at days 21 postpartum. In conclusion, the present study has shown that regression of the CL of pregnancy is accelerated in the period after parturition and effectively completed within three weeks postpartum.

• The Korean Journal of Physiology and Pharmacology
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• v.14 no.5
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• pp.265-272
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• 2010

Follicular cystic ovary (FCO) is one of the most frequently diagnosed ovarian diseases and is a major cause of reproductive failure in mammalian species. However, the mechanism by which FCO is induced remains unclear. Genetic alterations which affect the functioning of many kinds of cells and/or tissues could be present in cystic ovaries. In this study, we performed a comparison analysis of gene expression in order to identify new molecules useful in discrimination of bovine FCO with follicular cystic follicles (FCFs). Normal follicles and FCFs were classified based on their sizes (5 to 10 mm and $\geq25mm$). These follicles had granulosa cell layer and theca interna and the hormone $17{\beta}$-estradiol ($E_2$)/ progesterone ($P_4$) ratio in follicles was greater than one. Perifollicular regions including follicles were used for the preparation of RNA or protein. Differentially expressed genes (DEG) that showed greater than a 2-fold change in expression were screened by the annealing control primer (ACP)-based PCR method using $GeneFishing^{TM}$ DEG kits in bovine normal follicles and FCFs. We identified two DEGs in the FCFs: ribosomal protein L15 (RPL15) and microtubule-associated protein 1B (MAP1B) based on BLAST searches of the NCBI GenBank. Consistent with the ACP analysis, semi-quantitative PCR data and Western blot analyses revealed an up-regulation of RPL15 and a down-regulation of MAP1B in FCFs. These results suggest that RPL15 and MAP1B may be involved in the regulation of pathological processes in bovine FCOs and may help to establish a bovine gene data-base for the discrimination of FCOs from normal ovaries.

• Korean Journal of Pharmacognosy
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• v.37 no.1 s.144
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• pp.21-27
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• 2006

Yeast based estrogenicity assay is the simplest and useful for the assay and the discovery of novel estrogenic substances in natural specimens, The estrogen receptor(ER) modulation activity of 50% EtOH extracts of 101 traditional medicinal herbs was assessed using a recombinant yeast assay system with both a human estrogen receptor expression plasmid and a receptor plasmid. Among them, 14 species proved to be active. Pureariae Flos (flower of Puerraria thunbergiana BENTH.) had the highest estrogenic relative potency$(7.75{\times}10^{-3})$ $(EC_{50}=9.39\;{\mu}g/ml)$. The $EC_{50}$ value of $17{\beta}-estradiol$ used as the positive control was $0.073\;{\mu}g/ml)$ (Relative Potency=1.00). There results demonstrated that some of the traditional medical herb may be useful in the therapy of estrogen replacement.

• Korean Journal of Animal Reproduction
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• v.18 no.3
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• pp.207-216
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• 1994

본 연구는 돼지 난포란으로부터 생산된 수정란의 체외발달에 미치는 아미노산, 우태아혈청 (FBS)과 Leukemia Inhibitry Factor의 영향을 조사하였다. 돼지난포란은 도살된 돼지의 난소로부터 회수하여 39$^{\circ}C$, 5% CO2 배양조건하에서 1$\mu\textrm{g}$/ml FSH-p, 1$\mu\textrm{g}$/ml Estradiol-17$\beta$와 10%FBS가 첨가된 TCM-199 배양액내에서 42시간동안 성숙시켰다. 사출된 정자의 수정능 획득은 45와 90% Percoll density gradient법을 통한 원심분리에 의해 얻었으며, 이들 수정능획득된 정자를 성숙된 난포란이 함유된 배양액에 3$\times$105/ml의 농도로 주입하여 10$\pm$1시간동안 배양함으로서 체외수정을 유도하였다. 수정된 난포란은 ; 1) 10% FBS가 함유된 TCM-199, DMEM, mKRB 또는 CR1aa 배양액, 2) 아미노산 또는 10% FBS가 첨가된 CR1 배양액, 3) STO 세포 또는 mLIF (1,000 unit/ml) 첨가된 CR1aa(10%FBS) 배양액, 4) mLIF (1,000 unit/ml)를 수정 직후 또는 8-세포기 이후에 첨가된 CR1aa(10%FBS)의 네가지 배양조건에서 각각 분리 배양하였다. 그결과 체외수정란의 배반포까지 발달율은 아미노산과 10%FBS가 포함된 CR1 배양액에서 다른 배양액에서보다 양호하였고, 특히 8-세포기 이후에 mouse LIF를 첨가한 CR1aa(10% FBS) 배양액에서는 다른 배양조건보다 현저히 높은 결과를 보였으며, 부화 배반포까지도 배발달을 유도할 수 있었다. 따라서 돼지수정란의 발달에 있어서 배양액내에 아미노산과 FBS 및 mouse LIF의 첨가는 효과가 있으며, 특히 8-세포기 이후에 있어서 mouse LIF의 첨가는 돼지의 수정란을 배반포 이후의 단계에까지 발달시킬 수 있었다.

• Development and Reproduction
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• v.16 no.1
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• pp.9-18
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• 2012

The effects of the estrogenic compound 4-nonylphenol (4-NP) on vitellogenesis in juvenile olive flounder ($Paralichthys$ $olivaceus$) and rockfish ($Sebastes$ $schlegeli$) exposed continuously at 10, 50 and $100{\mu}g{\ell}^{-1}$ levels for 7 days were compared. The expression of VTG mRNA level and protein using specific probes were examined. The levels of plasma estradiol-17 ${\beta}(E_2)$ and testosterone (T) were assessed by radioimmunoassay (RIA). Plasma $E_2$ concentrations increased significantly in two female fish species exposed to $100{\mu}g{\ell}^{-1}$ of 4-NP over concentrations in control fish. Plasma T concentrations increased in $P.$ $olivaceus$. Four days after exposure, the level of VTG mRNA expression increased in $P.$ $olivaceus$ and $S.$ $schlegeli$ exposed to $20{\mu}g{\ell}^{-1}$ of 4-NP. In addition, plasma VTG protein expression was seen in $P.$ $olivaceus$ and $S.$ $schlegeli$. In $S.$ $schlegeli$ and $P.$ $olivaceus$ exposed to 4-NP, the changes were noticed mainly in hepatocytic vaculation after 7 days of exposure. Thus, 4-NP may disrupt vitellogenesis in immature fish both directly and indirectly via disrupted steroidogenesis and liver pathology. Immature $S.$ $schlegeli$ were the most sensitive to 4-NP exposure in vitellogenesis.