• Korean Journal of Fisheries and Aquatic Sciences
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• v.40 no.6
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• pp.374-379
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• 2007

Two experiments were designed to examine short-term effects of human chorionic gonadotropin (hCG), and long-term effects of gonadotropin-releasing hormone agonist (GnRHa), $17{\alpha}-hydroxyprogesterone$ (17P), and $17{\alpha},20{\beta}-dihydroxy-4-pregnen-3-one\;(17,20{\beta}P)$, alone or in combination, on milt production of the starry flounder Platichthys stellatus. In the first experiment, fish were injected with either 200 IU hCG/kg body weight or the same volume of marine fish Ringer's solution (MFRS). In the second experiment, each fish was implanted with a blank cholesterol pellet (control), $200\;{\mu}g$ GnRHa, $500\;{\mu}g$ 17P, or $100\;{\mu}g\;17,20{\beta}P/kg$ body weight alone or in combination. In the first experiment, hCG injection resulted in an increase in the expressible milt volume and a decrease in the spermatocrit (Sct). After pellet implantation in the second experiment, the milt volume was increased in males treated with GnRHa, GnRHa+17P, or $GnRHa+17,20{\beta}P$. On day 7 after hormone pellet implantation, the milt volume began to increase, and on day 14, the milt volume in the $GnRHa+500\;{\mu}g$ 17P group was significantly higher than that in the control group. Compared with the control group, the hormone pellet-treated groups had a significant reduction in the mean Sct and sperm concentration (Sc) at day 7 after pellet implantation, while there were no differences in total sperm number. The results suggest that increases in milt volume are generally associated with decreases in Sct and SC, suggesting that the main mechanism for the increase in milt volume was milt hydration.

• Natural Product Sciences
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• v.25 no.1
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• pp.49-58
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• 2019

Eleven steroid hormones (SHs: androstene-3,17-dione, estrone, ${\beta}$-estradiol, ${\alpha}$-estradiol, testosterone, dehydroepiandrosterone, $17{\acute{a}}$-hydroxyprogesterone, medroxyprogesterone, megestrol acetate, progesterone, and androsterone) were detected from New Zealand deer (Cervus elaphus var. scoticus) velvet antler (NZA, 鹿茸 ). A method for the quantification of eleven SHs was established by using ultraperformance liquid chromatography (UPLC)-MS/MS. The linearities ($R^2$ > 0.991), limits of quantification (LOQ values, 0.3 ng/mL to 23.1 ng/mL), intraday and interday precisions (relative standard deviation: RSD < 2.43%), and recovery rates (97.3% to 104.6%) for all eleven SHs were determined. In addition, a method for the quantification of three 7-oxycholesterols (7-O-CSs: 7-ketocholesterol, $7{\alpha}$-hydroxycholesterol, and $7{\beta}$-hydroxycholesterol) in the NZA was established by using an HPLC-photodiode array (PDA) method. The linearities ($R^2$ > 0.999), LOQ values (30 ng/mL to 350 ng/mL), intraday and interday precisions (RSD < 1.93%), and recovery rates (97.2% to 103.5%) for the three 7-O-CSs were determined. These quantitative methods are accurate, precise, and reproducible. As a result, it is suggested that the five steroid compounds of androstene-3,17-dione, androsterone, 7-ketocholesterol, $7{\alpha}$-hydroxycholesterol, and $7{\beta}$-hydroxycholesterol could be marker steroids of NZA. These methods can be applied to quantify or standardize the marker steroids present in NZA.

• Journal of The Korean Society of Inherited Metabolic disease
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• v.16 no.2
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• pp.70-78
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• 2016

21-hydroxylase deficiency (21-OHD), most common form of congenial adrenal hyperplasia, is categorized into classical forms, including the salt-wasting (SW) and the simple virilizing (SV) types, and nonclassical (NC) forms based on the severity of the disease. Newborn screening for 21-OHD has been performed in Korea since 2006. $17{\alpha}$-hydroxyprogesterone (17-OHP) is a marker for 21-OHD and is measured using a radioimmunoassay or a fluoroimmunoassay. Premature and low birth weight infants are likely to give false positive 17-OHP findings, therefore, cutoff values for these infants should be determined based on gestational weeks or birth weight. ACTH simulation test is helpful when the 17-OHP shows equivocal increase, and it is gold standard for diagnosis of NC type. Recently, liquid chromatography linked with tandem mass spectrometry was developed for rapid, highly specific, and sensitive analysis of multiple analytes. Molecular analysis of CYP21A2 is useful for confirming diagnosis of mild SV or NC type, predicting prognoses, and genetic counseling. In order to make newborn screening for 21-OHD more efficient, early detection of boy with SW type, early determination of girl with ambiguous genitalia, detection of NC type, and overcoming of false positive in premature and low birth weight infants should be considered. Above all, early treatment should be started when the patient is suspected as having 21- OHD clinically before confirming the diagnosis to prevent adrenal crisis. Here, author reviewed recent articles of guideline and proposed guideline for 21-OHD.

• Journal of Physiology & Pathology in Korean Medicine
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• v.25 no.3
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• pp.417-424
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• 2011

Ketoconazole (KET) is an antifungal drug with a broad spectrum of activity that also induces reproductive toxicity in humans and animals. KET inhibits C17-20 lyase which blocks the conversion of 17 ${\alpha}$-hydroxyprogesterone to androstenedione. The effect of Cuscutae semen(CS) extract against KET-induced testicular damage was evaluated in male rats. CS extract was administered orally (100 and 200 mg/kg) for 26 days. Three weeks after CS extract administration, KET was CS-administered intraperitoneally at a dose of 100 mg/kg once a day for 5 days. KET-induced reproductive toxicity was associated with clear reductions of the weights of testes and epididymides, sperm indices and serum testosterone levels. In addition, marked oxidative damage to testicular lipids and alterations of natural antioxidant enzymes were reported in association with KET toxicity. Most of the KET-induced effects were greatly decreased with the concomitant application of CS extract. This study suggests a protective role of Cuscutae semen extract that could be attributed to its antioxidant properties.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.47 no.6
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• pp.882-887
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• 2014

We studied oocyte steroidogenesis in blacktip grouper Epinephelus fasciatus ovarian follicles during vitellogenesis. Vitellogenic oocytes with average diameters of 0.45, 0.48 and 0.50 mm were incubated in vitro in the presence of $[^3H]17{\alpha}$-hydroxyprogesterone as a precursor. The steroid metabolites were analyzed using thin layer chromatography (TLC), high performance liquid chromatography (HPLC), and gas chromatography-mass spectrometry (GC/MS). The major metabolites in the vitellogenic oocytes were androstenedione ($A_4$), testosterone (T), estradiol-$17{\beta}$ ($E_2$), and estrone ($E_1$). The metabolites of androgen ($A_4$ and T) were higher in the 0.50-mm oocytes than in the 0.45- and 0.48-mm oocytes, while the estrogen metabolites (E2 and E1) were lower in the 0.50-mm oocytes. These results suggest that 0.50-mm oocytes are fully vitellogenic following initiation of the maturation process.

• Development and Reproduction
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• v.13 no.3
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• pp.163-172
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• 2009

Some organotin compounds such as butyltins and phenyltins are known to induce impo-sex in various marine animals and are considered to be endocrine disruptors. In this study, the effect of organotins on follicular steroidogenesis in amphibians was examined using ovarian follicles of Rana dybowskii and Rana catesbeiana. Isolated follicles were cultured for 6 or 18 h in the presence and absence of frog pituitary homogenate (FPH) or various steroid precursors, and the levels of product steroids in the culture media oassay. Among the butyltin compounds, tributyltin (TBT) strongly and dose-dependently inhibited the FPH-induced synthesis of pregnenolone ($P_5$) and progesterone ($P_4$) by the follicles. TBT also strongly suppressed the conversion of cholesterol to $P_5$ and partially suppressed the conversion of $P_5$ to $P_4$. A high concentration of dibutyltin (DBT) also inhibited steroidogenesis by the follicles while monobutyltin and tetrabutyltin had negligible effects. The toxic effect of TBT or DBT was irreversible and a short time of exposure (30 min) was enough to suppress steroidogenesis. All the phenyltin compounds significantly inhibited FPH-induced $P_5$ synthesis by the follicles. The effective dose of 50% inhibition by diphenyltin was $0.04\;{\mu}M$ and those of monophenyltin and triphenyltin were $0.24\;{\mu}M$ and $0.3\;{\mu}M$, respectively. However, none of the phenyltin compounds significantly suppressed the conversion of $P_4$ to $17{\alpha}$-hydroxyprogesterone ($17{\alpha}$-OHP) (by $17{\alpha}$-hydroxylase), $17{\alpha}$-OHP to androstenedione (AD) (by $C_{17-20}$ lyase), or AD to testosterone by the follicles. Taken together, the data show that among the steroidogenic enzymes, P450scc in the follicles is the most sensitive to organotin compounds and that an amphibian follicle culture system can be a useful screening model for endocrine disruptors.

• The Korean Journal of Zoology
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• v.39 no.3
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• pp.273-281
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• 1996

Previously, we have proposed a two-cell type model for follicular steroidogenesis inamphibians with Rana nigromacu lota. Present experiments were carried out to ascertain whether the model Is applicable to R. dybowskii. The role of theca layer were also reassessed by using granulosa cell-free pure theca layer (P-THEP). Theca/epithelium (THEP) layers, P-THEP layers, and granulosa cell enclosed-oocytes () were obtained from ovarian follicles of R. dybowskii by microdissection. Intact follicles (IFs) and different types of tissues were cultured for 6 hour in amphibian Ringer's m the presence or absence of FPII (0.05 gland/mi) or various steroid precursor (100 ng/ml). The amounts of product steroids converted by the components were measured by RIA. Exogenously added pregnenolone (P5) resulted in a marked increase in progesterone (P$_4$) by GCEOs (2143 pg/follicle) and IFs (2346 pg/follicle) but a smaller increase in P4 by THEP layer (495 pg/follicle). Addition of P$_4$ increased 17 a-hydroxyprogesterone (17 $\alpha$-OHP$_4$) levels by GCEOs (1118 pg/follicle) and IFs (1333 pg/follicle) but less by THEP layer (290 pg/follicle). However, much less amounts of P$_4$ or 17 $\alpha$-OHP$_4$ were producad by P-THEP layers than THEP in the presence of P5. Exogenous 1 7$\alpha$-OIIP$_4$ increased androstenedione (AD) levels by GCEOs (1415 pg/follicle) and IFs (561 pg/follicle) but not by THEP layers. In contrast, addition of AD resulted m a marked increase in testosterone (T) levels by TIIEP (2594 pg/follicle) and IFs (2223 pg/follide) but much less by GCEOs (339 pg/follicle). Exogenous T increased estradiol (E$_2$) levels by GCEOs (551pg/follicle) and IFs (887 pg/follicle), but not by THEP layer (<10 pg/follicle). Without addition of FPH or steroid precursors, very low or nondetectable levels of steroids were produced (< 20 pg/follicle) by all the types of follicular components examined. The data presented here indicate that the two-cell type model based on the study with R. nigromacu Iota is applicable to R. dybowskii and also suggest that the minor pathway, which convert P5 to 17$\alpha$-OHP$_4$, is not present in theca layer.

• Animal cells and systems
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• v.15 no.3
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• pp.189-196
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• 2011

We investigated the in vitro effects of nonylphenol (NP) and 3,3',4,4',5-pentachlorobiphenyl (PCB126) on steroidogenesis in redlip mullet, Chelon haematocheilus, oocytes. In experiment 1, we investigated the effects of NP and PCB126 on steroid production from exogenous steroid precursors. Vitellogenic oocytes (0.75 mm in diameter) were incubated with 10 and 100 ng/ml NP or PCB126 with $[^3H]17{\alpha}$-hydroxyprogesterone as a precursor. The major metabolites produced were androstenedione, testosterone (T), estrone, and estradiol-$17{\beta}$ ($E_2$). Both NP and PCB126 increased T production and decreased $E_2$ production, except for 100 ng/ml PCB126. In experiment 2, oocytes (0.65-0.75 mm in diameter) were exposed to NP and PCB126 at different concentrations (0.01, 0.1, 1, 10, and 100 ng/mL). After the incubation, T and $E_2$ production was measured by radioimmunoassay. NP inhibited $E_2$ production at concentrations of 0.01 and 0.1 ng/ml in 0.75-mm-diameter oocytes. NP at 1 and 100 ng/mL stimulated T production, but had no observable effect on $E_2$ production. PCB126 treatment did not affect $E_2$ production at any of the concentrations tested. NP alone at 0.1 ng/mL resulted in a significant decrease in $E_2$ production in 0.65-mm-diameter oocytes. PCB126 did not show any significant effects on either T or $E_2$ production at all concentrations tested. These results suggest that NP acts like an antiestrogen at lower concentrations (0.01-0.1 ng/ml) in vitellogenic oocytes of redlip mullet.

• Reproductive and Developmental Biology
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• v.41 no.1
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• pp.1-6
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• 2017

In all mammalian species, progesterone is essential to both the preparation for, and maintenance of, pregnancy. The $20{\alpha}$-hydroxysteroid dehydrogenase ($20{\alpha}$-HSD) enzyme predominantly converts progesterone into its biologically inactive form $20{\alpha}$-hydroxyprogesterone, thereby regulating its activity. Thus, to directly assess sexual maturation in the MediKinetics $micropig^{(R)}$, we analyzed the concentration of the steroid hormones progesterone and estradiol during the estrous cycle. Our results show that the progesterone level exhibited by the analyzed $micorpig^{(R)}$ was low at the beginning of the estrous cycle, and then abruptly increased to $30.32{\pm}10.0ng/mL$ and $46.37{\pm}11.0ng/mL$ by days 9 and 11 of the cycle, respectively. It reached the highest level $55.87{\pm}3.5ng/mL$ on day 13 of the estrous cycle, before decreasing to $46.58{\pm}13.1ng/mL$ and $10.0{\pm}7.6ng/mL$ by days 15 and 17 of the cycle, respectively. In contrast, the estradiol level was shown to be highest ($27.13{\pm}11.2ng/mL$) at the initiation of the estrous cycle, after which point it decreased to $13.29{\pm}6.5ng/mL$ and $10.94{\pm}5.9ng/mL$ by days 4 and 5 of the estrous cycle, respectively. By day 17 of the estrous cycle, the estradiol level decreased to $4.13{\pm}7.6ng/mL$. We anticipate that these results will provide useful information to enable the study of human ovulation and reproductive physiology using the MediKinetics $micoripig^{(R)}$ as a model system. We recommend further investigation to elucidate the functional mechanisms underlying the regulation of sexual maturation in the MediKinetics $micropig^{(R)}$.

• The Korean Journal of Zoology
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• v.39 no.3
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• pp.257-265
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• 1996

Previously, we demonstrated that estradiol (E$_2$) was produced by medium sized follicles of Rana nigromaculata and Rana dybowskii in vitro. Present experiments were carried out to determine when the growing follicles have obtained the ability to produce E$_2$. Follicles in different growth stages were isolated and cultured for 6 hours in the presence or absence of frog pituitary homogenates (FPH, 0.1 pituitary/2m1) or various steroid precursors (200 ng/2m1). levels of progesterone (P$_4$), 17 -hydroxyprogesterone (17$\alpha$-OHP), androstenedione (AD), testosterone Cr) or E$_2$ in the medium were measured by RIA. The smallest follicles failed to produce steroids, whereas the smaller follicles produced considerable amounts of steroids (211 pg/follicle), and the medium sized follicles produced a large amounts of steroids (1653 pg/follicle) in response to FPH. Addition of pregnenolone (P5) resulted in a marked increase in P$_4$ but not in other steroids by the smallest follicles whereas the treatment resulted in a marked increase in P$_4$, 17$\alpha$-OHP, AD, T and E$_2$by the smaller and medium follicles. When the amounts of steroids are calculated on the basis of unit surface area (pg/mm$_2$), the ability of the smallest follicles to produce P$_4$ from P5 was similar to those of smaller and medium sized follicles. However, the smallest follicles failed to metabolize P$_4$ to other steroids whereas the smaller and medium follicles did. Taken together, the data suggest that the smallest follicles do not response to FPH in terms of steroid production but they have capacity to convert P5 to P$_4$ and that the smaller follicles have potential to produce E$_2$ although much less efficient than medium sized follicles.