• Development and Reproduction
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• v.14 no.2
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• pp.115-121
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• 2010

The in vitro effects of nonylphenol (NP) and 2,2',4,6,6'-pentachlorobiphenyl (PCB104) on ovarian steroidogenesis of the ribbed gunnel, Dictyosoma burgeri were investigated. Oocytes taken during maturation stage were incubated with 100 ng/$m{\ell}$ of NP and PCB104 in the presence of exogenous precursor, $[^3H]-17{\alpha}$-hydroxyprogesterone ($[^3H]-17{\alpha}OHP$). Steroids were extracted from the media and the isolated oocytes, and the extracts were separated and identified by thin layer chromatography. The identities of the major metabolites were testosterone (T) and estradiol-$17{\beta}$ (E2). NP treatment inhibited production of E2 metabolite in the oocytes of 1.2, 1.3 and 1.4 mm although NP inhibited production of T metabolite at the oocytes of 1.1, 1.3 and 1.4 mm. PCB104 treatment inhibited production of T metabolite in the oocytes of all groups and E2 metabolite in the oocytes of 1.2, 1.3 and 1.4 mm. In conclusion, these results suggested that NP and PCB104 had an inhibitory effects on conversion of $[^3H]-17{\alpha}OHP$ to T and E2 during the oocyte maturation process of ribbed gunnel.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.48 no.4
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• pp.483-488
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• 2015

We studied oocyte steroidogenesis in the blackfin flounder Glyptocephalus stelleri as a region-specific species, in the East Sea of Korea during the spawning season. Maturing oocytes (0.76, 0.82, 0.88, and 0.91 mm in oocyte diameter) were incubated in vitro in the presence of [$^3H$] $17{\alpha}$-hydroxyprogesterone ($[^3H]17{\alpha}$-OHP) as a precursor. Steroid metabolites were extracted from the incubated medium and oocytes, and the extracts were separated and identified by thin-layer chromatography (TLC), high-performance liquid chromatography (HPLC) and gas chromatographymass spectrometry (GC/MS). The major metabolites produced from $[^3H]17{\alpha}$-OHP were androgens [androstenedione (A4) and testosterone (T)] and estrogens [$17{\beta}$-estradiol (E2) and estrone (E1)] and progestins [$17{\alpha},20{\alpha}$-dihydroxy-4-pregen-3-one ($17{\alpha}20{\alpha}P$) and $17{\alpha}20{\beta}$-dihydroxy-4-pregnen-3-one ($17{\alpha}20{\beta}P$)] in maturing oocytes. The metabolic rate of $17{\alpha}20{\beta}$ was elevated (29.04%) in oocytes measuring 0.88 mm (nucleus migration stage following the induction of germinal vesicle breakdown), but was very low in oocytes measuring 0.76, 0.82, and 0.91 mm (0.42, 0.67, and 2.62%, respectively). From these results, we suggest that $17{\alpha}20{\beta}P$ acts as a maturation-inducing steroid in the blackfin flounder.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.37 no.3
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• pp.192-196
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• 2004

The in vitro effect of bisphenol A (BPA) on ovarian steroidogenesis of the longchin goby (Chasmichthys dolichognathus) was investigated. Oocytes taken during the maturing phase (vitellogenic, fully vitellogenic or germinal vesicle breakdown stage) were incubated with BPA (100 ng/mL) in the presence of exogenous precursor $^{3}H-17\alpha\;hydroxyprogesterone\;(^{3}H-17\alphaOHP).$ Steroids were extracted from the media and the isolated oocytes, and the extracts were separated and identified by thin layer chromatography and gas chromatography-mass spectrometry. The identities of the major metabolites were progestogens $[17{\alpha}-hydroxy,20{\alpha}-dihydroprogesterone\;(17{\alpha}20{\alpha}OHP)\;and\;17{\alpha}-hydrxy,20{\beta}-dihydroprogesterone\;(17{\alpha}20{\beta}OHP),$ androgens [androstenedione (A4) and testosterone (T)] and estrogens [estrone $(E_1)\;and\;estradiol-17{\beta}(E_2)].$ BPA treatment inhibited production of estrogens in all the maturing phases and progestogens in the germinal vesicle migrating stage. Percentage yield of estrogens was decreased with increased yield of androgens. In conclusion, BPA had an inhibitory effect on the conversion of $^3H-17\alphaOHP$ to estrogens and progestogens. These results demonstrate that BPA can act either estrogenic or anti-estrogenic effects.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.28 no.4
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• pp.469-474
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• 1995

The effects of sperific inhibitors of $3\beta-hydroxy-\Delta^5-steroid\;dehydrogenase$ $(3\beta-HSD;\;an\;enzyme\;catalyzing\;conversion\;of \Delta^5\;steroids\;to\;\Delta^4 steroids),$ cyanoketone and trilostane, on $^3H-pregnenolone$ metabolism in isolated ovarian thecal layers have been investigated in vitro. At all doses of cyanoketone $(10^{-5}\;and\;10^{-4}\;M)$ and trilostane $(10^{-5}\;and\;10^{-4}\;M)$ $(3\beta-HSD$ enzyme activity that transforms pregnenolone to $17\alpha-hydroxyprogesterone$ was inhibited in the thecal layers. Trilostane appeared to be more efficient than cyanoketone. Trilostane at doses of $10^{-8},\;10^{-7},\;10^{-6},\;and\;10^{-5},\;M/ml$ caused a dose-response inhibition of $\Delta^4$ steroids accumulation in the medium from pregnenolone, but not completely blocked the conversion of $\Delta^5\;to\;\Delta^4$ steroids.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.40 no.3
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• pp.153-159
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• 2007

We studied oocyte steroidogenesis in relation to oocyte development in the greenling, Hexagrammos otakii, a marine multiple spawner. Vitellogenic and mature oocytes were incubated in vitro in the presence or absence of $[^3H]-17\;{\alpha}-hydroxyprogesterone$ as a precursor. The major metabolites were androgens [androstenedione $(A)_4)$ and testosterone (T)] and estrogens [$17\;{\beta}-estradiol\;(E_2)$ and estrone ($E_1$)] in vitellogenic oocytes. The metabolic rate of T was lower in 1.08 to 12-mm oocytes, while that of $E_2$ increased with oocyte size. The endogenous productions of T, $E_2$ and 17 ${\alpha}-hydroxy$, 20 ${\beta}-dihydroprogesterone\;(17{\alpha}20{\beta}OHP)$ were quantified using a radioimmunoassay in the non-precursor group. The endogenous levels of T and $E_2$ were highest in 1.08 to 12-mm oocytes and $17{\alpha}20{\beta}OHP$ was produced only in 1.90 to 95-mm oocytes. The relationship between oocyte size and steroidogenesis showed that 1.08 to 12-mm oocytes are full vitellogenic following induction of the maturation process. Moreover, $17{\alpha}20{\beta}OHP$ acts as a maturation inducing hormone in H. otakii.

• Animal cells and systems
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• v.10 no.4
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• pp.203-209
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• 2006

Azoles are widely used antifungal agents, which inhibit the biosynthesis of fungal cell-membrane ergosterol. In this study, using an amphibian follicle culture system, the effects of azoles on follicular steroidogenesis in frogs were examined. Itraconazole (ICZ), clotrimazole (CTZ) and ketoconazole (KCZ) suppressed pregnenolone ($P_5$) production by the follicles ($ED_{50};\;0.04_{\mu}M,\;0.33_{\mu} M,\;and\;0.91_{\mu}M$, respectively) in response to frog pituitary homogenates (FPH). However, fluconazole (FCZ), miconazole (MCZ) and econazole (ECZ) were not effective in the suppression of $P_5$ production. Not all the azoles examined suppressed the conversion of exogenous $P_5$ to progesterone ($P_4$) (by $3{\beta}$- HSD) or $P_4$ to $17{\alpha}$-hydroxyprogesterone ($17{\alpha}$-OHP) (by $17{\alpha}$-hydroxylase), or androstenedione (AD) to testosterone (T) (by $17{\beta}$-HSD). In contrast, CTZ, MCZ and ECZ in medium partially suppressed the conversion of $17{\alpha}$-OHP to AD (by C17-20 lyase) ($ED_{50};\;0.25{\mu} M,\;4.5{\mu}M,\;and\;0.7{mu}M$, respectively) and CTZ, KCZ, ECZ and MCZ strongly suppressed the conversion of exogenous T to estradiol ($E_2$) (by aromatase) ($ED_{50};\;0.02{\mu}M,\;8{\mu}M,\;0.07{\mu}M,\;0.8{\mu}M$, respectively). These results demonstrated that some azole agents strongly suppress amphibian follicular steroidogenesis and particularly, P450scc and aromatase are more sensitive to azoles than other steroidogenic enzymes.

• Perinatology
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• v.29 no.4
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• pp.170-174
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• 2018

Objective: Progesterone is used to prevent recurrent preterm delivery, however the molecular mechanisms of its effect are incompletely understood. The objective of this study was to determine the effect of progesterone on tumor necrosis factor $(TNF)-{\alpha}$-induced matrix metalloproteinase (MMP)-9 activity in human choriodecidual (CD) membranes. Methods: We collected CD membranes from women with uncomplicated term pregnancies who were scheduled for elective cesarean delivery (n=10). CD membranes ($1{\times}1cm$) were incubated in tissue culture media at $37^{\circ}C$. We pre-treated the CD membranes with progesterone (P4), $17{\alpha}$-hydroxyprogesterone caproate (17P), promegestone (R5020), or vehicle (ethanol) for 24 hours. The CD membranes were subsequently treated with $TNF-{\alpha}$ (with continued progesterone treatment) for 48 hours, then media was harvested for measuring MMP-9 activity by zymography and total protein was isolated from CD membrane tissues for MMP-9 expression by western blot analysis. Results: P4, 17P, and R5020 significantly reduced $TNF-{\alpha}$-induced MMP-9 activity in fetal membrane tissue samples (P=0.0078, P=0.0156, and P=0.0391, respectively) by zymography. Western blot analysis also showed decreased expression of MMP-9 in progesterone pretreated groups (P=0.0313). Conclusion: Progesterone reduces $TNF-{\alpha}$-induced MMP-9 activity in human CD membranes. These findings may provide further support for the role of progesterone in preventing preterm birth.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.32 no.5
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• pp.624-628
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• 1999

Annual plasma levels of vitellogenin and sex steroids were investigated in relation to the gonadal development for understanding the endocrine control of reproduction in spotted flounder, Verasper variegatus. The plasma vitellogenin level was highest, 6.36 mg/ml, in November when vitellogenesis was most active. The level, thereafter, decreased to 3.81 mg/ml in December with the initiation of spawning. On the other hand, estradiol-17 $\beta$ was highest, 2.7 ng/ml, in December, and rapidly decreased in January when spawning occurred. The decreased level of estradiol-17$\beta$, around 0.2 ng/ml, remained unchanged until May. The profiles of plasma testosterone were similar to those of estradiol-17$\beta$ in the fish, The plasma 17 $\alpha$-hydroxyprogesterone level was relatively low throughout the spawning period, but increased slightly with the initiation of ovarian development, In males, the plasma testosterone and 11-ketotestosterone were highest in December when spermiation actively proceeded, but rapidly decreased during the spawning period (January).

• Fisheries and Aquatic Sciences
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• v.12 no.4
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• pp.293-298
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• 2009

Several studies have reported that nonylphenol (NP) and 2,2', 4,6,6'-pentachlorobiphenyl (PCB104) exhibit estrogenic activity. To investigate the estrogenic potency of NP and PCB104 during oocyte maturation, fully vitellogenic oocytes (0.76 mm diameter in average) of yellow fin goby, Acanthogobius flavimanus, were exposed in vitro to these chemicals at different concentrations (0.1, 1, 10, 100, and 1,000 ng/mL) with the exogenous precursor $17\alpha$-hydroxyprogesterone ($17{\alpha}OHP$) 50 ng/mL in the presence or absence of human chorionic gonadotropin (HCG). The production of testosterone (T), estradiol-$17\beta$ (E2), and $17\alpha,20\beta$-dihydroxy-4-pregnen-3-one ($17\alpha20{\beta}OHP$) in response to NP or PCB104 were measured by radioimmunoassay. Steroid levels were also expressed as E2/T and E2/$17\alpha20{\beta}OHP$ ratios. In the absence of HCG, no significant differences in either NP or PCB104 treatment groups were observed. In the presence of HCG, NP treatment did not show significant differences in the production of T, E2, and $17\alpha20{\beta}OHP$ at any concentrations tested, but E2/T ratios were decreased at concentrations of 0.1, 1, 10, and 1,000 ng/mL compared with the control group. PCB104 decreased E2 production at concentrations of 0.1, 10, and 1000 ng/mL, but did not show significant differences in the production of T and $17\alpha20{\beta}OHP$ at any concentration tested. While E2/T ratios were decreased at PCB104 concentrations of 0.1, 1, 10, and 1,000 ng/mL, E2/$17\alpha20{\beta}OHP$ ratios were also decreased at 0.1, 10, and 1,000 ng/mL compared with the control. Results indicate that both NP and PCB104 appeared to have antiestrogenic effects during this phase.

• Development and Reproduction
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• v.16 no.1
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• pp.39-45
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• 2012

To verify the sex steroids which are involved in oocyte maturation of the blacktip grouper, $Epinephelus$ $fasciatus$, we incubated vitellogenic oocytes (0.41 and 0.50 mm in average diameter) in the presence of exogenous steroid precursor ($[^3H]17{\alpha}$-hydroxyprogesterone). Steroids were extracted, separated and identified by thin layer chromatography. The major metabolites produced were androstenedione, estradiol-$17{\beta}$, estrone and progestogens. Progestogen metabolites in the oocytes of 0.50 mm were more abundant than those of 0.41 mm. Also, we investigated the $in$ $vitro$ effects of human chorionic gonadotropin (HCG; 5, 50 and 500 $IU/m{\ell}$), $17{\alpha},20{\beta}$-dihydroxy-4-pregnen-3-one ($17{\alpha}20{\beta}P$) and $17{\alpha},20{\beta}$-trihydroxy-4-pregnen-3-one ($17{\alpha}20{\beta}21P$; 5, 50 and 500 $ng/m{\ell}$, respectively) on oocyte maturation. In the oocytes of 0.41 mm, treatment with 50 IU HCG stimulated GVBD ($55.30{\pm}1.20%$) compared with controls ($32.41{\pm}3.13%$, $p$<0.05). In the oocytes of 0.50 mm, treatment of $17{\alpha}20{\beta}P$ (50 and 500 $ng/m{\ell}$) stimulated GVBD ($50.13{\pm}2.52$ and $51.77{\pm}5.91%$, respectively) compared with controls ($36.81{\pm}2.89%$, $p$<0.05). Treatment with 500 IU HCG also stimulated GVBD ($49.59{\pm}5.15%$) compared with controls ($p$<0.05). Taken together, these results suggested that both HCG and $17{\alpha}20{\beta}P$ were effective on in vitro oocyte maturation and $17{\alpha}20{\beta}P$ may act as a maturation inducing hormone in blacktip grouper.