• The Korean Journal of Zoology
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• v.32 no.3
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• pp.281-289
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• 1989

The present study was disigned to determine the concentration of bioavailable steroid hormones in the atretic follicular fluid (FF). The concentradons of progesterone (P), testosterone (T), estradiol (E), androstenedione (A), and 5-$\alpha$ dihydrotestosterone (DIlT) were determined by the established methods of luminescent immunoassay (LIA) or radioimmunoassay (RIA). Concentrations of T, A and Diff in human FF from smail (< 6 mm). medium (8-15 mm), and large (> 15 mm) atretic follicles were significandy higher than those of normal ones (p < 0.01). However, the levels of T, A and DHT in smail atretic foflicle were significandy lower than those found in normal one. The concentrations of P in atretic FF from porcine small (< 3 mm), medium (4-6 mm), and large (> 7 mm) follicles were not different from that of normal ones. However, the concentration of E in atretic forncles of each group was significantly lower than that of normal group (p < 0.001 in each group). On the other hand, the percentages of bioavailable T (BI) in human FF were significandy (p <0.001) higher than those in normal groups. The BT in normal or atretic FF was more than 90 % of total T. The present result demonstrates that the bioavailable androgen, but not E levels in atretic follicles is higher than that of normal one, and that the atretic mechanism might be dependent on the ovarian forncle size in the developmental stage and on the animal model system. Moreover, the present study suggests that the steroids found in the FF are the bioavailable forms and the concentration of BT in FF could be used as one of the valuable criteria classifying the ovarian atretic follicle.

• Journal of Animal Science and Technology
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• v.44 no.6
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• pp.711-718
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• 2002

In order to investigate the effect of feeding browses on the incidence of abortion in pregnant Korean native female goats, 20 pregnant does(3${\sim}$4 month) were used as experimental animal and they grouped 4 treatments with 5 heads according to experimental diets. Each group was fed pine browse, pine browse silage, fermented pine browse, or oak browse silage. Browses intake a day was 0.36kg for oak browse silage, 0.28kg for pine browse, 0.24kg for pine browse silage, and 0.14kg for fermented pine browse. Salmonella and Fungi were not found in pine browses but they were found in oak browse silage in the amount of 7.93${\times}$$10^3$ and 11.1${\times}$$10^1$ cfu/g, respectively. E. Coliwas found 11.67${\times}$$10^1$cfu/g in oak browse silage. The incidence of abortion was 60% for fermented pine browse, 40% for pine browse silage, and 20% for pine browse feeding. Abortion did not occur by feeding oak browse silage. Progesterone concentration was similar each other regardless of normal delivery or abortion at delivery day but the concentration of estradiol was higher for normal delivery, concentration of cortisol was decreased until the delivery day in normal but increased in abortion does. The results were suggested.

• Journal of the Korean Society of Food Science and Nutrition
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• v.36 no.7
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• pp.825-831
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• 2007

Anti-proliferation effects of decursin from Angelica gigas Nakai were investigated in the MCF-7 cells treated with environmental hormones. The proliferation was decreased in a dose-dependent manner at the concentration over 20 ${\mu}g/mL$ in the MCF-7 cells treated with decursin of various concentrations. The environmental hormones such as $17{\beta}$-estradiol and bisphenol increased the growth of MCF-7 cells in the charcoal-treated FBS (cFBS) medium and the proliferation was the highest at 0.1 ${\mu}M$ among the tested hormone concentration. Decursin was predicted to inhibit the proliferation in a dose-dependent fashion at tested concentrations (1, 3, 10 or 30 ${\mu}g/mL$) in the MCF-7 cells added environmental hormones; however, the survival rate of the cells was lower than that of control cells that were not treated with decursin at 30 ${\mu}g/mL$ concentration. The chromatin condensation and apoptotic body were examined in the decursin treated cells cultured with the cFBS medium added environmental hormones. These results suggest that decursin decreased the proliferation through apoptosis in the MCF-7 cells added environmental hormones.

• Environmental Mutagens and Carcinogens
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• v.24 no.4
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• pp.198-205
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• 2004

Cytochrome P4501B1(CYP1B1) is known to be inducible by xenobiotic compounds such as policyclic aromatic hydrocarbon(PAH) and dioxins such as 2,3,7,8-tetrachloro-dibenzo-p-dioxin(TCDD). And these induction of CYP1B1 is also regulated by many categories of chemicals. In order to investigate the effects of several chemicals on CYP1B1 gene expression in Hepa-I and MCF-7 cells, 5' flanking DNA of human CYP1B1 was cloned into pGL3 basic vector containing luciferase gene, and then transfected into these cells. After treatment of chemicals, the luciferase activity was measured. CYP1B1 enzyme metabolize PAHs and estradiol. CYP1B1 metabolize estradiol to 4-hydrozyestradiol that is considered as carcinogenic metabolite. Recent industrialized industrialized society, human has been widely been exposed to widespread environmental contaminants such as PAHs(polycyclic aromatic hydrocarbon) that are originated from the imcomplete combustion of hydrocarbons. PAHs are known to be ligands of the AhR(aryl hydrocarbon receptor). Induction of cytochrome P4501B1(CYP1B1) in cell culture is widely used as a biomarker for PAHs. Therefore we have studied the effect of PAHs in the human breast cancer cells MCF-7 to evaluate bioactivity of PAHs. We have used the United State of America EPA selected 13 different PAHs, PAHs mixtures and extracts from environmental samples to evaluate the bioassay system. We examined effects of PAHs on the CYP1B1-luciferase reporter gene and CYP1B1 mRNA level. Benzo(k)fluoranthene and dibenzo(a, h)anthracene showed strong response to CYP1B1 promoter activity stimulation, and also CYP1B1 mRNAs increase in MCF-7 cells in a concentration-dependent manner. RT-PCR analysis indicated that PAHs significantly up-regulate the level of CYP1B1 mRNA. Some flavonoids such as genistein, daidzein, chrysin, naringenin and morin were also investigeted. These flavonoids decreased B(k)F infuced luciferase activity at low concentration. But, these flavonoids exhibited stimulatory effect at high concentration.

• Asian Pacific Journal of Cancer Prevention
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• v.17 no.4
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• pp.1851-1855
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• 2016

Tamoxifen is a pharmacological estrogen inhibitor that binds to the estrogen receptor (ER) in breast cells. However, it shows an estrogenic effect in other organs, which causes adverse drug reactions (ADRs). The sulfotransferase 1A1 (SULT1A1) enzyme encoded by the SULT1A1 gene is involved in estrogen metabolism. Previous research has suggested that the SULT1A1 copy number is linked with the plasma estradiol (E2) concentration. Here, a total of 34 premenopausal breast cancer patients, selected from the Thai Tamoxifen (TTAM) Project, were screened for their SULT1A1 copy number, plasma E2 concentration and ADRs. The mean age was $44.3{\pm}11.1years$, and they were subtyped as ER+/progesterone receptor (PR)+ (28 patients), ER+/PR- (5 patients) and ER-/PR- (1 patient). Three patients reported ADRs, which were irregular menstruation (2 patients) and vaginal discharge (1 patient). Most (33) patients had two SULT1A1 copies, with one patient having three copies. The median plasma E2 concentration was 1,575.6 (IQR 865.4) pg/ml. Patients with ADRs had significantly higher plasma E2 concentrations than those patients without ADRs (p = 0.014). The plasma E2 concentration was numerically higher in the patient with three SULT1A1 copies, but this lacked statistical significance.

• 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.

• Toxicological Research
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• v.16 no.2
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• pp.109-116
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• 2000

The purposes of our study were to optimize the conditions of the screening and testing methods for endocrine disruptors, to characterize these assays using several compounds with well-defined endocrine activity, and to compare the sensitivity between these assays currently undergoing validation. Two in vitro test systems, MCF-7 cells proliferation (E-screen assay) and competitive binding to estrogen receptors (ER) were selected to evaluate the estrogenic effects. 17$\beta$-Estradiol (E2) and diethylstilbestrol (DES) were used as a positive control in vitro test. Also, E2 and ethinyl estradiol (EE) were used as a positive control in vivo uterotrophic assay. In in vitro test, E2 and DES showed a strong estrogenic response at concentration of 1.0 nM. In uterotrophic assay, E2 (0.3 $\mu\textrm{g}$/kg) and EE (0.3 $\mu\textrm{g}$/kg) produced a significant increase in uterus and vagina weight in both immature and ovariectomized rats. Although we did not com-pared the specificity between in vivo and in vitro assays, these assay systems may serve as a good tool for endocrine disruptors screening methods. Our data indicate that these assay systems exhibit some difference in their sensitivity to the same estrogenic compounds. Therefore, as a first rapid screening assay for estrogenic activity qf unknown chemicals, at least two assay systems should probably be carried out with a view of high sensitivity and standardization conditions. Also, a careful validation tests are necessary to obtain a reasonable degree of reproducibility.

• Asian-Australasian Journal of Animal Sciences
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• v.24 no.11
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• pp.1529-1540
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• 2011

Silencing of a specific gene using RNAi (RNA interference) is a valuable tool for functional analysis of a target gene. However, information on RNAi for analysis of gene function in farm animals is relatively nil. In the present study, we have validated the interfering effects of siRNA (small interfering RNA) using both quantitative and qualitative gene silencing in buffalo granulosa cells. Qualitative gene knockdown was validated using a fluorescent vector, enhanced green fluorescence protein (EGFP) and fluorescently labeled siRNA (Cy3) duplex. While quantitatively, siRNA targeted against the luciferase and CYP19 mRNA was used to validate the technique. CYP19 gene, a candidate fertility gene, was selected as a model to demonstrate the technique optimization. However, to sustain the expression of CYP19 gene in culture conditions using serum is difficult because granulosa cells have the tendency to luteinize in presence of serum. Therefore, serum free culture conditions were optimized for transfection and were found to be more suitable for the maintenance of CYP19 gene transcripts in comparison to culture conditions with serum. Decline in fluorescence intensity of green fluorescent protein (EGFP) was observed following co-transfection with plasmid generating siRNA targeted against EGFP gene. Quantitative decrease in luminescence was seen when co-transfected with siRNA against the luciferase gene. A significant suppressive effect on the mRNA levels of CYP19 gene at 100 nM siRNA concentration was observed. Also, measurement of estradiol levels using ELISA (enzyme-linked immunosorbent assay) showed a significant decline in comparison to control. In conclusion, the present study validated gene silencing using RNAi in cultured buffalo granulosa cells which can be used as an effective tool for functional analysis of target genes.

• Asian-Australasian Journal of Animal Sciences
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• v.14 no.7
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• pp.922-928
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• 2001

There exists considerable evidence that steroid hormones are involved in the regulation of ovulation rate and oviductal development in poultry. However, the effect of steroid hormones on egg productivity of Korean Native Ogol Chicken (KNOC) has yet to be studied. Therefore, this study was performed to relate the expression of steroid hormones, especially progesterone ($P_4$) and estradiol ($E_2$), with egg productivity during the laying period. Egg production and egg weight of 70 KNOC were recorded from 20 to 60wk. Blood was taken every 10 wk and serum $P_4$ and $E_2$ were measured by radioimmunoassay. Based on egg productivity and steroid hormones levels up to 60 wk, chickens were divided into two groups, high and low. Compared to the low egg production group, a significantly higher expression of $P_4$ at 30 wk was detected in the high group. Moreover, egg production in the high $P_4$ group significantly differed from that in the low group at 30 wk. On the other hand, a Significant difference (p<0.05) in $E_2$ expression was found between high and low egg weight groups at 30 wk. Although a significant difference in egg weight between two groups by $E_2$ was not detected, the high $E_2$ group showed a higher level of egg weight than the low $E_2$ group except for 25 wk. In the comparison of ovary weight and small yellow follicle number, the group with high egg productivity and steroid concentration showed greater levels than the low group. Taken together, the results indicate that $P_4$ is related to egg productivity whereas expression of $E_2$ is associated with egg weight in KNOC.

• Journal of Veterinary Clinics
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• v.13 no.2
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• pp.114-122
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• 1996

The aim of this study was to assess the precision of the estimates of the time of estrous cycle, optimal breeding and ovulation derived by vaginal cytology. The thirteen Korea Jin-do dogs were examined the vaginal cytology, plasma estradiol-17$$\beta $ and progesterone assay during the estrous cycle. Day 0 was the day of the first male acceptance. The main change of vaginal cytology during the estrous cycle was the high proportion of anuclear cell and erythrocyte in proestrus, superficial cell, anuclear cell and erythrocyte in estrus, parabasal cell, large intermediate cell and leukocytes in diestrus, and parabasal cell and small intermediate cell in anestrus, respectively. These data indicated that vaginal cytology was reliable method for estimating estrous cycle in Korea Jin-do dogs. In the cell indices during estrus the maximum eosinoghilic index was $92.0{\pm}$2.6 (Mean{\pm} SEM$)% at Day 2 and the maximum cornification indez was $96.0{\pm}1.3%$ at Day 2, respectively. The eosinothilic indez and cornification indez of up to 70% were found at Day -1 to Day 5 and Day -6 to Day 8, and up to 80% at Day 1 to Day 4 and Day -4 to Day 6, respectively. From these data it was presumed that eosinophilic index was more reliable index for monitoring optimal breeding time than cornification indexm because eosinophilic index peak period was shorter than cornification indeX peak period and Day 2 was the day of ovulation. Therefore, optimal breeding time was the eosinophilic index peak period, more than 80% of eosinoghilic index. The $estradiol-17{\beta}$ peak, with 3 days delayed when progesterone concentration was $4.5{\pm}0.5 ng/ml$. These data estimated that the ovulation time was the day of eosinophilic index peak, Day 2. breeding time and pvulation time in Korea Jin-do dogs.