• Development and Reproduction
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• v.22 no.2
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• pp.175-182
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• 2018

In order to examine the effects of four different light spectra (white, red, green, and blue) on the oocyte maturation, the change of reproductive parameters, via brain-pituitary-gonad (BPG) axis in grass puffer, were investigated. After exposure four different light spectra for 7 weeks, the abundance of gonadotropin-releasing hormone (GnRH) mRNA which is a type of seabream (sbGnRH) and two different subunit of gonadotropin hormones mRNAs, follicle-stimulating hormone ($fsh{\beta}$) mRNA and luteinizing hormone ($lh{\beta}$) mRNA, were analyzed in the brain and pituitary. Histological analysis showed that the mature oocyte ratio in the green spectrum was higher than other light spectra-exposed groups. Gonadosomatic index (GSI) and oocyte developmental stage were also investigated in the gonad based on histological observations. GSI value with the presence of yolk stage oocytes was significantly increased in the green spectrum-exposed group when compared to that of the other light-exposed groups (white, red, and blue) (p<0.05). The abundances of sbGnRH mRNA and $fsh{\beta}$ mRNA in the green spectrum-exposed group were also significant higher than those of the other light spectra-exposed groups (p<0.05). These results indicate that the maturation of oocyte in grass puffer can be accelerated by exposure to the spectrum of green. To better understand the molecular mechanism for the maturation of oocyte in grass puffer, further study examining the relationship between oocyte development and its related genes is required.

• Proceedings of the KSAR Conference
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• 2000.10a
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• pp.10-12
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• 2000

Members of the glycoprotein family, which includes CG, LH, FSH and TSH, comprise two noncovalently linked $\alpha$- and $\beta$-subunits. Equine chorionic gonadotropin (eCG), known as PMSG, has a number of interesting and unique characteristics since it appears to be a single molecule that possesses both LH- and FSH-like activities in other species than the horse. This dual activity of eCG in heterologous species is of fundamental interest to the study of the structure-function relationships of gonadotropins and their receptors. CG and LH $\beta$ genes are different in primates. In horse, however, a single gene encodes both eCG and eLH $\beta$-subunits. The subunit mRNA levels seem to be independently regulated and their imbalance may account for differences in the quantities of $\alpha$ - and $\beta$ -subunits in the placenta and pituitary. The dual activities of eCG could be separated by removal of the N-linked oligosaccharide on the $\alpha$-subunit Asn 56 or CTP-associated O-linked oligosaccharides. The tethered-eCG was. efficiently secreted and showed similar LH-like activity to the dimeric eCG. Interestingly, the FSH-like activity of the tethered-eCG was increased markedly in comparison with the native and wild type eCG. These results also suggest that this molecular can implay particular models of FSH-like activity not LH-like activity in the eCG/indicate that the constructs of tethered molecule will be useful in the study of mutants that affect subunit association and/or secretion. A single-chain analog can also be constructed to include additional hormone-specific bioactive generating potentially efficacious compounds that have only FSH-like activity. The LH/CG receptor (LH/CGR), a membrane glycoprotein that is present on testicular Leydig cells and ovarian theca, granulosa, luteal, and interstitial cells, plays a pivotal role in the regulation of gonadal development and function in males as well as in nonpregnant and pregnant females. The LH/CGR is a member of the family of G protein-coupled receptors and its structure is predicted to consist of a large extracellular domain connected to a bundle of seven membrane-spanning a-helices. The LH/CGR phosphorylation can be induced with a phorbol ester, but not with a calcium ionophore. The truncated form of LHR also was down-regulated normally in response to hCG stimulation. In contrast, the cell lines expressing LHR-t63I or LHR-628, the two phosphorylation-negative receptor mutant, showed a delay in the early phase of hCG-induced desensitization, a complete loss of PMA-induced desensitization, and an increase in the rate of hCG-induced receptor down-regulation. These results clearly show that residues 632-653 in the C-terminal tail of the LHR are involved in PMA-induced desensitization, hCG-induced desensitization, and hCG-induced down-regulation. Recently, constitutively activating mutations of the receptor have been identified that are associated with familial male-precocious puberty. Cells expressing LHR-D556Y bind hCG with normal affinity, exhibit a 25-fold increase in basal cAMP and respond to hCG with a normal increase in cAMP accumulation. This mutation enhances the internalization of the free and agonist-occupied receptors ~2- and ~17-fold, respectively. We conclude that the state of activation of the LHR can modulate its basal and/or agonist-stimulated internalization. Since the internalization of hCG is involved in the termination of hCG actions, we suggest that the lack of responsiveness detected in cells expressing LHR-L435R is due to the fast rate of internalization of the bound hCG. This statement is supported by the finding that hCG responsiveness is restored when the cells are lysed and signal transduction is measured in a subcellular fraction (membranes) that cannot internalize the bound hormone.

• Korean Journal of Animal Reproduction
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• v.24 no.4
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• pp.357-364
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• 2000

Members of the glycoprotein family, which includes CG, LH, FSH and TSH, comprise two noncovalently linked $\alpha$- and $\beta$-subunits. Equine chorionic gonadotropin (eCG), known as PMSG, has a number of interesting and unique characteristics since it appears to be a single molecule that possesses both LH- and FSH-like activities in other species than the horse. This dual activity of eCG in heterologous species is of fundamental interest to the study of the structure-function relationships of gonadotropins and their receptors. CG and LH $\beta$ genes are different in primates. In horse, however, a single gene encodes both eCG and eLH $\beta$ -subunits. The subunit mRNA levels seem to be independently regulated and their imbalance may account for differences in the quantities of $\alpha$ - and $\beta$-subunits in the placenta and pituitary. The dual activities of eCG could be separated by removal of the N-linked oligosaccharide on the $\alpha$-subunit Asn 56 or CTP-associated O-linked oligosaccharides. The tethered-eCG was efficiently secreted and showed similar LH-like activity to the dimeric eCG. Interestingly, the FSH-like activity of the tethered-eCG was increased markedly in comparison with the native and wild type eCG. These results also suggest that this molecular can implay particular models of FSH-like activity not LH-like activity in the eCG/indicate that the constructs of tethered molecule will be useful in the study of mutants that affect subunit association and/or secretion. A single-chain analog can also be constructed to include additional hormone-specific bioactive generating potentially efficacious compounds that have only FSH-like activity. The LH/CG receptor (LH/CGR), a membrane glycoprotein that is present on testicular Leydig cells and ovarian theca, granulosa, luteal, and interstitial cells, plays a pivotal role in the regulation of gonadal development and function in males as well as in nonpregnant and pregnant females. The LH/CGR is a member of the family of G protein-coupled receptors and its structure is predicted to of a large extracellular domain connected to a bundle of seven membrane-spanning a-helices. The LH/CGR phosphorylation can be induced with a phorbol ester, but not with a calcium ionophore. The truncated form of LHR also was down-regulated normally in response to hCG stimulation. In contrast, the cell lines expressing LHR-t631 or LHR-628, the two phosphorylation-negative receptor mutant, showed a delay in the early phase of hCG-induced desensitization, a complete loss of PMA-induced desensitization, and an increase in the rate of hCG-induced receptor down-regulation. These results clearly show that residues 632~653 in the C-terminal tail of the LHR are involved in PMA-induced desensitization, hCG-induced desensitization, and hCG-induced down-regulation. Recently, constitutively activating mutations of the receptor have been identified that are associated with familial male-precocious puberty. Cells expressing LHR-D556Y bind hCG with normal affinity, exhibit a 25-fold increase in basal cAMP and respond to hCG with a normal increase in cAMP accumulation. This mutation enhances the internalization of the free and agoinst-occupied receptors ~2- and ~17- fold, respectively. We conclude that the state of activation of the LHR can modulate its basal and/or agonist-stimulated internalization. Since the internalization of hCG is involved in the termination of hCG actions, we suggest that the lack of responsiveness detected in cells expressing LHR-L435R is due to the fast rate of internalization of the bound hCG. This statement is supported by the finding that hCG responsiveness is restored when the cells are lysed and signal transduction is measured in a subcellular fraction (membranes) that cannot internalize the bound hormone.

• Korean Journal of Agricultural Science
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• v.12 no.1
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• pp.55-61
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• 1985

This study was carried out to investigate reproductive traits and sex hormone levels by growth in Korean native goats. Serum levels of FSH, LH, prolactin, estradiol-$17{\beta}$ and progesterone were determined every 15 days from the 70 days of age to 190 days by radioimmunoassay. The age and weight at sexual maturity were 183.6 days and 14.3 kg; the length of estrous cycle and estrus period were 20.3 days and 36.7 hours, respectively. The gestation period, litter size and body weight at birth were 148.4 days, 1.4 head and 1.8 kg, respectively. The levels of serum LH were highest with 3.93 mIU/ml at 70 days of age, thereafter decreased gradually to 1.21 mIU/ml at 190 days of age. The concentrations of FSH in serum were below 1.25 mIU/ml throughout the experimental period. The levels of prolactin were lowest with 3.09ng/ml at 85 days of age and highest with 4.65 ng/ml at 175 days of age. The serum levels of estradiol-$17{\beta}$ were increased with age, thus highest with 7.95 pg/ml at 190 days of age. The serum progesterone concentrations were maintained low levels (below 1.0 ng/ml) throughout the experimental period.

• Korean Journal of Animal Reproduction
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• v.14 no.4
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• pp.263-271
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• 1990

This study was carried out to investigate the factors affecting maturation in vitro of follicular oocytes in Korean Native Cattle. The bovine ovaries were obtained at a slaughter house and the follicular oocytes were recovered by aspirating the follicular fluid from the visible follicles of 3~6mm. The bovine oocytes were matured in vitro in TCM-199 containing FCS and hormones. The effects of TCM-199 salt type, number of oocytes per drop, incubation time and co-culture with granulosa cells on maturation of oocytes, were investigated. The results obtained are summarized as follows. 1. The maturation rates of follicular oocytes cultured for 22, 25 and 28 hours in Hank's TCM-199 or Earle's TCM-199 were 59.3, 59.6, 80% and 80.0, 90.0, 93.7%, respectively. The maturation rate of follicular oocytes in Earle's TCM-199 was faster and higher than in Hank's TCM-199(P<0.05). 2. The maturation rates of oocytes were 54.5, 62.5 and 62.0% when cultured the oocytes number 10, 20 and 40 per 200${mu}ell$ drop for 18 hours. 3. The maturation of follicular oocytes in the Korean Native Cattle was induced at 14 hours culture, by giving the maturation rate of 90.0% after 20 hours. 4. The maturation rates were 63.3% and 66.7%, respectigely when the oocytes were co-cultured with granulosa cells from medium-size follicles used immediately after recovery in the presence or absence of hormones added(0.02AU/ml FSH, 10$\mu\textrm{g}$/ml LH and 1$\mu\textrm{g}$/ml estradiol 17-$\beta$). When the oocytes were co-cultured with granulosa cells from medium-size follicles cultured for 3 days, the maturation rates were 20.8% and 76.2%, respectively(P<0.05). 5. The maturation rate were 88.0% and 70.0%, respectively when the oocytes were co-cultured with granulosa cells from large-size follicles used immediately after recovery in the presence or absence of hormones added(0.02AU/ml FSH, 10$\mu\textrm{g}$/ml LH and 1$\mu\textrm{g}$/ml estradiol 17-$\beta$)(P<0.05). When the hormones added(0.02AU/ml FSH, 10$\mu\textrm{g}$/ml LH and 1$\mu\textrm{g}$/ml estradiol 17-$\beta$)(P<0.05). When the oocytes were co-cultred with granulosa cells from large-size follicles cultured for 3 days, the maturation rates were 41.2% and 73.3%, respectively(P<0.05).

• Reproductive and Developmental Biology
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• v.34 no.1
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• pp.47-53
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• 2010

Glycoprotein hormones have a common $\alpha$-subunit that is involved in the signaling pathway together with G protein, adenylcyclase and cAMP induction; however, it is an unclear how this common structure is related to hormonal action. To determine the biological functions of the COOH-terminal amino acids in the $\alpha$-subunit of these glycoprotein hormones, a tethered-molecule was constructed by fusing the $NH_2$-terminus of the $\alpha$-subunit to the COOH-terminus of the $\beta$-subunit of equine chorionic gonadotropin (eCG). The following deletion mutants were created by PCR; Ile was inserted at position 96 to form ${\Delta}96$, Lys was substituted at position 95 to form ${\Delta}95$, His was inserted at position 93 to form ${\Delta}93$ and Tyr was substituted at position 87 to form ${\Delta}87$. Each mutant was transfected into CHO-K1 cells. Tethered-wt eCG, and ${\Delta}96$, ${\Delta}95$, and ${\Delta}93$ mutants were efficiently secreted into the medium but the ${\Delta}87$ mutant was not secreted. Interestingly, the RT-PCR, real-time PCR, and northern blot analyses confirmed that the RNA was transcribed in the ${\Delta}87$ mutant. However, the ${\Delta}87$ mutant protein was not detected in the medium or the intracellular fraction of the cell lysates. The LH- and FSH-like activities of the recombinant proteins were assayed in terms of cAMP production using rat LH/CG and rat FSH receptors. The metabolic clearance rate (MCR) was determined by injecting rec-eCG (2 IU) into the tail vein. The ${\Delta}95$ and ${\Delta}93$ mutants were completely inactive in both the LH- and FSH-like activity assays. The ${\Delta}96$ mutant showed slight activity in the LH-like activity assay. In comparison to the wild type, the activity of the ${\Delta}96$ mutant in the FSH-like activity assay was the highest among all the mutants. The MCR assay in which rec-eCG was injected showed a peak at 10 min in all the treatment groups, which disappeared 4 h after injection. These results imply a direct interaction between the receptor and the COOH-terminal region of the a-subunit. The data also reveal a significant difference in the mechanism by which the eCG hormone interacts with the rLH and rFSH receptors. The COOH-terminal region of the $\alpha$-subunit is very important for the secretion and functioning of this hormone.

• Clinical and Experimental Reproductive Medicine
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• v.31 no.2
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• pp.111-117
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• 2004

Objectives: To investigate the effect of LH receptor in folliculogenesis, we confirm the expression level of LH receptor (LH-R) mRNA in human granulosa cells (GCs) and its expression levels were analyzed by comparison to embryo developmental rate and pregnancy rate. Materials and Methods: GCs were obtained at the time of oocyte retrieval from the patients undergoing IVF-ET program. The patients were divided into two groups: Group I (n=20) is poor responder (retrieved oocyte(s)$\leq$3ea), Group II (n=80) is normal responder (retrieved oocytes>3ea). After the extraction of total RNA, semiquantitative RT-PCR was performed and the expression level of LH-R mRNA was normalized by $\beta$-actin. Statistical analysis was performed by using $X^2$ test, Student's t-test and Pearson correlation. Results: In Group II, the relative values of LH-R mRNA (0.680 vs. 0.463, p<0.005) and pregnancy rate (54.7% vs. 23.1%, p<0.05) were significantly higher than in Group I. Number of retrieved oocyte(s) was gradually increased when the expression of LH-R mRNA was increased (p<0.05). But the quality of retrieved oocyte and transferred embryo were not related with the expression of LH-R mRNA. When the pregnancy rate was compared with FSH only group and FSH combined with hMG group in the ovarian stimulation protocol, FSH combined with hMG group was significantly higher than FSH only group in Group I (37.5% vs. 0%), and the expression of LH-R mRNA was significantly higher in hMG combined group than FSH only group (p<0.05). Conclusion: Expression level of LH-R mRNA has important role in ovarian function related with the response to gonadotrophin in human folliculogenesis. Furthermore these data might provide the evidence that additional use of hMG is helpful to poor responders.

• Proceedings of the Korean Aquaculture Society Conference
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• 2003.10a
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• pp.25-25
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• 2003

어류의 번식활동은 뇌-뇌하수체-생식소로 이어지는 내분비기관이 주로 담당하고 있으며, 그 중에서 뇌하수체에서 만들어지는 생식선자극호르몬 (GTH)은 생식소 발달에 매우 중요한 역할을 한다. 한편, GTH는 생식소에서 생성되는 스테로이드호르몬에 의해 유전자의 발현이 조절된다. 본 연구에서는 산천어 (Oncorhynchus masou)의 GTH 유전자발현에 미치는 estradiol-17$\beta$ (E2)의 영향을 생체내에서 먼저 조사하고, 유전자발현조절의 메카니즘을 구명하기 위하여 에스트로젠 수용체 (ER)의 cDNA를 cloning하였다. 또한 내분비교란물질로 알려져 있으며 에스트로젠 수용체와의 유사결합작용이 알려진 노닐페놀 (NP)의 영향도 아울러 조사하였다. 미성숙 산천어를 E2 및 NP (각각 5 mg/kg 및 10 mg/kg 체중)로 처리하면 72 시간째에 뇌하수체내의 $\beta$-actin mRNA에 비해 GTH (FSH$\beta$, LH$\beta$) mRNA가 상대적으로 증가하는 경향을 나타내었다. 뇌하수체에서 분리한 ER의 기본적인 구조는 DNA결합도메인 (Zf-C4), 호르몬결합도메인 (hormone_rec), 기타 전사활성화도메인으로 구성되어 있었으며 (그림 1), 기존에 보고된 에스트로젠 수용체중에서 무지개송어 및 대서양연어의 ER과 각각 98%, 96%의 높은 상동성이 관찰되었다. 향후, 준비된 ER을 재료로 하여 GTH 유전자발현의 호르몬의존성을 조사할 예정이다.

• Journal of Veterinary Clinics
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• v.24 no.4
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• pp.563-567
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• 2007

This study evaluated the effect of ANTORIN R-10(pFSH), a commercially available follicle stimulating hormone on ovarian morphology, on follicular development and serum estradiol levels in rats. Immature female Wistar S/T rats(27 day old; 80-100 g B.wt) maintained under controlled environmental conditions($22{\pm}2^{\circ}C$; 50% humidity; 12 h light/12 h dark cycle) with free access to standard laboratory feed and tap water were utilized. Animals were allowed to acclimatize to the new environment for at least 2 weeks before being included in the experiment. Rats were randomly allotted to 5 groups(Control, SL 0.1AU, SH 0.2AU, TL 0.1AU and TH 0.2AU). ANTORIN R-10 was subcutaneously injected twice daily for 3 days. Twenty hours after hormone treatment, blood was collected to estimate the serum estradiol $17-\beta$ concentration. Immediately, all rats were sacrificed and the ovarian morphology, ovary weight and number of follicles were recorded. Ovaries were fixed for histomorphological examination. Higher standard and treatment groups were significantly increased on ovary weight and the number of follicles more than 1mm compared with lower standard and treatment. However, no difference revealed between standard and treatment groups. ANTORIN R-10 was similar effects of follicles development and maturation compared with House standard FSH.

• 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.