• Animal Bioscience
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• v.35 no.3
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• pp.399-409
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• 2022

Objective: Follicle-stimulating hormone (FSH) is the central hormone involved in mammalian reproduction, maturation at puberty, and gamete production that mediates its function by control of follicle growth and function. The present study investigated the mutations involved in the regulation of FSH receptor (FSHR) activation. Methods: We analyzed seven naturally-occurring mutations that were previously reported in human FSHR (hFSHR), in the context of equine FSHR (eFSHR); these include one constitutively activation variant, one allelic variant, and five inactivating variants. These mutations were introduced into wild-type eFSHR (eFSHR-wt) sequence to generate mutants that were designated as eFSHR-D566G, -A306T, -A189V, -N191I, -R572C, -A574V, and -R633H. Mutants were transfected into PathHunter EA-parental CHO-K1 cells expressing β-arrestin. The biological function of mutants was analyzed by quantitating cAMP accumulation in cells incubated with increasing concentrations of FSH. Results: Cells expressing eFSHR-D566G exhibited an 8.6-fold increase in basal cAMP response, as compared to that in eFSHR-wt. The allelic variation mutant eFSHR-A306T was not found to affect the basal cAMP response or half maximal effective concentration (EC₅₀) levels. On the other hand, eFSHR-D566G and eFSHR-A306T displayed a 1.5- and 1.4-fold increase in the maximal response, respectively. Signal transduction was found to be completely impaired in case of the inactivating mutants eFSHR-A189V, -R572C, and -A574V. When compared with eFSHR-wt, eFSHR-N191I displayed a 5.4-fold decrease in the EC₅₀ levels (3,910 ng/mL) and a 2.3-fold decrease in the maximal response. In contrast, cells expressing eFSHR-R633H displayed in a similar manner to that of the cells expressing the eFSHR-wt on signal transduction and maximal response. Conclusion: The activating mutant eFSHR-D566G greatly enhanced the signal transduction in response to FSH, in the absence of agonist treatment. We suggest that the state of activation of the eFSHR can modulate its basal cAMP accumulation.

• Journal of Marine Life Science
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• v.3 no.2
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• pp.74-80
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• 2018

Mercury (Hg) poses a threat to marine ecosystem due to continuous inflow from various industries and bioaccumulation to higher trophic level via food web. Mercury can adversely affect growth, development, reproduction and metabolism to aquatic organisms. In the present study, acute toxicity and oxidative stress markers (total glutathione content, and activities of GST, GR and GPx) were investigated in brackish water flea Disphanosoma celebensis exposed to HgCl₂ for 24 h. As results, Hg showed negative effect in survival of D. celebensis. 24 h-LC₅₀ value was determined as 0.589 mg/l (95% C.I. 0.521~0.655 mg/l). After exposure to Hg (0.08 and 0.4 mg/l) for 24 h, total glutathione content was significantly decreased, whereas GST, GPx and GR activities were enhanced. These findings indicate that Hg induced oxidative stress in D. celebensis, and oxidative stress markers may be involved in cellular defense against Hg - mediated toxicity. This study provides a better understanding of molecular mode of action of Hg toxicity in this specie and potent of molecular markers for heavy metal monitoring in marine ecosystem.

• Development and Reproduction
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• v.5 no.1
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• pp.39-45
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• 2001

Black rockfish Sebastes schlegeli is one of scopaenid fish giving birth to yolk-sac larvae and distributed around the coast of Korea and Japan, and is one of main species which is cultured in Korea. The difffrentiation of thyroid gland and the changes of thyroid hormones (THs) concentrations in the whole body during early development of this species were examined and the relationship between thyroid gland and their growth will be presented. Total length (TL) and body weight (BW) of the larva at the parturition were 6.3${\pm}$0.1 mm and 3.6${\pm}$0.1 mg, respectively. The larvae transformed to juvenile after 30th day after parturition. According to Hoshiai(1977), they had gown into stage Fl (TL 13.2${\pm}$0.9 mm, BW 46.5${\pm}$1.5 mg) at 50th day. The thyroid gland of black rockfish was first observed histologically in hatching larvae in mother fish. The larvae just after parturition have 1${\sim}$3 of the thyroid follicles diffrentiated between basibranchial bone and ventral aorta, at the base of the first gill arch. In this time, thyroid follic1e number (TFN),thyroid follicle diameter (TFD) and thyroid cell height (TCH) were 1.6${\pm}$0.8 pieces/inds., 18.1${\pm}$0.6 ${\mu}$m and 4.1${\pm}$0.2 ${\mu}$m, respectively. TFN and TFD at 50th day were increased to 32.5${\pm}$6.9 pieces/inds. and 41.5${\pm}$1.7 ${\mu}$m, respectively. These results indicate that they are related to the growth of black rockfish during early development. However, TCH indicates that the activity of thyroid gland appearedat special day, eg. 5, 20 and 50th day, suggesting that TCH may have some role in the physiological activity. L-thyroxine (T$_4$)concentration decreased sharply to 10 days after parturition, and at 25th day (metamorphosis stage) increased markedly to 3.44${\pm}$0.93ng/g fish. After this time, T$_4$ concentration decreased at 35th day, but then increasedagain to the highest concentration, 5.63${\pm}$0.70ng/g fish. 3,5,3'-triiodo-L-thyronine (T$_3$) concentration declined sharply from just after pafurition (4.96${\pm}$1.90 ng/g fish) to 5th day (0.30${\pm}$0.07 ng/g fish). However T$_3$ concentration increased markedly to 0.95${\pm}$0.11 ng/g fish at 30th day and then did not significantly change until 45th day, increased also sharply to 1.67${\pm}$0.23 ng/g fish at 50th day.

• Proceedings of the KSAR Conference
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• 2001.03a
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• pp.31-31
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• 2001

This study was to establish in uitro culture system of mouse preantral follicles and to obtain higher in vitro development rates and production of live young. Preantral follicles were obtained from 12-day-old FI mouse (C57BL $\times$ CBA) by enzymatical methods. Oocyte-granulosa cell complexes (OGCs) of preantral follicles were loaded on Transwell-COL insert and cultured in $\alpha$MEM supplemented with 5% FBS, 100 mIU/$m\ell$ FSH and 100 mIU/$m\ell$ hMG for IVG. IVM was performed in $\alpha$MEM supplemented 1.5 IU/$m\ell$ hCG for 18 hrs and IVF was carried out in Ml6 medium. Embryos were cultured in modified Ml6 medium supplemented 10% FBS for 4 days. The effect of the OGCs size on the nuclear/cytoplasmic maturation was significantly higher in 120-150 ${\mu}{\textrm}{m}$ (MII: 33.0%, $\geq$2-cell: 36.7%, $\geq$morula: 20.9%) than in 70-110 ${\mu}{\textrm}{m}$ (MII: 12.2%, $\geq$2-cell: 10.2%, $\geq$morula: 4.8%) (p<0.001). In period of the IVG days, the rate of $\geq$2-cell was significantly higher in 10 days(38.2%) than in 12 days (20.0%) (p<0.01). In period of IVF time, 9 hrs ($\geq$2-cell: 31.5%, $\geq$ morula: 14.3%) indicated significantly higher cytoplasmic maturation rate than 4 hrs ($\geq$2-cell: 17.5%, This study was to establish in vitro culture system of mouse preantral follicles and to obtain higher in vitro development rates and production of live young. Preantral follicles were obtained from 12-day-old FI mouse (C57BL $\times$ CBA) by enzymatical methods. Oocyte-granulosa cell complexes (OGCs) of preantral follicles were loaded on Transwell-COL insert and cultured in $\alpha$MEM supplemented with 5% FBS, 100 mIU/$m\ell$ FSH and 100 mIU/$m\ell$ hMG for IVG. IVM was performed in $\alpha$MEM supplemented 1.5 IU/$m\ell$ hCG for 18 hrs and IVF was carried out in Ml6 medium. Embryos were cultured in modified Ml6 medium supplemented 10% FBS for 4 days. The effect of the OGCs size on the nuclear/cytoplasmic maturation was significantly higher in 120-150 ${\mu}{\textrm}{m}$ (MII: 33.0%, $\geq$2-cell: 36.7%, $\geq$morula: 20.9%) than in 70-110 ${\mu}{\textrm}{m}$ (MII: 12.2%, $\geq$2-cell: 10.2%, $\geq$morula: 4.8%) (p<0.001). In period of the IVG days, the rate of $\geq$2-cell was significantly higher in 10 days(38.2%) than in 12 days (20.0%) (p<0.01). In period of IVF time, 9 hrs ($\geq$2-cell: 31.5%, $\geq$ morula: 14.3%) indicated significantly higher cytoplasmic maturation rate than 4 hrs ($\geq$2-cell: 17.5%, $\geq$morula: 4.8%) and 7 hrs ($\geq$2-cell: 20.4%, $\geq$morula: 6.1%) (p<0.01). However, there was no difference in cytoplasmic maturation between co-cultured preantral follicle ( $\geq$morula: 17.4%) and preantral follicle cultured in Ml6 ( $\geq$morula: 17.4%). 22 morula and blastocysts produced in above optimal condition were transferred to uterus of 2 pseudopregnant recipients, 1 recipient was pregnant and then born 1 live young. This result demonstrates that in vitro culture system of preantral follicles can be used efficiently as another method to supply mouse oocyte.morula: 4.8%) and 7 hrs (2-cell: 20.4%, $\geq$morula: 6.1%) (p<0.01). However, there was no difference in cytoplasmic maturation between co-cultured preantral follicle ( $\geq$morula: 17.4%) and preantral follicle cultured in Ml6 ( $\geq$morula: 17.4%). 22 morula and blastocysts produced in above optimal condition were transferred to uterus of 2 pseudopregnant recipients, 1 recipient was pregnant and then born 1 live young. This result demonstrates that in vitro culture system of preantral follicles can be used efficiently as another method to supply mouse oocyte.

• Journal of the Korean Society of Marine Environment & Safety
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• v.21 no.6
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• pp.655-661
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• 2015

This study intends to evaluate the effect of nitric acid($HNO_3$) spill accidents on the marine ecosystem, while $HNO_3$ is known as one of the typical HNS. For this purpose, we performed (1) the growth inhibition test by using phytoplankton(Skeletonema costatum), (2) acute and chronic toxicity test by using invertebrate(Brachionus plicatilis and Monocorphium acherusicum), (3) fish(Cyprinodon variegatus) and (4) bacteria(Vibrio fischeri). In these tests, we observed the (1) pH changes induced by the nitric acid spill and (2) changes in nitrate($NO_3$) concentration disassociated from nitric acid after the accident, respectively. The toxicity test result on pH changes induced by $HNO_3$ shows that the no observed effect concentration(NOEC), lowest observed effect concentration(LOEC) and 50 % effect concentration($72h-EC_{50}$) values of M. acherusicum are pH 7 (0.3 mM), pH 5(1.1 mM) and pH 5.2(1.4 mM), respectively, indicating that M. acherusicum is the most sensitive species. The chronic toxicity test (population growth rate test) on $NO_3{^-}$ of B. plicatilis show that the NOEC, LOEC and $96h-EC_{50}$ are 5.9 mM, 11.8 mM and 32.6 mM, respectively, indicating that B. plicatilis is the most sensitive species. In conclusion, toxic effecst on the marine organism caused by the nitric acid spill accident is determined to be so slightly except for the most adjacent area of the ship in pH scale and such concentration of nitrate, to the extent of directly influencing the survival and reproduction of the marine organism, is determined practically not to be applicable in the typical accidents in the sea.

• Korean Journal of Environment and Ecology
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• v.29 no.4
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• pp.580-589
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• 2015

The artificially grown forests of larch, planted in accordance with the nationwide afforestation policy in the 1970s, are located inside national parks. This study intended to induce a forestation system by which the forests develop into an ecologically healthy and broadleaved ecosystem with broad species diversity. For this, the aspects of natural regeneration of broadleaves from 2010 to 2013 after thinning by density (30%, 50%, and 70%) in 2009 were surveyed using the larch forest in the Woljeong Temple region inside Odaesan National Park. There were no trees that were larger than 2 cm in in diameter at breast height among the trees recently introduced between 2012 and 2013. A significant number of herbs have been introduced to the subsurface alongside young arboreal trees species such as Bumalda bladdernut, Acer triflorum, Cornus controversa etc and shrubs. However, many woody species did not survive the competition with herbs and repeated withering and regeneration. The number of woody species generated within the 30% cutting area was 440 species in 2013 and this figure has been increasing twofold each year. The number of woody plants within the 50% cutting area also showed an upward tendency and most plants did not survive in the competition with herbs and Sasa borealis and withered in only 1 ~ 2 years after generation. Unlike other thinning areas, the 70% cutting area showed 608 broadleaved trees, reflecting a decrease from 748 trees in 2012. This appeared to be attributed to the luxuriance of S. borealis and the sharp increase of fatsia following the inflow of total sunlight to the forest floor. Herbs were hardly generated due to the influence of S. borealis. Regarding the density for thinning at 50% or upper height, the forest treatment division shall maintain a proper density in the course of inducing artificial forestation of larch into natural broadleaved forests considering the luxuriance of sasa borealis and herbs due to the inflow of total sunlight to the forest floor.

• Korean Journal of Plant Taxonomy
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• v.46 no.2
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• pp.256-266
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• 2016

A total of 13,000 individuals of Dendrobium moniliforme (L.) Sw. artificially propagated in laboratories and greenhouses were restored in their natural habitat of Bogildo Island, Wandogun, in the southern part of Korea in June of 2013. The growing conditions of the individuals were monitored for two years. The parental individuals for the restoration were obtained from a wild population in southern Korea, from which seeds were produced via artificial crossings. These seeds were germinated and cultivated in growing media and two-year-old plants were then grown in greenhouse beds. The genetic diversity among the propagated individuals was confirmed by examining DNA sequences of five regions of the chloroplast genome and the nuclear ITS region. The diversity values were as high as the average values of natural populations. All propagated individuals were transplanted into two different sites on Bogildo by research teams with local residents and national park rangers. After restoration, we counted and measured the surviving individuals, vegetative propagated stems, and growth rates in June of both 2014 and 2015. There was no human interference, and 97% of the individuals survived. The number of propagules increased by 227% in two years. In contrast, the average length of the stems decreased during the period. In addition, different survival and propagation rates were recorded depending on the host plants and the restored sites. The shaded sides of rock cliffs and the bark of Quercus salicina showed the best propagation rates, followed by the bark of Camellia japonica. A few individuals of D. moniliforme successfully flowered, pollinated, and fruited after restoration. Overall, our monitoring data over two years indicate that the restored individuals were well adapted and vigorously propagated at the restored sites. In order to prevent human disturbance of the restored sites, a CCTV monitoring system powered by a solar panel was installed after the restoration. In addition, a human surveillance system is operated by national park rangers with local residents.

• Development and Reproduction
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• v.8 no.1
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• pp.57-64
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• 2004

This study was carried out to examine the expression of the circadian clock genes in the mouse ovary and testis at different developmental stages. Expression of Period1(Per 1), Period2(Per2), Period3(Per3), Cryptochrome1(Cry1), Cyptochrome2(Cry2), Clock Small and Prokineticin1 and Prokineticin2 receptor(Prok1r, Prok2r) genes in mouse ovary was explored by semiquantitative reverse transcription Polymerase chain reaction(RT-PCR) according to the developmental stage(post partum day; ppd 1, 7, 10, 21 and 35). Immunohistochemistry using PER1 antibody was also analyzed. The differential expression pattern of clock genes was presented according to stages of the mouse ovarian development (ppd 1, 7, 10, 21 and 35). In the cases of ovaries, at the starting point of follicle growth at ppd 7 and 10, the clock gene expression patterns were changed vastly. According to the developmental stages, the clock genes were highly expressed at ppd 7 and 10 in mouse testis also. Receptors for Prok2, the circadian output molecule of SCN, were also expressed in ovary at ppd 7 and in testis at ppd 1 and 7, respectively. Immnunohistochemical analysis of PER1 showed positive signals in the cytoplasm of oocytes and granulosa cells. The level or PER1 expression was increased in cells at the spermatogonia and the condensing spermatids. The expression pattern of Perl and localization of PER1 were showed similar patterns according to the developmental stages in ovary and testis. Taken together, it could be observed that the expression of clock genes was highly correlated with gonadal development and germ cell differentiation in mice. Therefore, in this study, circadian programming of the genes in the ovary and testis is strongly imposed across a wide range of core reproductive cycles and normal development of gametes. Although the existence of circadian genes is clearly investigated, further studies on the direct evidence is required for the understanding of the relationship between circadian genes and regulation of gonadal differentiation and germ cell development.

• Ocean and Polar Research
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• v.37 no.1
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• pp.49-59
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• 2015

Spatial variation in the reproductive effort of Manila clam Ruditapes philippinarum is often closely associated with variation in the seawater temperature and food availability, which determines gonad maturity and the quantity of gamates produced during spawning. Previous studies also have reported that severe infection by the protozoan parasite Perkinsus olseni exerts a negative impact on clam reproduction, retarding gonad maturation or decreasing the reproductive effort. In the present study, we investigated impacts of P. olseni infection on the reproductive condition of Manila clam during a spawning season. Histology revealed that 54% of female clams in Wando off the south coast were in spawning, while only 10% of the female from Gomso and 0% of the female from Seonjaedo in Gyeonggi bay off the west coast were engaged in spawning at the end of May in 2004. Ray's fluid thioglycollate media (RFTM) assay was applied to assess P. olseni infection and indicated that the infection intensity in Wando ($3,608,000{\pm}258,000cells/g$ wet tissue) was significantly higher than the levels in Gomso ($1,305,000{\pm}106,000cells/g$ wet tissue) and Seonjaedo ($1,083,000{\pm}137,000cells/g$ wet tissue, p < 0.001). The size of the ripe female follicle determined from histology was significantly smaller in Wando ($0.032mm^2$) compared to the sizes in Gomso ($0.059mm^2$) and Seonjaedo ($0.052mm^2$, p < 0.05). Accordingly, the number of ripe eggs in the follicle was significantly fewer among clams in Wando (14) compared to the numbers determined in Gomso (23) and Seonjaedo (22). The absolute quantity of egg in ripe clams from Wando (31.01 mg) was also significantly smaller than Seonjaedo (61.79 mg) and Gomso (133.3 mg). Quantity of total protein, carbohydrate, and lipid in the tissue in the Wando samples was significantly smaller than the quantities determined in Gomso and Seonjaedo (p < 0.001). The observed poor reproductive condition and proximate tissue composition of the females in Wando were, in part, explained by the extremely high level of the parasites, sapping the ability to store energy in the host tissues, which is used in tissue growth and the egg production.

• Development and Reproduction
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• v.11 no.3
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• pp.167-177
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• 2007

In the present study, we isolated three human adult stem cells including adipose tissue-derived stem cells(HAD), umbilical cord-derived stem cells(HUC), and amnion-derived stem cells(HAM) and analysed their characteristics. And we examined whether HAD could be used as therapeutical cells for the heart diseases. Both HAM and HUC appeared very similar morphology but HAD was different. Doubling time of HUC was most fast, but total doubling numbers of HUC was same with HAM. Total doubling numbers of HAD was much more than others. Expression patterns of genes and proteins of three human adult stem cells were very similar. Also they were differentiated into adipocytes, osteocytes, and chondrocytes. In addition, they expressed many cardiomyocyte-related genes. But expression pattern of genes is a little different. When HAD were cultivated in the presence or absence of various combinations of BMP and FGF after 5-azacytidine expose for 24 h, expression of Cmlc-1, and ${\alpha}1c$ genes was significantly increased. However, expression of troponin T, troponin I and Kv4.3 genes was not changed. Based on these observations, it is suggested that HAD, HUC, and HAM might be used as potentially therapeutical cells for clinical application.