• Journal of Embryo Transfer
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• v.20 no.3
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• pp.191-200
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• 2005

Many studies have shown that the development of mouse early 2-cell embryos in vitro is related with the intracellular $Ca^{2+}$ changes. In ICR strain mouse, the development of embryos arrests at early 2-cell stage, but the arrested early 2-cell embryos can be rescued by the addition of $Ca^{2+}$-related materials. Acetylcholine (ACh) increases intracellular $Ca^{2+}$ concentration ([$Ca^{2+}$]i) via the mAChR-PLC-IP3 pathway in mouse oocytes. We examined whether ACh rescues 2-cell block in mouse. In early 2-cell embryos, ACh increased [$Ca^{2+}$]i in a dose-dependent manner (p<0.001), and had an effect on rescue of 2-cell block and embryonic development. To identify the signal pathway involved in ACh-induced rescue of 2-cell block, we first applied an agonist of ACh receptor (AChR). Like ACh, carbachol increased intracellular $Ca^{2+}$ concentration ([$Ca^{2+}$]i) and atropine, an antagonist of ACh receptor, blocked the ACh-induced $Ca^{2+}$ increase. In $Ca^{2+}$-free medium, ACh also increased [$Ca^{2+}$]i, indicating that $Ca^{2+}$ increased by ACh is mainly released from the intracellular $Ca^{2+}$ store. The ACh-induced $Ca^{2+}$ increase was blocked by PLC inhibitor (U73122), ryanodine receptor (RyR) antagonist (dantrolene), and CaM KII inhibitor (KN-93), but not by IP3R antagonists (xestospongin C). These results show that ACh increases intracellular $Ca^{2+}$ concentration via mAChR/PLC/RyR, and this contributes to the rescue of 2-cell block.

• Journal of Animal Science and Technology
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• v.46 no.4
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• pp.571-584
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• 2004

It is known widely that granulosa cell apoptosis leads follicular atresia and macrophages exert their effects directly and/or indirectly from the initiation to the completion of follicular atresia by phagocytosis of apoptotic bodies and secretion of various cytokines. However, the site of initiation, propagation routes and the elimination methods of apoptotic bodies, and the time and methods of penetration of macrophages into the follicles are not known completely. Using pig(Yorkshire-breed) ovary, immunohistochemical studies with TUNEL for apoptotic bodies and pig macrophage monoclonal antibody 4E9 for macrophages, and light and transmission electron microscopic observations were performed. In the pig, follicular atresia began with the granulosa cell apoptosis, and the apoptosis of theca intema cells occured at the same time. The apoptosis of granulosa cells initiated randomly within the granulosa cell layer and propagated rapidly into the whole layer. Ultrastructura1ly, apoptotic granulosa cells showed characteristic pyknotic and deformed nucleus and intracytoplasmic vesicles. Apoptotic bodies were eliminated by intact granulosa cells and macrophages. Intact granulosa cells ingested apoptotic bodies transiently, soon after they fell into the apoptosis. Finally, apoptotic bodies and degenerated oocyte were phagocytosed by macrophages. Macrophages entered the ovarian follicle at the time of initiation of granulosa cell apoptosis, and migrated with the progression of apoptosis. By elimination of theca cells, macrophages contributed the completion of follicular atresia These results will provide valuable informations on the study of the interrelation between macrophage and ovarian follicular atresia.

• Applied Microscopy
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• v.39 no.1
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• pp.57-64
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• 2009

The trapping leaves of Drosera capture insects by secreting sticky mucilage from numerous glandular trichomes (GTs) that are developed on the leaf epidermis. The present study examines and compares the structural features of those trichomes in Drosera binata and D. pygmy with the use of light and electron microscopy. The study focuses primarily on the development and differentiation pattern of the GTs during growth. Upon examination, the upper and lower epidermis were readily distinguishable by the features of GTs in developing leaves. In particular, the GTs were dense in the upper epidermis and along the leaf margin. In D. binata, the capitate GTs with elongated stalk and sessile peltate GTs were found most commonly, whereas only capitate GTs with varying degrees of the stalk length were observed in D. pygmy. Up to ca. $2.2{\sim}3.4\;mm$ long capitate GTs were seen in the leaf margins of D. binata and ca. $3.7{\sim}4.2\;mm$ long GTs having racket-like head with adaxial hemispheric structures, otherwise known as tentacles, were noted in the leaf margin of D. pygmy. The peltate GTs were found to be distributed in the lower epidermis of D. binata. In both species, head cells were dense with cytoplasm containing high numbers of Golgi bodies, ER, mitochondria and small vesicles. Secretory materials accumulated within numerous small vacuoles, then fused together to form a single large vacuole, which serves as a secretory cavity. Flection movement of the marginal GTs and leaf blade GTs, and increased mucilage secretion from the head cells upon contact with prey during the capturing process are considered to be major factors in their active insectivorous mechanism. The findings of this study will be useful in comparisons to similar findings in other species that form adhesive trapping leaves, such as Drosophyllum or Pinguicula., further contributing a better understanding of the function and structure of the trapping leaves of carnivorous plants.

• Applied Microscopy
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• v.31 no.1
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• pp.101-108
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• 2001

Five kinds of neurosecretory cells (type-A, B, C, D and E) and neuropiles surrounding them were observed in the visceral ganglion and the right parietal ganglion of the African giant snail, Achatina fulica, by transmission electron microscopy. Type-A cells (diameter, $35{\mu}m$) are the most popular cells in the cortex of the two ganglions, which are of triangular or irregular forms. In their cytoplasm, there are found large granules of 1 fm in diameters and small round granules of about $0.1{\mu}m$ in diameters. Small granules are classified into the ones of high electron density and the others of middle electron density. Type-B cells (diameter, $19\times12{\mu}m$) are evenly distributed over various portions of cortex and medulla of the two ganglions. They are similar to type-A cells in shapes. The cytoplasm of type-B cells is crowded with high electron dense granules of about $0.1{\mu}m$. Round granules of about $0.7{\mu}m$ in diameters are also found but rarely. Type-C cells are the smallest cells whose sizes are about $8\times6{\mu}m$. Each of them contains a large nucleus of about $6\times5{\mu}m$. Its cytoplasm is full of electron dense granules of about $0.23{\mu}m$, each of which is artually an assembly of tiny granules of about $0.03{\mu}m$. Type-D cells are middle-size cells of about $28\times20{\mu}m$, which take ellipsoidal or irregular forms. They are found in the cortex more than in the medulla. Their cytoplasm looks dark due to the high electron density and, in it, two kinds of round granules whose sizes are $1.6{\mu}m$fm and $0.6{\mu}m$, respectively, are observed. Type-E cells are large cells of about $100\times50{\mu}m$, which are rarely found in the upper and middle portions of the two ganglions. The nucleus of the cell, which is very large $(70\times30{\mu}m)$ for the cytoplasm, contains electron dense round granules of diverse sizes (diameters, $1\sim0.2{\mu}m$). The surface of the cell protrudes filopodia of various forms and phagocytizes decrepit cells. Neuropiles are surrounding the neurosecretory cells. In nerve fibers, synaptic vesicles are observed, which are classified into six classes according to their electron densities , sizes and shapes.

• Applied Microscopy
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• v.29 no.3
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• pp.303-313
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• 1999

In this paper, five kinds of neurosecretory cells-light green (LG) cell, dark green (DG) cell, caudo-dorsal (CD) cell, blue green (BG) cell, and yellow (Y) cell- and neuropils in the cerebral ganglion of the African giant snail, Achatina fulica, were observed with an electron microscope. The following results were obtained. The LG cells are circular or ovoid in shape, and about $60{\mu}m$ in size. The nucleus and cytoplasm of the LG cell look light due to their electron-low density. Large granular chromatins are evenly developed in the karyolymph, where round nucleoli are also found. In the cytoplasm, electron -high dense round granules of $0.4{\mu}m$ in average size are crowded. The DG cells are ovoid in shape, and $50\sim20{\mu}m$ in size. These relatively electron-high dense cells were rarely found. In their cytoplasm, cell organelles such as rough endoplasmic reticulum and mitochondria are found together with electron -high dense round granules of $0.2{\mu}m$ in average size. The CD cells are ellipsoidal cells densely distributed in caudo-dorsal parts of the cerebral ganglion. They have large nuclei compared with the cytoplasm. The developed granular heterochromatins are observed in the karyolymph, and lots of small round granules of $0.12{\mu}m$ in average size in the cytoplasm. The 3G cells, rarely found around endoneurium of the cerebral ganglion, take the shapes of long ellipses. They look dark due to their electron -high density. In the cytoplasm, small round granules of $0.1{\mu}m$ in average size are found. The Y cells are the smallest among the neurosecretory cells($9\times6.6{\mu}m$ in size). They are found mostly between the medio-dorsal parts and the caudo-dorsal parts of the cerebral ganglion. In the cytoplasm, tiny round granules of $0.08{\mu}m$ in average size form a group. The neuropils are found in the middle of the cerebral ganglion. In the axon ending, round granules with electron -high density ($0.07\sim0.03{\mu}m$ in diameter) and lucent vesicles ($0.03{\mu}m$ in diameter) are found in large quantities. They are excreted in the state of exocytosome formed by the invagination of the limiting membrane of the axon ending.

• Applied Microscopy
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• v.40 no.2
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• pp.89-99
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• 2010

Fibroblasts are the most common cells in connective tissue and are responsible for the synthesis of extracellular matrix components. The fibrosis associated with chronic inflammation and injury may contribute to cholangiocarcinoma pathogenesis, particularly through an increase in extracellular matrix components, which participate in the regulation of bile duct differentiation during development. Mitochondria produce ATP through oxidative metabolism to provide energy to the cell under physiological conditions. Also, mitochondrial dysfunction and oxidative stress have been implicated in cellular senescence and aging. Alternations in mitochondrial structure and function are early events of programmed cell death or apoptosis and mitochondria appear to be a central regulator of apoptosis in most somatic cell. Clonorchis sinensis, one of the most important parasite of the human bile duct in East Asia, arouses epithelial hyperplasia and ductal fibrosis. Isolated fibroblast from the bile ducts of rats infected by C. sinensis showed increase of cytoplasmic process. In addition, decrease of cellular proliferation was observed in fibroblasts which was isolated from normal rat bile duct and then cultured in media containing C. sinensis excretory-secretory product. However, the effects of C. sinensis infection on the mitochondrial enzyme distribution is not clearly reported yet. Therefore, we investigated the structural change of C. sinensis infected bile duct and mitochondrial enzyme distribution of the cultured fibroblast isolated from the C. sinensis infected rat bile duct. As a result, C. sinensis infected SD rat bile ducts showed the features of chronic clonorchiasis, such as ductal connective and epithelial tissue dilatation, or ductal fibrosis. In addition, fibroblast in ductal connective tissue was damaged by physical effect of fibrotic tissue and chemical stimulation. Immunohistochemically detected mitochondrial electron transferase (ATPase, COXII, Porin) was decreased in C. sinensis infected rat bile duct and cultured fibroblast from infected rat bile duct. It can be hypothesized that the reason why number of electron transferase decrease in fibroblast isolated from the rat bile duct infected with C. sinensis is because dysfunction of electron transport system is occurred mitochondrial dysfunction, increase of ROS (reactive oxygen species) and apoptosis after chemical damage on the cell caused by C. sinensis infection. Overall, C. sinensis infection induces fibrotic change of ductal connective tissue, mutation of cellular metabolism in fibroblast and mitochondrial dysfunction. Consequently, ductal fibrosis inhibits fibroblast proliferation and decreases mitochondrial electron transferase on fibroblast cytoplasm. It was assumed that the structure of bile duct could not normalized and ductal fibrosis was maintained for a long period of time according to fibroblast metamorphosis and death induced by mitochondrial dysfunction.

• Journal of Life Science
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• v.27 no.10
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• pp.1191-1198
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• 2017

Vesicles and organelles are transported along microtubule and delivered to appropriate compartments in cells. The intracellular transport process is mediated by molecular motor proteins, kinesin, and dynein. Kinesin is a plus-end-directed molecular motor protein that moves the various cargoes along microtubule tracks. Kinesin 1 is first isolated from squid axoplasm is a dimer of two heavy chains (KHCs, also called KIF5s), each of which is associated with the light chain (KLC). KIF5s interact with many different binding proteins through their carboxyl (C)-terminal tail region, but their binding proteins have yet to be specified. To identify the interacting proteins for KIF5A, we performed the yeast two-hybrid screening and found a specific interaction with Ras-GTPase-activating protein (GAP) Src homology3 (SH3)-domain-binding protein 2 (G3BP2), which is involved in stress granule formation and mRNA-protein (mRNP) localization. G3BP2 bound to the C-terminal 73 amino acids of KIF5A but did not interact with the KIF5B, nor the KIF5C in the yeast two-hybrid assay. The arginine-glycine-glycine (RGG)/Gly-rich region domain of G3BP2 is a minimal binding domain for interaction with KIF5A. However, G3BP1 did not interact with KIF5A. When co-expressed in HEK-293T cells, G3BP2 co-localized with KIF5A and was co-immunoprecipitated with KIF5A. These results indicate that G3BP2, which was originally identified as a Ras-GAP SH3 domain-binding protein, is a protein that interacts with KIF5A.

• Korean Journal of Food Science and Technology
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• v.27 no.1
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• pp.112-118
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• 1995

The effect of various packaging methods on kimchi quality was investigated in order to develop the packaging techniques for preventing commercial kimchi products from inflation and explosion, due to fermentative gas evolved during storage and distribution. Kimchi was packaged in different methods; 1) atmospheric packaging(AP), 2) check valve packaging(CV), 3) double packaging(DP), and 4) vacuum packaging(VP). The quality of kimchi during storage at $10^{\circ}C$ was evaluated in terms of gas composition, free volume, pH, titratable acidity color index and sensory properties. The gas composition inside packages showed different curves according to the packaging methods. Due to fermentative gas accumulation in both AP and CV, $CO_2$ concentration increased by 2 stepwise pattern, while $O_2$, concentration decreased exponentially. In DP, $O_2$ concentration remained constant, but $CO_2$ concentration increased by 2 stepwise pattern and then decreased. In contrast, VP produced low $O_2$ and high $CO_2$ concentrations only at the end of storage. The free volume in both AP and CV showed typical sigmoidal curves similar to $CO_2$ concentration changes. It remained constant in DP, but started to increase at the late stage of storage in VP. There was no significant effect of packaging methods on pH changes of kimchi. In titratable acidity, DP maintained relatively higher than others. Regarding to the color change of crushed kimchi juice in all packages, L and b values decreased exponentially but a value remained constant during storage. Color index(L b/a) of crushed kimchi juice decreased exponentially and remained constant at the end of storage. The growth of lactic acid bacteria was VP, CV, AP, DP in increasing order. In sensory test, the sourness scores of DP were fairly higher than those of others, but the texture was not significantly affected by the packaging methods. The preference for kimchi showed VP>AP, CV>DP in order of score. In this study, it could be proposed to employ DP and VP method as the effective packaging techniques for preventing commercial kimchi products from inflation.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.17 no.5
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• pp.423-435
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• 1984

The reproductive cycle of the small filefish, Rudarius ercodes was investigated based on the annual variations of gonadosomatic index(GSI) and hepatosomatic index(HSI) by electronic and photic microscophy. The specimens used were collected at the coastal area of Benden island, Sizuokagen, Japan, from September 1982 to August 1983. GSI began to increase from March, starting season of longer daylength and higher water temperature, and reached the maximum value between June and August. It began to decrease from September with the lowest value appearing between November and February without any evident variation. The annual variations of HSI were not distinct in male filefish and were negatively related to GSI in female : HSI decreased in the summer season when the ovary was getting mature and reached the maximum in the winter season when the ovary was getting retrogressive. The ovary consisted of a pair of saccular structure with numerous ovarian sacs branched toward the median cavity. Oogonia divided and proliferated along the germinal epithelium of the ovarian sac. Young oocytes with basophile cytoplasm showed several scattering nucleoli along the nuclear membrane. when the oocytes growing to about 300 ${\mu}m$, nuclear membrane to disappear with nucleus migrating toward the animal pole. The regions of protoplasm were extremely confined within the animal hemisphere in which most of cytoplasms were filled with yolk materials and oil drops. After ovulation, residual follicles and growing oocytes remaining in the ovarian sacs degenerated. But perinucleatic young oocytes without follicles formed were not degenerated, and growing continuously still in the next year. Mitochondria and endoplasmic reticula in the cytoplasm remarkably increased with oocytes maturing and yolk accumulating. Those were considered to be functionally related to the yolk accumulation. Five or six layers of possible vitellogenin, oval-shaped disc structures with high electron density, appeared in the apex of follicular processes stretching to the microvilli pits of mature oocytes. Testis consisting of a pair of lobular structures in the right and left were united in the posterior seminal vesicle, Cortex of testis was composed of several seminiferous tubules, and medulla consisting of many sperm ducts connected with tubules. Steroid hormone-secreting cells with numerous endoplasmic reticula and large mitochondria of well developed cristae were recognized in the interstitial cells of the growing testis. Axial filament of spermatozoon invaginated deeply in the central cavity of the nucleus and the head formed U-shape with acrosome severely lacking, mitochondria formed large globular paranuclei at the posterior head, and microtubular axoneme of the tail represented 9+9+2 type. The annual reproductive cycles could be divided into five successive stages : growth(March to July), maturation(May to September), Spawning(mid May to early October) and resting stages(October to February). The spawning peak occurred from June to August.

• Restorative Dentistry and Endodontics
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• v.32 no.6
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• pp.498-513
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• 2007

Since it was reported that incipient enamel caries can be recovered, previous studies have quantitatively evaluated that enamel artificial caries have been, remineralized with fluoride showing simultaneously the increase of width of surface layer and the decrease of width of the body of legion. There is, however, little report which showed that remineralization could occur without fluoride. In addition, the observations on the change of hydroxyapatite crystals also have been scarcely seen. In this study, enamel caries in intact premolars or molars was induced by using lactic acidulated buffering solutions over 2 days. Then decalcified specimens were remineralized by seven groups of solutions using different degree of saturation(0.212, 0.239, 0.301, 0.355) and different pH(5.0, 5.5, 6.0) over 10 days. A qualitative comparison to changes of hydroxyapatite crystals after fracturing teeth was made under SEM(scanning electron microscopy) and AFM(atomic force microscopy). The results were as follows: 1. The size of hydroxyapatite crystals in demineralized area was smaller than the normal ones. While the space among crystals was expanded, it was observed that crystals are arranged irregularly. 2. In remineralized enamel area, the enlarged crystals with various shape were observed when the crystals were fused and new small crystals in intercrystalline spaces were deposited. 3. Group 3 and 4 with higher degree of saturation at same pH showed the formation of large clusters by aggregation of small crystals from the surface layer to the lesion body than group 1 and 2 with relatively low degree of saturation at same pH did. Especially group 4 showed complete remineralization to the body of lesions. Group 5 and 6 with lower pH at similar degree of saturation showed remineralization to the body of lesions while group 7 didn't show it. Unlike in Group 3 and 4, Group 5 and 6 showed that each particle was densely distributed with clear appearance rather than crystals form clusters together.