• Journal of Life Science
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• v.24 no.7
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• pp.798-803
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• 2014

Cathepsin D (CtsD), an aspartyl peptidase, is involved in apoptosis, resulting in the release of cytochrome C from mitochondria in cells. Here, we investigated microRNA regulation of CtsD expression in 3T3-L1 cells First, we observed the expression of CtsD in cells in response to doxorubicin (Dox). As expected, the level of CtsD mRNA was increased in 3T3-L1 cells exposed to Dox in a dose-dependent manner. Cellular viability of ectopically expressed CtsD cells was also decreased. Next, we used the miRanda program to search for particular microRNA targeting CtsD. MiR-145 was selected as a putative controller for CtsD because miR-145 had a high mirSVR score. In a reporter assay, the luciferase activity of cells containing the CtsD 3'-UTR region was decreased in cells transfected with miR-145 mimic compared to that of a control. The level of CtsD expression was down-regulated in preadipocytes ectopically expressing miR-145 and up-regulated by an miR-145 inhibitor. Cells also suppressed miR-145 expression when exposed to Dox. The miR-145 inhibitor reduced the cellular viability of 3T3-L1 cells. Taken together, these data suggest that miR-145 regulates CtsD-mediated cell death in adipocytes. These findings may have valuable implications concerning the molecular mechanism of CtsD-mediated cell death in obesity, suggesting that CtaD could be a useful therapeutic tool for the prevention and treatment of obesity by regulating fat cell numbers.

• Journal of Life Science
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• v.24 no.7
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• pp.777-783
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• 2014

Cathepsin D (CtsD), an aspartyl peptidase, is involved in apoptosis, resulting in the release of cytochrome C from mitochondria in cells. Here, we investigated microRNA regulation of CtsD expression in 3T3-L1 cells. First, we observed the expression of CtsD in cells in response to doxorubicin (Dox). As expected, the level of CtsD mRNA increased in 3T3-L1 cells exposed to Dox in a dose-dependent manner. The cellular viability of ectopically expressed CtsD cells was decreased. Next, we used the miRanda program to search for particular microRNA targeting CtsD. MiR-145 was selected as a putative controller of CtsD because it had a high mirSVR score. In a reporter assay, the luciferase activity of cells containing the CtsD 3'-UTR region decreased in cells transfected with a miR-145 mimic compared to that of a control. The level of CtsD expression was down-regulated in preadipocytes ectopically expressing miR-145 and up-regulated by an miR-145 inhibitor. Cells also suppressed miR-145 expression when exposed to Dox. The miR-145 inhibitor reduced the cellular viability of 3T3-L1 cells. Taken together, these data suggest that miR-145 regulates CtsD-mediated cell death in adipocytes. These findings may have valuable implications concerning the molecular mechanism of CtsD-mediated cell death in obesity, suggesting that CtsD could be a useful therapeutic tool for the prevention and treatment of obesity by regulating fat cell numbers.

• Journal of Life Science
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• v.27 no.5
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• pp.545-552
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• 2017

Julbernardia globiflora, a tropical African tree widespread in Miombo woodland, has been used in folk medicine for the treatment of depression and stomach problems. However, the antioxidative and anticancer activities of J. globiflora remain unclear. The objective of this study is to evaluate the antioxidative and anticancer effects of methanol extract of J. globiflora (MEJG) and the molecular mechanism of its anticancer activity in human colon carcinoma HT29 cells. MEJG exhibited significant antioxidative effect with an $IC_{50}$ (concentration at 50% inhibition) value of $1.23{\mu}g/ml$ measuring by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay and inhibited cell proliferation in a dose-dependent manner in HT29 cells. We found that MEJG induced apoptosis of HT29 cells with the increase of apoptotic cells and apoptotic bodies using Annexin V staining and 4,6-diamidino-2-phenylindole (DAPI) staining, respectively. The MEJG treatment showed the increase of Fas, a death receptor, and Bax, a pro-apoptotic protein, and the decrease of Bcl-2, an anti-apoptotic protein, resulting in the release of cytochrome c from the mitochondria into the cytosol and activation of caspase-3, -8 and -9. The apoptotic effects of MEJG were confirmed by cleavage of poly (ADP-ribose) polymerase (PARP). Collectively, these results suggest that MEJG may exert the anticancer effect in HT29 cells by inducing apoptosis via both the intrinsic and extrinsic pathways.

• Journal of Life Science
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• v.27 no.11
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• pp.1340-1348
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• 2017

Sanguinarine is a benzophenanthridine alkaloid derived from the roots of Sanguinaria canadensis L., which is used for the purpose of treating various diseases. Although studies of anticancer activities have been performed using various cancer cell lines, the phenomenon of inducing apoptosis in cancer cells by using sanguinarine requires more research. Therefore, this study investigated the anti-cancer activities and related mechanisms of sanguinarine used with Hep3B human hepatocellular carcinoma cells in terms of the regulation of apoptosis. Sanguinarine inhibited the proliferation of Hep3B cells in a concentration-dependent manner, which was associated with the induction of apoptosis. Sanguinarine also increased the activity of caspase-3, which is a typical effector caspase, and the activities of caspase-8 and caspase-9, which are key when initiating extrinsic and intrinsic apoptosis pathways, respectively. In addition, sanguinarine increased the expression of death receptor-related genes and pro-apoptotic BAX, which belongs to the Bcl-2 family, while suppressing the expression of anti-apoptotic Bcl-2. Sanguinarine promoted the truncation of Bid and enhanced the release of cytochrome c from the mitochondria to the cytoplasm due to a loss of mitochondrial membrane potential. Furthermore, the reduction of a survival rate that was induced by sanguinarine and the induction of apoptosis disappeared with the inhibition of artificial caspase activity. Therefore, the results of the study indicated that sanguinarine-induced apoptosis in Hep3B cells involves both extrinsic and intrinsic pathways; such apoptosis is a caspase-dependent phenomenon.

• Journal of Radiation Protection and Research
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• v.43 no.2
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• pp.66-74
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• 2018

Background: Tumor response to anticancer therapies can much be influenced by microenvironmental factors. In this study, we determined the effect of these microenvironmental factors on DNA methylation using A549 human lung adenocarcinoma cell line. Materials and Methods: We subjected A549 cells to various conditions mimicking tumor microenvironment including hypoxia, acidosis (sodium lactate), oxidative stress ($H_2O_2$), bystander effect (supernatant from doxorubicin (Dox)-treated or irradiated cells), and immune cell infiltration (supernatant from THP-1 or Jurkat T cells). Genomic DNA was isolated from these cells and analyzed for DNA methylation. Clonogenic cell survival, gene expression, and metabolism were analyzed in cells treated with some of these conditions. Results and Discussion: We found that DNA methylation level was significantly decreased in A549 cells treated with conditioned media from Dox-treated cells or Jurkat T cells, or sodium lactate, indicating an active transcription. To determine whether the decreased DNA methylation affects radiation sensitivity, we exposed cells to these conditions followed by 6 Gy irradiation and found that cell survival was significantly increased by sodium lactate while it was decreased by conditioned media from Dox-treated cells. We further observed that cells treated with conditioned media from Dox-treated cells exhibited significant changes in expression of genes including BAX and FAS (involved in apoptosis), NADPH dehydrogenase (mitochondria), EGFR (cellular survival) and RAD51 (DNA damage repair) while sodium lactate increased cellular metabolism rather than changing the gene expression. Conclusion: Our results suggest that various tumor microenvironmental factors can differentially influence DNA methylation and hence radiosensitivity and gene expression in A549 cancer cells.

• Applied Microscopy
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• v.2 no.1
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• pp.7-15
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• 1972

The morphological and fine structural changes during the oogensis of Clonorchis sinensis were studied on the developing ovums in the ovary and ootype with electron microscope. Adult worms were removed from the hepar of the which and previously infected with metacercariae of Clonorchis sinensis. The ovary including the Mehlis' glands and an ootype from adult worm was prefixed for 1-2 hours in 1.25% glutaraldehyde buffered with 0.2M cacodylate at PH 7.2, secondarily fixed for 30 minutes in potassium bicromate and postfixed for an hour in 1% osmic acid buffered with 0.4M cacodylate at PH 7.2. After fixation tissues were dehydrated in an alcohol series, embedded in Epon 812 from propylene oxide and stained with saturated uranyl acetate and $Pn(NO_3)_2$ solution. Material was examined with a Hitachi HS-7S electron microscope. The periphery of the ovary, except for the posterior region, is made up of oogonia. As the oogonia divide they proliferate primary oocytes toward the central part of the ovary. After a period of growth the primary oocyte leaves the ovary and is penetrated by a sperm in the ootype. Sperm penetration immediately activates the primary oocyte to resume its meiotic activity. After the oocytes meiotic activity is completed, the pronuclei fuse to form a single cleavage nucleus which possesses two nucleoli. As the oocytes develop their cytoplamic materials are abundant; small mitochondria are abundant and often their profiles are more unmerous in one part of the cytoplasm than elsewhere; the granular endoplasmic reticulum becomes alveolar-sac form after it leaves the ovary it becomes stratified form. The reticulate Golgi apparatus is apparent in the developed oocyte. A little of cortical granules are distributed inside of the plasma membrane I oogonia and large quantity of cortical granules are arranged just inside of the plasma membrane of the primary oocyte and after fertilization they are disappeared with broken out.

• Applied Microscopy
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• v.35 no.2
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• pp.65-72
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• 2005

The ultrastructure of submandibular gland was examined in the lesser white-toothed shrew, Crocidura suaveolens. The submandibular gland of C. suaveolens was a mixed gland composed of serous and mucous acinar cells. Secretory granules from the acini were discharged through the salivary ducts into the oral cavity. Serous and mucous acinar cells had well developed rough endoplasmic reticulum and mitochondria and large amount of granules. In case of serous acinar granules, an immature granule was formless and had only dense specks, and a matured granule was a complete round type delimiting by a single membrane and had a homogeneous dense center with dense specks on the border. In case of mucous acinar granules, while an immature granule was a round type and had an only homogeneous matrix and an indistinct limiting membrane, a mature granule was an even round type having a variety of pattern with several dense bands into the homogeneous matrix and had a distinct membrane. Therefore, a mature mucous acinar granule of C. suaveolens was not only distinct from those of the other mammalian species to have a variety of pattern but also from those of C. lasiura to have an even round type. A great serous-like secretory granules and Myelin-like body were observed in the cytoplasm and lumen of granular duct cells. Myelin-like body, a characteristic structure only reported in salivary gland of three shrews, was discharged from secretory cell into lumen by the manner of exocytosis which has little differences from discharging manner of secretory granules.

• Applied Microscopy
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• v.35 no.2
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• pp.57-64
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• 2005

The ultrastructure and histochemical characteristics of the submandibular gland was examined in the big white-toothed shrew, Crocidura lasiura. A submandibular gland of Crocidura lasiura was a mixed gland composed of serous and mucous acinar cells. Secretory granules from the acini were discharged through the intercalated duct, the granular duct and the striated duct into the oral cavity. Serous and mucous acinar cells and granular duct cells had large amount of rough endoplasmic reticulum, free ribosome and prominent Golgi apparatus at the basal cytoplasm of the cell, and many granules at the apical cytoplasm. Oval type serous granules had a homogeneously pale round shape of bead at the center. Mucous granules were distinct from those of the other mammalian species having variety patterns with several dense bands into homogeneous pale matrix. A serous-like secretory granules and myelin-like body were observed in the cytoplasm and the lumen of granular duct cells. The myelin-like body is a characteristic structure only reported in the salivary glands of two shrews, Suncus murinus and C. dsinezumi. Striated duct cell had numerous well-developed mitochondria but secretory granule was not shown at all.

• Journal of Veterinary Clinics
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• v.26 no.6
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• pp.586-590
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• 2009

Horsehair worms (Chordodes koreensis) develop as parasites in the bodies of grasshoppers, crickets, cockroaches, and some beetles. Chordodes koreensis is an accidental parasite of humans, livestock, or pets and poses no public health threat. The male of Chordodes koreensis in the later larval stage from canine vomitus was investigated by the scanning and transmission electron microscopy. In cross sections, the body wall is composed of four components namely epicuticle, cuticle, epidermis, and muscle layers. The parenchymal tissue fills the rest of the body and surrounds the visceral organs such as intestine, and ventral nerve cord but testes were not found. The epicuticle is a thin superficial layer whose surface shows rows of polygonal elevations called areoles. The cuticle has 17 layers of collagenous fibers spirally wound about the long axis of the worm. The section through the cuticle reveals the layers of large fibers cut obliquely lengthwise, alternating with layers of fibers sectioned obliquely crosswise. The layers of large fiber formed a double helix about longitudinal axis of the worm. The epidermis is a single layer. The muscles were interrupted by the nervous lamella in the only midventral portion. The medulla of muscle plate is composed of lightly stained cytoplasm, mitochondria, weakly developed endoplasmic reticulum, and glycogen granules. Between the medulla of a cell and the plasmalemma lies a broad cortical zone of myofilaments. The circular muscles are absent. The characteristic feature of the cytoplasm is that there was no content in peripheral mesenchyme, but was an abundance of large clear vacuoles which give the cytosome a foamy appearance. The nucleus of mesenchyme is not easily identified in our specimens.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.33 no.5
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• pp.403-407
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• 2000

Ultrastructure and function of testis somatic cells in freshwater prawns Macrobrachium nipponense were studied. The paired testes of the prawn were elongated, united at their anterior end, which lay between the dorsal surface of the hepatopancreas and the heart. Each testis consisted of a large number of seminiferous cords compactly held together by connective tissue. A seminiferous cord was composed of an outer layer of simple squamous epithelium, a basement membrane, the closely packed germ cells and sustentacular cells of the germinal ridge, and an inner layer of simple cuboidal epithelial cells. Leydig cell-like cells in an angular areas filling the space of the seminiferous cords were observed. The nuclei of leydig cell-like cells were characterized by a distinct nucleolus. The simple squamous epithelial layer was composed of flattened cells tying on a basement membrane. The nuclei of the flattened cells were often overlapped in a layer, and the cytoplasm of the cells was observed just near the nuclei. The sustentacular cells were complex in morphology. These cells had relatively small cell bodies from which long cytoplasmic extensions ramified reached the space of germ cells in the germinal ridge. The nuclei of sustentacular cells usually exhibited angular profiles and located most commonly at the periphery of the cords. Cells of simple cuboidal epithelium located between germinal ridge and lumen of seminiferous cord, and part of the cells were adjacent to basal lamina, The cuboidal epithelial cells contained numerous mitochondria, the well-developed rER, the well-developed Golgi complex, and irregularly shaped nuclei. Transition vesicles appeared on the cis side of the Golgi complex. The large vesicles on the trans side of the complex appeared to fuse to form a membrane-bound structure. A number of pits on the cell apex suggested exocytotic activity for secretion of the sperm supporting matrix.