• Title/Summary/Keyword: Cell Disruption

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Inhibition of Langerhans cell function by UVB radiation

  • Okamoto, Hiroyuki;Mizuno, Kana;Horio, Takeshi
    • Journal of Photoscience
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    • v.9 no.2
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    • pp.190-193
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    • 2002
  • The functional disruption of Langerhans cells (LC) by UVB radiation is involved in antigen-specific immunosuppression of contact hypersensitivity. We tested whether UVB radiation inhibits the endocytotic activity of LC, which leads to impaired subsequent migration and maturation. Human monocyte-derived LC that took up lucifer yellow (L Y) or FITC-dextran (Fd) exclusively migrated in response to 6Ckine and matured. Exposing LC to 10-40 mJ/cm$^2$ of UVB radiation reduced their endocytotic activity in fluid phase pinocytosis (measured by uptake of LY) and in receptor-mediated endocytosis (measured by uptake of Fd). Membrane ruffling and CD32 expression were also suppressed by UVB radiation. UVB-irradiated, endocytosing LC had less movement towards 6Ckine, expressed less CD54 and CD86, and had less effective stimulatory activity in allo-MLR than nonirradiated, endocytosing LC. Endocytosis up-regulated TNF-$\alpha$ production by LC, but prior UVB radiation inhibited this enhancement. The finding that impaired endocytosis of LC by UVB radiation inhibits subsequent migration and maturation was also confirmed in murine epidermal cells obtained from unirradiated and 2OmJ/cm$^2$ of UVB-irradiated skin.

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A Preparative Purification Process for Recombinant Hepatitis B Core Antigen Using Online Capture by Expanded Bed Adsorption Followed by Size-Exclusion Chromatography

  • Ho, Chin Woi;Tan, Wen Siang;Chong, Fui Chin;Ling, Tau Chuan;Tey, Beng Ti
    • Journal of Microbiology and Biotechnology
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    • v.19 no.4
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    • pp.416-423
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    • 2009
  • Hepatitis B core antigen(HBcAg) is an important serological marker used in the diagnosis of hepatitis B virus(HBV) infections. In the current study, a fast and efficient preparative purification protocol for truncated HBcAg from Escherichia coli disruptate was developed. The recombinant HBcAg was first captured by anion exchange expanded bed adsorption chromatography integrated with a cell disruption process. This online capture process has shortened the process time and eliminated the "hold-up" period that may be detrimental to the quality of target protein. The eluted product from the expanded bed adsorption chromatography was subsequently purified using size-exclusion chromatography. The results showed that this novel purification protocol achieved a recovery yield of 45.1% with a product purity of 88.2%, which corresponds to a purification factor of 4.5. The recovered HBcAg is still biologically active as shown by ELISA test.

Development and Validation of the Custom Human cDNA Microarray (KISTCHIP-400) for Monitoring Expression of Genes involved in Hormone Disruption

  • Kim, Youn-Jung;Chang, Suk-Tai;Yun, Hye-Jung;Jeon, Hee-Kyung;Ryu, Jae-Chun
    • Proceedings of the Korea Society of Environmental Toocicology Conference
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    • 2003.05a
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    • pp.180-180
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    • 2003
  • Transcript profiling is a particularly valuable tool in the field of steroid receptor biology, as these receptors are ligand-activated transcription factors and therefore exert their initial effects through altering gene expression in responsive cells. Also, an increased awareness of endocrine disrupting chemicals (EBCs) and their potential to affect wildlife and humans has produced a demand for practical screening methods to identify endocrine activity. Here we developed an in-house cDNA microarray, named KISTCHIP-400, with 401 clones, hormone related genes, factors, and ESTs, based on public database and research papers. Theses clones contained estrogen, androgen, thyroid hormone St receptors, sex hormone signal transduction & regulation, c-fos, c-myc, ps2 gene, metabolism related genes etc. And to validate the KISTCHIP-400, we investigated gene expression profiles with reference hormones, 10$\^$-8/ M 17be1a-estradiol, 10$\^$-7/ M testosterone, 10$\^$-7/ M progesterone, and thyroxin in MCF-7 cell line. Although it is in first step of validation, low doses and combinations of EDCs need to be tested. Our preliminary results that indicate the developed microarray may be a useful laboratory tool for screening EDCs and elucidating endocrine disrupting mechanism.

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Improvement of Photoheterotrophic Hydrogen Production of Rhodobacter sphaeroides by Removal of B800-850 Light-Harvesting Complex

  • KIM EUI-JIN;YOO SANG-BAE;KIM MI-SUN;LEE JEONG K.
    • Journal of Microbiology and Biotechnology
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    • v.15 no.5
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    • pp.1115-1119
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    • 2005
  • The photoheterotrophic $H_2$ production of Rhodobacter sphaeroides was significantly increased through disruption of the genes coding for uptake hydrogenase and poly-${\beta}$-hydroxybutyrate (PHB) synthase (Lee et al., Appl. Microbiol. Biotechnol. 60: 147-153, 2002). In this work, we further removed the B800-850 light-harvesting (LH) complex from the strain and found an increase in $H_2$ production at the light-saturating cell growth (${\ge}10$ Watts $[W]/m^2$). Neither the mutant nor the wild-type produced more $H_2$ at the brighter light. Accordingly, light does not appear to be limited for the $H_2$ production by the presence of B800-850. However, increase in the level of the spectral complexes resulted in decrease of $H_2$ production. Thus, although the B875 is essential for light harvesting, the consumption of cellular energy for the synthesis of B800-850 and the surplus LH complexes may reduce the energy flow into the $H_2$ production of R. sphaeroides.

Fertilization Processes in Porcine Oocytes Following Intracytoplasmic Injection of Porcine and Mouse Spermatozoa

  • Lee, Youn-Jeung;Kim, Bong-Ki;Park, Chang-Sik;Kim, Nam-Hyung
    • Proceedings of the KSAR Conference
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    • 2001.03a
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    • pp.12-12
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    • 2001
  • To get insight into the nature of foreign mitochondria and syngamy during mammalian fertilization we compared fertilization processes in porcine oocytes following microinjection of porcine or mouse spermatozoa. Pronuclear movement, sperm mitochondria, and DNA synthesis were imaged with propidium iodide, mitotracker, and BrdU under confocal laser scanning microscope. Intracytoplasmic injection of either porcine or mouse spermatzoon activated porcine oocytes without additional parthengenetic stimulation. Foreign mitochondria in either mouse or porcine sperm midpiece were introduced into porcine oocytes following sperm injection, but rapidly disappeared from the actively developing porcine oocytes. BrdU experiment showed new DNA synthesis in porcine oocytes following injection of mouse spermatozoon or sperm head. At 24 h after injection of mouse isolated sperm head or a spermatozoon, mitoic metaphase was seen in oocyte, but they did not go to normal cell division (Table). These results suggest that pronuclear formation, foreign mitochondria disruption, DNA synthesis and syngamy formation during fertilization are not species specific processes.(Table Omitted).

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Gene Cloning and Expression of Cephalosporin-C Deacetylase from Bacillus sp. KCCM10143

  • Choi, Duk-Ho;Kim, Young-Duk;Chung, Il-Sun;Lee, Sang-Hun;Kang, Sang-Mo;Kwon, Tae-Jon;Han, Kum-Soo
    • Journal of Microbiology and Biotechnology
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    • v.10 no.2
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    • pp.221-226
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    • 2000
  • Cephalosporin-C deacetylase (CAH) catalyzes the deacetylation of cephalosporin derivatives. A novel gene encoding the CAH from Bacillus sp. KCCM10143 was cloned and sepuenced. The uncleotide sequence contained an open reading frame encoding a polypeptide consisting of 217 amino acids and a molecular weight of 24 kDa which was in good agreement with the value obtained by sodium dodecylsulfate-polyacrylamide gel electrophoresis. An expression plasmid was constructed by inserting the CAH gene into the region of the pTrc99A expression vector. An active from of the CAH protein was expressed in the soluble fraction obtained after cell disruption. in fermentation using a 5-1 jar fementer, the transformant E. coli JM109 (pDST654) produced 4.12 U of CAH per ml of culture during 16 h of incubation.

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The Nuclear Orphan Receptor NR4A1 is Involved in the Apoptotic Pathway Induced by LPS and Simvastatin in RAW 264.7 Macrophages

  • Kim, Yong Chan;Song, Seok Bean;Lee, Sang Kyu;Park, Sang Min;Kim, Young Sang
    • IMMUNE NETWORK
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    • v.14 no.2
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    • pp.116-122
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    • 2014
  • Macrophage death plays a role in several physiological and inflammatory pathologies such as sepsis and arthritis. In our previous work, we showed that simvastatin triggers cell death in LPS-activated RAW 264.7 mouse macrophage cells through both caspase-dependent and independent apoptotic pathways. Here, we show that the nuclear orphan receptor NR4A1 is involved in a caspase-independent apoptotic process induced by LPS and simvastatin. Simvastatin-induced NR4A1 expression in RAW 264.7 macrophages and ectopic expression of a dominant-negative mutant form of NR4A1 effectively suppressed both DNA fragmentation and the disruption of mitochondrial membrane potential (MMP) during LPS- and simvastatin-induced apoptosis. Furthermore, apoptosis was accompanied by Bcl-2-associated X protein (Bax) translocation to the mitochondria. Our findings suggest that NR4A1 expression and mitochondrial translocation of Bax are related to simvastatin-induced apoptosis in LPS-activated RAW 264.7 macrophages.

Calcifying Epithelioma of Malherbe(Pilomatricoma) in Children (소아의 Malherbe씨 석회화 상피종(Pilomatricoma))

  • Nam, Jung-Hyun;Jung, Poong-Man
    • Advances in pediatric surgery
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    • v.2 no.2
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    • pp.88-93
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    • 1996
  • A clinical analysis was made of 42 pilomatricoma in 34 children treated, from January 1980 to August 1996 at the Department of Pediatric Surgery, Hanyang University. The gender ratio was 13 to 21 with female preponderance. The average age at presentation was 7 years 3 months(range 8 months to 19 years old). The time average interval from onset to excision was 9.8 months. The predilection site was neck, upper extremities, face, trunk and lower extremities in the decreasing order of frequency. The masses were less than 3cm in diameter and there was no correlation between age at presentation and size of the mass. Of the six children with multiple lesions, two presented with multiple lesions initially and four presented with second lesion metachronously after excision of the primary lesions. Two girls had cysticercosis and von Recklinghausen's disease respectively in association with a pilomatricoma. All lesions were encapsulated subcutaneous masses which were clearly distinguished from surrounding tissue, and 4 of them had pigmentation, ulceration, skin disruption and hemorrhagic changes in the overlying skin. Microscopically the mass was composed of basophilic cells and eosinophilic shadow cells. Calcification, foreign body giant cell, ossification, focal cystic, inflammatory or necrotic changes were also seen.

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Lung interstitial cells during alveolarization

  • Choi, Chang-Won
    • Clinical and Experimental Pediatrics
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    • v.53 no.12
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    • pp.979-984
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    • 2010
  • Recent progress in neonatal medicine has enabled survival of many extremely low-birth-weight infants. Prenatal steroids, surfactants, and non-invasive ventilation have helped reduce the incidence of the classical form of bronchopulmonary dysplasia characterized by marked fibrosis and emphysema. However, a new form of bronchopulmonary dysplasia marked by arrest of alveolarization remains a complication in the postnatal course of extremely low-birth-weight infants. To better understand this challenging complication, detailed alveolarization mechanisms should be delineated. Proper alveolarization involves the temporal and spatial coordination of a number of cells, mediators, and genes. Cross-talk between the mesenchyme and the epithelium through soluble and diffusible factors are key processes of alveolarization. Lung interstitial cells derived from the mesenchyme play a crucial role in alveolarization. Peak alveolar formation coincides with intense lung interstitial cell proliferation. Myofibroblasts are essential for secondary septation, a critical process of alveolarization, and localize to the front lines of alveologenesis. The differentiation and migration of myofibroblasts are strictly controlled by various mediators and genes. Disruption of this finely controlled mechanism leads to abnormal alveolarization. Since arrest in alveolarization is a hallmark of a new form of bronchopulmonary dysplasia, knowledge regarding the role of lung interstitial cells during alveolarization and their control mechanism will enable us to find more specific therapeutic strategies for bronchopulmonary dysplasia. In this review, the role of lung interstitial cells during alveolarization and control mechanisms of their differentiation and migration will be discussed.

Molecular Detection of Verotoxigenic Escherichia coli (VTEC) from Animal Feces for Screening VTEC-shedders

  • Kobayashi, Y.;Sato, M.;Taguchi, H.;Koike, S.;Nakatsuji, H.;Tanaka, K.
    • Asian-Australasian Journal of Animal Sciences
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    • v.17 no.3
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    • pp.423-427
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    • 2004
  • Seventy-six animals including cattle, sheep, horses, 6 species of zoo animals were employed for collection of fresh feces in order to detect verotoxigenic Esherichia coli (VTEC) by safe, quick and sensitive PCR-based molecular methods. Bacterial cell disruption with bead-beating followed by bacterial DNA purification with hydroxyapatide chromatography and gel filtration allowed DNA preparation from animal feces with high recovery and purity. A mountain goat was firstly shown by PCR and sequencing to shed verotoxin 2 gene (vt2) that was used to generate vt2 probe and second primer set for nested PCR to attempt more sensitive detection. Most sensitive nested PCR revealed that 45% of tested cattle and 47% of tested zoo animals were VTEC-positive, while least sensitive normal PCR detected VTEC from none of these animals except a mountain goat. Moderately sensitive detection by PCR in combination with hybridization suggested that the VTEC density varied between the VTEC-positive cattle.