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Murrayafoline A Induces a G0/G1-Phase Arrest in Platelet-Derived Growth Factor-Stimulated Vascular Smooth Muscle Cells

  • Han, Joo-Hui;Kim, Yohan;Jung, Sang-Hyuk;Lee, Jung-Jin;Park, Hyun-Soo;Song, Gyu-Yong;Nguyen, Manh Cuong;Kim, Young Ho;Myung, Chang-Seon
    • The Korean Journal of Physiology and Pharmacology
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    • v.19 no.5
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    • pp.421-426
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    • 2015
  • The increased potential for vascular smooth muscle cell (VSMC) growth is a key abnormality in the development of atherosclerosis and post-angioplasty restenosis. Abnormally high activity of platelet-derived growth factor (PDGF) is believed to play a central role in the etiology of these pathophysiological situations. Here, we investigated the anti-proliferative effects and possible mechanism(s) of murrayafoline A, a carbazole alkaloid isolated from Glycosmis stenocarpa Guillamin (Rutaceae), on PDGF-BB-stimulated VSMCs. Murrayafoline A inhibited the PDGF-BB-stimulated proliferation of VSMCs in a concentration-dependent manner, as measured using a non-radioactive colorimetric WST-1 assay and direct cell counting. Furthermore, murrayafoline A suppressed the PDGF-BB-stimulated progression through $G_0/G_1$ to S phase of the cell cycle, as measured by [$^3H$]-thymidine incorporation assay and cell cycle progression analysis. This anti-proliferative action of murrayafoline A, arresting cell cycle progression at $G_0/G_1$ phase in PDGF-BB-stimulated VSMCs, was mediated via down-regulation of the expression of cyclin D1, cyclin E, cyclin-dependent kinase (CDK)2, CDK4, and proliferating cell nuclear antigen (PCNA), and the phosphorylation of retinoblastoma protein (pRb). These results indicate that murrayafoline A may be useful in preventing the progression of vascular complications such as restenosis after percutaneous transluminal coronary angioplasty and atherosclerosis.

An experimental study of particle deposition from high temperature gas-particle flows (고온의 기체 입자 유동으로부터 입자부착 현상에 관한 실험적 연구)

  • 김상수;김용진
    • Transactions of the Korean Society of Mechanical Engineers
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    • v.11 no.3
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    • pp.501-508
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    • 1987
  • Experimental studies of particle (TiO$\_$2/) deposition from the laminar hot gas particle flow (about 1565K) onto the cold wall surface (about 1215K-1530K) were carried out by the 'real time' laser light reflectivity method (LLRM) and the photographs of scanning electron microscope(SEM). The LLRM was used for the measurement of thermophoretic deposition rates of small particles (d$\_$p/<3.mu.m), and the photographs of SEM were used for determining what factors control the collection of particles having diameters ranging from 0.2 to 30 microns. Two phenomena are primarily responsible for transport of the particles across the laminar boundary layers and deposition: (1) particle thermophoresis (i.e. particles migration down a temperature gradient), and (2) particle inertial impaction, the former effect being especially larger factor of the particle deposition in its size over the range of 0.2 to 1 microns. And also, this study indicates that thermophoresis can be important for particles as large as 15 microns. Beyond d$\_$p/=16.mu.m, this effect diminishes and the inertial impaction is taken into account as a dominant mechanism of particle deposition. The results of present experiments found to be in close agreement with existing theories.

Modification of Efficient Vitrification Method by Using Open Pulled Straw (OPS) and EM Grid as Vehicles in Human Embryonic Stem Cell (인간 배아 줄기세포의 OPS와 Grid를 이용한 유리화 동결법의 효율성 비교)

  • 박규형;최성준;김희선;오선경;문신용;차광렬;정형민
    • Journal of Embryo Transfer
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    • v.18 no.3
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    • pp.179-186
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    • 2003
  • Human embryonic stem (hES) cell lines have been derived from human blastocysts and are expected to have far-reaching applications in regenerative medicine. The objective of this study is to improve freezing method with less cryo-injuries and best survival rates in hES cells by comparing various vitrification conditions. For the vitrifications, ES cells are exposed to the 4 different cryoprotectants, ethylene glycol (EG), 1,2-propanediol (PROH), EG with dime-thylsulfoxide (DMSO) and EG with PROH. We compared to types of vehicles, such as open pulled straw (OPS) or electron microscopic cooper grids (EM grids). Thawed hES cells were dipped into sequentially holding media with 0.2 M sucrose for 1 min, 0.1 M sucrose for 5 min and holding media for 5 min twice and plated onto a fresh feeder layer. Survival rates of vitrified hES cells were assessed by counting of undifferentiated colonies. It shows high survival rates of hES cells frozen with EG and DMSO (60.8%), or EG and PROH(65.8%) on EM grids better than those of OPS, compared to those frozen with EG alone (2.4%) or PROH alone (0%) alone. The hES cells vitrified with EM grid showed relatively constant colony forming efficiency and survival rates, compared to those of unverified hES cells. The vitrified hES cells retained the normal morphology, alkaline phosphates activity, and the expression of SSEA-3 and 4. Through RT-PCR analysis showed Oct-4 gene expression was down-regulated and embryonic germ layer markers were up-regulated in the vitrified hES cells during spontaneous differentiation. These results show that vitrification method by using EM grid supplemented with EG and PROH in hES cells may be most efficient at present to minimize cyto-toxicity and cellular damage derived by ice crystal formation and furthermore may be employed for clinical application.

Recent Development of Rapid and Automation Technology for Food Microbiological Examination

  • Hiroshi Kurata
    • Proceedings of the Korean Society of Food Hygiene and Safety Conference
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    • 1996.06a
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    • pp.33-33
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    • 1996
  • Interests in the field of rapid methods and automation in microbiology have been growing steadily on an international scale in recent years. International meetings concerned this problem have been held in elsewhere in the world countries since the past twenty years. But, unfortunately in the field of microbial examination in food hygiene, this problem have not yet been developed so much as in the field of clinical microbiology. Today, I would like to introduce you here present aspects of rapid and automation technologies, those which are manly carrying in milk and meats industries. My illustration will be given recent improved technologies using automatic apparatus and instruments along with process of microbial count procedure. Recent direct microbiological counting system (ChemeScan \ulcorner) as real time ultrasensitive analysis created by Cheminex Ltd., France is now most evolutional instrument to provide direct microbial counts, down to one cell, within 30 minutes. The results from these evaluations how a good correlation between the ChemScan system and the standard plate count method. This system will be successful application for not only in the field of pharmacology but also food microbiology. In addition, current identification of microbes by sophisticated instruments suitable for food microbiology, one of which Biology is manual system (BIOLOG\ulcorner), provides reference-level capability at a modes price. For the manual system, the color reactions in the microplate are read by eye and manually keyed into personal computer. Species identification appears on the computer screen within seconds, along with biotype patterns, a list of closely related species, and other useful statistics. In present this is useful application for microbial ecology and epidemiological survey. RiboPrinter system newly produced by DuPont is now focusing among microbiologists in the world, and is one of the biggest microbial characterization system using a DNA-based approach. The technology analyzer is bacterial culture for its genetic fingerprint or riboprint pattern. Finally Bio-cellTracer system for automatic measurement of fungal growth and Fukitori-Maseter, a Surface Hygiene Monitoring Kit by using swabe procedure in food processing environment are briefly illustrated in this presentation.

Improvement of Hong-Ou-Mandel Interference Visibility by Using a Single-Mode Optical-Fiber Photon Collector (단일모드 광섬유 집광기를 이용한 Hong-Ou-Mandel 간섭 가시도 향상)

  • Han, Sung-Wook;Kim, Heonoh;Seo, Joo Yeon;Kim, Myung-Whun
    • New Physics: Sae Mulli
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    • v.68 no.12
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    • pp.1374-1377
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    • 2018
  • We achieved 95% visibility in the Hong-Ou-Mandel interference experiment while we achieved only 56% visibility in a previous report. We used a 120 mW 405 nm single-mode continuous wave laser, a 5-mm-thick type-1 ${\beta}$-barium borate single crystal, standard Hong-Ou-Mandel interferometer optics, two avalanche photodiode single-photon counters, and a homemade coincidence counting unit. The photon collection unit was the key difference between the present study and the previous study. In the present experiment, we used single-mode optical fibers for photon collection, which suppressed accidental coincidence between-different mode photons by acting as a spatial filter because of its core size being much smaller than a multi-mode fiber.

Activating transcription factor 4 aggravates angiotensin II-induced cell dysfunction in human vascular aortic smooth muscle cells via transcriptionally activating fibroblast growth factor 21

  • Tao, Ke;Li, Ming;Gu, Xuefeng;Wang, Ming;Qian, Tianwei;Hu, Lijun;Li, Jiang
    • The Korean Journal of Physiology and Pharmacology
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    • v.26 no.5
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    • pp.347-355
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    • 2022
  • Abdominal aortic aneurysm (AAA) is a life-threatening disorder worldwide. Fibroblast growth factor 21 (FGF21) was shown to display a high level in the plasma of patients with AAA; however, its detailed functions underlying AAA pathogenesis are unclear. An in vitro AAA model was established in human aortic vascular smooth muscle cells (HASMCs) by angiotensin II (Ang-II) stimulation. Cell counting kit-8, wound healing, and Transwell assays were utilized for measuring cell proliferation and migration. RT-qPCR was used for detecting mRNA expression of FGF21 and activating transcription factor 4 (ATF4). Western blotting was utilized for assessing protein levels of FGF21, ATF4, and markers for the contractile phenotype of HASMCs. ChIP and luciferase reporter assays were implemented for identifying the binding relation between AFT4 and FGF21 promoters. FGF21 and ATF4 were both upregulated in Ang-II-treated HASMCs. Knocking down FGF21 attenuated Ang-II-induced proliferation, migration, and phenotype switch of HASMCs. ATF4 activated FGF21 transcription by binding to its promoter. FGF21 overexpression reversed AFT4 silencing-mediated inhibition of cell proliferation, migration, and phenotype switch. ATF4 transcriptionally upregulates FGF21 to promote the proliferation, migration, and phenotype switch of Ang-II-treated HASMCs.

Circ-SNX27 sponging miR-375/RPN1 axis contributes to hepatocellular carcinoma progression

  • Chao Zheng;Jin Liang;Shoude Yu;Hua Xu;Lin Dai;Dan Xu
    • The Korean Journal of Physiology and Pharmacology
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    • v.27 no.4
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    • pp.333-344
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    • 2023
  • Hepatocellular carcinoma (HCC) is a prevalent malignant tumor with high fatality. It has yet to be reported whether circ-SNX27 can affect the progression of HCC. This study attempted to analyze circ-SNX27's precise role and underlying mechanisms in HCC. HCC cell lines and tumor specimens from HCC patients were analyzed using quantitative real-time PCR and Western blotting to quantify the expressions of circ-SNX27, miR-375, and ribophorin I (RPN1). Cell invasion and cell counting kit 8 experiments were conducted for the evaluation of HCC cell invasion and proliferation. Caspase-3 Activity Assay Kit was utilized to gauge the caspase-3 activity. Luciferase reporter and RNA immunoprecipitation assays were executed to ascertain the relationships among miR-375, circ-SNX27, and RPN1. To determine how circ-SNX27 knockdown affects the growth of HCC xenografts in vivo, tumor-bearing mouse models were constructed. Elevated expressions of circ-SNX27 and RPN1 as well as a reduced miR-375 expression were observed among HCC cells and HCC patient tumor specimens. Knocking-down circ-SNX27 in HCC cells abated their proliferative and invasive abilities but raised their caspase-3 activity. Moreover, the poor levels of circ-SNX27 inhibited HCC tumor growth among the mice. Circ-SNX27 enhanced RPN1 by competitively binding with miR-375. Silencing miR-375 in HCC cells promoted their malignant phenotypes. Nonetheless, the promotive effect of miR375 silencing was reversible via the knockdown of circ-SNX27 or RPN1. This research demonstrated that circ-SNX27 accelerated the progression of HCC by modulating the miR-375/RPN1 axis. This is indicative of circ-SNX27's potential as a target for the treatment of HCC.

Knockdown of GCF2/LRRFIP1 by RNAi Causes Cell Growth Inhibition and Increased Apoptosis in Human Hepatoma HepG2 Cells

  • Li, Jing-Ping;Cao, Nai-Xia;Jiang, Ri-Ting;He, Shao-Jian;Huang, Tian-Ming;Wu, Bo;Chen, De-Feng;Ma, Ping;Chen, Li;Zhou, Su-Fang;Xie, Xiao-Xun;Luo, Guo-Rong
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.6
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    • pp.2753-2758
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    • 2014
  • Background: GC-binding factor 2 (GCF2) is a transcriptional regulator that represses transcriptional activity of the epidermal growth factor receptor (EGFR) by binding to a specific GC-rich sequence in the EGFR gene promoter. In addition to this function, GCF2 has also been identified as a tumor-associated antigen and regarded as a potentially valuable serum biomarker for early human hepatocellular carcinoma (HCC) diagnosis. GCF2 is high expressed in most HCC tissues and cell lines including HepG2. This study focused on the influence of GCF2 on cell proliferation and apoptosis in HepG2 cells. Materials and Methods: GCF2 expression at both mRNA and protein levels in HepG2 cells was detected with reverse transcription (RT) PCR and Western blotting, respectively. RNA interference (RNAi) technology was used to knock down GCF2 mRNA and protein expression. Afterwards, cell viability was analyzed with a Cell Counting Kit-8 (CCK-8), and cell apoptosis and caspase 3 activity by flow cytometry and with a Caspase 3 Activity Kit, respectively. Results: Specific down-regulation of GCF2 expression caused cell growth inhibition, and increased apoptosis and caspase 3 activity in HepG2 cells. Conclusions: These primary results suggest that GCF2 may influence cell proliferation and apoptosis in HepG2 cells, and also provides a molecular basis for further investigation into the possible mechanism at proliferation and apoptosis in HCC.

Chemical Age Dating of Zircon and Monazite by E1ectron Microprobe (전자현미분석기를 이용한 저어콘 및 모나자이트의 화학적 연대 측정법)

  • 이석훈
    • The Journal of the Petrological Society of Korea
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    • v.10 no.3
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    • pp.179-189
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    • 2001
  • The determination of trace concentration of U, Th and Pb was carried out for chemical dating of zircon and monazite by electron microprobe. Detection limit and error range should be considered to measure characteristic X-rays of M-line from those minerals, which are low in the ionization of atom and low peak intensity in the spectrum. The element of U, Th and Pb were simultaneously measured with 3 spectrometers equipped with PET crystal to reduce a total counting time and error due to drift of instrumental operating condition. Detection limit could be improved from increase of the peak/background ratio through adjusting pulse height analyzer about 1000 mv baseline. Under permissible maximum analytical conditions, theoretical detection limit of U, Th and Pb is down to 30 ppm (99% confidence level). The analytical result was maintained at a relative error $\pm$10% ($2{\sigma}$) in 800 ppm Pb, $\pm$5% ($2{\sigma}$) in 2330 ppm U and $\pm$10% ($2{\sigma}$) in dating from a single measurement of zircon at 15 keV and 100 nA. However, for the precise dating of zircon and monazite, if it is considered a 3 $\mu\textrm{m}$ spatial resolution, <100 ppm ($3{\sigma}$) detection limit and <$\pm$10% ($2{\sigma}$) relative error, optimum analytical conditions are given as 15~20 keV accelerating voltage, 100~200 nA beam current and 300~1200 sec total counting time. To reduce material damage by high current, there is need to be up to 3~5 $\mu\textrm{m}$ of electron beam diameter, or to use arithmetic average of multiple measuring at a shorter counting time. A younger or relatively low concentration rocks can be dated chemically by lower detection limit and improved precision resulted from increase of current and measuring time.

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APEX-1 Regulates Cell Proliferation through GDNF/GFRα1 Signaling (APEX-1은 GDNF/GFRα1 시그널을 통해 세포증식을 조절한다)

  • Kim, Hong-Beum;Hariharasudhan, Gurusamy;Youn, Cha-Kyung
    • Journal of Life Science
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    • v.23 no.10
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    • pp.1183-1191
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    • 2013
  • Human apurinic/apyrimidinic endonuclease (APEX-1) is a multifunctional protein that is capable of repairing abasic sites and single-strand breaks in damaged DNA. In addition, it serves as a redox-modifying factor for a number of transcription factors. Identifying the transcriptional targets of APEX-1 is essential for understanding how it affects various cellular outcomes. Expression array analysis was used to identify glial cell-derived neurotropic factor receptor ${\alpha}1$ ($GFR{\alpha}1$), which is an encoding receptor for the glial cell-derived neurotropic factor (GDNF) family, the expression of which is induced by APEX-1. A target of GDNF/$GFR{\alpha}$ signaling, c-Src (Tyr418) was strongly phosphorylated by GNDF in the APEX-1 expressing cells. Moreover, GDNF initiated cell proliferation, measured by counting the number of cells, in the APEX-1 expressing cells. Importantly, the down-regulation of APEX-1 by siRNA caused a marked reduction in the $GFR{\alpha}1$ expression level, and it reduced the ability of GDNF to phosphorylate c-Src (Tyr418) and stimulate cell proliferation. These results demonstrate an association between APEX-1 and GDNF/$GFR{\alpha}$ signaling and suggest a potential molecular mechanism for the involvement of APEX-1 in cell survival and proliferation.