• Title/Summary/Keyword: Medium-term assay

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Chemopreventive Effects of Ginseng on Rat Carcinogenesis

  • Wanibuchi Hideki;Ichihara Toshio;Morimura Keiichirou;Fukushima Shoji
    • Proceedings of the Ginseng society Conference
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    • 2002.10a
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    • pp.277-287
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    • 2002
  • The chemopreventive effects of ginseng on rat carcinogenesis models were investigated, In the present study, the inhibitory effects of white and red ginseng on tumor development were examined using medium-term liver, initiation and medium-term multi-organ carcinogenicity bioassay systems. No modifying potential of the ginsengs was evident in terms of the numbers or areas of glutathione S-transferase placental form (GST -P)-positive foci, which is a marker of preneoplastic lesion in rat livers. However, white ginseng, but not red ginseng was found to decrease the incidences of adenocarcinoma of the small intestine and colon in the medium-term multi-organ carcinogenesis model. These results indicate that white ginseng may have inhibitory effects on progression stage of rat intestinal carcinogenesis, but the influence is not strong. Ginseng is unlikely to have promoting or inhibitory effects in other organs under the present type of experimental conditions. Possible application on ginseng for chemoprevention of colon cancer in humans, can be concluded given the lack of obvious adverse effects.

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Genetic polymorphism analysis of somatic embryo-derived plantlets of Cymbopogon flexuosus through RAPD assay

  • Bhattacharya, S.;Dey, T.;Bandopadhyay, T.K.;Ghosh, P.D.
    • Plant Biotechnology Reports
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    • v.2 no.4
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    • pp.245-252
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    • 2008
  • The genetic status of somatic embryo-derived plantlets of Cymbopogon flexuosus was examined by randomly amplified polymorphic DNA (RAPD) analysis. Auxins such as 2, 4-dichlorophenoxyacetic acid (2, 4-D) (1-4 mg/l) were used in Murashige and Skoog (MS) medium for induction of calli from rhizomatous explants of Cymbopogon flexuosus. Optimum calli were induced on MS medium supplemented with 2, 4-dichlorophenoxyacetic acid (2, 4-D) (3.5 mg/l) alone or in combination with $N^6-benzyladenine$ (2 mg/l). Somatic embryogenesis was achieved from long term calli when cultured on MS medium containing 2, 4-dichlorophenoxyacetic acid (2, 4-D) (2 mg/l) along with $N^6-benzyladenine$ (BA) (1-2 mg/l). Regeneration was achieved when freshly induced embryogenic calli were sub-cultured on MS medium supplemented with $N^6-benzyladenine$ (3 mg/l) alone. Long-term cultured embryos showed profuse minute rooting on regeneration medium supplemented with N6 -benzyladenine (3 mg/l). Microshoots were rooted in the presence of indole-butyric acid (IBA) (2 mg/l). DNA samples from the mother plant and 18 randomly selected regenerated plants from a single callus were subjected to RAPD analysis with 6 arbitrary decamer primers for the selection of putative somaclones. A total of 64 band positions were scored, out of which 19 RAPD bands were polymorphic. From genetic similarity coefficient based on RAPD band data sharing, it was found that the majority of the clones were almost identical or more than 92% similar to the mother plant, except CL2 and CL9 (66%) which showed highest degree of genetic change with CL2 and CL9 showing presence of two non-parental bands each.

MEDIUM-TERM LIVER BIOASSAY FOR RAPID DETECTION OF CARCINOGENS

  • Hasegawa, Ryohei;Shirai, Tomoyuki;Ito, Nobuyuki
    • Proceedings of the Korean Society of Applied Pharmacology
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    • 1993.04a
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    • pp.91-91
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    • 1993
  • The vast number of compounds which have been introduced into our environment in recent years is far beyond our capacity to assess for carcinogenicity using expensive, life-time carcinogenicity studies in rodents. Several in vitro short term assays which are very rapid and inexpensive have therefore been applied as mass screening methods. However, increasing evidence has indicated that mutagenicity results do not always correlate with long-term carcinogenicity. Therefore, the existence of discrepancies dictates the necessity of suitable in vivo rapid assay systems which could bridge the gap between mutagenicity and conventional long-term testing.

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Isolation and Characterization of Trophoblast Stem Cells-like Cells Derived from Human Term Placenta

  • Na, Kyu-Hwan;Shin, Kyung-Seon;Choi, Jong-Ho;Cha, Dong-Hyun;Kim, Gi-Jin
    • Development and Reproduction
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    • v.14 no.3
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    • pp.155-162
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    • 2010
  • The trophectoderm is one of the earliest cell types to differentiate in the forming placenta. It is an important for the initial implantation and placentation during pregnancy. Trophoblast stem cells (TBSCs) develop from the blastocyst and are maintained by signals emanating from the inner cell mass. However, several limitations including rarity and difficulty in isolation of trophoblast stem cells derived from blastocyst still exist. To establish a model for trophoblast differentiation, we isolated TBSCs from human term placenta ($\geq$38 weeks) and characterized. Cell cycle was analyzed by measuring DNA content by FACS analysis and phenotype of TBSCs was characterized by RT-PCR and FACS analysis. TBSCs have expressed various markers such as self-renewal markers (Nanog, Sox2), three germ layer markers (hNF68, alpha-cardiac actin, hAFP), trophoblast specific markers (CDX-2, CK7, HLA-G), and TERT gene. In FACS analysis, TBSCs isolated from term placenta showed that the majority of cells expressed CD13, CD44, CD90, CD95, CD105, HLA-ABC, cytokeratin 7, and HLA-G. Testing for CD31, CD34, CD45, CD71, vimentin and HLA-DR were negative. TBSCs were shown to decrease the growth rate when cultured in conditioned medium without FGF4/heparin as well as the morphology was changed to a characteristic giant cell with a large cytoplasm and nucleus. In invasion assay, TBSCs isolated from term placenta showed invasion activities in in vivo using nude mice and in vitro Matrigel system. Taken together, these results support that an isolation potential of TBSCs from term placenta as well as a good source for understanding of the infertility mechanism.

Expression Patterns of Cancer Stem Cell Markers During Specific Celecoxib Therapy in Multistep Rat Colon Carcinogenesis Bioassays

  • Salim, Elsayed I;Hegazi, Mona M;Kang, Jin Seok;Helmy, Hager M
    • Asian Pacific Journal of Cancer Prevention
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    • v.17 no.3
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    • pp.1023-1035
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    • 2016
  • The purpose of this study was to investigate the role of colon cancer stem cells (CSCs) during chemically-induced rat multi-step colon carcinogenesis with or without the treatment with a specific cyclooxygenase-2 inhibitor drug (celecoxib). Two experiments were performed, the first, a short term 12 week colon carcinogenesis bioassay in which only surrogate markers for colon cancer, aberrant crypt foci (ACF) lesions, were formed. The other experiment was a medium term colon cancer rat assay in which tumors had developed after 32 weeks. Treatment with celecoxib lowered the numbers of ACF, as well as the tumor volumes and multiplicities after 32 weeks. Immunohistochemical proliferating cell nuclear antigen (PCNA) labeling indexes LI (%) were downregulated after treatment by celecoxib. Also different cell surface antigens known to associate with CSCs such as the epithelial cell adhesion molecule (EpCAM), CD44 and CD133 were compared between the two experiments and showed differential expression patterns depending on the stage of carcinogenesis and treatment with celecoxib. Flow cytometric analysis demonstrated that the numbers of CD133 cells were increased in the colonic epithelium after 12 weeks while those of CD44 but not CD133 cells were increased after 32 weeks. Moreover, aldehyde dehydrogenase-1 activity levels in the colonic epithelium (a known CSC marker) detected by ELISA assay were found down-regulated after 12 weeks, but were up-regulated after 32 weeks. The data have also shown that the protective effect of celecoxib on these specific markers and populations of CSCs and on other molecular processes such as apoptosis targeted by this drug may vary depending on the genetic and phenotypic stages of carcinogenesis. Therefore, uncovering these distinction roles of CSCs during different phases of carcinogenesis and during specific treatment could be useful for targeted therapy.

Microbiological Stability Test of Biodisel (바이오디젤의 미생물 안정성 조사)

  • 박근형;정해영;채희정
    • Journal of the Korea Academia-Industrial cooperation Society
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    • v.4 no.4
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    • pp.387-390
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    • 2003
  • The microbiological stability was examined by detecting the growth of microorganisms in the closed bottles under anaerobic long-term storage at $25^{\circ}C$ and 35$^{\circ}C$. Microbial growth was examined by a microscope and total cell number on a plate medium was quantitatively measured. There was no observed microbial growth in biodiesel for 90 days.

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Effects Of $Interferon-{\gamma}$ On The Biological Activity Of Mouse Osteoblast MC3T3/E1 Cells In Culture (($Interferon-{\gamma}$)가 마우스 조골세포의 생물학적 활성에 미치는 영향에 관한 연구)

  • Lee, Kwan-Hoon;Kim, Jung-Geun;Chung, Chin-Hyung
    • Journal of Periodontal and Implant Science
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    • v.26 no.1
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    • pp.216-229
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    • 1996
  • Interferon(IFN) is a sort of glycoproteins that are produced by activated lymphocyte, monocyte and fibroblast. IFN has anti-viral effects, immuno-defensive mechanism and regulating properties to the several kinds of cells that includes affect on the bone formation and resorption. The effect of IFN on the osteoclast & other tissue cells has been studied in a number of researchers with the limited reports on the osteoblast. The purpose of this study was to evaluate the effects of IFN on the osteoblastic function. The MC3T3/El cell(Mouse osteoblast) was incubated in ${\alpha}-minimum$ essential medium containing 10% FBS. To detect the cytotoxic effect of $IFN-{\gamma}$ on osteoblast, the cells were cultured in 96-well plate to which $IFN-{\gamma}$ of various concentrations were added for 2 days. After staining with trypan blue, total cells and living cells were counted under microscope. To determine the activity of alkaline phosphataset(ALP), various concentrations of $IFN-{\gamma}$ were treated to culture medium, and biochemical assay was performed. $IFN-{\gamma}$ and $IFN-{\gamma}$ plus cycloheximide were added to culture medium separately and then ALP activity were determined. To detect the effect of the $IFN-{\gamma}$ on the bone formation of osteoblast, long-term culture was performed, and calcified nodule formation were observed using von Kossa's staining. After the addition of $IFN-{\gamma}$ with various concentrations to the medium, no cytotoxic effect of $IFN-{\gamma}$ was detected at any concentration. The significant increase in ALP activity of osteoblast were found the concentration of $IFN-{\gamma}$ 500-2500U/ml and the culture time of 24-48 hours respectively. The enhancement of ALP activity by $IFN-{\gamma}$ of osteoblast was decreased significantly by the treatment of cycloheximide. After long-term culture of osteoblast, the nodule formation was found to be increased in number and density by the addition of 500 U/ml $IFN-{\gamma}$. These results suggest that $IFN-{\gamma}$ was affected on the bone formation of osteoblast. Forthemore this kind of study or $IFN-{\gamma}$ to osteoblast will be held continuously.

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Effects of Mito-TEMPO on the survival of vitrified bovine blastocysts in vitro

  • Jeong, Jae-Hoon;Yang, Seul-Gi;Park, Hyo-Jin;Koo, Deog-Bon
    • Journal of Animal Reproduction and Biotechnology
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    • v.36 no.4
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    • pp.299-306
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    • 2021
  • Vitrification methods are commonly used for mammalian reproduction through the long-term storage of blastocyst produced in vitro. However, the survival and quality of embryos following vitrification are significantly low compared with blastocyst from in vitro production (IVP). This study evaluates that the survival of frozen-thawed bovine embryos was relevant to mitochondrial superoxide derived mitochondrial activity. Here we present supplementation of the cryopreservation medium with Mito-TEMPO (0.1 µM) induced a significant (p < 0.001; non-treated group: 56.8 ± 8.7%, reexpanded at 24 h vs Mito-TEMPO treated group: 77.5 ± 8.9%, re-expanded at 24 h) improvement in survival rate of cryopreserved-thawed bovine blastocyst. To confirm the quality of vitrified blastocyst after thawing, DNA fragmentation of survived embryos was examined by TUNEL assay. As a result, TUNEL positive cells rates of frozen-thawed embryos were lower in the Mito-TEMPO treated group (4.2 ± 1.4%) than the non-treated group (7.1 ± 3.5%). In addition, we investigated the intracellular ROS and mitochondrial specific superoxide production using DCF-DA and Mito-SOX staining in survived bovine embryos following vitrification depending on Mito-TEMPO treatment. As expected, intracellular ROS levels and superoxide production of vitrified blastocysts after cryopreservation were significantly reduced (p < 0.05) according to Mito-TEMPO supplement in freezing medium. Also, mitochondrial activity measured by MitoTracker Orange staining increased in the frozen-thawed embryos with Mito-TEMPO compared with non-treated group. These results indicate that the treatment of Mito-TEMPO during cryopreservation might induce reduction in DNA fragmentation and apoptosis-related ROS production, consequently increasing mitochondrial activation for developmental capacity of frozen-thawed embryos.

THE EVALUATION OF PERIODONTAL LIGAMENT CELLS OF RAT TEETH AFTER LOW-TEMPERATURE PRESERVATION UNDER HIGH PRESSURE (고압-저온 보관에 따른 쥐 치아 치주인대세포의 활성도 평가)

  • Chung, Jin-Ho;Kim, Jin;Choi, Seong-Ho;Kim, Eui-Seong;Park, Ji-Yong;Lee, Seung-Jong
    • Restorative Dentistry and Endodontics
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    • v.35 no.4
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    • pp.285-294
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    • 2010
  • The purpose of this study was to evaluate the viability of periodontal ligament cells of rat teeth after low-temperature preservation under high pressure by means of MTT assay, WST-1 assay. 12 teeth of Sprague-Dawley white female rats of 4 week-old were used for each group. Both side of the first and second maxillary molars were extracted as atraumatically as possible under tiletamine anesthesia. The experimental groups were group 1 (Immediate extraction), group 2 (Slow freezing under pressure of 3 MPa), group 3 (Slow freezing under pressure of 2 MPa), group 4 (Slow freezing under no additional pressure), group 5 (Rapid freezing in liquid nitrogen under pressure of 2 MPa), group 6 (Rapid freezing in liquid nitrogen under no additional pressure), group 7 (low-temperature preservation at $0^{\circ}C$ under pressure of 2 MPa), group 8 (low-temperature preservation at $0^{\circ}C$ under no additional pressure), group 9 (low-temperature preservation at $-5^{\circ}C$ under pressure of 90 MPa). F-medium and 10% DMSO were used as preservation medium and cryo-protectant. For cryo-preservation groups, thawing was performed in $37^{\circ}C$ water bath, then MTT assay, WST-1 assay were processed. One way ANOVA and Tukey HSD method were performed at the 95% level of confidence. The values of optical density obtained by MTT assay and WST-1 were divided by the values of eosin staining for tissue volume standardization. In both MTT and WST-1 assay, group 7 ($0^{\circ}C$/2 MPa) showed higher viability of periodontal ligament cells than other group (2-6, 8) and this was statistically significant (p < 0.05), but showed lower viability than group 1, immediate extraction group (no statistical significance). By the results of this study, low-temperature preservation at $0^{\circ}C$ under pressure of 2 MPa suggest the possibility for long term preservation of teeth.

THE EFFICACY OF PROGRAMMED CRYO-PRESERVATION UNDER PRESSURE IN RAT PERIODONTAL LIGAMENT CELLS (압력 저속 냉동 방법의 쥐 치아 치주인대세포 보존 효율 평가)

  • Lee, Young-Eun;Kim, Eui-Seong;Kim, Jin;Han, Seung-Hoon;Lee, Seung-Jong
    • Restorative Dentistry and Endodontics
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    • v.34 no.4
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    • pp.356-363
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    • 2009
  • The purpose of this study was to evaluate the viability of periodontal ligament cells in rat teeth using slow cryo-preservation method under pressure by means of MTT assay and WST-1 assay. Eighteen teeth of Sprague-Dawley white female rats of 4 week-old were used for each group. Both sides of the first and second maxillary molars were extracted as atraumatically as possible under Tiletamine anesthesia. The experimental groups were group 1 (Immediate control), group 2 (Cold preservation at $4^{\circ}C$for 1 week), group 3 (Slow freezing), group 4 (Slow freezing under pressure of 3 MPa). F-medium and 10% DMSO were used as preservation medium and cryo-protectant. For cryo-preservation groups, thawing was performed in $37^{\circ}C$water bath, then MTT assay and WST-1 assay were processed. One way ANOVA and Tukey method were performed at the 95% level of confidence. The values of optical density obtained by MTT assay and WST-1 were divided by the values of eosin staining for tissue volume standardization. In both MTT and WST-1 assay, group 4 showed significantly higher viability of periodontal ligament cells than group 2 and 3 (p < 0.05), but showed lower viability than immediate control group. By the results of this study, slow cryo-preservation method under pressure suggests the possibility for long term cryo-preservation of the teeth.