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Apoptosis and Development of Porcine Parthenogenetic Embryos Activated and Cultured in Different Condition (활성화 및 배양조건이 돼지 단위발생란의 발달 및 Apoptosis에 미치는 영향)

  • Hwang In-Sun;Seo Jin-Sung;Cheong Hee-Tae;Im Gi-Sun
    • Reproductive and Developmental Biology
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    • v.30 no.1
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    • pp.65-70
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    • 2006
  • This study investigated apoptosis and in vitro development of parthenogenetic preimplantation porcine embryos. In vitro matured oocytes for $42{\sim}44h$ were used. Apoptotic cell death was analyzed by using a terminal deoxynucleatidyl transferase mediated deoxyuridine 5-triphosphate nick-end tabling (TUNEL) assay. In experiment 1, oocytes were activated with two electric pulses (CH) of 1.2 kV/cm for $30{\mu}sec$ (E), E + 6-dimethylaminopurine (6-DMAP) or E + cycloheximide (CH) and cultured in PZM-3 under 5% $CO_2$ in air at $38.5^{\circ}C$. In experiment 2, oocytes were activated by E and cultured in PZM-3 or NCSU-23 under a gas atmosphere of 20% $O_2$ ($5%\;CO_2$, in air) or 5% $O_2$ $(5%\;CO_2,\;5%\;O_2\;90%\;N_2)\;at\;38.5^{\circ}C$. Oocytes activated with E+6-DMAP or E+CH showed higher blastocyst rates (36.3% and 32.5%) compared to E alone (27.7%). The frequency of apoptosis according to treatments were 5.3%, 7.7% and 7.1% respectively. Oocytes activated with E alone showed lower (P<0.05) frequency of apoptosis compared to other groups. In experiment 2, parthenotes cultured in PZM-3 showed slightly higher blastocyte rates (28.2% and 29.7%) compared to NCSU-23 (22.6% and 24.4%) regardless of atmosphere. Blastocysts generated in PZM-3 showed lower (P<0.05) apoptosis rate under 20% $O_2$ (9.2% vs 16.9%), whereas those in NCSU-23 had slightly lower apoptosis rate under 5% $O_2$ (14.0% vs 18.4%). This result represents that activation method and culture condition could affect the frequency of apoptosis as well as in vitro developmental rate.

Effect of Fusion Procedure on the Development of Embryos Produced by Somatic Cell Nuclear Transfer in Hanwoo (Korean Cattle) (한우에서 융합방법이 체세포 핵이식 수정란의 발달에 미치는 영향)

  • Im, G.S.;Yang, B.S.;Park, S.J.;Chang, W.K.;Park, C.S.
    • Korean Journal of Animal Reproduction
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    • v.24 no.4
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    • pp.365-373
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    • 2000
  • The purpose of this study was to investigate the effects of the fusion pulses and fusion media on fusion rate and the development of embryos produced by somatic cell nuclear transfer in Hanwoo (Korean cattle). Nuclear donor cumulus and fetal fibroblast cells were cultured in Dulbecco's modified Eagle medium supplemented with 10% fetal bovine serum at 38.5$^{\circ}C$ in a humidified atmosphere of 5% $CO_2$in air. The in vitro matured oocytes were enucleated and then the isolated donor cells were introduced. The cumulus cell and cytoplast were fused using one pulse of 70 volts for 40$mutextrm{s}$, two pulses of 70 volts for 40$mutextrm{s}$ and one pulse of 180 volts for 15$mutextrm{s}$. The fetal fibroblast cell and cytoplast were fused using one pulse of 180 volts for 15$mutextrm{s}$ or 30$mutextrm{s}$. The cumulus cell and cytoplast were fused using mannitol and Zimmerman cell fusion medium (ZCFM) as a fusion medium. The fused embryos were activated after the fusion with 10 $\mu$M calcium ionophore for 5 min and 2 mM 6-dimethyl- aminopurine for 3 h. The nuclear transfer embryos were cultured in 500 ${mu}ell$ well of modified CR1aa supplemented with 3 mg/$m\ell$ BSA in th $\varepsilon$ four well dish cove red with mineral oil. After 3 days culture, culture medium was changed into modified CRlaa medium containing 1.5 mg/$m\ell$ BSA and 5% FBS for 4 days. The incubation environment was 5% $CO_2$, 5% $O_2$, 90% $N_2$ at 38.5$^{\circ}C$. When the cumulus cells were fused with enucleated oocytes by three different fusion pulses, one pulse of 180 volts for 15 $mutextrm{s}$ yielded the highest fusion rate and developmental rate to blastocyst among the pulses (P<0.05). When the fetal fibroblast cells were fused with enucleated oocytes, one pulse of 180 volts for 30$mutextrm{s}$ yielded significantly higher fusion rate compared with that for 15 $mutextrm{s}$(P<0.05). The present result indicates that the fusion rate between karyoplast and cytoplast was affected by the cell type and the optimal fusion condition was different according to cell type or size. When the fusion was conducted by the use of mannitol and ZCFM, the fusion rate was 71.2% and 65.8%, respectively. The developmental rates to blastocyst were 37.8% and 39.8%, respectively. There was no significant difference between two fusion media in the developmental rate of cumulus cell nuclear transfer embryos. These results indicate that optimal electric current should be selected according to cell type.

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Usefulness of $^{201}Tl$ Myocardial Perfusion SPECT in Prediction of Left Ventricular Remodeling following an Acute Myocardial Infarction (급성심근경색 후 발생하는 좌심실 재구도 예측에 대한 $^{201}Tl$ 심근관류 SPECT의 운용성)

  • Yoon, Seok-Nam;Park, C.H.;Hwang, Kyung-Hoon
    • The Korean Journal of Nuclear Medicine
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    • v.34 no.1
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    • pp.30-38
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    • 2000
  • Purpose: We investigated the role of myocardial perfusion SPECT in prediction of ventricular dilatation and the role of revascularization including thrombolytic therapy and PTCA in prevention of ventricular dilatation after an acute myocardial infarction (AMI). Materials and Methods: We performed dipyridamole stress, 4 hour redistribution, and 24 hour reinjection Tl-201 SPECT in 16 patients with AMI two to nine days after attack. Perfusion and wall motion abnormalities were quantified by perfusion index (PI) and wall motion index (WMI). Left ventricular ejection fraction (LVEF), WMI and ventricular volume were measured within 1 week of AMI and after average of 6 months. According to serial changes of left ventricular end-diastolic volume (LVEDV), patients were divided into two groups. We compared WMI, PI and LVEF between the two groups. Relationships among degree of volume, stress-rest PI, WMI, CKMB, Q wave, LVEF and revascularization were analysed using multivariate analysis. Results: Only initial rest perfusion index was significantly different between the two groups (p<0.05). While initial LVEF, stress PI, CKMB, trial of revascularization procedure, presence of Q wave and WMI were not significantly different between the two groups. Eight of 16 patients (50%) showed LV dilatation on follow-up echocardiography. Three of 3 patients (100%) who did not undergo revascualrization procedure documented LV dilatation. And only 5 (38%) of the remaining 13 patients who underwent revascularization revealed LV dilatation. There was no difference in infarct location between the two groups. By multivariate linear regression analysis in patients only undergoing revascularization, rest perfusion index was the only significant factor. Conclusion: Myocardial perfusion SPECT performed prior to revascularization was useful in prediction of LV dilatation after an AMI. Rest perfusion index on myocardial perfusion plays as a significant predictor of left ventricular dilatation after AMI. And revascularization appears to be a valuable procedure in alleviating LV dilatation after AMI with or without viable myocardium in a limited number of patients studied retrospectively.

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Eco-friendly remediation and reuse for coastal dredged materials using a bioaugmentation technology (생물증강법을 이용한 오염해양준설토의 환경친화적 정화 및 재활용)

  • Kim, In-Soo;Ha, Shin-Young;Koh, Sung-Cheol
    • Korean Journal of Microbiology
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    • v.51 no.4
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    • pp.374-381
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    • 2015
  • Occurrences of coastal dredged materials are ever increasing due to port construction, navigational course maintenance and dredging of polluted coastal sediments. Ocean dumping of the coastal dredged materials has become virtually prohibited as London Treaty will be enacted as of the year 2012. It will be necessary to treat and recycle the dredged materials that may carry organic pollutants and heavy metals in a reasonable and effective process: collection of the dredged materials, liquid and solid separation, and treatment of organic compounds and heavy metals. In this study we have developed a continuous bioreactor system that can treat a mixture of silt and particulate organic matter using a microbial consortium (BM-S-1). The steady-state operation conditions were: pH (7.4-7.5), temperature ($16^{\circ}C$), DO (7.5-7.9), and salt concentration (3.4-3.7%). The treatment efficiencies of SCOD, T-N and T-P of the mixture were 95-96%, 92-99%, and 79-97%. The system was also effective in removal of heavy metals such as Zn, Ni, and Cr. Levels of MLSS during three months operation period were 11,000-19,000 mg/L. Interestingly, there was little sludge generated during this period of operation. The augmented microbial consortium seemed to be quite active in the removal of the organic component (30%) present in the dredged material in association with indigenous bacteria. The dominant phyla in the treatment processes were Proteobacteria and Bacteroidetes while dominant genii were Marinobacterium, Flaviramulus, Formosa, Alteromonadaceae_uc, Flavobacteriaceae_uc. These results will contribute to a development of a successful bioremediation technology for various coastal and river sediments with a high content of organic matter, inorganic nutrients and heavy metals, leading to a successful reuse of the polluted dredged sediments.

Inverse Agonists at $A_1$ Adenosine Receptors in Rat Cerebral Cortex (흰쥐의 뇌의$A_1$ 아데노신 수용체에 작용하는 역효현제에 관한 연구)

  • Park, Kyung-Sun;Yang, Wan-Suk;Kim, Kyung-Hwan
    • The Korean Journal of Pharmacology
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    • v.32 no.1
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    • pp.23-29
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    • 1996
  • According to the traditional receptor model, competitive antagonists share with agonists the ability to bind to a common site on receptors, but they are different from agonist in that they cannot trigger the biological response-i.e., they lack intrinsic efficacy. Recent findings extend the model by indicating that not all antagonists display an intrinsic efficacy of zero but that some display 'inverse agonism'. In the present study we studied the inverse agonism at $A_1$ adenosine receptors in membranes prepared from rat cerebral cortex. Eight commercially available $A_1$ adenosine receptor antagonists (CGS-15943, ADPX, CPT, DPCPX, DPX, N-0840, PACPX and 8-PT) were screened for inverse agonism by measuring the extent of $[^{35}S]guanosine-5'-({\gamma}-thio)$ triphosphate $([^{35}S]GTP_{\gamma}S)$ binding to G proteins. The agonist-induced stimulation of $[^{35}S]GTP_{\gamma}S$ bindings was completely blocked in the presence of $A_1$ adenosine receptor antagonists. Under optimal conditions, two types of antagonists could be distinguished. Seven antagonists including DPCPX decreased the basal $[^{35}S]GTP_{\gamma}S$ binding in the absence of agonist, displaying inverse agonist activity. One (CGS-15943) had no effect on the basal bindings. N-ethylmaleimide treatment reduced the basal bindings as well as agonist-mediated stimulation of $[^{35}S]GTP_{\gamma}S$ bindings, indicating that a substantial amount of this binding reflects an activated state of the C proteins. In good agreement with these findings, 0.1 mM GTP decreased the apparent affinity of the receptors for the agonist PIA, increased that for DPCPX, and had no effect on that for CGS-15943.

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Le Lien de Causalit$\acute{e}$ et la Prescription Extinctive en Mati$\acute{e}$re de Responsabilit$\acute{e}$ M$\acute{e}$dicale en Droit Français (프랑스법상 의사책임분야(醫事責任分野)에서의 인과관계와 소멸시효론에 관한 소고)

  • Park, Soo-Gon
    • The Korean Society of Law and Medicine
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    • v.15 no.2
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    • pp.367-397
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    • 2014
  • Dans le contentieux m$\acute{e}$dical, il est difficile de prouver l'existence d'un lien de causalit$\acute{e}$ entre la faute m$\acute{e}$dicale et les pr$\acute{e}$judices subis par la victime. R$\acute{e}$partir bien de façon raisonnable la charge de la preuve du lien causal est alors une des questions pr$\acute{e}$occup$\acute{e}$es par la doctrine cor$\acute{e}$enne. La Cour supr$\hat{e}$me cor$\acute{e}$enne semble toutefois facilliter l'indemnisation des victimes dans les cas o$\grave{u}$ la responsabilit$\acute{e}$ du m$\acute{e}$decin a ${\acute{e}}t{\acute{e}}$ mise en cause, et cela en admettant des fois une solution op$\acute{e}$rant un renversement de la charge de la preuve du lien causal. Une telle attitude a ${\acute{e}}t{\acute{e}}$ m$\hat{e}$me affirm$\acute{e}$e dans un arr$\hat{e}$t rendu r$\acute{e}$cemment en cas de dommage caus$\acute{e}$ par le fait du produit de sant$\acute{e}$, notamment pour le cas de contamination virale par voie de transfusion. La Cour a $\acute{e}$galement reconnu que l'action se pr$\acute{e}$scrit $\grave{a}$ partir du moment de la consolidation du pr$\acute{e}$judice. Aux termes de cette $\acute{e}$tude, on pourra constater que le juge français reconna$\hat{i}$t aussi l'assouplissement de la charge de la preuve du lien de causalit$\acute{e}$ en mati$\acute{e}$re d'action m$\acute{e}$dicale. Il faudra toutefois souligner que le ph$\acute{e}$nom$\acute{e}$ne ne soit pas g$\acute{e}$n$\acute{e}$ralis$\acute{e}$ en droit français, d'autant plus que la pr$\acute{e}$somption de l'existence de la causalit$\acute{e}$ en la mati$\grave{e}$re a $\acute{e}$t$\acute{e}$ admise de mani$\grave{e}$re restrictive par la l$\acute{e}$gislation sp$\acute{e}$cifique. Tel $\acute{e}$tait notamment le cas pour les accidents de la contamination par le virus du sida ou de l'h$\acute{e}$patite C survenus apr$\grave{e}$s la transfusion. En d$\acute{e}$finitive, on peut dire qu'en droit français, le principe est maintenu en cas de manquement $\grave{a}$ une obligation de r$\acute{e}$sultat n$\acute{e}$e du contrat m$\acute{e}$dical, tandis que la Cour de cassation admet parfois en mati$\grave{e}$re de droit commun de la responsabilit$\acute{e}$ contractuelle la pr$\acute{e}$somption de causalit$\acute{e}$ en cas d'inex$\acute{e}$cution des obligations de r$\acute{e}$sultat. En fait, la pr$\acute{e}$somption de causalit$\acute{e}$ dans le contentieux m$\acute{e}$dical pourra mener les m$\acute{e}$decins $\grave{a}$ se diriger vers les traitements d$\acute{e}$fensifs. Cette situation peut m$\hat{e}$me conduire $\grave{a}$ emp$\hat{e}$cher le d$\acute{e}$veloppement de la science m$\acute{e}$dicale, enfin $\grave{a}$ une situation d$\acute{e}$savantageuse aux patients. Il y a alors lieu de se m$\acute{e}$fier des int$\acute{e}$r$\hat{e}$ts d$\acute{e}$s$\acute{e}$quilibr$\acute{e}$s entre le m$\acute{e}$decin et le patient. De ce point de vue, on peut estimer que le droit français donne des suggestions aux juristes cor$\acute{e}$ens dans la recherche des solutions plus ad$\acute{e}$quates en ce qui concerne la charge de la preuve du lien causal en mati$\acute{e}$re de responsabilit$\acute{e}$ m$\acute{e}$dicale.

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Effect of Submergence and Air Exposure of the Shoot on Growth, Nutrient Uptake and Photosynthesis in Monochoria vaginalis Presl. (물달개비 경엽(莖葉)의 침수여부(沈水與否)에 따른 생장(生長), 양분흡수(養分吸收) 및 광합성(光合成) 비교(比較))

  • Soh, C.H.;Yang, K.S.;Kwon, Y.W.
    • Korean Journal of Weed Science
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    • v.16 no.1
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    • pp.14-20
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    • 1996
  • Growth, nutrient uptake and photosynthesis as affected by submersion of shoot in pickerel weed (Monochoria vaginalis Presl.) were determined. The shoots of pickerel weeds in hydroponic culture were subjected to the submerged or emerged condition at 3- or 5-leaf stage for 8 or 10 days. Under submerged condition, growth in plant height was enhanced, but leaf number, leaf area, fresh and dry weight were reduced compared to those under the emerged condition. Similar responses in growth to submergence were obtained with the pickerel weeds rooted in the soil. Under submergence, chlorophyll content increased during the first 2 days, but thereafter remarkably decreased at 3-leaf stage and after the first 4 days at 5-leaf stage. Compared to the emerged condition, uptakes of $NH_4\;^+$-N, $NO_3\;^-$-N, $P_2O_5$ and $K^+$ were reduced, but uptakes of $Ca^{++}$ and $Mg^{++}$ increased under the submerged condition. Photosynthetic rate of shoot under water, measured by $CO_2$electrode, showed the maximum by 210 ${\mu}$moles $HCO_3\;^-$/g F.W. at the 8th day after submergence(DAS) at 3-leaf stage and 320 ${\mu}$moles $HCO_3\;^-$/g F. W. at 6 DAS at 5-leaf stage. These results indicate that pickerel weeds grow much better when the shoot is air-exposed and are less tolerable to submergence at 3 leaf-stage than at 5-leaf stage.

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A Study on the Chitin and Protein Contents in Shells of 5 Marine Crustaceans (5종(種)의 해산(海産) 갑각류각피(甲殼類殼皮)에서의 Chitin 및 단백질함량(蛋白質含量)에 관한 연구(硏究))

  • Lee, Mee-Sook;Seo, Jung-Sook;Mo, Su-Mi
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.13 no.3
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    • pp.307-312
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    • 1984
  • The dried pure shells deprived of soft tissues were subjected to analysis of chitin-protein complexes from 5 species of marine crustaceans, including 2 species of crabs and 3 species of shrimps. The protein fractions were obtained from chitin-protein complexes under the varying conditions of extractions and the crude chitin was prepared from the shells by the sulfurous acid process. The crude chitin was purified through the extraction with several organic solvents such as dimethyl-acetamide, N-methylpyrrolidone. The purified chitin was also examined using the phase contrast microscope. Total protein contents of the shells were diverse, showing 9.6% for Portunus trituberculatus, 3.1%, Charybdis bimaculata, 9.4%, Penaeus japonicus, 10.9%, Metapenaeus intermedius and 5.8%, Squilla oratoria. Covalently bound protein varied with species from 2.1% for Charybdis bimaculata to 9.9% for Metapenaeus intermedius. The puified chitin contents of the shells were shown to 21.1% for Portunus tritube rculatus, 6.2%, Charybdis bimaculata, 20.2%, Penaeus japonicus, 27.1%, Metapenaeus intermedius and 25.5%, Squilla oratoria. Exceptionally low analytical value obtained with Charybdis bimaculata are supposed to be due to the very young subjects. The ratios of chitin to covalently bound protein in the shells were various such as 2.7 to 1 for Portunus trituberculatus, Penaeus japonicus and Metapenaeus intermedius, 3.1 to 1, Charybdis bimaculata and 6.1 to 1, Squilla oratoria. The microscope finding of the purified chitin showed the filamentous form in all the specimen.

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A study on the ecological habitat and protection of natural Sorbus commixta forest at Mt. Seorak (설악산(雪嶽山)에 분포(分布)하는 마가목 천연림(天然林)의 생태환경(生態環境)과 보호(保護)에 관(關)한 연구(硏究))

  • Shin, Jai Man;Kim, Tong Su;Han, Sang Sup
    • Journal of Forest and Environmental Science
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    • v.3 no.1
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    • pp.1-9
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    • 1983
  • The purpose of this study was to elucidate the ecophysiological habitat of natural Sorbus commixta forest at Mt. Seorak. The results obtained were as follows: 1. The Sorbus commixta trees mainly distributed from 900m to 1,500m altitude. In there, the warm index(WI) was about 42$3.2{\times}10^3$ to $9.2{\times}10^3$, cation exchange capacity(CEC) was 13.7 to 19.5mg/100g, N content 0.21 to 0.39%, $P_2O_5$ content was 22.6 to 38.7ppm, and pH value was 5.6 to 5.8 respectively. 4. The upper crown trees in Sorbus commixta communities were Abies nephrolepis, Taxus cuspidata, Betula platyphylla var. japonica, Quercus${\times}$grosseserrata, Acer mono, Prunus sargentii, Carpinus cordata, Tilia amurensis, and the under crown trees were Rhododendron brachycarpum, Acer pseudo-sieboldianum, Thuja olientalis, Corylus heterohpylla, Philadelphus schrenckii, Rhododendron schlippenbachii, Rhododendron mucronulatum, and Magnolia sieboldii. 5. The stand densities were 1,156 trees/ha at 1,160m and 3,600 trees/ha at 1,300m respectively. The coverages by the DBH basal area were 0.37 at 1,160m and 0.31 at 1,300m respectively, and the vegetation coverages by the crown projection area were 2.04 at 1,160m and 1.61 at 1,300m respectively. 6. The light extinction coefficient(k) in Beer-Lambert's law, showed the distance, F(z), from top canopy to aboveground, was 0.17. 7. The water relations parameters of Sorbus commixta shoot were obtained by the pressure chamber technique. The osmotic pressure, ${\pi}_o$, at maximum turgor was -16.2 bar, and VAT pressure was 14.5bar. The osmotic pressure, ${\pi}_p$, at incipient plasmolysis was -19.4bar. The relative water contents at incipient plasmolysis were 83.1% ($v_p/v_o$) and 87.1%($v_p/w_s$;$w_s$, total water at maximum turgor). 8. The bulk modulus of elasticity(E) of shoot was about 69.6. The total symplasmic water to total water in shoot was 67.7%, and the apoplastic water to total water was 32.3%.

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Expression of TIMP1, TIMP2 Genes by Ionizing Radiation (이온화 방사선에 의한 TIMP1, TIMP2 유전자 발현 측정)

  • Park Kun-Koo;Jin Jung Sun;Park Ki Yong;Lee Yun Hee;Kim Sang Yoon;Noh Young Ju;Ahn Seung Do;Kim Jong Hoon;Choi Eun Kyung;Chang Hyesook
    • Radiation Oncology Journal
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    • v.19 no.2
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    • pp.171-180
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    • 2001
  • Purpose : Expression of TIMP, intrinsic inhibitor of MMP, is regulated by signal transduction in response to genotoxins and is likely to be an important step in metastasis, angiogenesis and wound healing after ionizing radiation. Therefore, we studied radiation mediated TIMP expression and its mechanism in head and neck cancer cell lines. Materials and Methods : Human head and neck cancer cell lines established at Asan Medical Center were used and radiosensitivity $(D_0)$, radiation cytotoxicity and metastatic potential were measured by clonogenic assay, n assay and invasion assay, respectively. The conditioned medium was prepared at 24 hours and 48 hours after 2 Gy and 10 Gy irradiation and expression of TIMP protein was measured by Elisa assay with specific antibodies against human TIMP. hTIMP1 promoter region was cloned and TIMP1 luciferase reporter vector was constructed. The reporter vector was transfected to AMC-HN-1 and -HN-9 cells with or without expression vector Ras, then the cells were exposed to radiation or PMA, PKC activator. EMSA was peformed with oligonucleotide (-59/-53 element and SP1) of TIMP1 promoter. Results : $D_0$ of HN-1, -2, -3, -5 and -9 cell lines were 1.55 Gy, 1.8 Gy, 1.5 Gt, 1.55 Gy and 2.45 Gy respectively. n assay confirmed cell viability, over $94\%$ at 24hrs, 48hrs after 2 Gy irradiation and over 73% after 10 Gy irradiation. Elisa assay confirmed that cells secreted TIMP1, 2 proteins continuously. After 2 Gy irradiation, TIMP2 secretion was decreased at 24hrs in HN-1 and HN-9 cell lines but after 10 Gy irradiation, it was increased in all cell lines. At 48hrs after irradiation, it was increased in HN-1 but decreased in HN-9 cells. But the change in TIMP secretion by RT was mild. The transcription of TIMP1 gene in HN-1 was induced by PMA but in HN-9 cell lines, it was suppressed. Wild type Ras induced the TIMP-1 transcription by 20 fold and 4 fold in HN-1 and HN-9 respectively. The binding activity to -59/-53, AP1 motif was increased by RT, but not to SP1 motif in both cell lines. Conclusions : We observed the difference of expression and activity of TIMPs between radiosensitive and radioresistant cell line and the different signal transduction pathway between in these cell lines may contribute the different radiosensitivity. Further research to investigate the radiation response and its signal pathway of TIMPs is needed.

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