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Establishment of Mouse Embryonic Stem Cell-like Cells from In Vitro Fertilized Embryos (체외수정 생쥐 배아에서의 배아 줄기세포 확립)

  • Shin, Yong-Moon;Park, Yong-Bin;Kim, Hee-Sun;Oh, Sun-Kyung;Chun, Dae-Woo;Suh, Chang-Suk;Choe, Young-Min;Kim, Jung-Gu;Lee, Jin-Yong;Kim, Seok-Hyun
    • Clinical and Experimental Reproductive Medicine
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    • v.29 no.1
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    • pp.1-12
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    • 2002
  • Objective: In order to acquire the technique for the establishment of human embryonic stem cells (ESe) derived from the human frozen-thawed embryos produced in IVF-ET program, this study was performed to establish mouse ESC derived from the in vitro fertilized embryos. Materials and Methods: After Fl hybrid (C57BL female $\times$ CBA mael) female mice were superovulated with PMSG and hCG treatment, their oocytes were retrieved and inseminated, and the fertilized embryos were cultured for 96-120 hours until the expected stages of blastocysts were obtained. To isolate the inner cell mass (ICM), either the blastocysts were treated with immunosurgery, or the whole embryos were cultured for 4 days. Isolated ICMs were then cultured onto STO feeder cell layer, and the resultant ICM colonies were subcultured with trypsin-EDTA treatment. During the subculture process, ESC-like cell colonies were observed with phase contrast microscopy. To identify ESC in the subcultured ESC-like cell colonies, alkaline phosphatase activity and Oct-4 (octamer-binding transcription factor-4) expression were examined by immunohistochemistry and RT-PCR, respectively. To examine the spontaneous differentiation, ESC-like cell colonies were cultured without STO feeder cell layer and leukemia inhibitory factor (LIF). Results: Seven ESC-like cell lines were established from ICMs isolated from the in vitro fertilized embryos. According to the developmental stage, the growth of ICMs isolated from the expanded blastocysts was significantly better than that of ICMs isolated from the hatched blastocysts (80.3% vs. 58.7%, p<0.05). ESC-like cell colonies were only obtained from ICMs of expanded blastocysts. However, the ICMs isolated from the embryos treated with immunosurgery were poorly grown and frequently differentiated during the culture process. The established ESC-like cell colonies were positively stained with alkaline phosphatase and expressed Oct-4, and their morphology resembled that observed in the previously reported mouse ESC. In addition, following the extended in vitro culture process, they maintained their expression of cell surface markers characteristic of the pluripotent stem cells such as alkaline phosphatase and Oct-4. When cultured without STO feeder cell layer and LIF, they were spontaneously differentiated into the various types of cells. Conclusion: The findings of this study suggest that the establishment of mouse ESC can be successfully derived from the in vitro fertilized embryos. The established ESC-like cells expressed the cell surface markers characteristic of the pluripotent stem cells and spontaneously differentiated into the various types of cells.

Sedimentary Characteristics and Evolution History of Chenier, Gomso-Bay tidal Flat, Western Coast of Korea (황해 곰소만 조간대에 발달한 Chenier의 퇴적학적 특성과 진화)

  • 장진호;전승수
    • 한국해양학회지
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    • v.28 no.3
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    • pp.212-228
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    • 1993
  • A chenier, about 860 m long, 30 to 60 m wide and 0.6∼1.6 m high, occurs on the upper muddy tidal flat in the Gomso bay, western coast of Korea, It consists of medium to fine sands and shells with small amounts of subangular gravels. Vertical sections across the chenier show gently landward dipping stratifications which include small-scale cross-bedded sets. the most probable source of the chenier is considered to be the intertidal sandy sediments. Vibracores taken along a line transversing the tidal flat reveal that the intertidal sand deposits are more than 5 m thick near the low-water line and become thinner toward the chenier. The most sand deposits are undertrain by tidal muds which occur behind the chenier as salt marsh deposits. C-14 age dating suggests that the sand deposits and the chenier are younger than about 1,800 years B.P. The chenier has originated from the intertidal sand shoals at the lower to mid sand flat, and has continuously moved landward. A series of aerial photographs (1967∼1989) reveal that intertidal sand shoals (predecessor of the western part of chenier) on the mid flat have continuously moved landward during the past two decades and ultimately attached to the eastern part of the chenier already anchored at the present position in the late 1960s. Repeated measurements (four times between 1991 and 1992) of morphological changes of the chenier indicate that the eastern two thirds of the chenier, mostly above the mean high water, has rarely moved whereas the western remainder below the mean high water, has moved continuously at a rate of 0.5 m/mo during the last two years (1991∼1992). This displacement rate has been considerably accelerated up to 1.0 m/mo in winter, and during a few days of typhoon in the summer of 1992 the displacement amounted to about 8∼11 m/mo for the entire chenier. these facts suggest that macro-tidal currents, coupled with winter-storm waves and infrequent strong typhoons, should play a major role for the formation and migration of chenier after 1,800 B.P., when the sea level already rose to the present position and thereafter remained constant.

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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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Effect of Dietary Betaine and Energy Levels on Liver Fats and Cholesterol in Laying Hens (사료내 비태인과 에너지 수준이 산란계의 간지방과 콜레스테롤에 미치는 영향)

  • Park J. H.;Park S. Y.;Ryu K. S.
    • Korean Journal of Poultry Science
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    • v.32 no.3
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    • pp.149-156
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    • 2005
  • Two experiments were conducted to examine the effects of betaine intake on blood and yolk cholesterol, abdominal fat, liver fat, tissue triglyceride(TG) and liver HMG-CoA reductase In laying hens. In Expt. 1, a total of 72 ISA-brown laying hens were individually assigned into four treatments from 18 to 21 weeks old. Com-soybean meal based diet were fed with the addition of 0, 300, 600 and 1,200ppm. In Expt. 2, 72 ISA-brown laying hens were housed into individual cage to evaluate the effect of dietary betaine(0, 600ppm) and energy(ME, 2,800, 2,900kca/kg) from 70 to 74 weeks. Serum total cholesterol, LDL and HDL-cholesterol and TG concentration in blood of hens fed betaine tended to increase compared to those of the control, but were not significantly different. However, betaine supplementation showed a statistically significant decrease in yolk cholesterol(P<0.05). There was no significant difference in abdominal fat among the treatments. Liver fats and 7c of birds 130 betaine was decreased compared with control. Serum total cholesterol, LDL-cholesterol and triglyceride concentration were significantly inc.eased by ffeding a diet containing 600ppm betaine in Expt. 2(P<0.05), but were not influenced by the dietary energy levels. Yolk cholesterol, abdominal fat and HMG-CoA reductase activity were affected neither by dietary energy nor betaine level.

A Study on Fractionation and Characterization of Water-Soluble Natural Fe-Chelates From Garbage Compost and Activated Sewage Sludge (활성오니(活性汚泥) 및 진개퇴비중(塵芥堆肥中) 수용성(水溶性) 철(鐵) 킬레이트의 분리(分離)와 특성(特性)에 관(關)한 연구(硏究))

  • Park, Nae-Joung;Lindsay, W.L.
    • Applied Biological Chemistry
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    • v.18 no.4
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    • pp.194-202
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    • 1975
  • This study was conducted to study the properties of the water-soluble natural chelating agents from garbage compost and activated sewage sludge responsible for Fe chelation, which is closely associated with the effectiveness in correcting iron chlorosis in plant. The water-soluble fraction of these materials was fractionated by menas of Sephadex gel filtration and the fractions of Fe chehates were traced by radioactive $^{59}Fe$. The fractions were examined by ultraviolet and infrared. spectroscopy and stability constants for Fe. The water-soluble fraction from garbage compost was separated by Sephadex G-25 into approximately four fractions. Most of the added $^{59}Fe$ was associated with fraction I, which appeared at the void volume. Further fractionation by Sephadex G-50 indicated that the molecular weight of water-soluble chelating agents is in the approximate range of 5000 to 10,000. The water-soluble fraction from activated sewage sludge gave six fractions by Sephadex G-25. Most of the added $^{59}Fe$ was found in the fraction I,II, and III, The molecular weights of most chelating agents associated with $^{59}Fe$ appeared to be less than 5,000 and those of fraction I that appeared at the void volume was in the range of 5,000 to 1,000. Discrepancy between radio activity count and UV absorption indicated the heterogeneity of the fractions obtained by Sephadex gel filtration. Ultraviolet absorption spectra of all fractions separated by Sephadex G-25 and containing chelating agents showed no differences. Fraction IV and V of sewage extract showed absorption maxima and shifting similar to nucleic acid components suggesting the presence of decomposition products of nucleic acid. Similarity fraction VI contained phenolic type amino acid groups. Fraction I of compost extract contained most of the added $^{59}Fe$ and showed weak but extra definite absorption in the 1230, and $1270cm^{-1}$ region, suggesting that extra oxygen groups in polyphenolic structure were probably involved in Fe chelation. In sewage extract, fraction I,II, and III in which most of the $^{59}Fe$ was found, showed strong definite polypeptide absorption in the region of $1540cm^{-1}$ due to NH deformation and C-N stretching of amide groups in the peptidebond. These extra functional groups in fraction I, II, and III appeared to be associated with Fe chelation. The other fractions, not associated with $^{59}Fe$, still have carboxyl and hydroxyl groups, suggesting that these functional groups in these water extracts may not independently form the Fe chelates. Precipitation of ferric hydroxide precluded measuring the stability constants for Fe-chelates. However, the formation constants for Zn chelates as log K values for compost extract and sewage extract at pH 4.0 from which the strength of chelation with Fe could be presumed, were 8.23, and 9.75, respectively, indicating strong complexation with metals. The chelating capacity of compost extract containing 6.5 g organic matter per liter was 0.82 mM, and that of sewage extract containing 5.3 g per liter was 0. 64 mM.

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Effects of Barley Straw Management Practices on Greenhouse Gases(GHGs) Emission During Rice Cultivation in Rice-barley Double Cropping System (벼보리 이모작 재배에서 보리짚 처리 방법이 벼재배시 온실가스 배출에 미치는 영향)

  • Ko, Jee-Yeon;Lee, Jae-Saeng;Jung, Ki-yul;Choi, Young-Dae;Ramos, Edwin P;Yun, Eul-Soo;Kang, Hwang-Won;Park, Seong-Tae
    • Korean Journal of Soil Science and Fertilizer
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    • v.41 no.1
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    • pp.65-73
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    • 2008
  • Because main barley straw management is changing these days from off-fields to burning that may relate to air quality concerning the global warming, this study was conducted to investigate the effects of barley-straw management practices on greenhouse gas emissions during rice cultivation in rice-barley double cropping system. The treatments were barley straw burning, off-field usage of barley straw and incorporation of barley straw in paddy fields. Laboratory experiment showed that burning of barley straw at the rate of $4.5Mg\;ha^{-1}$ emitted GHGs in the amounts of 4,607, 19.5, and $0.9kg\;ha^{-1}$ of $CO_2$, $CH_4$, and $N_2O$, respectively. During the rice cultivation of the rice-barley double cropping system, the highest GHG emission by evaluated close-static chamber method was observed from the soil incorporation of barley straw with 387 and $1.0kg\;ha^{-1}$ of $CH_4$ and $N_2O$, respectively. The GHGs emissions from the barley straw burning and off-field usage treatments were 233 and $160kg\;ha^{-1}$ for $CH_4$ and 0.80 and $0.79kg\;ha^{-1}$ for $N_2O$, respectively. The barley straw burning treatment showed the greatest GHGs emission among barley straw management practices in rice-barley double cropping system when considering GHGs emissions both during burning and from paddy fields during the cropping seasons. As a result, the GHGs emissions recorded in the barley straw incorporation to soil and off-field usage treatments were 22.4 and 66.8%, respectively, less than sum of GHGs emissions from the burning of barley straw and from paddy fields during rice cultivation.

Effect of Vegetation Types on the Distribution of Soil Invertebrates (식생유형이 토양무척추동물 분포에 미치는 영향)

  • Kim, Myung-Hyun;Bang, Hea-Son;Han, Min-Su;Hong, Hey-Kyoung;Na, Young-Eun;Kang, Kee-Kyung;Lee, Jeong-Taek;Lee, Deog-Bae
    • Korean Journal of Environmental Agriculture
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    • v.28 no.2
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    • pp.125-130
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    • 2009
  • The aim of this study was to investigate whether differences in the distribution of soil invertebrates among different vegetation types (forest, reservoir, and crop land types) in rural area. A total of 18 orders and 137 species were collected by pitfall traps. Species numbers were the lowest (33 species) at the Chamaecyparis obtusa plantation (St. 6). On the forest sites, the individual number of Hymenoptera was the most abundant, and Acari and Coleoptera was the relatively more abundant than the other sites. On the reservoir sites (Salix chaenomeloides community), the individual number of Collembola was the most abundant, and Diptera was the relatively more abundant than the other sites. On the crop land sites, the individual numbers of Collembola, Hymenoptera, and Araneae were the relatively more abundant than the other orders. The density of Araneae was higher in the reservoir and crop land sites than in the forest sites. From a point of view of biodiversity, although the diversity index(H') was the highest in the mixed broad-leaved forest type (St. 2) with Quercus serrata and Q. acutissima, and the lowest in the upland levee of crop land(St. 11), there was no significant difference among the habitat or vegetation types. According to the community analysis, the soil invertebrates could be divided into 4 groups, the mixed broad-leaved forest type (A group), the plantation or pure forest type (B group), the reservoir type (C group), and the crop land type (D group).

Chondrogenesis of Mesenchymal Stem Cells Derived from Human Umbilical Cord Blood (사람 제대혈 유래 간엽줄기세포로부터 연골세포 분화)

  • Koh, Phil-Ok;Cho, Jae-Hyun;Nho, Kyoung-Hwan;Cha, Yun-Im;Kim, Young-Ki;Cho, Eun-Hae;Lee, Hee-Chun;Jung, Tae-Sung;Yeon, Seong-Chan;Kang, Kyung-Sun;Lee, Hyo-Jong
    • Journal of Veterinary Clinics
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    • v.26 no.6
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    • pp.528-533
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    • 2009
  • In the current study, the mesenchymal stem cells (MSCs) isolated and propagated from the human umbilical cord blood (UCB) were tested for their capabilities of differentiation into chondrocytes in vitro. The mesenchymal progenitor cells (MPCs) collected from UCB were cultured in a low glucose DMEM medium with 10% FBS, L-glutamine and antibiotics. The human MSC colonies were positively stained by PAS reaction. When the immunophenotypes of surface antigens on the MSCs were analyzed by fluorescence-activated cell sorter (FACS) analysis, these cells expressed positively MSC-related antigens of CD 29, CD44, CD 90 and CD105, whereas they did not express antigens of CD14, CD31, CD34, CD45, CD133 and HLA-DR. Following induction these MSCs into chondrocytes in the chondrogenic differentiation medium for 3 weeks or more, the cells were stained positively with safranin O. We clearly confirmed that human MSCs were successfully differentiated into chondrocytes by RT-PCR and immunofluorescent stain of type-II collagen protein. These data also indicate that the isolation, proliferation and differentiation of the hUCB-derived MSCs in vitro can be used for elucidating the mechanisms involved in chondrogenesis. Moreover this differentiation technique can be applied to developing cell-based tissue regeneration or repair damaged tissues.

Plant Community Structure of Abies holophylla Community from Sinseongam to Jungdaesa in Odaesan National Park (오대산국립공원 신성암~중대사 전나무림 식물군집구조 특성)

  • Kim, Dong-Wook;Han, Bong-Ho;Kim, Jong-Yup;Yeum, Jung-Hun
    • Korean Journal of Environment and Ecology
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    • v.29 no.6
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    • pp.895-906
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    • 2015
  • This study was carried out to the structure of plant community from Sinseongam to Jungdaesa in Odaesan National Park, furthermore, it seeks to curate the basic data for planning of the Abies holophylla's forest management in Odaesan National Park. In order to identify the current ecological environment, this study explored the actual vegetation as primary research and set to twenty plots(i.e. $400m^2$) for analysing detailed structure of plant communities. The research methodology was qualitative analysis, therefore it used TWINSPAN and DCA analysis tools. Especially, TWINSPAN performed well in several comparisons of classification techniques, DCA is one of the ordination technique showed that the plant communities. The plant community was analysed classification and ordination by TWINSPAN and DCA, moreover it was analysed the structure of plant community such as importance percentage of woody species, DBH class distribution, the index of diversity and rate of sample tree growth. The main vegetation was A. holophylla-Quercus mongolica forest and Deciduous broad-leaved forest in the communities where located in low altitude and valley, whereas main vegetation where located in high altitude and slope was Q. mongolica forest. The research site's plant communities were classified four groups. In all of communities, A. holophylla was dominant species in main canopy layer, furthermore, the three communities (community I, II, III) are growing up next generation of A. holophylla excluding community IV. The communities (community I, II, III) can be sustained current status which dominates the A. holophylla communities, simultaneously, there might be expanded the Deciduous broad-leaved communities by Carpinus cordata, Betula schmidtii and so on. While, it showed that the community IV tended to be weaken the forces of A. holophylla, therefore the community IV can be transferred to C. cordata-Deciduous broad-leaved communities in the future. The age of sample trees was 79~128(i.e. A. holophylla), 75~87(i.e. Pinus koraiensis) and 190 years(i.e. Ulmus davidiana var. japonica). The index of Shannon's Species diversity (H') were ranged from 0.3889 to 1.3332 in the communities.

Antioxidant Activity of Different Parts of Lespedeza bicolor and Isolation of Antioxidant Compound (싸리나무(Lespedeza bicolor) 부위별 추출물의 항산화 활성 및 항산화물질 분리)

  • Lee, Jae-Hak;Jhoo, Jin-Woo
    • Korean Journal of Food Science and Technology
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    • v.44 no.6
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    • pp.763-771
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    • 2012
  • In this study, total antioxidant properties of extracts from different parts of Lespedeza bicolor were determined using techniques of measuring 1,1-diphenyl-2-picryl hydrazyl/2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)-radical scavenging activity and total phenolic contents. The total antioxidant activities of leaf, stem and root extracts from various solvents (water, 50, 70, 100% ethanol, and hot-water) indicated that 50 and 70% ethanol extracts have high radical scavenging activities and phenolic contents. A systematic approach was used to determine the total antioxidant activity of different solvent fractions of the Lespedeza bicolor extracts, partitioning with chloroform, ethyl acetate, n-butanol, and water, and the ethyl acetate fraction was found to have the strongest antioxidant activity. Antioxidant assay-guided isolation was carried out to isolate potential antioxidant compounds. The ethyl acetate fraction of the leaf extract was subjected to silica gel, LH-20 and RP-18 column chromatography successively, and afforded compound 1, which was identified as eriodictyol by NMR and MS analysis, after which its antioxidant activity was determined.