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Studies on Feed Formulation to Reduce Methane Emission and Optimal Rearing Density for Environmental-Friendly Hanwoo Production (친환경 한우 생산을 위한 메탄 저감용 사료 배합 및 적정 사육밀도에 관한 연구)

  • Shin, Jong-Suh;Choi, Byoung-Yang;Kim, Mi-Jung;Kim, Sung-Gi;Ra, Changsix
    • Journal of Animal Environmental Science
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    • v.17 no.sup
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    • pp.7-20
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    • 2011
  • This study was conducted to analyze the mixing ratio of raw feed materials for the methane mitigation and also to identify the minimum rearing density for improving the productivity of beef calves as eco-friendly fodder. Raw materials used in this study for the formulation of feed for methane reduction were crushed corn and alfalfa along with other 21 species. In addition, to investigate the appropriate rearing density, 12 Hanwoo calves with average weight of 150 kg was selected and experiment was conducted for four months. Methane gas emission (Bo) is about 3-4 times less in TMR 4 compared to TMR 1, 2 and 3. Feed price calculated for TMR 4 ration was also affordable. In addition, all TMRs showed a normal ruminal pH. Disappearance rate was observed to be lower in TMR 4 as compared to TMR 1, 2 and 3, but methane production decreased by 24 to 37%. The result showed improved total body weight, average daily gain and feed conversion ratio in rearing low-density ($18m^2/head$), and general treatment ($9m^2/head$) compared to overcrowding treatment ($6m^2/head$). In addition, blood components (total protein, glucose, AST, ALT and GGT factors) involved in health and disease treatments and health-related nutrition metabolism are lower in the low-density and general treatment compared to the high density treatment. Postural development (development of body size) i.e., weight, height and width significantly increased in the low and general density treatment compared to high density treatment. Especially excellent improvement was observed in low-density treatment than the general treatment. Moisture content, colonic bacteria and coccidium are higher in low and high density treatments than in the general treatment. The adequacy for beef rearing density is considered to be more desirable in an area more than $6m^2/head$. In conclusion, present study suggests that possibility of methane reduction through adjusting mixed feed ration. Also, rearing density is also an important factor in the growth and development of beef calves.

The Study on Musculoskeletal Symptoms and it's Related Factors in Radio-Technologists (방사선사의 근골격계 증상과 유해 요인에 관한 연구)

  • Lee, Hyang-Seob;Han, Man-Seok
    • Journal of radiological science and technology
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    • v.31 no.3
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    • pp.239-247
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    • 2008
  • In order to study the occurrence of symptoms of musculoskeletal disorders of radio-technologists employed at metropolitan general hospitals and the factors that influence such occurrence, standardized questionnaire by NIOSH that was modified and supplemented to be suitable for conditions in Korea was used. Answers collected from 143 radio-technologists in two weeks from June 13, 2007 were analyzed and the results are as follows. Factor that influence symptoms of musculoskeletal disorders by area were analyzed through multiple logistic regression analysis and the results found that in the neck area, risk increased as the burdening work category 2(Korea ministry of labor)(OR=3.94) and burdening work category 9(Korea ministry of labor)(OR=4.72) increased. In the shoulder region, risk increased as burdening work category 2(Korea ministry of labor)(OR=5.36), burdening work category 7(Korea ministry of labor)(OR=3.90), and burdening work category 9 (Korea ministry of labor)(OR=5.76) increased. In the arm/hand/wrist regions, risk increased as burdening work category 2 (Korea ministry of labor) (OR=6.91), and burdening work category 9 (Korea ministry of labor)(OR=3.76) increased. In the lower back region, risk increased as burdening work category 2 (Korea ministry of labor) (OR=3.06), and burdening work category 8 (Korea ministry of labor)(OR=8.14) increased. In the leg/knees/foot regions, risk increased as burdening work category 2 (Korea ministry of labor) (OR=3.63), and burdening work category 9 (Korea ministry of labor)(OR=2.96) increased. Conclusively, in factors that influence musculoskeletal disorder symptoms in radio-technologists, influence of subjective health conditions, total work experience, experience in current division, and burdening work category 2, 7, 8, and 9 (Korea ministry of labor) were most significant. Therefore, for preventive management, in addition to ergonomic and educational intervention for correcting improper posture during work, efforts for break time adjustment and stress reduction is needed, and encouragement and support for regular exercise is needed.

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Effects of ischemic preconditioning, KATP channel on the SOD activation and apoptosis in ischemic reperfused skeletal muscle of rat (허혈양상화와 KATP 통로가 허혈후 재관류된 흰쥐의 골격근육에서 SOD 활성 및 apoptosis에 미치는 영향)

  • Abn, Dong-choon;Paik, Doo-jin;Yang, Hong-hyun
    • Korean Journal of Veterinary Research
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    • v.39 no.5
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    • pp.878-895
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    • 1999
  • Ischemic preconditioing (IPC), i.e., a preliminary brief episode of ischemia and reperfusion, has been shown to reduce the cell damage induced by long ischemia and reperfusion. Superoxide radical which is produced during reperfusion after ischemia was recognized as a factor of the ischemic injury and it is dismutated into $H_2O_2$ and $O_2$ by two types of intracellular superoxide dismutase (SOD), Cu,Zn-SOD in cytoplasm and Mn-SOD in mitochondria. Recently oxygen free radicals are suggested to induce the apoptosis, however mechanism of the reduced apoptosis by ischemic preconditioing was unknown, while many studies performed in mammalian heart indicated that ATP-sensitive $K^+$ ($K_{APT}$) channel activation related with the protective effects. The aim of present study is to investigate 1) whether IP upregulate the Cu,Zn-SOD and Mn-SOD activities, and 2) whether ischemic preconditioning decreases apoptosis via $K_{APT}$ channel activation in timely reperfused skeletal muscle after long ishemia. The experimental animals, Sprague-Dawley rats weighing 250~300g, were divided into 8 groups; 1) control group, 2) ischemic preconditioning only groups, 3) pinacidil, a $K_{APT}$ channel opener, treatment only groups, 4) glibenclamide, a $K_{APT}$ channel blocker, treatment only groups, 5) ischemia groups, 6) ischemia after IPC groups, 7) ischemia and pinacidil treatment groups, and 8) IP and ischemia after glibenclamide pretreatment groups. Animals of the control group were administered with the vehicle (DMSO) alone. Pinacidil (1mg/kg) was administered intravenously 5 minutes after initiation of ischemia, and glibenclamide (0.5mg/kg) was injected intravenously 20 minutes before IPC. In rats that were ischemic preconditioned, the left common iliac artery was occluded for 5 minutes followed by 5 minutes of reperfusion by three times using vascular clamp. Ischemia was done by occlusion of the same artery for 4 hours. The specimens of left rectus femoris muscle were obtained immediately (0 hour), 12 hours, 24 hours after drug administrations, IP or ischemia and reperfusion. The immunoreactivities of SOD and its alterations were observed by use of sheep antihuman Cu,Zn-SOD and Mn-SOD antibodies on the $10{\mu}m$ cryosections. The incidencies of apoptosis were observed by TUNEL methods with in situ apoptosis detection kit on $6{\mu}m$ paraffine section. The results obtained were as follows : 1. After IPC, immunoreactivities of Cu,Zn-SOD mainly in the small-sized fibers were increased by 24 hours, that of Mn-SOD at 0 hour and 24 hours. 2. No significant changes in immunoreactivities of SOD was observed in the pinacidil and in the glibenclamide treatment only groups, and in the ischemia only groups. 3. The immunoreactivities of the Cu,Zn-SOD were increased in the ischemia after IPC groups and the ischemia and pinacidil treatment groups. 4. The immunoreactivities of the Cu,Zn-SOD in the IPC and ischemia after glibenclamide pretreatment groups were not increased except for the 12 hours reperfusion group. But, Mn-SOD immunoreactivities were increased in the 0 hours, 12 hours and 24 hours after reperfusion. 5. In the control group, the IPC only groups, and the pinacidil treatment only groups, negative or trace apoptotic reactions were observed, but the positive apoptotic reaction occured in the glibenclamide treatment groups. 6. Moderate or many number of apoptosis were revealed in the ischemia groups, and also the IPC and ischemia after glibenclamide pretreatment group except for 12 hours and 24 hours after reperfusion. However, the incidence of apoptosis was decreased in the ischemia after IPC groups and in the ischemia and pinacidil treatment groups. 7. There is a coincidence between the increase of Cu,Zn-SOD immunoreactivities and the decrease of apoptosis in the presence of ischemia and reperfusion. These results suggest that the protective effects of ishemic preconditioing may related to the SOD activation, and the ischemic preconditioning decreases the apoptosis partially via $K_{APT}$ channel activation in timely reperfused rat skeletal muscle. It is also suggested that inhibition of apoptosis by IPC may related with the SOD activation.

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The Study on Change in Sex-Related Knowledge and Attitude through Sex Education : focusing on the 1st grade students in girls' junior high schools (성교육 실시에 따른 성지식, 성태도 변화 연구 -1학년 여중생을 대상으로-)

  • 계수연;문인옥
    • Korean Journal of Health Education and Promotion
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    • v.16 no.2
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    • pp.137-155
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    • 1999
  • The purpose of this study was to assess the effect of sex education on knowledge and attitude related to sex. The subjects were taken from by 199 students in 3 classes from 1st grade in H girl's junior high school as the study group, and 2 classes from 1st grade in S girl's junior high school as control group. During the survey period(September 21, 1998 to September 30, 1998), 6 times in terms of one-hour class for sex education were taught to the study group. A pre-test was executed on September 19, 1998 and the post-test on September 30. The findings were as follows. 1. According to the research, 20.1% of the subjects have experienced sex education from parents and 89.9% from teacher. They have mostly obtained the sex-related information from teachers(59.8%), following movie, radio, TV, or video tape(40.7%), friends(35.2%), reading materials such as books, cartoons, news papers and magazines(31.7%), parents(15.6%), siblings(7.0%), PC(1.5%) and telephone service(1.5%). 2. 27.1% of the subjects reported that they had sex-related worry concerning from friendship with the opposite sex, following physiological phenomenon(31.5%), sex violence(11.1%), physical characteristics(7.4%), VD and contraception(5.6%), sexual impulse(5.6%), pregnancy and delivery(5.5%), and sexual behaviour(3.7%). The research showed that the adolescents usually solved their problems through the consultation with theifriends(44.4%). However, 16.7% of the subjects were turned out not to request any solution. The other minor routes to settle their problems were written materials such as books, magazines(13.0%), parents(13.0%), movie, radio, TV, or video tape(5.5%), acquainted female elders(3.7%) and teachers(3.7%). 3. The most interesting part regarding sex was the friendship with the opposite sex(61.8%), following adolescent's emotion(55.8%), physiological differences between two genders(52.8%), AIDS(48.7%), VD(46.7%), pregnancy(45.2%), contraception(45.2%), abortion(41.7%), intercourse(41.7%), masturbation(41.2%), sex violence(41.2%) and genital structure and secondary sexual characteristics(28.6%). 4. In regard to characteristics of the subjects influencing sex-related knowledge, the higher educational career of mother, living with at least either parent and the experience of sex education by teachers were statistically significant factors(p〈0.05). 5. In regard to characteristics of the subjects influencing attitudes toward sex, the experience of sex education by parents or teachers was a statistically significant factor(p〈0.05). 6. The analysis of knowledge score comparing results before and after sex education showed that control group's score decreased from 12.5 to 12.44 while the study group's score increased from 12.33 to 21.31, which was statistically significant(p〈0.001). 7. The analysis of the attitude scores before and after sex education showed that the control group's score slightly increased from 55.57 to 56.36, while the study group's score increased from 54.79 to 61.95, which was statistically significant(p〈0.001). 8. The level of sex-related concerns of the study group after sex education marked both the increase in some items and the decrease in others. 9. Most instructive session among the sex education was the third “to be a good friend to the opposite sex”(27.0%).

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A Study for Continue and Decline of Abies koreana Forest using Species Distribution Model - Focused in Mt. Baekwun Gwangyang-si, Jeollanam-do - (종 분포 모형을 이용한 구상나무림의 지속 및 쇠퇴에 관한 연구 - 전라남도 광양시 백운산을 중심으로 -)

  • Cho, Seon-Hee;Park, Jong-young;Park, Jeong-Ho;Lee, Yang-Geun;Mun, Lee-man;Kang, Sang-Ho;Kim, Gwang-Hyun;Yun, Jong-Guk
    • Journal of Korean Society of Forest Science
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    • v.104 no.3
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    • pp.360-367
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    • 2015
  • The present study investigated the habitats of Korean fir trees (Abies koreana E. H. Wilson) on Mt. Baekwun (Baekwun-san), determined the current distribution, quantified the contribution of biological and non-biological environmental factors affecting the distribution, derived actual and potential habitats, presented a plan for the establishment of protected areas, applied RCP 8.5 climate change scenario to analyze the effects of climate change on the future distribution of Korean fir trees, and predicted future potential habitats. According to the results of the study, 3,325 Korean fir trees (DBH >= 2.5 cm) inhabited Mt. Baekwun, and their distribution area was approximately 150 ha. Populations of Korean fir trees were confirmed to exist at an altitude of 900 m above sea level and were distributed up to 1,200 m. Based on potential distribution, areas appropriate for habitation by Korean fir trees were analyzed to be 450 ha, three times the current distribution area, with a focus on Sang Peak (Sang-bong), Eokbul Peak (Eokbul-bong), Ddari Peak (Ddari-bong), and Dosol Peak (Dosol-bong). The forest stands near Sang Peak, the main peak, were evaluated as those with the most appropriate potential for the habitation of Korean fir trees, and populations of the trees tended to prefer the northern slope rather than the southern slope. When climate change scenario RCP 8.5 was applied and future potential distribution was analyzed, the habitats were expected to decrease in area to 20 ha by 2050, with a focus on Sang Peak, and areas appropriate for habitation were predicted not to exist by 2080. Judging from such results, as global warming accelerates, the habitats of Korean fir trees are clearly expected to move from lowlands to highlands.

Factors Influencing the Meaning of Life for Middle-aged Women (중년여성의 삶의 의미와 영향요인)

  • Park, Geum-Ja
    • Women's Health Nursing
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    • v.8 no.2
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    • pp.232-243
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    • 2002
  • The purpose of this study was to examine factors influencing the meaning of life for middle-aged women. The subjects for the study were 190 middle-aged women who live in Busan. Data were collected from May 25 to June 20, 1999. The instruments for this study were as follows : the meaning of life scale developed by Jung D. R(1978) ; the self-esteem scale developed by Ro, E. Y, Kwon, J. H.(1997) ; the volunteer activity scale developed by Park G. J.(1999), the marital satisfaction scale developed by Choi G. Y.(1999) ; and the health status scale developed by Kim S. Y. (1991). SPSS PC+ was utilized for data analysis. Data were analysed according to frequency and percentage, mean and standard deviation, t-test, ANOVA, Sheffe' s test and Pearson' s correlation coefficients and stepwise multiple regression. The reliability of the tools was tested by Cronbach's $\alpha$ : and it showed reliability scores of 0.88(for the meaning of life tool), 0.81(for the self-esteem tool), 0.94(for the marital satisfaction tool) and 0.78(for the health status tool). The results were as follows: 1) The scores showed that middle-aged women had a moderate level of meaning of life, with a mean of 30.31 and standard deviation of 6.24 (with values ranging from 12.00 to 48.00). The scores showed that middle-aged women had a moderate levels of self-esteem, with a mean of 28.92 and standard deviation of 4.67 (with values ranging from 10.00 to 40.00). The scores showed that middle-aged women had a very low level of volunteer activity, with a mean of 0.87 and standard deviation of 0.93(with values ranging from 0.00 to 1.00). The scores showed that middle-aged women had a moderate level of satisfaction in their marriages, with a mean of 31.99 and standard deviation of 7.84(with values ranging from 12.00 to 48.00). The scores showed that middle-aged women had a moderate level of health status, with a mean of 6.63 and standard deviation of 1.57(with values ranging from 3.00 to 9.00). 2) There was a statistically significant difference of the meaning of life of middle-aged women according to their religion(F=8.930, p=.000), christian ($31.94{\pm}5.96$) had more meaning th life than buddhists ($28.40{\pm}6.23$). 3) There was a statistically significant correlations between the subject' s self-esteem and the her meaning of life(r=0.477, p=0.000), the subject' s level of volunteer activity and meaning of life(r=.428, p=.000), her level of marital satisfaction and meaning of life(r=.417, p=.000), and her level of health status and meaning of life(r=.261, p=.000) among these middle-aged women. 4) Self- esteem was the highest factor influencing the level of meaning of life in middle-aged women. 40.0% of the total variance of levels of meaning of life by was dependent on self-esteem, volunteer activity, marital satisfaction and health status. In conclusion, the higher the self-esteem and levels of volunteer activity, marital satisfaction, and health status of middle-aged women, the higher the meaning of life for them. Therefore, it is necessary to elevate levels of the self-esteem and volunteer activity, marital satisfaction and health status in order to help middle-aged women have more meaning of life.

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Relative Contribution rate on Soil Physico-chemical Properties Related to Fruit Quality of 'Hongro' Apple (사과 '홍로' 품종의 과실 품질에 미치는 토양이화학성의 상대적 기여도)

  • Kim, Seung-Heui;Park, Seo-Jun;Han, Jeom-Wha;Cho, Jung-Gun;Choi, Hyeong-Suk;Lim, Tae-Jun;Yun, Hea-Keun
    • Journal of Bio-Environment Control
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    • v.21 no.2
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    • pp.102-107
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    • 2012
  • This study was carried out to investigate the optimum soil environmental conditions of ten contents on production of high quality fruit in 'Hongro' apple. The soil and fruit characteristics were analyzed at total 60 orchards in major apple producing areas such as Chungju, Moonkyeung, Yeongju, Andong, Yeosan and Yeongcheon (10 orchards an area). The soil environmental factors affected fruit weight were the highest relative contribution in saturated hydraulic conductivity of 33.3%. The cation was 24.6%, the bulk density, soil texture and solid phase were also high as relative contribution. The fruit weight was influenced by soil physical properties more than soil chemical properties. The soil environmental factors affected sugar content were highest soil texture of 21.9%, and the CEC and bulk density were low as relative contribution. The fruit coloring was the highest relative contribution in phosphate of 55.9%. While saturated hydraulic conductivity and organic matter content were low. The coloring was influenced by soil chemical properties more than soil physical properties. Fruit coloring was high influenced over 70% by soil physical properties. Finally, relative contribution on fruit quality related with sugar content, fruit weight, and coloring were high influenced by cultivation layer depth of 25.8%, soil texture 22.2%, and soil pH of 21.0% but bulk density and solid phase were low relative contribution. The fruit growth and soil chemical properties in 'Hongro' apple were very closely related. Therefore, orchard soil management to produce high quality fruit was very importance drainage management and organic matter application. We concluded that scientific soil management is possible by quanlifiable of soil management factors.

The Study on the Buttons (centering around 19th-20th Centuries) (단추에 관한 연구 -19, 20세기를 중심으로-)

  • 이영란
    • Journal of the Korean Society of Costume
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    • v.22
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    • pp.263-276
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    • 1994
  • The achievement of notable social reoforms attained during the period of 19th and 20th centuries needlessly speaking remodelded the social environmental into several different patterns such as :1) high industrialization 2) propensity to consume 3) up graded overall social stands. Accordingly the industrial world of the but-tons too established the mess production syhstem by breaking from convention of hand-craft work of 17th century. The raw materials used in the production line on buttons during the 20th century are almost all-kind of materials one can possibly named including cheap plastic which enabled production lines to produce cheaper but higher productivities of the buttons being produced, The design (incused design) used in the 19-20h centuries are : men landscape, sports features, birds, livestocks, bugs, or geomatric features, tec, 1, The classification o f the buttons by materials Techniques shapes colors marking (Incused design) used in the productionof buttons in the England United States of America Laska Italy france Denmark Japan and India are categolizzed as : natural raw materials and syntetical resines. 1) Of the natural raw materials used are : Matal Enamel Iodine Agate, Coral, Green jade(Jasper) Granite, Wood, Ivory, Horn and bone etc. 2) The sythetical resin used in the button in-dustries are : Artificial jewell glass Acrylic material Styroform Celluloid and Nylon etc. 2. The thecnique quoted in producing buttons are hand craft work inlay work precision casting press mosic dye etching, processing, engraving and embossed carving etc. 3. The major designs used in the buttons in -dustries are : Round shape however elliptical column angular and edge shape often used. 4. The colors used are : The multi-colors were highly used than mono-colored materials such as : Adjoining Color and Contrast Color. The highest consideration to be considered in choosing the colors for the buttons are harmonization and matching factor with the garment or dresses to be wore. 5. The major design(incused design) on the buttons are embodiment and the design were also used in order of abstractive-combination abstractive with has offers much surprising. The button industries during the 19th and 20th centuries were not only the determination factors those can judge the value of self-pride of Nation and which were far beyond the in-dustrial arts in those days but also highly refelected and influenced by cultural sense ideology and self-pride of the Nation of those period. The followings are details of the role of the buttons categolized in the order of functional ornamental and symbolical aspects : 1. The functional role : The functional role of the buttons were simply designed for dress how-ever the buttons beyond from this role of function now a days. 2. The ornamental role : The ornamental role of he button beyond from this role of the button were effectuated by : 1) shape materials colors 2) technique locations size and design (incused design) 3) The ramaterials used for buttons shall not be over looked because it is highly depends on the taste sense and combination of harmony with the garment to be wore. 4) The color of the buttons are made well contrasted with the color of garments just as in the case of other artistical area such as matchs with the color of garment of contrast with brigtness of colors contrasted as complementary color and so and so. 5) The technique being adoped are: precision casting press handcraft inlay work etching mosic etc,. Since the buttons are no longer a simple catching devise used to fasten together the different part of the dress but now it has formed own and occupied the independent role in the garment or dresses location can be de-termined and varying depending on the ideas of designers. The size of the buttons has no specific limits, However the variation has widely dependined on the entire circumperence rhythm contrast harmonization of the garments. 3. The symbolical role : Since the button is no longer a just a simple devise for catching and fastening device used fastening together the different part of the garments but now were built a independent area as major part of the Garment and well reflected all kinds of occupations political background cultural as-pect etc. on the buttons. The design of buttons in the western circles are more simplified but they are polished looks and their techniques of manufacturing are comination of both machanis and handcraft. The colors used in the buttons are pretty well harmonized with garment(dress). Almost all kind of materials can be used in the but-tons however materials used in the buttons are : Bone of livestocks ivory, turtle shell are no longer used because the prevention of cruely of animal. On the contraly the level of buttons indus-try of Korea is far to reach and catch up with the level of western circles. It is highly suggested therefore the but-tons industrial field of Republic of Korea shall place and encouragement in producing beter industrial environment of the buttons based on the traditional and cultural aspect of republic of Korea to produce both manufacturing of qulified and best designed and colored buttons.

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The Effect of Exercise Training on Aβ-42, BDNF, GLUT-1 and HSP-70 Proteins in a NSE/ APPsw-transgenic Model for Alzheimer's Disease. (지구성 운동이 NSE/APPsw 알츠하이머 질환 생쥐의 인지능력, Aβ-42, BDNF, GLUT-1과 HSP-70 단백질 발현에 미치는 영향)

  • Eum, Hyun-Sub;Kang, Eun-Bum;Lim, Yea-Hyun;Lee, Jong-Rok;Cho, In-Ho;Kim, Young-Soo;Chae, Kab-Ryoung;Hwang, Dae-Yean;Kwak, Yi-Sub;Oh, Yoo-Sung;Cho, Joon-Yong
    • Journal of Life Science
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    • v.18 no.6
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    • pp.796-803
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    • 2008
  • Mutations in the APP gene lead to enhanced cleavage by ${\beta}-$ and ${\gamma}-secretase$, and increased $A{\beta}$ formation, which are closely associated with Alzheimer's disease (AD)-like neuropathological changes. Recent studies have shown that exercise training can ameliorate pathogenic phenotypes ($A{\beta}-42$, BDNF, GLUT-1 and HSP70) in experimental models of Alzheimer's disease. Here, we have used NSE/APPsw transgenic mice to investigate directly whether exercise training ameliorates pathogenic phenotypes within Alzheimer's brains. Sixteen weeks of exercise training resulted in a reduction of $A{\beta}-42$ peptides and also facilitated improvement of cognitive function. Furthermore, GLUT -1 and BDNF proteins produced by exercise training may protect brain neurons by inducing the concomitant expression of genes that encode proteins (HSP-70) which suppress stress induced neuron cell damages from APPsw transgenic mice. Thus, the improved cognitive function by exercise training may be mechanistically linked to a reduction of $A{\beta}-42$ peptides, possibly via activation of BDNF, GLUT-1, and HSP-70 proteins. On the basis of the evidences presented in this study, exercise training may represent a practical therapeutic management strategy for human subjects suffering from Alzheimer's disease.

Development of Assay Methods for Enterotoxin of Escherichia coli Employing the Hybridoma Technology (잡종세포종기법을 이용한 대장균의 장독소 측정법 개발)

  • Kim, Moon-Kyo;Cho, Myung-Je;Park, Kyung-Hee;Lee, Woo-Kon;Kim, Yoon-Won;Choi, Myung-Sik;Park, Joong-Soo;Cha, Chang-Yong;Chang, Woo-Hyun;Chung, Hong-Keun
    • The Journal of the Korean Society for Microbiology
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    • v.21 no.1
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    • pp.151-161
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    • 1986
  • In order to develop sensitive and sepcific assay methods for E. coli heat labile enterotoxin(LT) hybridoma cell lines secreting LT specific monoclonal antibody were obtained. LT was purified from cell lysate of E. coli O15H11. The steps included disruption of bacteria by French pressure, DEAE Sephacel ion exchange chromatography, Sephadex G200 gel filtration, and second DEAE Sephacel ion exchange chromatography, successively. Spleen cells from Balb/c mice immunized with the purified LT and $HGPRT^{(-)}$ plasmacytomas, $P3{\times}63Ag8.V653$ were mixed and fused by 50% (w/v) PEG. Hybrid cells were grown in 308 wells out of 360 wells, and 13 wells out of them secreted antibodies reacting to LT. Among these hybridoma cell 1G8-1D1 cell line was selected since it had produced high-titered monoclonal antibody continuously. By using culture supernatant and ascites from 1G8-1D1 cells the monoclonal antibody was characterized, and an assay system for detecting enterotoxigenic E. coli was established by double sandwich enzyme-linked immunosorbent assay (ELISA). The following results were obtained. 1. Antibody titers of culture supernatant and ascites from 1G8-1D1 hybridoma cells were 512, and 102, 400, respectively by GM1-ELISA and its immunoglobulin class was IgM. 2. The maximum absorption ratio of 1G8-1D1 cell culture supernatant to LT was 90% at $300\;{\mu}g/ml$ of LT concentration. LT concentration shown at 50% absorption ratio was $103.45{\mu}g$ and the absorption ratio was decreased with tile reduction of LT concentration. This result suggests that monoclonal antibody from 1G8-1D1 hybridoma cell bound with LT specifically. 3. The reactivities of 1G8-1D1 cell culture supernatant to LT and V. cholerae enterotoxin(CT) were 0.886 and 0.142(O.D. at 492nm) measured by the GM1-ELISA, indicating 1G8-1D1 monoclonal antibody reacted specifically with LT but not with CT. 4. The addition of 0.1ml of ascites to 0.6mg and 0.12mg of LT decreased the vascular permeability factor to 41% and 44% respectively, but it did not completely neutralize LT. 5. By double sandwich ELISA using monoclonal antibody, as little as 75ng of the purified LT per ml could be detected. 6. The results by assay of detecting LT in culture supernatants of 14 wild strains E. coli isolated from diarrhea patients by the double sandwich ELISA were almost the same level as those by reverse passive latex agglutination.

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