노화가 인체 중간엽 줄기세포로부터 조골세포로의 증식 및 분화에 미치는 영향

백기현,태현정,오기원,이원영,조정기,권순용,강무일,차봉연,이광우,손호영,강성구,김춘추 대한내분비학회 2003 Endocrinology and metabolism Vol.18 No.3

연구배경: 일반적으로 골다공증과 연관된 위험인자로는 연령, 폐경, 약물, 불충분한 칼슘섭취, 만성질환 및 운동부족 등이 있는데, 특히 노화가 진행할수록 골밀도가 감소하는 것은 잘 알려져 있다. 노화와 관련하여 진행되는 골소실은 조골세포 및 전구조골세포의 기능적 결핍에 의한 골형성의 감소가 주요한 요인으로 여겨지고 있다. 그 동안 연령이 조골모 세포의 양과 조골모 세포로부터 성숙조골세포로의 분화 및 증식에 미치는 영향에 대한 일부 보고들이 있었으나 아직 일치된 견해는 없는 형편이다. 방법: 다양한 연령의 사람으로부터 골수를 채취, 중간엽 줄기세포가 포함된 단핵세포를 분리한 후 조골세포로 분화하기 좋은 조건하에서 배양하였다. 대상군은다시 젊은군과 노령군으로 구분하여 다양한 변수를 비교 분석하였다. 일차배양에서는 CFU-F를 계수하여 골수내 중간엽 줄기세포의 수를 추산하였고, 칼슘측정을 통하여 기질의 무기화 정도를 비교하였다. 계대배양후 이차배양에서는 시기별로 알카리성 포스파타제 활성도를 측정하고 오스테오칼신 mRNA의 발현을 관찰하여 젊은군과 노령군 사이의 증식능 차이를 비교하였다. 또한 이차배양 시기별로 MTT 측정을 하여 양군간에 증식능 차이가 있는지 알아보았다. 결과: 1. 일차배양 15일째에 평균 CFU-F의 수는 젊은군에서 유의하게 많았다(젊은군 148.3±28.9, 노령군 54.3±9.1, p=0.02). CFU-F의 평균면적은 젊은군에서 넓은 경향을 보였으나 통계적으로 유의하지는 않았다. 2. 일차배양 17일 경과 후 양군간에 기질 칼슘 침착정도는 유의한 차이를 보이지 않았다(젊은군 103.6±50.6, 노령군: 114.0±56.5, p=NS). 3. 이차배양 10일째에 젊은군에서 알카리성 포스파타제 활성도가 고령군에 비해 유의하게 높았다(젊은군: 935.5±115.0 U/mg, 노령군: 578.4±115.7U/mg,p.0.05). 고령군에서는 시간 경과에 따른 변화가 미약했으며 전반적으로 알카리성 포스파타제의 활성도가 젊은군에 비해 낮았다. 4. 이차배양도중 오스테오칼신 mRNA의 발현은 배양시기별로 젊은군에 비해 고령군에서 더 낮은 경향을 관찰할 수 있었다. 5. 이차배양 10일과 15일에 젊은군에서 노령군보다세포증식이 유의하게 증가된 양상을 보였다(10앓 젊은군 0.73±0.05, 노령군 0.58±0.04, p=0.05, 15일; 젊은군 0.80±0.05, 노령군 0.70±0.03, p=0.05).결론: 이상의 연구에서 저자들은 노령군에서 젊은군보다 골수 내 중간엽줄기세포의 수가 적고, 노령군에서 유래한 전구조골세포의 성숙조골세포로의 증식 및 분화가 젊은군 보다 감소해 있는 것을 관찰할 수 있었다. Background: Osteoblasts originate from osteoprogenitor cells in bone marrow stroma, termed mesenchymal stem cells (MSCs) or bone marrow stromal cells. Each MSC forms colonies (colony forming units-fibroblasts [CFL-Fs]) when cultured ex vivo. There are some reports about the age-related changes of the number and osteogenic potential of osteoprogenitor cells, but any relationship has not been clearly established in humans. In this study, we counted MSCs using CFU-Fs count and examined the proliferative capacity and differentiation potential of osteoprogenitor cells. Finally, we analyzed how these parameters varied with donor age. Methods: Bone marrow was obtained from the iliac crest of young (n=6, 27.2±8.6 years old) and old (n= 10, 57.4k6.7 years old) healthy donors. Mononuclear cells, including MSCs, were isolated and cultured in osteogenic medium. In primary culture, we compared the colony-forming efficiency of MSCs between the two groups and determined the matrix calcification. When primary culture showed near confluence, the cells were subcultured. Alkaline phosphatase activity, osteocalcin expression by RT-PCR and proliferative potential by MTT assay were examined by the time course of secondary culture. Results: At the 15th day of primary culture, the mean number of CFU-Fs was significantly higher in the younger donors (young: 148.3±28.9, old: 54.3±9.1, p=0.02) and the mean size of CFL-Fs was also larger in the younger donors than the older donors. However, matrix calcification was not different between the two groups (young: 103.6±50.6, old: 114.0±56.5, p=NS). In secondary culture, alkaline phosphatase activities were significantly lower in the older donors. The younger donors showed peak alkaline phosphatase activity at day 10, while the older donors didn't showed a remarkable peak (young: 935.5±115.OU/mg, old: 578.4±115.7U/mg, p<0.05). Total cell number as a proliferative index increased progressively during the secondary culture and a significantly greater cell number was noted in the younger donors. Osteocalcin expression was generally upregulated in the younger donors, but this was not statistically significant. Conclusion: Our study shows that the number of osteoprogenitor cells is decreased during aging and that the proliferative capacity and differentiation potential of osteoprogenitor cells seem to be reduced during aging (J Kor SOC Endocrinol 18:296-305, 2003).

돼지고기육종의 방사선 유래 휘발성 물질 분석

차용준,박지영,김훈,김소정,정연정,소양순,유영재 창원대학교 생활과학연구소 2000 생활과학연구 Vol.4 No.-

Bacillus cereus KCTC 3674 produced at least two kinds of extracellular proteases. Two major bands of the protease activity with molecular weights of approximately 100 and 38 kDa were obtained after gelatin-SDS-PAGE. The production of extracellular proteases occurred during the logarithmic growth phase and was greatest when cultures reached the stationary growth phase. On the other hand, the growth of B. cereus KCTC 3674 and the production of its extracellular proteases were severely inhibited by the addition of protonophore carbonylcyanide m-chlorophenylhydrazone (CCCP). Thus, it was concluded that the growth of B. cereus KCTC 3674 and the production of its extracellular pretenses were strongly affected by a H+ electrochemical potential.

Tomato 中의 Lycopenen의 檢索

車榮華,鄭順子 梨花女子大學校 藥學硏究所 1961 梨花藥學硏究 Vol.- No.2

퉁퉁마디로부터 염에 의하여 유도되는 Aldolase 유전자의 분리 및 발현분석

차준영,네티 엘마와티,김순길,이증주,임채오,정우식,이곤호,손대영 Plant molecular biology and biotechnology research 2003 Plant molecular biology and biotechnology research Vol.2003 No.-

Soil salinity is one of the most serious abiotic stresses limiting the productivity of agricultural crops. To cope with salt stress, plants respond with physiological, developmental and biochemical changes, including the synthesis of a number of proteins and the induction of gene expression. Salicornia herbacea is a halophytic plant that grows in salt marches and on muddy seashores. In order to understand the biochemical and molecular mechanisms of salt tolerance in S. herbacea, we isolated several genes that involved in the salt tolerance by mRNA differential display.Here we report the cloning of a cDNA encoding fructose-1, 6-bisphosphate aldolase, named ShADL, which is 1293 bp long and contains an open reading frame consisted of 359 amino acids with calculated molecular mass of 39 kDa. ShADL protein showed 86% identity with Arabidopsis and 78% with aldolase of common ice plant. Northern blot analysis revealed that the transcript of ShADL gene was increased dramatically depending on the NaCI concentrations.

PCR 방법에 의한 Renin cDNA의 증폭합성과 클로닝을 통한 유전자 발현

차영주,이숙영,김성준,박영순 조선대학교 부설 유전생물공학연구소 1991 유전생물공학연구지 Vol.1 No.1

본 실험은 생쥐 악하선의 renin mRNA로부터 cDNA를 합성하였으며 분자 육종을 통하여 rennin cDNA의 유전자 발현을 유도하여 얻은 결과는 다음과 같다. 생쥐 악하선으로부터 총RNA를 GIT/CsCl 방법에 의하여 분리하고 oligo(dT) cellulose와 poly(U) sepharose 4B matrix를 이용하여 mRNA를 분리하였다. Renin cDNA로부터 primer Ⅰ과 primer Ⅱ를 합성하여 polymerase chain reaction(PCR)방법으로 cDNA를 합성하고 증폭하였다. cDNA를 pUC19 플라스미드에 ligation시켜 재조합 플라스미드(pUCR1)를 제조하였다. pUCR1을 E. coli JM103 균주에 형질전환시켰으며 rennin 유전자를 갖는 형질전환체를 Southern blot analysis 방법에 의하여 확인하였다. 형질전환체인 E. coli JM103에서 renin 유전자의 발현물질인 분자량이 45,000되는 rennin 단백질을 확인하였다. These experiments were designed to investigate the molecular cloning of renin cDNA synthesized from renin mRNA of mouse submaxillary gland, and thus to characterize the renin protein expressed by the renin cDNA. The results obtained were as follows: The total RNA was isolated from the mouse submaxillary gland by the method of GIT/CsCl, and the mRNA was separated by using oligo(dT) cellulose and poly(U) sepharose 4B matrix. Primer Ⅰ and primer Ⅱ was synthesized from renin cDNA, and cDNA was transcribed reversely and amplified by the method of polymerase chain reaction(PCR). The recombinant plasmid(pUCR1) was constructed by ligating the cDNA into the pUC19 plasmid of E. coli vector, and then pUCR1 was transformed into E. coli(JM103) and the recombinant plasmid was identified with southern blot analysis. The renin gene expression was identified with polyacrylamide gel electrophoresis and the molecular weight of renin was 45,000.

쇠고기육중의 방사선 유래 휘발성물질의 분석

차용준,박지영,김훈,박승영,조우진,송양순,유영재 창원대학교 생활과학연구소 2000 생활과학연구 Vol.4 No.-

This study was conducted to identify irradiation-derived volatile components, having positive-correlation with irradiation doses, which may play roles as maker materials for detecting post-irradiation in pork meat. The volatile components of irradiated(0, 1, 3, 5 and 10 kGy doses) pork(Belly) meat was analyzed by liquid liquid continuous extraction(LLCE)/GC/MSD methods. By the results of linear regression analysis between irradiation doses and volatile component amounts in fresh pork, 3 compounds including γ-octalactone(r=0.82), 1-octen-3-ol(r=0.90) and butanoic acid(r=0.81) had high correlations with irradiation doses.

Cimetidine과 Phenobarbital 이 Acetaminophen에 依한 急性 肝毒性에 미치는 影響에 關한 硏究

車淳道,姜大榮 충남대학교 의과대학 지역사회의학연구소 1988 충남의대잡지 Vol.15 No.1

In an attempt to elucidate the biochemical mechanism and efffects of phenobarbital and cimetidine on acetaminophen-induced hepatic necrosis, the present study in rats was undertaken to evaluate the relation between serum transaminase and pathologic change of the liver at 3, 6, 12 and 24 hours after acetaminophen administration with pretreatment of phenobarbital and cimetidine. The results were as follows: 1. Twenty--four hour mortalities in acetaminophen 600mg/kg administrated groups are 3..3% for the control group, 3.3% for cimetidine pretreatment group and 16.6% for the phenobarbital pretreatment group, respectively. In acetaminophen 1200mg/kg administrated groups, the mortalities are 33.3%, 6.6% and 43%, respectively. 2. The sGOT activity is increased at 3 hours after acetaminophen administration, and is in proportion to the dose of the drugs. In animals pretreated with cimetidine the elevation of the sGOT is decreased, but is increased in phenobarbital preated groups. The change of the sGPT is similiar to the sGOT, and reveal striking elevation at 24 hours after acetaminophen administration. 3. In animals receiving acetaminophen 600mg/kg, the necrotic change of the liver is not observed in the cimetidine pretreated group, but present in the phenobarbital pretreated group at 12 hours afterr acetaminophen administration. 4. In animals receiving acetaminophen 1200mg/kg, necrotic change is well demonstrated in the experimental and control group at 12 hours and 24 hours after acetaminohen administration. The severity of necrotic change is mild in the cimetidine pretreated group but severe in the phenobarbital. pretreated group in contrast to the control group. In summary, the results obtained by the present study indicate the severity of hepatic necrosis in the acetaminophen overdose treated rats is in proportion to the dose, increased by Phenobarbital and decreased by cimetidine.

DMBA에 의해 유도된 생쥐 생식기 종양의 세포학적 연구

차동수,한상원,권장연,정인배,이영진,한혁동,정순희,등영건 연세대학교 원주의과대학 1993 원주의대논문집 Vol.6 No.1

STZ 유발 당뇨쥐에서 시간에 따른 생리적 항산화계의 변화

이순재,양정아,김성옥,최정화,신주영,채영미,차복경 대구효성가톨릭대학교 응용과학연구소 1997 응용과학연구논문집 Vol.5 No.-

The purpose of this study was to investigate change of physiological antioxidative system according to the time in streptozotocin-induced diabetic rats. Sprague-Dawley male rats weighing 150±10gm were randomly assigned to one normal and five STZ-induced diabetic groups. Diabetic groups were classified to experimental period. Diabetes was experimentally induced by intravenous injection of 55 mg/kg of body weight of STZ in citrate buffer(PH 4.3) after 6 weeks feeding of three experimental diets. Animals were sacrificed at 0, 3, 6, 12, 18th days of diabetic states. 1. Activity of superoxide dismutase(SOD) in liver was increased in that of diabetic mellitus(DM) groups at 3th day but that of DM groups was not significant from 6th day according to period. Glutathion peroxidase(GPX) was significantly decreased in DM groups from 6th day 2. Reduced glutathione(GSH) contents in liver significantly decreased in diabetic groups from 3rd day compared with those of DM groups according to the period. Oxidized glutathione(GSSG) was higher from 6th day. GSH/GSSG ratio was significantly lower than that of normal group from 3rd day to all experimental period 3. Contents of vitamin E in liver of DM groups were significantly decreased compared with that of normal group from 6th day. 4. Lipid peroxide(LPO) contents in liver of DM groups were significantly increased compared with that of normal group from 3rd day. The present results indicate that STZ-induced diabetic rats were reduced by antioxidative defense system and taken by peroxidate damage in tissue compared with normal group from 3rd day or 6rd day after injection STZ. It lended to acceleration all diabetic groups but no significance according to the experimental time.

IMF시기 중소건설업체의 경쟁력 요소 분석과 경영전략에 관한 硏究 : 光州, 全南지역의 건설會社를 中心으로

강영순,차강석,김용수 中央大學校 建設環境硏究所 1999 環境科學硏究 Vol.10 No.-

This study aims at suggesting competition factors and management strategies necessary for survival and continuous growth of small and medium-size construction companies during IMF period. For this study, both literature and questionnaire survey have been performed, figuring out major competition factors and management strategies before and after IMF period. For the questionnaire survey, small and medium-size construction companies located in Kwangju city and Chounnam province are utilized The results of this study are as follows: 1) The major competition factors for the well-managed small and medium-size construction firms after IMF period are surveyed as following order ; ability of CEO, project performance, risk control, marketing, procurement, and strategic planning capability. 2) The degree of significance in terms of management strategies for the firms depicted above are discovered as following order ; quality enhancement and pursuing high value-added products, betterment andrationalization of management system, strenthening technical development, training and utilization of human resources, diversification and conversion of business structure, and drive forwards internationalization.