• Title/Summary/Keyword: enzyme assay

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A Study on the Determinants of Liporotein(a) Level - Primary, Middle, and High School Teachers in Seoul - (혈액내 Lipoprotein(a) 농도의 결정요인에 관한 연구 - 서울시 일부 초, 중, 고교 교사들을 대상으로 -)

  • 최현오;이용욱;나승식;이후장
    • Journal of Food Hygiene and Safety
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    • v.13 no.2
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    • pp.155-163
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    • 1998
  • The purpose of this study was to determine the association between blood lipids, fibrinogen, fasting blood sugar, lifestyle-related factors and lipoprotein(a). This study was performed with 140 healthy adults (male:80, female:60) among the teachers in primary, middle, and high school in Seoul in November 1996. Lipoprotein(a) concentration was measured with an enzyme-linked-immunosorbents assay(ELISA), and the results showed that the distribution was highly skewed and the average concentrations of male and female were $16.94{\pm}12.70\;mg/dL\;and\;17.92{\pm}7.58\;mg/dL$, respectively. The percentage of subject with lipoprotein(a) concentrations higher than 30 mg/dL was 26.4%. Univariate analysis showed significant association between lipoprotein(a) and age, alcohol, meat, and NaCl consumption (p<0.05). In multivariate analysis, lipoprotein(a) correlated positively with age (p<0.05) and meat consumption (p<0.01). These data suggest that lifestyle-related factors may affect the lipoprotein(a) concentration regarded as a risk factor of atherosclerosis disease.

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The Role of Specific IgG in Phagocytosis of Actinobacillus(Haemophilus) Actinomycetemcomitans Y4 by Human Neutrophils (다형핵 백혈구의 Actinobacillus actinomycetemcomitans Y4 균주 탐식시 특이항체의 역할)

  • Kim, Jin-Myung;Chung, Chong-Pyoung;Lee, Young-Hee;Lee, Jin-Yong
    • The Journal of the Korean Society for Microbiology
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    • v.21 no.4
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    • pp.441-446
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    • 1986
  • Previous studies have demonstrated that phagocytosis of encapsulated bacteria needs the opsonization of bacteria with immunoglobulin and complement. Several investigators have studied the role of specific antibody to the bacteria. The purpose of this study is to investigate the role of specific anti-Actinobacillus actinomycetemcomitans Y4($A{\alpha}Y4$) antibody, which was obtained from the immunized rabbit serum for phagocytosis of $A{\alpha}Y4$ by PMNL. For this study, specific and nonspecific IgG were separated from the sera of the rabbits and PMNL were isolated from 15 healthy adults. By an enzyme-linked immunosorbent assay, the results showed that the binding capacity of anti-$A{\alpha}Y4$ IgG to $A{\alpha}Y4$ was much higher than that of nonspecific IgG; 0.75 and 0.14(O.D. at 400nm), respectively. The oxygen consumption of PMNL, phagocytizing $A{\alpha}Y4$ which was opsonized with specific $A{\alpha}Y4$ IgG(37.13 nmol/min/$1{\times}10^7$ PMNL), was significantly higher than that with nonspecific IgG(27.95 nmol/min/$1{\times}10^7$ PMNL, p<0.01). In immunofluorescence microscopic examination, the difference between the numbers of the ingested $A{\alpha}Y4$ opsonized with specific anti-$A{\alpha}Y4$ IgG and nonspecific IgG reached to statistically significant level; $184{\pm}11.4$ and $133.2{\pm}8.3$ per 100 PMNL, p<0.05. These results suggest that specific anti-$A{\alpha}Y4$ IgG has a significant role in PMNL phagocytosis of encapsulated $A{\alpha}Y4$ and also it can be available to adopt this system to develop anti-capsular antibody to $A{\alpha}Y4$ for enhancing and emphasizing the phagocytic activity against this bacterium.

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Allopurinol Decreases Liver Damage Induced by Dermal Scald Burn Injury (피부 화상으로 유도된 간 손상에서 Allopurinol의 효과)

  • Cho, Hyun-Gug;Yoon, Chong-Guk;Park, Won-Hark
    • Applied Microscopy
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    • v.31 no.1
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    • pp.37-47
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    • 2001
  • In order to investigate a pathogenesis of liver damage induced by skin burn, thermal injury was induced by scald burn on entirely dorsal surface in rats (total burn surface area $20\sim25\%$) except for inhalated injury. At 5 and 24 h after scald burn, biochemical assay and morphological changes in serum and liver tissue were examined. Skin burn increased liver weight (% of body weight, p<0.05) and the activity of serum aniline amino-transferase (ALT, p<0.05), in addition, the activity of xanthine oxidase (XO), an enzyme of oxygen free radical generating system, was elevated (p<0.01) in serum, but not in skin and in liver. Postburn treatment of allopurinol intraperitoneally decreased liver weight, serum ALT activity and serum XO activity. Scald burn induced ultrastructurally swelling of endoplasmic reticulum, ribosome detachment, accumulation of lipid, dilatation of bile canaliculi and intercellular space, neutrophil infiltration, activation of Kupffer's cells and degeneration of hepatocytic microvilli. Futhermore , thermal injury decreased not only the protein concentration in plasma but also the number of intravascular leukocytes, that indicates induction of edema formation with protein exudation and inflammation by neutrophil infiltration into the internal organs. However allopurinol injection after burn inhibited post burn ultrastructural changes. These data suggest that acute dermal scald burn injury leads to liver damage, that is related to elevation of xanthine oxidase activity in serum. Xanthine oxidase may be a key role in the pathogenesis of liver damage induced by skin burn.

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Enhanced Antioxident Effect by over Expression of Tomato β-carotene Hydroxylase Gene (ChyB) Using Agrobacterium-infiltration in Tobacco Plant (Agro-infiltration을 이용한 토마토 β-carotene hydroxylase 유전자(ChyB) 과발현 및 담배식물체의 항산화 효과 증진)

  • Choi, Yoon-Jeong;Yoon, Kyung-Young;Yun, Hae-Keun;Suh, Sang-Gon;Moon, Yong-Sun
    • Horticultural Science & Technology
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    • v.29 no.3
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    • pp.267-272
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    • 2011
  • Several reports indicated that astaxanthin and zeaxanthin have more active anticancer activity than pro-vitamin A carotenes. ${\beta}$-carotene hydroxylase is a key enzyme to synthesize zeaxanthin and astaxanthin in carotenoids biosynthesis pathway. We isolated the ChyB gene encoding ${\beta}$-carotene hydroxylase from tomato leaves. The ChyB gene (1.5Kbp) fragment was cloned into the binary vector and designated to pIG121-ChyB-tom. Agrobacterium-mediated infiltration was used for transient assay in Nicotiana benthamiana. Leaf samples were collected 0, 1, 2, 3 days after infiltration (DAI). RT-PCR result showed that the expression of ${\beta}$-carotene hydroxylase transcripts was not detected in control (0DAI), but its expression was detected after 1 DPI and increased later on. When the activity of ${\beta}$-carotene hydroxylase was measured, the 1,1-diphenyl-pricryl hydrazyl (DPPH) radical scavenging activity (27%) at 2 DAI was significantly higher than that (21%) at 0 DAI. These results indicated that anti-oxidant activity dramatically increased at 2 DAI in tobacco leaves was due to over expression of tomato ${\beta}$-carotene hydroxylase. These results can be the foundation to develop tomato cultivars with high oxy-carotenoids content using the ChyB gene transformation.

Production and Characterization of Monoclonal Antibodies Specific to PAT Protein Expressed in Genetically Modified Herbicide-Resistance Maize (제초제 내성 유전자 변형 옥수수 중 PAT단백질에 특이한 단크론성 항체의 생산과 특성 확인)

  • Kim, Sol-A;Lee, Jeong-Eun;Shim, Won-Bo;Kang, Sung-Jo;Chung, Duck-Hwa
    • Journal of Food Hygiene and Safety
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    • v.33 no.3
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    • pp.193-199
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    • 2018
  • In this study, PAT protein of genetically modified maize was prepared from the recombinant E. coli strain BL21 (DE3), and mice were immunized with the recombinant PAT protein. After cell fusion and cloning, two hybridoma cells (PATmAb-7 and PATmAb-12) were chosen since the monoclonal antibodies (Mabs) produced by them were confirmed to be specific to PAT protein in the indirect enzyme-linked immunsorbent assay (ELISA) and western blot tests. There were no cross-reactions of either Mabs to other GM proteins or to the extracts of non-GM maize. The ELISA based on the PATmAb-7 can sensitively detect 0.3 ng/g PAT protein in corn. These results indicate that the developed Mabs can be used as bio-receptors in the development of immunosensors and biosensors for the rapid and simple detection of GM corn adulterated in foods.

Evaluation of stress-induced analgesia in acupuncture analgesic effect - An Approach on diameters of acupuncture needles and acupuncture point needlings - (침진통 효과에서 스트레스에 의한 진통 효과 개입 여부 평가 - 침굵기, 침자극 부위를 중심으로 -)

  • Jeong, Byeong-Ju;Choe, Il-Hwan;Shin, Hee-Sup;Lim, Sabina
    • Korean Journal of Acupuncture
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    • v.25 no.3
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    • pp.65-80
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    • 2008
  • Objective : In recent years, many investigators have questioned whether the analgesic effect of acupuncture is simply related to the stress-induced analgesia (SIA). However, there has been lack of studies on this issue. In this study, the stress levels induced by manual acupunctures are compared with the stress in animal experiment models. The experiments have been carried out with Sprague Dawley (SD) rats. Method : For stress level evaluation, Hot plate test has been used. Maximum Possible Effect (MPE) has been measured by checking the pre-test time and post-test time. Cortisol and corticosterone concentrations in serum were measured by enzyme-linked immunosorbent assay (ELISA). Results : In the hot plate test, MPE values of post-test time were significantly decreased after 10 minutes than after 5 minutes. Therefore, optimal time interval was chosen as 10 minutes. There was significant difference of MPE values between Suspension group and all other treatment groups. However, there were no significant differences of MPE values between Sham group and all other treatment groups. However, MPE values showed tendency to decrease when acupuncture needle diameter increased. MPE values of ST040, ST040(lido), NAP040(lido) groups were markedly decreased than that of Suspension group, while that of NAP040 group was substantially increased than that of Sham group increased in acupoint and nonacupoint models. Serum cortisol concentrations of treatment groups were not significantly different from that of Suspension and Sham groups. Serum corticosterone concentration of 0.25 mm group was substantially increased than that of compared with Sham group. Serum cortisol and corticosterone concentrations of treatment groups were not significantly different from those of Suspension and Sham groups in acupoint and nonacupoint models. Conclusion : From hot plate test and serum stress hormones concentrations, it is found that manual acupuncture treatment induces negligible stress or SIA on ST36. And the stress induced by manual acupuncture is more closely related to acupuncture point needlings than diameters of acupuncture needles.

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Promoter Cloning of Human SETDB1 Gene Utilizing Bioinformatic Programs (생물정보 프로그램을 활용한 SETDB1 유전자 프로모터 클로닝)

  • Noh, Hee-Jung;Kim, Keun-Cheol
    • Journal of Life Science
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    • v.24 no.1
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    • pp.1-7
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    • 2014
  • Eukaryotic gene expression is an important process, which is initiated by several transcription factors and RNA polymerases that occupy the promoter region of genomic DNA. Although there are many experiments to identify the promoter region in a gene, it is time and labor consuming to finalize it. In this study, we utilized bioinformatic programs, including Ensembl, NCBI, and CpG plots, to identify the cloning promoter region in SETDB1 genomic DNA. We performed PCR amplification to obtain the SETDB1 promoter on an approximately 2 kb region upstream from the TSS named SETDB1-P1. The PCR product was ligated with TA cloning vectors, and we confirmed the insert size using restriction enzyme digestion. Sequentially, the insert was subcloned into a pGL3-luc vector to produce pGL3-SETDB1- P1-luc and then confirmed by DNA sequencing. We also obtained a fragmented PCR product called P2 and P3 and performed a luciferase assay using pGL3-SETDB1-P1-luc transfection. We found that several anticancer drugs, including taxol, 4-FU, and doxorubicin, decreased the promoter activity of SETDB1. We obtained consistent data on the regulation of SETDB1 gene expression after anticancer drug treatment using Western blot analysis and RT-PCR. Our results suggest that promoter cloning of the human SETDB1 gene utilizing bioinformatics is a very useful and timesaving approach to study gene expression.

Anti-inflammatory Effect of Gagam-GongJin-dan in mouse peritoneal macrophages (마우스 복강대식세포에서 가감공진단(加減拱辰丹)의 항염증 효과)

  • Kim, Hong-Jun;Kim, Young-Sik;Mok, Ji-Ye;Jeong, Seung-Il;Hwang, Sung-Yeoun;Cho, Jung-Keun;Jang, Seon-Il
    • Herbal Formula Science
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    • v.19 no.1
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    • pp.207-217
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    • 2011
  • Objectives : In a previous study, we have shown that Gagam-Gongjin-Dan(GGD) has an inhibitory effect on the ovalbumin-induced immune responses and a hepatoprotective effect on actaminophen-induced liver injury in Balb/c Mice. However, the possible anti-inflammatory effect of GGD extract for inflammatory mediators was not reported. Therefore, the purpose of this study was to investigate an inhibitory effects of GGD extract against lipopolysaccharides(LPS) induced inflammatory mediators in mouse peritoneal macrophages. Methods : GGD extract was prepared by extracting with methanol for 7 days. The extract was freeze-dried following filtration through vacuum distillation system. Accumulated nitrite, an oxidative product of nitric oxide(NO), was measured in the culture medium by the Griess reaction. The levels of prostaglandin $E_2(PGE_2)$, interleukin-$1{\beta}$(IL-$1{\beta}$), tumor necrosis factor-${\alpha}$(TNF-${\alpha}$) were measured by enzyme-linked immunosorbent assay. The expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2(COX-2) were measured by Western blot analysis. Results : GGD extract (50-$400\;{\mu}g$/ml) per se had no cytotoxic effect in LPS-stimulated peritoneal macrophages. GGD extract dose-dependently reduced NO, $PGE_2$, IL-$1{\beta}$ and TNF-${\alpha}$ production and COX-2 activity caused by stimulation of LPS. The levels of iNOS and COX-2 protein expressions were markedly suppressed by the treatment with GGD extract in a dose dependent manner. Conclusions : These results suggest that GGD extract has an anti-inflammatory effect against LPS-induced inflammatory mediators in peritoneal macrophages, these properties may contribute to inflammation disease care.

Anti-inflammatory effect of ethanol extract from Ecklomia cava on gram-positive bacteria(Peptidoglycan)-induced macrophages (그람-양성균(Peptidoglycan)으로 유도한 대식세포에서 감태 에탄올 추출물의 항염증 효과)

  • Kang, Ok-Hwa;Kim, Sung-Bae;Keum, Joon-Ho;Mun, Su-Hyun;Kim, Yong-Sik;An, Byung-Kwan;An, Hyeon-Jin;Kwon, Dong-Yeul
    • Herbal Formula Science
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    • v.19 no.1
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    • pp.195-205
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    • 2011
  • Objectives : Ecklonia cava is brown alga(Laminariaceae) which grows is sea, it has antioxidant, diarrhea and anticoagulant effect. In this study, the effect of ethanol extract of Ecklonia cava (EC) on peptidoglycan(PGN) -induced NO production in RAW 264.7 cells was investigated. Methods : In the present study, IL-6 production was measured using the enzyme-linked immunosorbent assay(ELISA), prostaglandin $\E_2$($\PGE_2$) production was measured using the EIA kit, and inducible NO synthase(iNOS), cyclooxygenase-2(COX-2), and mitogen-activated protein kinase(MAPK) activation, as determined by western blot analysis and reverse transcription -polymerase chain reaction(RT-PCR). Results : EC inhibited PGN-induced NO and IL-6 production. Consistent with these observations, the protein expression of iNOS and COX-2 were inhibited by EC. Moreover, EC suppressed the phosphorylation of extracellular signal-regulated kinase(ERK) 1/2 in PGN-induced RAW 264.7. Conclusions : These results suggest that EC has inhibitory effects on PGN-induced $\PGE_2$, NO, and IL-6 production, as well as the expressions of iNOS and COX-2 in the murine macrophage. These inhibitory effects occur through blockades on the MAPKs phosphorylation.

PRODUCTION OF GM-CSF AND TGF-${\beta}1$ IN IRRADIATED HUMAN GINGIVAL FIBROBLASTS CULTURED WITH LIPOPOLYSACCHARIDE (Lipopolysaccharide로 자극시킨 방사선 조사 치은 섬유아 세포에서 granulocytemacrophage colony-stimulating factor와 transforming growth factor-${\beta}1$ 생성)

  • Kim, Hong-Sik;Lee, Seong-Geun;Kim, Kwang-Hyuk;Kim, Uk-Kyu;Kim, Jong-Ryoul;Chung, In-Kyo;Yang, Dong-Kyu
    • Journal of the Korean Association of Oral and Maxillofacial Surgeons
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    • v.28 no.3
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    • pp.169-174
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    • 2002
  • Purpose: Irradiation in the oral cancer patients causes early and late complications such as intraoral mucositis and fibrosis, with a various expression of GM-CSF and TGF-${\beta}1$. The purpose of this study was to investigate the production of GM-CSF and TGF-${\beta}1$ by the irradiated human gingival fibroblasts cultivated with lipopolysaccharide. Materials and Methods: Irradiated (total dose, 60 Gy) human gingival fibroblasts were incubated with LPS. Culture supernatants that were collected at 24, 48, and 72 hours were assessed for GM-CSF and TGF-${\beta}1$ by enzyme-linked immunosorbent assay. Results: 1. GM-CSF production in nomal gingival fibroblasts was increased with incubation time, but decreased with incubation time in irradiated gingival fibroblasts. GM-CSF production in both normal and irradiated gingival fibroblasts induced with LPS was higher than the control. 2. TGF-${\beta}1$ production in normal gingival fibroblasts was decreased after 24 hours, but, it was increased until 48 hours in irradiated gingival fibroblasts. TGF-${\beta}1$ production in normal gingival fibroblasts exposed with LPS was higher than the control. Conversely, It was lower than the control in irradiated gingival fibroblasts exposed with LPS. Conclusion: This indicates that irradiation in gingival fibroblasts may play an important role in radiation-induced intraoral mucositis and fibrosis. However, LPS decreases the production of TGF-${\beta}1$ in the irradiated gingival fibroblasts.