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Mechanism underlying NO-induced apoptosis in human gingival fibroblasts

  • Hwang, In-Nam;Jeong, Yeon-Jin;Jung, Ji-Yeon;Lee, Jin-Ha;Kim, Kang-Moon;Kim, Won-Jae
    • International Journal of Oral Biology
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    • v.34 no.1
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    • pp.7-14
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    • 2009
  • Nitric oxide (NO) acts as an intracellular messenger at the physiological level but can be cytotoxic at high concentrations. The cells within periodontal tissues, such as gingival and periodontal fibroblasts, contain nitric oxide syntheses and produce high concentrations of NO when exposed to bacterial lipopolysaccharides and cytokines. However, the cellular mechanisms underlying NO-induced cytotoxicity in periodontal tissues are unclear at present. In our current study, we examined the NO-induced cytotoxic mechanisms in human gingival fibroblasts (HGF). Cell viability and the levels of reactive oxygen species (ROS) were determined using a MTT assay and a fluorescent spectrometer, respectively. The morphological changes in the cells were examined by Diff-Quick staining. Expression of the Bcl-2 family and Fas was determined by RT-PCR or western blotting. The activity of caspase-3, -8 and -9 was assessed using a spectrophotometer. Sodium nitroprusside (SNP), a NO donor, decreased the cell viability of the HGF cells in a dose- and time-dependent manner. SNP enhanced the production of ROS, which was ameliorated by NAC, a free radical scavenger. ODQ, a soluble guanylate cyclase inhibitor, did not block the SNP-induced decrease in cell viability. SNP also caused apoptotic morphological changes, including cell shrinkage, chromatin condensation, and DNA fragmentation. The expression of Bax, a member of the proapoptotic Bcl-2 family, was upregulated in the SNP-treated HGF cells, whereas the expression of Bcl-2, a member of the anti-apoptotic Bcl-2 family, was downregulated. SNP augmented the release of cytochrome c from the mitochondria into the cytosol and enhanced the activity of caspase-8, -9, and -3. SNP also upregulated Fas, a component of the death receptor assembly. These results suggest that NO induces apoptosis in human gingival fibroblast via ROS and the Bcl-2 family through both mitochondrial- and death receptor-mediated pathways. Our data also indicate that the cyclic GMP pathway is not involved in NO-induced apoptosis.

Determination of Niacin in Infant Formula by Solid-phase Clean-up and HPLC with Photodiode Array Detector (고체상 정제 및 HPLC/PDA에 의한 영유아식 중 나이아신의 분석)

  • Hong, Jee-Eun;Kim, Mi-Ran;Cheon, Sang-Hee;Chai, Jung-Young;Park, Eun-Ryong;Mun, Chun-Sun;Gwak, In-Shin;Kim, Ok-Hee;Lee, Kwang-Ho
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.38 no.3
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    • pp.359-363
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    • 2009
  • This study was performed to establish a rapid and simple analytical method for niacin (nicotinic acid and nicotinamide) using HPLC. A pretreatment method for the extraction and clean-up of niacin in infant formula sample and an instrumental condition for HPLC were optimized. Niacin was extracted by 5 mM hexanesulfonate with ultrasonication for 30 min. For the clean-up of the sample, the extract was applied to a HLB cartridge, and then niacin was eluted from the cartridge using 5 mL of 80% methanol after washing with 5 mL of n-hexane. The total recoveries were $83{\sim}104%$ and relative standard deviation were in the range of $1.5{\sim}3.5%$ during the extraction and clean-up process. Niacin was determined by gradient elution with sodium hexanesulfonate/methanol buffer as a mobile phase and a photodiode array detector (260 nm). It showed a high linearity between the content of niacin and the peak area ($r^2$=1.000) in the range of $0.02{\sim}10.0$ mg/L of nicotinic acid and nicotinamide. The detection limit was 0.02 mg/L (0.2 mg/kg in the sample). The method was successfully applied for the determination of niacin in infant formula. Total niacin contents were in the range of $53.5{\sim}140.3$ mg/kg.

Prognostic Factor Analysis of Overall Survival in Gastric Cancer from Two Phase III Studies of Second-line Ramucirumab (REGARD and RAINBOW) Using Pooled Patient Data

  • Fuchs, Charles S.;Muro, Kei;Tomasek, Jiri;Van Cutsem, Eric;Cho, Jae Yong;Oh, Sang-Cheul;Safran, Howard;Bodoky, Gyorgy;Chau, Ian;Shimada, Yasuhiro;Al-Batran, Salah-Eddin;Passalacqua, Rodolfo;Ohtsu, Atsushi;Emig, Michael;Ferry, David;Chandrawansa, Kumari;Hsu, Yanzhi;Sashegyi, Andreas;Liepa, Astra M.;Wilke, Hansjochen
    • Journal of Gastric Cancer
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    • v.17 no.2
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    • pp.132-144
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    • 2017
  • Purpose: To identify baseline prognostic factors for survival in patients with disease progression, during or after chemotherapy for the treatment of advanced gastric or gastroesophageal junction (GEJ) cancer. Materials and Methods: We pooled data from patients randomized between 2009 and 2012 in 2 phase III, global double-blind studies of ramucirumab for the treatment of advanced gastric or GEJ adenocarcinoma following disease progression on first-line platinum- and/or fluoropyrimidine-containing therapy (REGARD and RAINBOW). Forty-one key baseline clinical and laboratory factors common in both studies were examined. Model building started with covariate screening using univariate Cox models (significance level=0.05). A stepwise multivariable Cox model identified the final prognostic factors (entry+exit significance level=0.01). Cox models were stratified by treatment and geographic region. The process was repeated to identify baseline prognostic quality of life (QoL) parameters. Results: Of 1,020 randomized patients, 953 (93%) patients without any missing covariates were included in the analysis. We identified 12 independent prognostic factors of poor survival: 1) peritoneal metastases; 2) Eastern Cooperative Oncology Group (ECOG) performance score 1; 3) the presence of a primary tumor; 4) time to progression since prior therapy <6 months; 5) poor/unknown tumor differentiation; abnormally low blood levels of 6) albumin, 7) sodium, and/or 8) lymphocytes; and abnormally high blood levels of 9) neutrophils, 10) aspartate aminotransferase (AST), 11) alkaline phosphatase (ALP), and/or 12) lactate dehydrogenase (LDH). Factors were used to devise a 4-tier prognostic index (median overall survival [OS] by risk [months]: high=3.4, moderate=6.4, medium=9.9, and low=14.5; Harrell's C-index=0.66; 95% confidence interval [CI], 0.64-0.68). Addition of QoL to the model identified patient-reported appetite loss as an independent prognostic factor. Conclusions: The identified prognostic factors and the reported prognostic index may help clinical decision-making, patient stratification, and planning of future clinical studies.

Competitive Extraction of Chlorinated Solvents by Headspace SPME GC/FID (Headspace SPME GC/FID를 이용한 Chlorinated Solvents의 경쟁적 추출효과에 관한 연구)

  • An, Sangwoo;Kim, Youngju;Chun, Sukyoung;Lee, Sijin;Park, Jaewoo;Chang, Soonwoong
    • Journal of the Korean GEO-environmental Society
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    • v.11 no.5
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    • pp.61-67
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    • 2010
  • In this study, Solid-phase microextraction (SPME) with GC/FID was studied as a possible alternative to liquid-liquid extraction for the analysis of chlorinated solvents (PCE and TCE) and these by-products (cis-DCE, VC, and Ethylene). Experimental parameters affecting the SPME process (such as kind of fibers, adsorption time, desorption time, volume ratio of sample to headspace, salt addition, and magnetic stirring) were optimized. Experimental parameters such as CAR/PDMS, adsorption time of 20 min, desorption time of 5 min at $250^{\circ}C$, headspace volume of 50mL, sodium chloride (NaCl) concentration of 25% combined with magnetic stirring were selected in optimal experimental conditions for analysis of chlorinated solvents and these by-products. The general affinity of analytes to CAR/PDMS fiber was high in the order PCE>TCE>cis-DCE>VC>Ethylene. The linearity of $R^2$ for chlorinated solvents and these by-products was from 0.912 to 0.999 when analyte concentrations range from $10{\mu}g/L$ to $500{\mu}g/L$, respectively. The relative standard deviation (% RSD) were from 2.1% to 3.6% for concentration of $500{\mu}g/L$ (n=5), respectively. Finally, the limited of detection (LOD) observed in our study for chlorinated solvents and these by-products were from $0.5{\mu}g/L$ to $10{\mu}g/L$, respectively.

Preparation of Insoluble Dietary Fiber from Forest Waste and Its Physiological Function in Rat Fed High Cholesterol Diets

  • Chai, Young-Mi;Lim, Bu-Kug;Lee, Jong-Yoon;Rhee, Soon-Jae
    • Preventive Nutrition and Food Science
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    • v.7 no.1
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    • pp.78-87
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    • 2002
  • This study investigated the production of insoluble dietary fiber using forest waste and the dietary effect of manufactured insoluble fiber on physiological function in rat fed high cholesterol duets. Insoluble dietary fiber was prepared from the wood chips of oak (Quercus mongolica). The best condition for steam-explosion treatment for the preparation of insonuble dietary fiber was 25 kg/cm$^3$pressure for 6 minutes. In the chemical analysis of insoluble dietary fiber pretreated by 1% sodium hydroxide solution with steam-exploded wood, $\alpha$-cellulose content was 61.7% in the insoluble dietary fiber which contained 7.6% residual lignin. In order to compare insoluble dietary fiber with commercial $\alpha$-cellulose of physiological function, Sprague-Dawley male rats weighing 100$\pm$10 g were randomly assigned to one normal diet and five high cholesterol diet containing 1% cholesterol. The high cholesterol diet groups were classified as fiber free diet (FF group), 5% commercial $\alpha$-cellulose diet (5C group), 10% commercial $\alpha$-cellulose (l0C group), 5% insoluble dietary fiber dict (5M group), and 10% insoluble dietary fiber (10M group). The rats were fed ad libidum for 4 weeks. Food intake, weights gain, and food efficiency ratio in high cholesterol groups were higher than those of normal group, but there were no significant differences between the experimental groups. There were not any significant differences in the weights of livers, kidneys and small intestine of insoluble dietary fiber supplemented groups, but weight of cecum in all insolube dietary fiber group were significantly higher than those of FF group. A gstrointestinal transit time was decreased by supplementation of insoluble dietary fiber. Weight and water contents of feces in the insoluble dietary fiber supplemented groups were significantly higher than those of the FF group. There were not any significant differences in the activities of the glutamic pyruvic transaminase (GPT) and glutamic oxaloacetic transaminase (GOT) between the experimental groups. In conclusion, the manufactured insoluble dietary fiber and commercial insoluble fiber have the same physiological effects. The preparation method of the insoluble dietary fiber from the oak chips suited its purpose.

Development of Seasoned Semi-Dried Oyster (조미 반건조 굴 가공품의 개발)

  • Kim, Hye-Suk;Heu, Min-Soo;Kim, Jin-Soo
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.35 no.10
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    • pp.1475-1483
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    • 2006
  • This study was carried out to examine the optimum processing conditions of seasoned semi-dried oyster, and to investigate its food component characteristics. Three types of semi-dried oyster were prepared: semi-dried oyster prepared without seasoning and coating (C), seasoned semi-dried oyster without coating (S) and seasoned semi-dried oyster coated with alginate (SA). SA was high in moisture (48.6%), while low in lipid (2.8%), and crude protein (25.9%) compared to those of C and S. Hardness and sensory scores of SA were $209.8g/cm^2$ and $3.9\sim4.5$ points, respectively. Total amino acid content (24,299 mg/100 g) of SA was lower than that (27,181 mg/100 g) of C, and the major amino acids were aspartic acid, glutamic acid, leucine and Lysine. The major fatty acids of SA were 16:0 (25.5%) as saturates and EPA (23.5%) and DHA (9.3%) as polyenes. Calcium and phosphorus contents of SA were 42.6 mg/100 g and 245.5 mg/100 g, respectively.

Purification and Some Characteristics of the Proteolytic Enzyme in Fruitbody of Neungee [Sarcodon aspratus (Berk.) S. Ito] (능이 [Sarcodon aspratus (Berk.) S. Ito]중 단백질(蛋白質) 가수분해(加水分解) 효소(酵素)의 정제(精製) 및 성질(性質)에 관하여)

  • Lee, Tae-Kyoo
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.15 no.3
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    • pp.276-285
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    • 1986
  • This study was undertaken to investigate the characteristics of the proteolytic enzyme extracted from Neungee mushroom [Sarcodon aspratus (Berk.) S. Ito]. The enzyme was purified by using Tris-acryl CM-cellulose ion exchange, gel filtration on Ultrogel AcA 54, Hydroxy apatite column chromatography and preparative isoelectic focusing. The specific activity of the purified enzyme increased 8 times as compared with that of the crude enzyme. The enzyme was homogeneous on polyacrylamide gel electrophoresis (PAGE). The optimum pH was 10.1, indicating the enzyme to be alkaline protease and the optimum temperature was $57^{\circ}C$. The enzyme was stable at temperatures lower than $50^{\circ}C$and at pH values ranging from 4.0 to 10.8. However, the enzyme activity decreased by 26 and 65% at 60 and $65^{\circ}C$, respectively, when incubated for 30 minutes. The enzyme activity was activated by $Mn^{++}$ and inhibited by $Cu^{++}$ and $Hg^{++}$. The enzyme was consisted of monomer and its molecular weight estimated to be about 30,100 when determined by sodium dodecyl sulfate PAGE. Isoelectric point of the enzyme was determined to be 9.80.

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Synthesis, Characterizations, and Applications of Metal-Ions Incorporated High Quality MCM-41 Catalysts (고품질 금속 이온 첨가 MCM-41 분자체 촉매의 제법, 특성화 및 응용 반응)

  • Lim, Steven S.;Haller, Gary L.
    • Korean Chemical Engineering Research
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    • v.51 no.4
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    • pp.443-454
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    • 2013
  • Various metal ions (transition and base metals) incorporated MCM-41 catalysts can be synthesized using colloidal and soluble silica with non-sodium involved process. Transition metal ion-typically $V^{5+}$, $Co^{2+}$, and $Ni^{2+}$-incorporated MCM-41 catalysts were synthesized by isomorphous substitution of Si ions in the framework. Each incorporated metal ion created a single species in the silica framework, single-site solid catalyst, showing a substantial stability in reduction and catalytic activity. Radius of pore curvature effect was investigated with Co-MCM-41 by temperature programmed reduction (TPR). The size of metallic Co clusters, sub-nanometer, could be controlled by a proper reduction treatment of Co-MCM-41 having different pore size and the initial pH adjustment of the Co-MCM-41 synthesis solution. These small metallic clusters showed a high stability under a harsh reaction condition without serious migration, resulting from a direct anchoring of small metallic clusters to the partially or unreduced metal ions on the surface. After a complete reduction, partial occlusion of the metallic cluster surface by amorphous silica stabilized the particles against aggregations. As a probe reaction of particle size sensitivity, carbon single wall nanotubes (SWNT) were synthesized using Co-MCM-41. A metallic cluster stability test was performed by CO methanation using Co- and Ni-MCM-41. Methanol and methane partial oxidations were carried out with V-MCM-41, and the radius of pore curvature effect on the catalytic activity was investigated.

The Effect of pH on Citric Acid Leaching of Soil Contaminated with Heavy Metals (중금속(重金屬) 오염토양(汚染土壤)의 구산(枸酸) 침출(浸出)에 대한 pH의 영향(影響))

  • Jung, Kyungbae;Park, Hongki;Yoo, Kyoungkeun;Park, Jay Hyun;Choi, Ui Kyu
    • Resources Recycling
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    • v.22 no.5
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    • pp.13-19
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    • 2013
  • The effect of pH on the citrate leaching behavior of heavy metal ion was investigated to develop an eco-friendly process for removing heavy metals from soil contaminated with copper, zinc, and lead. The leaching tests were performed using citrate solution with pH adjusted by mixing citric acid and sodium citrate under the following leaching conditions: particle size, under $75{\mu}m$; temperature, $50^{\circ}C$; citrate concentration, $1kmol/m^3$; pulp density, 5%; shaking speed, 100 rpm; leaching time, 1 hour. The difference of pH before and after the leaching test was not observed, and this result indicates the direct effect of hydrogen ion concentration on the leaching of metals was insignificant. The removal ratios of copper, zinc, and lead from the contaminated soil decreased with increasing pH. The thermodynamic calculation suggests that the leaching behaviors of metal ions were determined by two reactions; one is the reaction to form complex ions between heavy metal ions and citrate ion species, and the other is the reaction to form metal hydroxide between heavy metal ions and hydroxide ion.

A Study on the Solubilization of $\alpha$-Chymotrypsin Using AOT Reverse Micelles; Effects of pH and salts (AOT 역미셀을 이용한 $\alpha$-chymotrypsin의 가용화에 대한 연구;pH와 염의 영향)

  • 노선균;강춘형
    • KSBB Journal
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    • v.15 no.6
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    • pp.664-669
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    • 2000
  • Micellar aggregates are known to be useful for the selective isolation of biologically active materials such as amino acids, proteins, and enzymes from crude mixtures sparsely dispersed in water. In this study, the effects of pH, salt type and its concentration on the solubilization of $\alpha$-chymotrypsin into the organic micellar phase, which consisted of AOT (sodium 야(2-ethylhexy)sulfosuccinate) and iso-octane, were comprehensively examined. It was found that maximum extraction efficiency was attained at a pH below the isoelectric point of $\alpha$-chymotrypsin; at pH=5.0 for NaCl and KCl, and at pH=7.0 for $CaCl_2$and $MgCl_2$. In order to avoid complications stemming from the precipitationof protein at low pH interfaces, the protein concentrations in the organic and aqueous phases were directly measured. The size of the micelle water pool was estimated by measuring the molar ratio of the surfactant to the water, W(sub)o. The resulting values of W(sub)o were nearly constant at 30 and 19 for NaCl and KCl, respectively, and were independent of pH. The addition of 1:2 salts like $MgCl_2$and $CaCl_2$ led to much lower, but a constant value of, W(sub)o than the 1:1 salts.

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