Background: TRAIL (TNF-related apoptosis inducing ligand) is a newly identified member of the TNF gene family which appears to have tumor-selective cytotoxicity due to the distinct decoy receptor system. TRAIL has direct access to caspase machinery and induces apoptosis regardless of p53 phenotype. Therefore, TRAIL has a therapeutic potential in lung cancer which frequently harbors p53 mutation in more than 50% of cases. However, it was shown that TRAIL also could activates $NF-{\kappa}B$ in some cell lines which might inhibit TRAIL-induced apoptosis. This study was designed to investigate whether TRAIL can activate $NF-{\kappa}B$ in lung cancer cell lines relatively resistant to TRAIL-induced apoptosis and inhibition of $NF-{\kappa}B$ activation using proteasome inhibitor MG132 which blocks $I{\kappa}B{\alpha}$ degradation can sensitize lung cancer cells to TRAIL-induced apoptosis. Methods: A549 (wt p53) and NCI-H1299 (null p53) lung cancer cells were used and cell viability test was done by MTT assay. Apoptosis was confirmed with Annexin V assay followed by FACS analysis. To study $NF-{\kappa}B$-dependent transcriptional activation, a luciferase reporter gene assay was used after making A549 and NCI-H1299 cells stably transfected with IgG ${\kappa}-NF-{\kappa}B$ luciferase construct. To investigate DNA binding of $NF-{\kappa}B$ activated by TRAIL, electromobility shift assay was used and supershift assay was done using anti-p65 antibody. Western blot was done for the study of $I{\kappa}B{\alpha}$ degradation. Results: A549 and NCI-H1299 cells were relatively resistant to TRAIL-induced apoptosis showing only 20~30% cell death even at the concentration 100 ng/ml, but MG132 ($3{\mu}M$) pre-treatment 1 hour prior to TRAIL addition greatly increased cell death more than 80%. Luciferase assay showed TRAIL-induced $NF-{\kappa}B$ transcriptional activity in both cell lines. Electromobility shift assay demonstrated DNA binding complex of $NF-{\kappa}B$ activated by TRAIL and supershift with p65 antibody. $I{\kappa}B{\alpha}$ degradation was proven by western blot. MG132 completely blocked both TRAIL-induced $NF-{\kappa}B$ dependent luciferase activity and DNA binding of $NF-{\kappa}B$. Conclusion: This results suggest that inhibition of $NF-{\kappa}B$ can be a potentially useful strategy to enhance TRAIL-induced tumor cell killing in lung cancer.
Kim, Hyung-Jo;Jang, Ho-Sun;Lee, Soo-Min;Kim, Joo-Hyung;Shin, Jin-Ho;Kim, Heung-Tae
The Korean Journal of Pesticide Science
/
v.14
no.1
/
pp.65-71
/
2010
This study was performed to investigate the inhibiting activity of etridiazole and thiophanate-mthyl on mycelial growth of Rhizoctonia solani and Pythium ultimum and the effect of inoculum density of each pathogen on the control efficacy of the mixture of etridiazole and thiophanate-methyl in a seedling assay test. In mycelial growth inhibition test, $EC_{50}$ values of etridiazole and thiophanate-methyl against R. solani were 15.87 and 9.34 ${\mu}g\;mL^{-1}$, while those were 0.2 and more than $500\;{\mu}g\;mL^{-1}$ against P. ultimum, respectively. Controlling activity of the mixture of etridiazole and thiophanate-methyl against damping-off of pepper and cucumber, caused by R. solani and P. ultimum was tested in a greenhouse. With the inoculation of R. solani, disease incidences of seedling in pepper were 73% and 95% at 0.5% and 1.0% of inoculum concentration, while in cucumber those were 55% and 62% at 1.0% and 2.0%. When P. ultimum was inoculated into soil by 2.0% of inoculum concentration, those in pepper and cucumber were 66.7% and 96.8%, respectively. The efficacy of the mixture was somehow affected by the concentration of R. solani. While each control value of the mixture was 94.4% and 90.7% in pepper and cucumber at low inoculum concentration (0.05%), the efficacy of the fungicide decreased in pepper and cucumber by 70.7% and 72.9% at high concentration of R. solani (0.1% in pepper and 0.5% in cucumber). However, the control value of the mixture was 100% in pepper and cucumber, irrespective of the inoculum concentration of P. ultimum, however, the increase of inoculum concentration in soil did not result in the decrease of the fungicide efficacy.
Selenium is an essential micronutrient for normal body function and functions as an essential constituent of selenoproteins. This study was carried out to investigate effect of selenium on the formation of colonic aberrant crypt foci (ACF) and tumor formation in a mouse model. Five-week old ICR mice were acclimated for one week and fed different selenium diet (0.02, 0.1, and 0.5 ppm) for 12 weeks. Animals received three intraperitoneal injections of azoxymethane (10 mg/kg B.W. in saline for 3 weeks), followed by 2% dextran sodium sulfate in the drinking water for a week. There were four experimental groups, including a normal control group and three different selenium levels groups. After sacrifice, the total numbers of aberrant crypt (AC) and ACF were measured in the colonic mucosa after methylene blue staining. The number of tumors was noted for tumor incidence. Liver selenium concentration was measured using ICP-AES method. Gutathione peroxidase (GPx) activity was determined using a GPx assay kit in the liver and colon. TUNEL assay and proliferating cell nuclear antigen (PCNA) staining were performed to examine the cell apoptosis and cell proliferation, respectively. Immunohistochemistry of $\beta$-catenin was also performed on the mucous membrane tissue of colon. The activity of GPx in the liver and colon was decreased in the selenium-deficient diet group while it was increased in the selenium-overloaded diet group. Apoptotic positive cells were increased in the selenium-overloaded diet group but decreased in the selenium-deficient diet group. PCNA staining area was decreased in the selenium-overloaded diet group. In addition, the $\beta$-catenin protein level in the selenium-deficient diet group was increased but decreased in the selenium-overloaded diet group. These results indicate that dietary selenium might exert a modulating effect on colon cancer by inhibiting the development of ACF and colon tumor formation in this mouse model.
The antioxidant and anti-inflammatory effects of Corni fructus extracts (CEF, EtOAc extraction; CBF, buthanol extraction; CWF, water extraction) were investigated. The total phenolics of CEF (173.3 mg TAE/g) were significantly higher than those of CWF (26.7 mg TAE/g) and CBF (94.8 mg TAE/g). DPPH and ABTS free radical scavenging activity of CEF (DPPH: $RH_{50}$; $25.1{\mu}g/mL$, ABTS: $RC_{50}$; $36.1{\mu}g/mL$) showed even higher than that of BHA and ${\alpha}-tocopherol$ used as positive control. All three Corni fructus extracts in the concentration of $1{\sim}100{\mu}g/mL$ were effective inhibitors of NO and prostaglandin $E_2$ ($PGE_2$). NO production was inhibited 71.3~92.2% by CEF, 76.8~85.5% by CBF and 74.4~96.9% by CWF, respectively. CEF, CBF and CWF ($1{\sim}100{\mu}g/mL$) inhibited also pro-inflammatory cytokines like $TNF-{\alpha}$, $IL-1{\beta}$ and IL-6 very effectively. $TNF-{\alpha}$ was inhibited up to 51.2% by CWF and $IL-1{\beta}$ was inhibited up to 67.1% by CEF. IL-6 was best inhibited by CEF up to 58.9%. This study suggested the potential of Corni fructus for use as an excellent antioxidant substance and inflammatory inhibiting mediators. Therefore CEF, CBF and CWF Corni fructus extracts may be used for therapeutic approach to various inflammatory diseases.
The objective of this study was to investigate the antioxidative capacity of ethanol extracts from Rumex crispus L. The concentration of R. crispus L. extract at which DPPH radical scavenging activity was inhibited by 50% was 2.15 mg/mL, which was lower than that of ${\alpha}$-tocopherol (0.43 mg/mL), as compared to 100% by pyrogallol as a reference. Total antioxidant status was examined by total antioxidant capacity against ABTS radical reactions. Total antioxidant capacities of R. crispus L. extract at concentrations of 0.1 and 1 mg/mL were 0.47 and 2.33 mM Trolox equivalents, respectively, which were higher than those of ${\alpha}$-tocopherol. Superoxide scavenging activities of R. crispus L. extract at concentrations of 0.1 and 1 mg/mL were 21.5 and 78.9%, respectively, which were not significantly (p>0.05) different from those of catechin. Oxygen radical absorbance capacities of R. crispus L. extract at concentrations of 20 and 100 ${\mu}g/mL$ were 62.5 and 156.4 ${\mu}M$ Trolox equivalents, respectively, which were lower than those of ascorbic acid. Cupric reducing antioxidant capacities of R. crispus L. extract at concentrations of 0.1 and 1 mg/mL were 0.28 and 1.88 mM Trolox equivalents, which were similar or significantly (p<0.05) higher than those of ${\alpha}$-tocopherol, respectively. R. crispus L. extract prevented supercoiled DNA strand breakage induced by hydroxyl radical and peroxyl radical. Total phenolic contents of R. crispus L. extract at concentrations of 0.5 and 5 mg/mL were 0.58 and 3.85 mM gallic acid equivalents, respectively. R. crispus L. extract at concentration of 0.1 and 0.5 mg/mL inhibited 0.2 mM tert-butyl hydroperoxide-induced cytotoxicity by 38.5 and 63.5%, respectively, in HepG2 cell culture system. Thus, strong antioxidant and cytotoxicity-inhibiting effects of R. crispus L. extract seem to be due to, at least in part, the prevention from free radicals-induced oxidation as well as high levels in total phenolic contents.
Journal of the Korean Society of Food Science and Nutrition
/
v.41
no.10
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pp.1371-1377
/
2012
Biological activities of Korean Panax ginseng 55% ethanol extract (KPGE) were investigated. The measured total polyphenol content of KPGE was 357.45 mg/100 g. KPGE showed the highest ${\alpha},{\alpha}$-diphenyl-${\beta}$-picrylhydrazyl (DPPH) and 2,2'-azino-bis-3-ethylbenzo-thiazoline-6-sulfonic acid (ABTS) radical scavenging activities of 80% and 86% at 1,000 ${\mu}g/mL$, respectively. DPPH and ABTS radical scavenging activities significantly increased in a KPGE concentration-dependent manner. SOD-like activity of KPGE (1, 10, and 100 ${\mu}g/mL$) increased from 22% up to 33% at KPGE concentrations of 500 and 1,000 ${\mu}g/mL$. KPGE treatment significantly suppressed the generation of pro-inflammatory mediators, including nitric oxide (NO), prostaglandin $E_2$ ($PGE_2$), and cytokines (tumor necrosis factor-alpha: TNF-${\alpha}$, interleukin-6: IL-6, interleukin-$1{\beta}$: IL-$1{\beta}$), in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. KPGE demonstrated strong anti-inflammatory activity that reduced NO and $PGE_2$ production in LPS-stimulated RAW 264.7 cells. Even low concentrations of KPGE (1 and 10 ${\mu}g/mL$) reduced $PGE_2$ and NO production in RAW 264.7 macrophages without LPS-stimulation, respectively. At concentrations of 100, 500, and 1,000 ${\mu}g/mL$, TNF-${\alpha}$, IL-$1{\beta}$ and IL-6 production were significantly suppressed. The results of our study suggest the potential of Korean Panax ginseng as an excellent antioxidant substance for inhibiting inflammatory mediators. Therefore, Korean Panax ginseng (KPGE) may be used as a therapeutic approach to various inflammatory diseases.
Anti-diabetic effects of extracts and fractions of Sasa borealis (SB), white lotus roots (LR) and leaves (LL), and their mixture were determined in 3T3-L1 adipocytes and Min6 cells by investigating insulin-sensitizing activity and glucose-stimulated insulin secretion, respectively. SB, LR, LL, and mixture of SB, LR, and LL (3 : 2 : 3) were extracted using 70% ethanol, and m mixture extract was fractionated by XAD-4 column chromatography with serial mixture solvents of methanol and water. Fractional extractions were utilized for anti-diabetic effect assay. SB and LR extracts increased insulin-stimulated glucose uptake, but not as much as mixture of SB, LR, and LL. Significant insulin-sensitizing activities of 20 and 80% methanol fractions of SB, LR, and LL mixture extract were observed in 3T3-L1 adipocytes, giving 0.5 or $5\;{\mu}g/mL$ each fraction with 0.2 nM insulin to attain glucose uptake level similar to that attained by 10 nM insulin alone. Similar to pioglitazone, peroxisome proliferators-activated $receptor-{\gamma}\;(PPAR-{\gamma})$ agonist, 20 and 80% methanol fractions increased adipocytes by stimulating differentiation from fibroblasts and triglyceride synthesis. LL extract and 20, 60, and 80% methanol fractions of the mixture suppressed ${\alpha}-amylase$ activity, but did not modulate insulin secretion capacity of Min6 cells in both low and high glucose media. These data suggest 20 and 80% methanol tractions contain potential insulin sensitizers with functions similar to that of $PPAR-{\gamma}$ agonist. Crude extract of SB, LR, and LL mixture possibly improves glucose utilization by enhancing insulin-stimulated glucose uptake and inhibiting carbohydrate digestion without affecting insulin secretion in vivo.
We investigated to analyze total flavonoid content and fatty acid composition of Stachys sieboldii Miq root. In order to determine antioxidant and anti-inflammatory effects of fractions from S. sieboldii Miq. root, we conducted 1.1-Diphenyl-2-picryhydrazyl (DPPH) and 2,2'-Azino-bis (3-ethylbenothiazoline-6-sulfonic acid) diammonium salt radical cation (ABTS) assays for antioxidant and measured nitric oxide (NO) production induced by lipopolysaccharide (LPS) in RAW 264.7 cells. In addition, we examined an inhibitory effect of fractions from S. sieboldii Miq. root on smad signaling induced by transforming growth factor (TGF) β. Among the fractions, n-butanol (n-BuOH) fraction showed the highest flavonoid content (16.67 mg/g), followed by n-Hexane, water and 85% aqueous methanol (85% aq. MeOH) fractions. The fatty acid composition of S. sieboldii Miq. root was in the following order : n-6 fatty acids (54.3%) > n-3 fatty acids (21.2%) > saturated fatty acids (19.7%) > n-9 fatty acids (3.6%). As a result of the antioxidant efficacy, the DPPH and ABTS assays showed that n-BuOH fraction had higher scavenging activity compared to other fractions. Inhibitory effect on NO production showed that all fractions decreased LPS-induced NO production, indicating an anti-inflammatory activity of S. sieboldii Miq. root. 85% aq. MeOH and water fractions showed a higher efficacy in inhibiting transforming growth factor (TGF) β induced smad signaling. From the results, it suggests that food processing products using S. sieboldii Miq. root will be developed as a functional food for promoting health.
Won Yeoung Choi;Sung Min Park;Ra Hye Kim;Hyoung Jin Lee;Jung No Lee;Hwa Sun Ryu
Journal of the Society of Cosmetic Scientists of Korea
/
v.50
no.2
/
pp.179-192
/
2024
In this study, six types of natural products, Prunus tomentosa (P. tomentosa), Akebia quinata (A. quinata), Prunus armeniaca (P. armeniaca), Smallanthus sonchifolius (S. sonchifolius), Citrus japonica (C. japonica), and Citrus australasica (C. australasica), were used to verify the effect of improving sleep and skin barriers by stress relief. As a result of the experiment, the production of cortisol, a stress hormone, was significantly inhibited by the P. tomentosa, C. australasica, A. quinata, and C. japonica among the six natural products. In addition, the expression of GAD67, a GABA-producing enzyme involved in sleep regulation, showed a significant increase in P. tomentosa purified water extract and C. australasica 50% ethanol extract, and the extract by each P. tomentosa solvent was found to have the highest total polyphenol content. Based on the results, the P. tomentosa extract with the highest activity was finally selected, and subsequent experiments were conducted. Among each P. tomentosa solvent extract, the DPPH radical scavenging activity was the highest in the 30% ethanol extract, and purified water extract increased GABA production and skin barrier factors filaggrin and claudin-1 expression the highest. HPLC analysis confirmed quercitrin as the main component of P. tomentosa extract, and quercitrin content by extraction solvent was high in the order of 30% ethanol > purified water > 70% ethanol > 50% ethanol. Quercitrin inhibited the production of cortisol in a concentration-dependent manner, significantly increasing GAD67 expression and GABA production, which had been reduced by cortisol. From the results of this study, it has been demonstrated that P. tomentosa can be used as a cosmetic material to help improve sleep and strengthen skin barriers by relieving stress.
Kim, Young;Kim, Se Kyu;Kim, Hyung Jung;Chang, Joon;Ahn, Chul Min;Kim, Sung Kyu;Chang, Yoon Soo
Tuberculosis and Respiratory Diseases
/
v.58
no.2
/
pp.120-128
/
2005
Background : Insulin-like growth factor binding protein (IGFBP)-3 regulates non-small cell lung cancer(NSCLC) cell proliferation in vitro and in vivo by inhibiting IGF-mediated signaling pathways. To have better strategies for the treatment of lung cancer, we analyzed the combining effects of adenovirus expressing IGFBP-3 (Ad5CMV-BP3) and SCH66336, a farnesyl transferase inhibitor (FTI) designed to block Ras-mediated proliferative signaling pathways. Methods : To measure the combining effects of Ad5CMV-BP3 and SCH66336 on the proliferation of NSCLC cells, human NSCLC cell lines (H1299, H596, A549, H460, and H358), SCH66336, recombinant adenovirus expressing IGFBP-3 (Ad5CMV-BP3) and athymic nude mice were used in these experiments. Results : The combination of Ad5CMV-BP3 and SCH66336 produced a synergistic enhancement in antiproliferative effects over a range of clinically achievable concentrations in a variety of NSCLC cell lines. Furthermore, we observed a significant reduction in growth of NSCLC xenograft induced in athymic nude mice. Conclusion : In conclusion, this study demonstrated for the first time that the FTI SCH66336 synergizes with IGFBP-3 and enhances its apoptotic activity in NSCLC cells in vitro and in vivo. The combined treatment of Ad5CMV-BP3 and SCH66336 raises the possibility of using this regimen in clinic for the treatment of NSCLC.
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