Background : Activation of the transcription factor NF-${\kappa}B$ has been shown to protect cells from tumor necrosis factor-alpha, chemotherapy, and radiation-induced apoptosis. NF-${\kappa}B$-dependent cIAP expression is a major antiapoptotic mechanism for that. NF-${\kappa}B$ activation and cIAP expression in A549 lung cancer cells which is relatively resistant to radiation-induced cell death were investigated for the mechanism of radioresistance. Materials and methods : We used A549 lung cancer cells and Clinac 1800C linear accelerator for radiation. Cell viability test was done by MTT assay. NF-${\kappa}B$ activation was tested by luciferase reporter gene assay, Western blot for $I{\kappa}B{\alpha}$ degradation, and electromobility shift assay. For blocking ${\kappa}B$, MG132 and transfection of $I{\kappa}B{\alpha}$-superrepressor plasmid construct were used. cIAP expression was analyzed by RT-PCR and cIAP2 promoter activity was performed using luciferase assay system. Results : MTT assay showed that cytotoxicity even 48 hr after radiation in A549 cells were less than 20%. Luciferas assay demonstrated weak NF-${\kappa}B$ activation of $1.6{\pm}0.2$ fold compared to PMA-induced $3.4{\pm}0.9$ fold. Radiation-induced $I{\kappa}B{\alpha}$ degradation was observed in Western blot and NF-${\kappa}B$ DNA binding was confirmed by EMSA. However, blocking NF-${\kappa}B$ using MG132 and $I{\kappa}B{\alpha}$-superrepressor transfection did not show any sensitizing effect for radiation-induced cell death. The result of RT-PCR for cIAP1 & 2 expression was negative induction while TNF-${\alpha}$ showed strong expression for cIAP1 & 2. The cIAP2 promoter activity also did not show any change compared to positive control with TNF-${\alpha}$. Conclusion : We conclude that activation of NF-${\kappa}B$ does not determine the intrinsic radiosensitivity of cancer cells, at least for the cell lines tested in this study.
The synthesis of methanol from carbon dioxide and hydrogen was studied for various compositions of Cu/ZnO catalyst system. Effect of the composition ratio of CuO and ZnO on the catalytic activity in the above reaction and the relationship between the activity and the characteristics of the catalysts were explained from the result of surface area measurements, SEM, XRD, and XPS. The major products of the reaction were methanol and carbon monoxide. The selectivity to methanol increased with increase of the copper oxide content in the catalyst up to CuO: ZnO = 30:70 weight ratio, and decreased rapidly when the content is above 70%. SEM and BET measurements, indicate that this point corresponds to the increasing point of the catalyst crystallite size and the decreasing point of the surface area. As to the Cu/Cu + Zn atomic ratio, the surface concentration of copper measured by XPS decreased remarkably when the copper oxide content in catalyst was higher than 50%. All the unreduced catalysts had almost same binding energy of Cu(2P3) level, but the binding energy for $Cu(2P^3)$ level of reduced catalysts was lowered than that of calcined catalysts. The surface copper species which was in the maximum amount when the CuO:ZnO composition in the catalyst was 30:70, existed as zero valent copper. This result agreed with the experimental result that the highest rate of methanol formation was observed when the CuO content in the catalyst was 30%. It was postulated that these reduced catalysts performed with a relatively strong basicity because the formation rate of acetone was higher than that of propylene in isopropanol decomposition as measured in a pulse type reactor.
Purpose : A new virtual simulation technique for craniospinal irradiation (CSI) that uses a CT-simulator was developed to improve the accuracy of field and shielding placement as well as patient positioning. Materials and Methods : A CT simulator (CT-SIM) and a 3-D conformal radiation treatment planning system (3D-CRT) were used to develop CSI. The head and neck were immobilized with a thermoplastic mask while the rest of the body was immobilized with a Vac-Loc. A volumetric image was then obtained with the CT simulator. In order to improve the reproducibility of the setup, datum lines and points were marked on the head and body. Virtual fluoroscopy was performed with the removal of visual obstacles, such as the treatment table or immobilization devices. After virtual simulation, the treatment isocenters of each field were marked on the body and on the immobilization devices at the conventional simulation room. Each treatment fields was confirmed by comparing the fluoroscopy images with the digitally reconstructed radiography (DRR) and digitally composited radiography (DCR) images from virtual simulation. Port verification films from the first treatment were also compared with the DRR/DCR images for geometric verification. Results : We successfully performed virtual simulations on 11 CSI patients by CT-SIM. It took less than 20 minutes to affix the immobilization devices and to obtain the volumetric images of the entire body. In the absence of the patient, virtual simulation of all fields took 20 min. The DRRs were in agreement with simulation films to within 5 mm. This not only reducee inconveniences to the patients, but also eliminated position-shift variables attendant during the long conventional simulation process. In addition, by obtaining CT volumetric image, critical organs, such as the eyes and the spinal cord, were better defined, and the accuracy of the port designs and shielding was improved. Differences between the DRRs and the portal films were less than 3 m in the vertebral contour. Conclusion : Our analysis showed that CT simulation of craniospinal fields was accurate. In addition, CT simulation reduced the duration of the patient's immobility. During the planning process. This technique can improve accuracy in field placement and shielding by using three-dimensional CT-aided localization of critical and target structures. Overall, it has improved staff efficiency and resource utilization by standard protocol for craniospinal irradiation.
Lee Min-wha;Lee Tai-hee;Ahn Bong-whan;Park Byung-ju;Yang Sung-yeul
Proceedings of the Ginseng society Conference
/
1984.09a
/
pp.83-88
/
1984
It is now well established that the rodenticide Vacor (N-3-pyridyl-mehtyl-N'-p-nitropheny-lurea) causes a hyperglycemia in human and rats. It is also reported that there are some components (DPG-3) in ginseng radix which cause hypoglycemic effect on alloxan diabetic mice. In the present study, attempts were made to demonstrate in Vacor-poisoned rats the hypo-glycemic activity of red ginseng component(RGC), which was extracted by Kimura's DPG-3 extraction procedure and found to be effective for lowering a hyperglycemia in alloxan-diabetic rats. Vacor in a dose of $LD_{50}$ (10mg/kg) produced a glucose intolerance with a paradoxical moderate increase in blood immunoreactive insulin and derangement in glucose metabolism of epididymal adipocytes in rats. Although RGC (20mg/kg, i.p.) did not exert any significant influence on a hyperglycemia induced by large lethal doses (25mg/kg) of Vacor ingestion, it improved the LDso Vacor-induced glucose intolerance and caused a further increase in blood insulin levels in Vacor-poisoned rats. The administration of RGC (20mg/kg, i.p.) normalized Vacor-induced depression of glucose metabolism and lipogenesis in the epididymal adipocytes with an improvement of reduced responses to insulin of adipocytes from Vacor-poisoned rats. These results suggest that some red ginsneng components contained in RGC fraction normalize the depressed peripheral glucose unitlization and insulin response and eventually lead to an improvement of abnormal glucose tolerance developed in rats poisoned with small doses of Vacor.
This study was performed to develop the maufacturing processes of Makgeolli using red ginseng starch (RGS). After the fermentation of RGS with koji, nuruk, and yeast, the different temperature effects on the number of the yeast cells, the content of organic acid, free sugars, and total acid, and pH were investigated. There were no changes in the composition of the yeast cell number and content of organic acid amd during 20 days at $4^{\circ}C$. The content of free sugars (sucrose, glucose and mannose) and the pH value of red ginseng Makgeolli decreased during storage at $4^{\circ}C$. This meant that the total acid content and pH value increased after organic acid was produced from fermentation. Therefore, red ginseng Makgeolli is highly acidic and sour. Since high acidity helps improve storage conditions, so this developed red ginseng Makgeolli is considered safe for consumption. Furthermore, the total content of ginsenoside was 2.47 mg/mL, which was differentiate Makgeolli using red ginseng starch, with others. Therefore, new red ginseng Makgeolli is rich in organic acid, free sugars, and ginsenoside. As a result, its storage, taste, and flavor improved.
In this study, we investigated the effect of fermentation conditions on the amylolytic and proteolytic activities of Aspergillus luchuensis strain 74-5 and Aspergillus oryzae strain 75-2, which are used in the preparation of the starter culture, for Takju (Korean traditional rice wine). The starter culture was optimized using different conditions, such as inoculum size, inoculation temperature, and incubation time. The enzyme activities under each condition were measured. In the A. luchuensis strain 74-5 starter culture, the ${\alpha}-amylase$ and glucoamylase activities increased, however the activity of acidic protease decreased as the diluent to starter culture ratio increased. In the A. oryzae 75-2 starter culture, all enzyme activities were maintained at a higher level even at 5% inoculation ratio. Higher enzyme activities were observed in the middle range of inoculation temperature (35, $40^{\circ}C$), than in the lower range (20, $30^{\circ}C$). Enzyme activity in the starter culture varied with incubation time, however it was the highest at 144 and 120 hr, respectively, for A. luchuensis strain 74-5 and A. oryzae strain 75-2. The spore count of the starter culture was approximately $2{\times}10^7$ during fermentation, out of which contamination by aerobic bacteria was about $3{\times}10^3$. The results suggested that the starter culture of each strain could be used as an inoculum for fermentation. However, we needs to conduct further research for the selection of suitable diluting agents as well as drying methods to reduce the contamination by aerobic bacteria, while retaining the enzyme activity.
Kim, Mi Jeong;Kim, Ka Hye;Kim, Moon Jeong;Kim, Jin Ik;Choi, Hye Jung;Moon, Ja Young;Joo, Woo Hong;Kim, Dong Wan
Journal of Life Science
/
v.26
no.9
/
pp.991-998
/
2016
NF-κB acts as a critical transcription factor for the survival of cells via the induction of antiapoptotic genes. Constitutive activation of NF-κB in many types of solid tumors suggests that the inhibition of NF-κB might prevent or inhibit tumorigenesis. Although a number of studies demonstrated that Hsp70 regulated NF-κB activity, the exact mechanism is not clear. This study investigated the functional relationship of Hsp70 and IKKγ in the regulation of NF-κB activation using expression plasmids of components of the IKK complex. Wild-type and deletion mutants of IKKγ were expressed together with Hsp70, and the combined regulatory effect of Hsp70 and IKKγ on NF-κB activation was assayed. Hsp70 suppressed the activation of NF-κB in a reporter plasmid assay. Hsp70 also suppressed the phosphorylation and degradation of IκBα. The suppressive effect of Hsp70 on NF-κB activation was synergistically elevated by IKKγ. The N-terminal IKKβ binding site, C-terminal leucine zipper, and zinc finger domains of IKKγ were not necessary for the suppressive effect. Furthermore, Hsp70 and IKKγ synergistically suppressed the induction of COX-2 expression by lipopolysaccharides in RAW264.7 cells. These results suggest that overexpression of Hsp70 and IKKγ may be a strategic method for inhibition of NF-κB and related diseases.
Although Euonymus alatus (EA) has been used as traditional medicine for cancer treatment, exact substances involved in curing of the disease are not yet known. Free radical scavenging and reactive oxygen species (ROS) removal activities of aqueous extract components isolated from winged stem of EA in animal cell line were investigated. Aqueous extract of EA (AEEA) was fractionated by ultrafiltration. All fractions mainly consisted of polysaccharide (44.8%), protein (2.1%), small amounts of phenol compounds and organic acids. Antioxidant activity of AEEA increased depending on concentration fractions, as determined by 1,1-diphenyl-2-picrylhydrazyl method. ROS removal activity was visualized in Chinese hamster ovary cell line using laser scanning confocal microscope, and AEEA activity increased in order of F IV>F III>F I>F II. These results suggest AETA has bioactive carbohydrates with potentials as functional foods and antioxidants.
Mutations in the cystathionine ${\beta}$-synthase (CBS) gene cause homocystinuria, the most frequent inherited disorder in sulfur metabolism. CBS is the unique enzyme using both heme and pyridoxal 5-phosphate (PLP) for activity. Among the reported 140 mutations, one of the most common disease-causing alterations in human CBS is G307S mutation. To investigate the pathogenic mechanism of G307S by spectroscopic methods, we engineered the full length and the truncated G247S mutation of yeast CBS that is corresponding mutation to human G307S. Yeast CBS does not contain heme and thus gives a merit to study the spectroscopic properties. The UV-visible spectra of the purified full length and the truncated G247S yeast CBSs showed the total absence of PLP in the protein. The absence of PLP in G247S mutation was also confirmed by the PLP-cyanide adduct formation experiment, which was conducted by the incubation of the purified enzyme with KCN. The adducts were detected using a circular dichroism (CD) and a spectrofluorimeter. Radio isotope activity assay of full length and truncated G247S proteins also gave no activity. Our yeast G247S mutation data suggested that G307S might make the distortion of the active site so that cofactor PLP and substrate can not fit inside the active site. Our yeast CBS study addressed the reason why the G307S mutation in human CBS makes the enzyme inactive that consequently leads to severe clinical phenotype.
A functional screen of 60,672 fosmid metagenomic clones amplified from marine sediment obtained from the Dokdo islets in Korea identified the gene EstES1, whose product, EstES1, displayed lipolytic properties on tributyrin-supplemented media. EstES1 is a 576 amino acid protein with a predicted molecular weight of 59.4 kDa including 37 N-terminal leader amino acids. EstES1 exhibited the highest sequence similarity (44%) to a carboxylesterase found in Haliangium ochraceum DSM14365. Phylogenetic analysis indicated that EstES1 belongs to a currently uncharacterized family of lipases. Within the conserved domain, EstES1 retains the catalytic triad that consists of the consensus penta-peptide motif, GESAG. EstES1 demonstrated a broad substrate specificity toward the long acyl group of ethyl esters (C2-C12), and its optimal activity was recorded toward p-Nitrophenyl butyrate (C4) at pH 9.0 and $40^{\circ}C$ (specific activity of 255.4 U/mg). The enzyme remained stable in the ranges of $60-65^{\circ}C$ and pH 9.0-10.5 and in the presence of methanol, ethanol, isopropanol, and dimethyl sulfoxide. Therefore, EstES1 has potential for use in industrial applications involving high temperature, organic solvents, and/or alkaline conditions.
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