• Animal cells and systems
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• 제9권2호
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• pp.79-85
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• 2005

Oxidized abasic residues arise as a major class of DNA damage by a variety of agents involving free radical attack and oxidation of deoxyribose sugar components. 2-deoxyribonolactone (dL) is a C1'-oxidized abasic lesion implicated in DNA strand scission, mutagenesis, and covalent DNA-protein cross-link (DPC). We show here that mammalian cell-free extract give rise to stable DPC formation that is specifically mediated by dL residue. When a duplex DNA containing dL at the site-specific position was incubated with cell-free extracts of Po ${\beta}-proficient$ and -deficient mouse embryonic fibroblast cells, the formation of major dL-mediated DPC was dependent on the presence of DNA polymerase (Pol) ${\beta}$. Formation of dL-specific DPC was also observed with histones and FEN1 nuclease, although the reactivity in forming dL-mediated DPC was significantly higher with Pol ${\beta}$ than with histones or FEN1. DNA repair assay with a defined DPC revealed that the dL lesion once cross-linked with Pol ${\beta}$ was resistant to nucleotide excision repair activity of cell-free extract. Analysis of nucleotide excision repair utilizing a model DNA substrate containing a (6-4) photoproduct suggested that excision process for DPC was inhibited because of DNA single-strand incision at 5' of the lesion. Consequently DPC mediated by dL lesion may not be readily repaired by DNA excision repair pathway but instead function as unusual DNA damage causing a prolonged DNA strand break and trapping of the major base excision repair enzyme.

• Archives of Pharmacal Research
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• 제24권5호
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• pp.455-465
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• 2001

The highly potent cytotoxic DNA-DNA cross-linker consists of two cyclopropa[c]pyrrolo[3,4-3]indol-4(5H)-ones insoles [(+)-CPI-I] joined by a bisamido pyrrole (abbreviated to "Pyrrole"). The Pyrrole is a synthetic analog of Bizelesin, which is currently in phase II clinical trials due to its excellent in vivo antitumor activity. The Pyrrole has 10 times more potent cytotoxicity than Bizelesin and mostly form DNA-DNA interstrand cross-links through the N3 of adenines spaced 7 bp apart. The Pyrrole requires a centrally positioned GC base pair for high cross-linking reactivity (i.e., $5^1$-T$AT_2$A*-$3^1$), while Bizelesin prefers purely AT-rich sequences (i.e., $5^1$-T$AT_4$A*-$3^1$, where /(equation omitted) represents the cross-strand adenine alkylation and A* represents an adenine alkylation) (Park et al., 1996). In this study, the high-field $^1$H-NMR and rMD studies are conducted on the 1 1-mer DNA duplex adduct of the Pyrrole where the 5′(equation omitted)TAGTTA*-3′sequence is cross-linked by the drug. A severe structural perturbation is observed in the intervening sequences of cross-linking site, while a normal B-DNA structure is maintained in the region next to the drug-modified adenines. Based upon these observations, we propose that the interplay between the bisamido pyrrole unit of the drug and central C/C base pair (hydrogen-bonding interactions) is involved in the process of cross-linking reaction, and sequence specificity is the outcome of those interactions. This study suggests a mechanism for the sequence specific cross-linking reaction of the Pyrrole, and provides a further insight to develop new DNA sequence selective and distortive cross-linking agents.

• Asian Pacific Journal of Cancer Prevention
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• 제14권10호
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• pp.5741-5745
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• 2013

Background: Heat-shock protein70 (HSP70) are intracellular protein chaperones, with emerging evidence of their association with various diseases. We have previously reported significantly elevated plasma-HSP70 (pHSP70) in pancreatic cancer. Current methods of pHSP70 isolation are ELISA-based which lack specificity due to cross-reactivity by similarities in the amino-acid sequence in regions of the protein backbone resulting in overestimated HSP70 value. Materials and Methods: This study was undertaken to develop a methodology to capture all isoforms of pHSP70, while further defining their tyrosine and serine phosphorylation status. Results: The methodology included gel electrophoresis on centrifuged supernatant obtained from plasma incubated with HSP70 antibody-coupled beads. After blocking non-specific binding sites, blots were immunostained with monoclonal-antibody specific for human-HSP70, phosphoserine and phosphotyrosine. Conclusions: Our novel immunocapture approach has distinct advantages over the commercially available methods of pHSP70 quantification by allowing isolation of molecular aggregates of HSP70 with additional ability to precisely distinguish phosphorylation state of HSP70 molecules at serine and tyrosine residues.

• 한국환경독성학회:학술대회논문집
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• 한국환경독성학회 2003년도 추계국제학술대회
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• pp.13-13
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• 2003

Methylmercury (MeHg) is a ubiquitous environmental toxicant that can be exposed to humans by ingestion of contaminated food including fish and bread. MeHg has been suggested to exert its toxicity through its high reactivity to thiols, generation of arachidonic acid and reactive oxygen species (ROS), and elevation of intracellular $Ca^{2+}$ levels (［$Ca^{2+}$］$_{i}$). However, the precise mechanism has not been fully defined. Here we show that phosphatidylcholine-specific phospholipase C (PC-PLC) is a critical pathway for MeHg-induced toxicity. MeHg activated the acidic form of sphingomyelinase (A-SMase) and group IV cytosolic phospholipase $A_2$ ($cPLA_2$) downstream of PC-PLC, but these enzymes as well as protein kinase C were not linked to MeHg's toxicity. Furthermore, MeHg produced ROS, which did not cause the toxicity. However, D6O9, an inhibitor of PC-PLC, significantly reversed the toxicity in a time- and dose-dependent manner in MDCK and SH-5YSY cells. Addition of EGTA to culture media resulted in partial decrease of [$Ca^{2+}$］$_{i}$ and partially blocked cell death. In contrast, D609 completely prevented cell death with parallel decreases in diacylglycerol and ［$Ca^{2+}$］$_{i}$. Together, our findings indicated that MeHg-induced toxicity was caused by elevation of ［$Ca^{2+}$]$_{i}$ through activation of PC-PLC. The toxicity was not attributable to the signaling pathways such as $cPLA_2$, A-SMase, and PKC, or to the generation of ROS.

• 농약과학회지
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• 제8권4호
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• pp.265-271
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• 2004

제초제 flumioxazine의 가수분해 반응성을 분자 궤도(MO)론적으로 검토한 결과, pH 5.0 이하의 산성에서는 $A_{AC}1$형의 반응 메커니즘으로 1,2-dicarboximino group의 carbonyl oxygene 원자$(O_{21})$에 대하여 hydronium ion $(H_3O^+)$에 의한 양성자화$(SH^+)$가 일반 산-촉매반응(general acid catalysis)에 따른 전하조절(charge-control) 반응이 일어난다. pH 8.0이상의 염기성에서는 $B_{AC}2$형의 반응 메커니즘으로 hydroxide anion $(OH^-)$에 의한 특정 염기-촉매반응(specific base catalysis)에 따른 궤도조절(orbital-control) 반응이 일어난다. 그리고 pH $5.0\sim8.0$ 사이에서 두 반응은 경쟁적으로 일어나 친핵성 첨가-제거반응$(Ad_{N-E})$으로 진행된다.

• 한국감성과학회:학술대회논문집
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• 한국감성과학회 2000년도 추계학술대회 논문집
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• pp.73-80
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• 2000

The paper is addressed to the topic of physiological response-specificity in disgust induced by visual stimulation. The purpose of this study was to evaluate reproducibility of physiological reactivity pattern during disgust elicited by the International Affective Pictures System (IAPS) in 2 experiments. Twenty-nine subjects participated in the first experiment with 3 visual stimulation sessions with disgust-eliciting slides (3 slides in each 1 min long session). In the second experiment disgust-eliciting slides from the IAPS were presented to 42 subjects in 2 sessions (one slide for 1 min). Spectral power of frontal EEG, skin conductance (SCL, SCR and NS.SCR), heart rate(HR), heart period variability(HPV) and respiration rate were recorded. Visual stimulation evoked 1:.n deceleration, higher power of high frequency component of HPV, increased SCL and NS.SCR frequency, frontal slow alpha blocking and moderate increase in fast beta power in most of the sessions in both experiments. However in the second experiment the EEG pattern associated with disgust showed inconsistent shifts in fast alpha and slow beta bands, but was marked by higher power of theta activity. Our data in both experiments emphasizes presence of disgust-specific profiles of autonomic and at the less extent EEG responses in visual stimulation context. Discussed are potential behavioral mechanisms leading to observed physiological manifestations in disgust elicited by visual stimulation. The results support the consideration that disgust is an withdrawal type negative valence emotion associated with relatively low autonomic arousal (low HR, low amplitude SCRs with relatively high NS.SCR frequency) and moderate EEG activation signs. Obtained data showed more consistent reproducibility of disgust-specific autonomic rather than EEG response patterns during visual stimulation design.

• 한국산업보건학회지
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• 제22권4호
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• pp.309-315
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• 2012

Objectives: Animal husbandry workers could be exposed to various work hazards including toxic gases, chemicals such as pesticides or organic dust. Immunological evaluation focusing on respiratory allergic hypersensitivity occurrence was under-taken for swine farm workers as a part of the study on immunologic status of dairy barn, swine confinement, and poultry farm workers. Materials and Methods: Peripheral bloods were collected from 25 workers at the year of 2001 and 12 workers at the year of 2012 from swine farms located at Gyeonggi province, Korea. Seven adults not involved with animal husbandry were recruited at the year of 2001 from the same residential area as the swine farm workers'. Level of plasma IgE and 20 respiratory allergen-specific IgE were evaluated using commercially available ELISA kit. Results: Plasma IgE level was approximately five-fold higher in the swine farm workers regardless of the sampling year than the control subjects. Plant allergens from outdoor environments such as golden rod, pigweed, Russian thistle, or ragweed were the major allergens with positive reaction(allergen specific IgE${\geq}$0.7 IU/mL) for the swine farm workers at 2001 year. Meanwhile, house dust mite(Dermatophagoides farinae, D. pteronyssinus) and cockroach, typical indoor allergens in Korea, were the major respiratory allergens for the swine farm workers at 2012 year. Conclusions: Overall, even though our results are primitive, the results suggest that immunological function of swine farm workers could be modulated toward type-2 reactivity.

• Food Science and Biotechnology
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• 제18권5호
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• pp.1273-1278
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• 2009

This study aimed to evaluate the potential allergenicity of Cry proteins in insect-resistant genetically modified (GM) maizes (Bt11, MON810, and MON863) using serum screening tests. Serum samples were obtained from Korean children (0-15 years old) with allergic symptoms who had positive maize-specific IgE. The levels of serum specific IgE was measured by the Phadia ImmunoCAP system and considered as positive when they are 0.35 kU/L or higher. Cry proteins (Cry1Ab in Bt11, mCry1Ab in MON810, and Cry3Bb1 in MON863) were expressed in Escherichia coli and purified for serum screening. The reactivity of purified Cry proteins was confirmed by IgE immunoblots in 50 patients (maize-sensitized patients). There was no reaction between Cry proteins and sera from maize-sensitized patients. Our results suggest that these Cry proteins are not likely to cause allergic reactions. Further studies using more sera from patients with true clinical allergies are needed to evaluate the potential allergenicity of novel proteins in GM maize.

• 농촌의학ㆍ지역보건
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• 제17권1호
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• pp.46-55
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• 1992

A systematic study was conducted to identify and isolate a serologically pertinent antigen with high specific activity and low cross reactivity from P.westermani antigen. Differential centrifugation of the homogenate yield three particulate and one soluble fractions ; the $480{\times}G$ pellet(Pw1), the $7650{\times}G$ pellet(Pw2) treated with n-butanol(Pw3), and $100000{\times}G$ supernatnat(Pw4). Comparison of antigenicity of these antigens, based upon differential centrifugation, to that of saline extract of P. westermani worm(SEP) was performed by SDS-PAGE and immunoblot techniques. The results obtained were as follows : 1) The ratio of absorbance value of ELISA against paragonimiasis positive pool sera to that of negative sera was highest when using Pw3 as antigen and that was lowest using Pwl. 2) Silver stained and SDS-PAGE of each antigen showed 34 and 13Kd band as common antigen band, but Pw2 didn't show clear band. 3) By immunoblot 55 and 34Kd bands using SEP and Pw4 showed strong positive reaction without cross reaction with sera from other helmenthic infections. Using Pw3, 10Kd band was observed as specific band. In conclusion, Pw3($100000{\times}G$ pellet urea soluble, treated with n-butanol) and Pw4($100000{\times}G$ supernatant) were usable for ELISA and immunoblot technique.

• 한국식품과학회지
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• 제30권6호
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• pp.1409-1414
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• 1998

Zearalenone에 특이한 단크론성 항체를 개발하기위하여 형질세포종세포인 myeloma cell $(P3{\times}63Ag.V653)$과 zearalenone-oxime-bovine serum albumin (BSA) 결합체를 항원으로 면역시킨 BALB/c female mice의 비장세포를 융합시켜 hybridoma cell을 생산하였다. 그들의 항체가를 indirect competitive ELISA법으로 측정한 결과 zearalenone에 대하여 높은 친화력을 갖는 5세포주를 얻었다. 그중 Z-2-M26 hybridoma cell line이 생산하는 단크론성항체는 특히 zearalenone과 민감하게 반응하였고, ${\alpha}-zearalenol$과도 약간의 교차반응을 보였으나 ${\beta}-zearalenol,\;{\alpha}-zearalenol,\;{\beta}-zearalenol$ 및 DON과는 거의 반응하지 않았다. 따라서 본 실험에서 개발된 항체는 앞으로 zearalenone에 대한 민감하고 정량적인 효소면역측정법의 개발을 위한 중요한 면역시약으로 사용될 것으로 생각한다.