• Title/Summary/Keyword: Indicator assay

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Neutrophil oxidative burst as a diagnostic indicator of IgG-mediated anaphylaxis

  • Won, Dong Il;Kim, Sujeong;Lee, Eun Hee
    • BLOOD RESEARCH
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    • v.53 no.4
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    • pp.299-306
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    • 2018
  • Background IgG-mediated anaphylaxis occurs after infusion of certain monoclonal antibody-based therapeutics. New in vitro tests are urgently needed to diagnose such reactions. We investigated whether allergens trigger neutrophil oxidative burst (OB) and if neutrophil OB occurs due to allergen-specific IgG (sIgG). Methods Neutrophil OB was measured by dihydrorhodamine 123 flow cytometry using a leukocyte suspension spiked with a very small patch of the allergen crude extract, Dermatophagoides farinae (Der f). The mean fluorescence intensity ratio of stimulated to unstimulated samples was calculated as the neutrophil oxidative index (NOI). Results The Der f-specific NOI (Der f-sNOI) showed a time-dependent increase after Der f extract addition. At 15 min activation, higher Der f-sIgG levels were associated with lower Der f-sNOI values in 31 subjects (P<0.05). This inverse relationship occurs due to the initial blocking effect of free Der f-sIgG. Additionally, neutrophil OB was nearly absent (Der f-sNOI of -1) in two cases: a subject with undetectable Der f-sIgG levels and washed leukocyte suspensions deprived of Der f-sIgG. Conclusion Allergens can trigger neutrophil OB via preexisting allergen-sIgG. Neutrophil OB can be easily measured in a leukocyte suspension spiked with the allergen. This assay can be used to diagnose IgG-mediated anaphylaxis.

Protective Effect of Paeoniae Radix Alba against UVB Photodamage ( UVB 광손상에 대한 백작약의 보호 효과)

  • Sook Jahr Park;Jong Rok Lee
    • Herbal Formula Science
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    • v.31 no.3
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    • pp.183-191
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    • 2023
  • Objective : UVB damages skin health by causing skin redness and intense inflammation, sunburn, and skin cancer. Paeoniae Radix Alba has been used to relieve gynecological symptoms, muscle spasms, and skin ailments. This study was conducted to confirm whether it has a protective effect against UVB photodamage. Methods : Ethanol extract of Paeoniae Radix Alba (PRA) was prepared by extracting 100 g Paeoniae Radix Alba in 1 L of ethanol for 48 h. Apoptosis was monitored by the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and expression levels of apoptosis indicator proteins, and tyrosinase activity was measured with a colorimetric commercial kit. Results : In human keratinocyte HaCaT cells, PRA reduced UVB-induced cell death through apoptosis by inhibiting PARP cleavage and caspase-3 and -9. UVB-induced increase in cellular reactive oxygen species (ROS) was suppressed by PRA pretreatment. PRA also showed dose-dependent ABTS and DPPH radical scavenging activities. Furthermore, the inhibitory effect of tyrosinase activity by PRA was confirmed. Conclusion : These results demonstrated the protective role of PRA in UVB photodamage of human keratinocytes, mainly due to its antioxidant and antiapoptotic properties. We also suggest that PRA can be considered as an effective natural agent to prevent skin photodamage.

Gene Expression of Exposure to Mineral Trioxide Aggregate(MTA) on Dental Pulp Cells (Mineral Trioxide Aggregate(MTA)에 의한 치수세포의 유전자 발현변화)

  • Choi, Yu-Seok;Lee, Nan-Young;Lee, Sang-Ho
    • Journal of the korean academy of Pediatric Dentistry
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    • v.35 no.1
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    • pp.30-38
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    • 2008
  • Dental pulp cells are assumed to possess the capacity to elaborate both bone and dentin matrix under the pathological conditions following tooth injury. The purpose of this study is to examine the effects of mineral trioxide aggregate (MTA) on various gene expression regarding dentinogenesis and cell viability assay in cultured primary human dental pulp cells. The author also examined the effects of this material on cellular alkaline phosphatase activity as a potential indicator of dentinogenesis. For gene expression on MTA, reverse transcriptase polymerase chain reaction was performed using primer sets of glyceraldehyde-3-phosphate dehydrogenase, type I collagen, alkaline phosphatase(ALP), osteonectin, and dentin sialoprotein after 2 and 4 days. Cell viability assay showed that the proportion of MTA-treated pulp cells which had been exposed for 5 days to MTA was higher than that of the control cells. Among the genes investigated in this study, ALP and osteonectin(SPARC) were increased in MTA treated group than in control. These findings suggest that this dental pulp culture system may be useful in the future as a model for studying the mechanisms underlying dentin regeneration after the treatment with MTA. Exposure to MTA material would not induce cytotoxic response in the dental pulp cells. In addition, MTA could influence the behavior of human pulp cells by increasing the ALP activity and SPARC synthesis.

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Effects of Antibacteria and Adhesive Inhibition of Scutellaria baicalensis Extract on Streptococcus mutans (황금(Scutellaria baicalensis) 추출물에 의한 Streptococcus mutans의 항균 및 부착억제 효과)

  • Paek, Jong-Yoon;Kim, Young-Hyun;Kwon, Hyun-Jeoung;Kim, Eun-Nim;Kim, Wan-Jong;Han, Man-Deuk
    • Journal of dental hygiene science
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    • v.8 no.4
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    • pp.367-373
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    • 2008
  • The natural products are used to be development of new antibacterial substances against human pathogenic bacteria. Adherence to the tooth surface by S. mutans is an important step in initiation of dental caries. This study was to examine antibacterial activity and anti-adhesive effect of Scutellaria baicalensis extract against S. mutans. Extracts of S. baicalensis were tested for antimicrobial activities by paper disc methods and radial diffusion assay methods, and bacterial adherence assay using 3 type of hydroxyapatite. The antibacterial level of ethyl acetate extract, IPK-3 on the growth of S. mutans was 125 mg/ml of minimum inhibitory concentration (MIC). The maximum growth of S. mutans in medium added with IPK-3 extract (50 mg/ml) was delayed to 30 hr, while the highest at 24 hr in control medium. The pH values of the control medium was 5.63 at 18 hr, but the media supplemented with IPK-3 extract was pH 6.50 at 12 hr. In adhesive inhibition assay, S. mutans was labelled with the fluorescent indicator DAPI and measured with fluorescence microscope. Adhesion of S. mutans on hydroxyapatite beads was inhibited by IPK-3 extracts. These results suggest that S. baicalensis extract can be used as an effective material for antibacterial activity and adhesive inhibition against S. mutans.

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Mouse model system based on apoptosis induction to crypt cells after exposure to ionizing radiation (방사선에 전신 조사된 마우스 음와 세포의 아포토시스 유도를 이용한 생물학적 선량 측정 모델 개발 연구)

  • Kim, Tae-Hwan
    • Korean Journal of Veterinary Research
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    • v.41 no.4
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    • pp.571-578
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    • 2001
  • To evaluate if the apoptotic fragment assay could be used to estimate the dose prediction after radiation exposure, we examined apoptotic mouse crypt cells per 1,000 cells after whole body $^{60}Co$ $\gamma$-rays and 50MeV ($p{\rightarrow}Be^+$) cyclotron fast neutron irradiation in the range of 0.25 to 1 Gy, respectively. The incidence of apoptotic cell death rose steeply at very low doses up to 1 Gy, and radiation at all doses tigger rapid changes in crypt cells in stem cell region. These data suggest that apoptosis may play an important role in homeostasis of damaged radiosensitive target organ by removing damaged cells. The curve of dose-effect relationship for the data of apoptotic fragments was obtained by the linear-quadratic model $y=0.18+(9.728{\pm}0.887)D+(-4.727{\pm}1.033)D^2$ ($r^2=0.984$) after $\gamma$-rays irradiation, while $y=0.18+(5.125{\pm}0.601)D+(-2.652{\pm}0.7000)D^2$ ($r^2=0.970$) after neutrons in mice. The dose-response curves were linear-quadratic, and a significant dose-response relationship was found between the frequency of apoptotic cell and dose. These data show a trend towards increase of the numbers of apoptotic crypt cells with increasing dose. Both the time course and the radiation dose-response curve for high and low linear energy transfer (LET) radiation modalities were similar. The relative biological effectiveness (RBE) value for crypt cells was 2.072. In addition, there were significant peaks on apoptosis induction at 4 and 6h after irradiation, and the morpholoigcal findings of the irradiated groups were typical apoptotic fragments in crypt cells that were hardly observed in the control group. Thus, apoptosis in crypt cells could be a useful in vivo model for studying radio-protective drug sensitivity or screening test, microdosimetric indicator and radiation-induced target organ injury. Since the apoptotic fragment assay is simple, rapid and reproducible in the range of 0.25 to 1 Gy, it will also be a good tool for evaluating the dose response of radiation-induced organ damage in vivo and provide a potentially valuable biodosimetry for the early dose prediction after accidental exposure.

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The Effect of Red Cabbage (Brassica oleracea L. var. capitata f. rubra) Extract on the Apoptosis in Human Breast Cancer MDA-MB-231 Cells (적양배추 추출물이 인체 유방암 세포 MDA-MB-231의 세포사멸에 미치는 영향)

  • Nam, Mi Kyung;Kang, Keum Jee
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.42 no.1
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    • pp.8-16
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    • 2013
  • We investigated the effect of red cabbage extract (RCE) on cell death in MDA-MB-231 human breast cancer cells. Cells were cultured in the presence 1.0, 1.5, and 2.0 mg/mL concentrations of RCE for 24 hours. MTT assays demonstrated that mitochondrial dehydrogenase activities decreased in a dose-dependent manner in cells (p<0.05). In contrast, the proportion of dual staining with Hoechst 33342/ethidium bromide (EtBr) for cell death increased in a dose-dependent manner in cells (p<0.05). Flow cytometry assays revealed that cell death caused by an apoptotic program increased in a dose-dependent (p<0.05). Also, increased ROS accumulation in cells, as revealed by DCF-DA staining, was observed in a dose-dependent fashion (p<0.05). The apoptosis suppressor gene Bcl-2 decreased significantly at the mRNA level. Pro-apoptotic genes Bax and caspase-3, genes that are related to the last stage of apoptosis significantly increased. The Bcl-2/Bax ratio which is an important indicator of apoptosis, was found to have significantly decreased dose dependence. These results taken together indicate that the effect of red cabbage extract induces cell death in MDA-MB-231 human breast cancer cells.

Matrix metalloproteinase-8 and substance P levels in root canal exudates of nonvital teeth (근관 삼출액에서의 기질금속단백분해효소-8과 substance P의 수준에 관한 연구)

  • Shin, Su-Jung;Lee, Woo-Cheol;Lee, Jae-Il;Baek, Seung-Ho;Kum, Kee-Yeon;Shon, Won-Jun;Bae, Kwang-Shik
    • Restorative Dentistry and Endodontics
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    • v.36 no.3
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    • pp.196-202
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    • 2011
  • Objectives: The aim of this study was to investigate levels of matrix metalloproteinase-8 (MMP-8) and substance P (SP) in root canal exudates during root canal treatment (RCT) of nonvital, painful teeth. Materials and Methods: Patients scheduled for nonsurgical RCT were prospectively selected; the study was performed after obtaining informed consent from the patients and was approved by the Institutional Review Board for Clinical Research of Gangnam Severance Hospital, Yonsei University (3-2008-0118). Canal exudates samples were collected using sterilized paper points from teeth scheduled for RCT across three different time periods. MMP-8 and SP levels were measured using enzyme-linked immunosorbent assay (ELISA). Data were analyzed using a mixed model analysis and the Pearson correlation analysis (p < 0.05). Results: MMP-8 and SP levels in GCF were decreased during RCT (p < 0.0001), and they showed a weak positive correlation to each other (p < 0.05). Patients' subjective pain levels and the response from percussion test were significantly related to SP level. Conclusions: This study demonstrated that periradicular inflammation endodontic origin can elevate SP and MMP-8 levels in root canal exudates. Interestingly, SP level of canal exudates showed a possibility of being used as an indicator of pain due to periapical pathosis.

Multiple-biometric Attributes of Biomarkers and Bioindicators for Evaluations of Aquatic Environment in an Urban Stream Ecosystem and the Multimetric Eco-Model (도심하천 생태계의 수환경 평가를 위한 생지표 바이오마커 및 바이오인디케이터 메트릭 속성 및 다변수 생태 모형)

  • Kang, Han-Il;Kang, Nami;An, Kwang-Guk
    • Journal of Environmental Impact Assessment
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    • v.22 no.6
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    • pp.591-607
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    • 2013
  • The objectives of the study were to evaluate the aquatic environment of an urban stream using various ecological parameters of biological biomarkers, physical habitat quality and chemical water quality and to develop a "Multimetric Eco-Model" ($M_m$-E Model) for the ecosystem evaluations. For the applications of the $M_m$-E model, three zones including the control zone ($C_Z$) of headwaters, transition zone ($T_Z$) of mid-stream and the impacted zone ($I_Z$) of downstream were designated and analyzed the seasonal variations of the model values. The biomarkers of DNA, based on the comet assay approach of single-cell gel electrophoresis (SCGE), were analyzed using the blood samples of Zacco platypus as a target species, and the parameters were used tail moment, tail DNA(%) and tail length (${\mu}m$) in the bioassay. The damages of DNA were evident in the impacted zone, but not in the control zone. The condition factor ($C_F$) as key indicators of the population evaluation indicator was analyzed along with the weight-length relation and individual abnormality. The four metrics of Qualitative Habitat Evaluation Index (QHEI) were added for the evaluations of physical habitat. In addition, the parameters of chemical water quality were used as eutrophic indicators of nitrogen (N) and phosphorus (P), chemical oxygen demand (COD) and conductivity. Overall, our results suggested that attributes of biomarkers and bioindicators in the impacted zone ($I_Z$) had sensitive response largely to the chemical stress (eutrophic indicators) and also partially to physical habitat quality, compared to the those in the control zone.

Selection of Lactic Acid Bacteria with Antibacterial Activity for Extension of Kimchi Shelf-life (김치의 저장성 향상을 위한 항균활성 우수 유산균 선발)

  • Choi, Hak-Jong;Kim, Yu Jin;Lee, Na Ra;Park, Hae Woong;Jang, Ja Young;Park, Sung-Hee;Kang, Miran;Kim, Hyun Ju;Lee, Jong-Hee;Lee, Jong-Hoon;Pyun, Yu-Ryang;Kim, Tae-Woon
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.43 no.2
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    • pp.328-332
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    • 2014
  • A survey was conducted on the isolation of lactic acid bacteria with antibacterial activity to extend kimchi shelf-life. Antibacterial activity was tested against bacteria associated with acidification of kimchi, including Lactobacillus plantarum, Pediococcus pentosaceus, and Lactobacillus sakei, using agar-well diffusion assay. Two isolates from kimchi were identified as Lactococcus lactis subsp. lactis and Lactobacillus brevis by 16S rRNA gene sequence analysis and API 50 CHL assay, and they showed antibacterial effects against indicator strains. The isolates displayed acid tolerance at pH 3.5, salt tolerance in 5% NaCl, and growth at $4^{\circ}C$. These result imply that the selected strains might be used to extend kimchi shelf-life as a potential starter.

Verification of Cardiac Electrophysiological Features as a Predictive Indicator of Drug-Induced Torsades de pointes (약물의 염전성 부정맥 유발 예측 지표로서 심장의 전기생리학적 특징 값들의 검증)

  • Yoo, Yedam;Jeong, Da Un;Marcellinus, Aroli;Lim, Ki Moo
    • Journal of Biomedical Engineering Research
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    • v.43 no.1
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    • pp.19-26
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    • 2022
  • The Comprehensive in vitro Proarrhythmic Assay(CiPA) project was launched for solving the hERG assay problem of being classified as high-risk groups even though they are low-risk drugs due to their high sensitivity. CiPA presented a protocol to predict drug toxicity using physiological data calculated based on the in-silico model. in this study, features calculated through the in-silico model are analyzed for correlation of changing action potential in the near future, and features are verified through predictive performance according to drug datasets. Using the O'Hara Rudy model modified by Dutta et al., Pearson correlation analysis was performed between 13 features(dVm/dtmax, APpeak, APresting, APD90, APD50, APDtri, Capeak, Caresting, CaD90, CaD50, CaDtri, qNet, qInward) calculated at 100 pacing, and between dVm/dtmax_repol calculated at 1,000 pacing, and linear regression analysis was performed on each of the 12 training drugs, 16 verification drugs, and 28 drugs. Indicators showing high coefficient of determination(R2) in the training drug dataset were qNet 0.93, AP resting 0.83, APDtri 0.78, Ca resting 0.76, dVm/dtmax 0.63, and APD90 0.61. The indicators showing high determinants in the validated drug dataset were APDtri 0.94, APD90 0.92, APD50 0.85, CaD50 0.84, qNet 0.76, and CaD90 0.64. Indicators with high coefficients of determination for all 28 drugs are qNet 0.78, APD90 0.74, and qInward 0.59. The indicators vary in predictive performance depending on the drug dataset, and qNet showed the same high performance of 0.7 or more on the training drug dataset, the verified drug dataset, and the entire drug dataset.