Current protocols in toxicology最新文献_第10页

Evaluation of Toxicity in Mouse Bone Marrow Progenitor Cells 小鼠骨髓祖细胞毒性评价

Current protocols in toxicology Pub Date : 2016-02-01 DOI: 10.1002/0471140856.tx1809s67

Peace C. Ezeh, Huan Xu, Shu Chun Wang, Sebastian Medina, Scott W. Burchiel

引用次数: 7

Associating Changes in the Immune System with Clinical Diseases for Interpretation in Risk Assessment 免疫系统变化与临床疾病在风险评估中的关联解释

Current protocols in toxicology Pub Date : 2016-02-01 DOI: 10.1002/0471140856.tx1801s67

Jamie C. DeWitt, Dori R. Germolec, Robert W. Luebke, Victor J. Johnson

引用次数: 14

PCR-Based Analysis of Mitochondrial DNA Copy Number, Mitochondrial DNA Damage, and Nuclear DNA Damage 基于pcr的线粒体DNA拷贝数、线粒体DNA损伤和核DNA损伤分析

Current protocols in toxicology Pub Date : 2016-02-01 DOI: 10.1002/0471140856.tx2011s67

Claudia P. Gonzalez-Hunt, John P. Rooney, Ian T. Ryde, Charumathi Anbalagan, Rashmi Joglekar, Joel N. Meyer

引用次数: 72

Measuring p66Shc Signaling Pathway Activation and Mitochondrial Translocation in Cultured Cells 培养细胞p66Shc信号通路激活和线粒体易位的测定

Current protocols in toxicology Pub Date : 2015-11-04 DOI: 10.1002/0471140856.tx2506s66

Mariusz R. Wieckowski, Cláudia M. Deus, Renata Couto, Monika Oparka, Magdalena Lebiedzińska-Arciszewska, Jerzy Duszyński, Paulo J. Oliveira

{"title":"Measuring p66Shc Signaling Pathway Activation and Mitochondrial Translocation in Cultured Cells","authors":"Mariusz R. Wieckowski, Cláudia M. Deus, Renata Couto, Monika Oparka, Magdalena Lebiedzińska-Arciszewska, Jerzy Duszyński, Paulo J. Oliveira","doi":"10.1002/0471140856.tx2506s66","DOIUrl":"10.1002/0471140856.tx2506s66","url":null,"abstract":"The adaptor protein p66Shc links membrane receptors to intracellular signaling pathways, with downstream consequences on mitochondrial metabolism and reactive oxygen species production. Moreover, p66Shc has also been implicated in cancer development, progression, and metastasis. Increased phosphorylation of serine 36 residue of p66Shc very often correlates with oxidative stress–associated pathologies. The pro-oxidative role of p66Shc also appears to be involved in chemical toxicity, being an important component of stress responses triggered by xenobiotics. Here, we present a protocol that can be used: (a) for isolation of mitochondrial, cytosolic, and mitochondrial-associated membrane fractions from adherent cells lines; (b) to perform p66Shc detection with specific antibodies in order to monitor its translocation between different cellular compartments in response to the oxidative stress; and (c) to modulate the p66Shc pathway with the use of pharmacological approaches or gene-silencing methods. © 2015 by John Wiley & Sons, Inc.","PeriodicalId":72743,"journal":{"name":"Current protocols in toxicology","volume":"66 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2015-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/0471140856.tx2506s66","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"50794179","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Caenorhabditis elegans as a Model for Toxic Effects of Nanoparticles: Lethality, Growth, and Reproduction 秀丽隐杆线虫作为纳米颗粒毒性效应的模型:致死性、生长和繁殖

Current protocols in toxicology Pub Date : 2015-11-04 DOI: 10.1002/0471140856.tx2010s66

Laura L. Maurer, Ian T. Ryde, Xinyu Yang, Joel N. Meyer

{"title":"Caenorhabditis elegans as a Model for Toxic Effects of Nanoparticles: Lethality, Growth, and Reproduction","authors":"Laura L. Maurer, Ian T. Ryde, Xinyu Yang, Joel N. Meyer","doi":"10.1002/0471140856.tx2010s66","DOIUrl":"10.1002/0471140856.tx2010s66","url":null,"abstract":"The nematode Caenorhabditis elegans is extensively utilized in toxicity studies. C. elegans offers a high degree of homology with higher organisms, and its ease of use and relatively inexpensive maintenance have made it an attractive complement to mammalian and ecotoxicological models. C. elegans provides multiple benefits, including the opportunity to perform relatively high-throughput assays on whole organisms, a wide range of genetic tools permitting investigation of mechanisms and genetic sensitivity, and transparent bodies that facilitate toxicokinetic studies. This unit describes protocols for three nanotoxicity assays in C. elegans: lethality, growth, and reproduction. This unit focuses on how to use these well-established assays with nanoparticles, which are being produced in ever-increasing volume and exhibit physicochemical properties that require alteration of standard toxicity assays. These assays permit a broad phenotypic assessment of nanotoxicity in C. elegans, and, when used in combination with genetic tools and other assays, also permit mechanistic insight. © 2015 by John Wiley & Sons, Inc.","PeriodicalId":72743,"journal":{"name":"Current protocols in toxicology","volume":"66 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2015-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/0471140856.tx2010s66","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"50793658","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 19

Use of Ciliogenesis to Detect Aneugens: The Role of Primary Cilia 用纤毛发生检测纤毛:初级纤毛的作用

Current protocols in toxicology Pub Date : 2015-11-02 DOI: 10.1002/0471140856.tx0313s66

Kathyayini V. Divi, Yvona Ward, Miriam C. Poirier, Ofelia A. Olivero

{"title":"Use of Ciliogenesis to Detect Aneugens: The Role of Primary Cilia","authors":"Kathyayini V. Divi, Yvona Ward, Miriam C. Poirier, Ofelia A. Olivero","doi":"10.1002/0471140856.tx0313s66","DOIUrl":"10.1002/0471140856.tx0313s66","url":null,"abstract":"Primary cilia arise from the centrosomes of quiescent or post-mitotic cells, and serve as sensory organelles that communicate mechanical and chemical stimuli from the environment to the interior of the cell. Cilium formation may, therefore, become a useful end point signaling exposure to genotoxins or aneugens. Here we have used the aneugen, zidovudine (AZT), an antiretroviral drug that induces DNA replication arrest and centrosomal amplification (>2 centrosomes per quiescent cell), to evaluate cilia formation in retinal epithelial (pigmented) cells. Since cilia are derived from centrosomes, and aneugens can induce centrosomal amplification, the production of multiple cilia arising from multiple centrosomes may reveal the aneugenic nature of the agents. Cells were exposed to AZT to induce centrosomal amplification, cultured without serum to allow the centrioles to develop cilia, and immunostained to visualize cilia and centrosomes. Nuclear DNA was stained with DAPI. Preliminary observations suggest that cells with multiple centrosomes are able to generate extra cilia. © 2015 by John Wiley & Sons, Inc.","PeriodicalId":72743,"journal":{"name":"Current protocols in toxicology","volume":"66 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2015-11-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/0471140856.tx0313s66","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"50789230","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Immune Cell Phenotyping Using Flow Cytometry 流式细胞术免疫细胞表型分析

Current protocols in toxicology Pub Date : 2015-11-02 DOI: 10.1002/0471140856.tx1808s66

A. Graham Pockley, Gemma A. Foulds, Julie A. Oughton, Nancy I. Kerkvliet, Gabriele Multhoff

{"title":"Immune Cell Phenotyping Using Flow Cytometry","authors":"A. Graham Pockley, Gemma A. Foulds, Julie A. Oughton, Nancy I. Kerkvliet, Gabriele Multhoff","doi":"10.1002/0471140856.tx1808s66","DOIUrl":"10.1002/0471140856.tx1808s66","url":null,"abstract":"Fluorescent immunophenotyping uses fluorescently-conjugated antibodies to identify, characterize and quantify distinct subpopulations of cells within heterogeneous single-cell populations, either in the context of tissue (using fluorescence and imaging microscopy) or in a single-cell suspension (using multiparameter imaging microscopy, imaging cytometry, and/or flow cytometry). Flow cytometry is an optical, laser-based technology which analyzes the physical and fluorescent properties of cells in suspension in real-time as they flow through the instrument. This approach has a number of advantages over other techniques that can be used for characterizing cell populations in single-cell suspensions, in that it can nonsubjectively interrogate up to millions of cells and acquire data on the presence of different cell subpopulations and phenotypical changes within these populations in seconds. This unit describes basic procedures for the direct and indirect immunofluorescent staining of surface and intracellular proteins that are expressed by lymphoid cells which have been isolated from tissues or blood. Protocols for the resolution of dead cells and for the fixation of cells are also included. This unit also provides essential information relating to the selection and titration of antibodies, fluorochrome choice, spectral overlap and compensation, the use of controls, and the standardization of data acquisition and analysis. It also highlights new technologies and platforms that can be used to interrogate the presence of cell subpopulations and their phenotype to an even greater depth. © 2015 by John Wiley & Sons, Inc.","PeriodicalId":72743,"journal":{"name":"Current protocols in toxicology","volume":"66 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2015-11-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/0471140856.tx1808s66","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"50792988","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 28

Seahorse Xfe24 Extracellular Flux Analyzer-Based Analysis of Cellular Respiration in Caenorhabditis elegans 基于海马Xfe24细胞外通量分析仪的秀丽隐杆线虫细胞呼吸分析

Current protocols in toxicology Pub Date : 2015-11-02 DOI: 10.1002/0471140856.tx2507s66

Anthony L. Luz, Latasha L. Smith, John P. Rooney, Joel N. Meyer

{"title":"Seahorse Xfe24 Extracellular Flux Analyzer-Based Analysis of Cellular Respiration in Caenorhabditis elegans","authors":"Anthony L. Luz, Latasha L. Smith, John P. Rooney, Joel N. Meyer","doi":"10.1002/0471140856.tx2507s66","DOIUrl":"10.1002/0471140856.tx2507s66","url":null,"abstract":"Mitochondria are critical for their role in ATP production as well as multiple nonenergetic functions, and mitochondrial dysfunction is causal in myriad human diseases. Less well appreciated is the fact that mitochondria integrate environmental and intercellular as well as intracellular signals to modulate function. Because mitochondria function in an organismal milieu, there is need for assays capable of rapidly assessing mitochondrial health in vivo. Here, using the Seahorse XFe24 Extracellular Flux Analyzer and the pharmacological inhibitors dicyclohexylcarbodiimide (DCCD, ATP synthase inhibitor), carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP, mitochondrial uncoupler), and sodium azide (cytochrome c oxidase inhibitor), we describe how to obtain in vivo measurements of the fundamental parameters [basal oxygen consumption rate (OCR), ATP-linked respiration, maximal OCR, spare respiratory capacity, and proton leak] of the mitochondrial respiratory chain in the model organism Caenorhabditis elegans. © 2015 by John Wiley & Sons, Inc.","PeriodicalId":72743,"journal":{"name":"Current protocols in toxicology","volume":"66 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2015-11-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/0471140856.tx2507s66","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"50794207","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 49

High-Throughput Assays for Assessing Mitochondrial Dysfunction Caused by Compounds that Impair mtDNA-Encoded Protein Levels in Eukaryotic Cells 评估真核细胞中损害mtdna编码蛋白水平的化合物引起的线粒体功能障碍的高通量测定

Current protocols in toxicology Pub Date : 2015-08-07 DOI: 10.1002/0471140856.tx0311s48

Sashi Nadanaciva, James Murray, Casey Wilson, David F. Gebhard, Yvonne Will

{"title":"High-Throughput Assays for Assessing Mitochondrial Dysfunction Caused by Compounds that Impair mtDNA-Encoded Protein Levels in Eukaryotic Cells","authors":"Sashi Nadanaciva, James Murray, Casey Wilson, David F. Gebhard, Yvonne Will","doi":"10.1002/0471140856.tx0311s48","DOIUrl":"10.1002/0471140856.tx0311s48","url":null,"abstract":"Compounds that impair the synthesis of either mitochondrial DNA (mtNDA) or mtDNA-encoded proteins reduce the levels of 13 proteins essential for oxidative phosphorylation, leading to a decrease in mitochondrial ATP production. Toxicity caused by these compounds is seldom identified in 24 to 72 hr cytotoxicity assays due to the low turnover rates of both mtDNA and mtDNA-encoded proteins. Here, we describe three high-throughput screening assays that detect compounds that affect mtDNA-encoded protein levels. All three assays measure the levels of two proteins, one a mtDNA-encoded protein synthesized on mitochondrial ribosomes and the other, a nuclear DNA-encoded protein synthesized on cytosolic ribosomes. The first assay measures the levels of these two proteins by quantitative image analysis and requires a high-content imaging system. The second assay is an in-cell immunoassay that utilizes infrared dyes for detection of the two proteins and, thus, requires a LI-COR Odyssey system. The third assay is an in-cell immunoassay that utilizes colorimetric detection of the two proteins and requires an absorbance microplate reader. Curr. Protoc. Toxicol. 48:3.11.1-3.11.17. © 2011 by John Wiley & Sons, Inc.","PeriodicalId":72743,"journal":{"name":"Current protocols in toxicology","volume":"48 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2015-08-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1002/0471140856.tx0311s48","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29866990","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 6

Alkaline Comet Assay for Assessing DNA Damage in Individual Cells 评估单个细胞DNA损伤的碱性彗星试验

Current protocols in toxicology Pub Date : 2015-08-07 DOI: 10.1002/0471140856.tx0312s65

Xinzhu Pu, Zemin Wang, James E. Klaunig

引用次数: 79