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Automation of a Liver Genotoxicity Assay

肝脏基因毒性测定的自动化

基本信息

批准号：
9038484
负责人：
Jeffrey C Bemis
金额：
$ 17.26万
依托单位：
LITRON LABORATORIES, LTD.
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-03-01 至 2016-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9038484
关键词：
Affect Age Animal Testing Animals Antibodies Apoptotic Automation Benchmarking Biological Assay Bone Marrow Cell Nucleus Cells Chemicals Comet Assay DNA DNA Damage DNA Double Strand Break Data Detergents Development Diethylnitrosamine Dose Dyes Erythrocytes Evaluation Experimental Designs Exposure to Family suidae Female Flow Cytometry Fluorescence Microscopy Frequencies Genetic Goals Harvest Hepatocyte Histopathology Japan Kinetics Label Laboratories Liver Manuals Measurement Measures Metabolism Methodology Methods Micronucleus Tests Microscopy Mitotic Modeling Mutagenicity Tests Mutagens Mutation Organ Performance Pharmaceutical Preparations Pharmacologic Substance Play Procedures Process Rattus Reagent Reporting Research Design Rodent Safety Sampling Schedule Scheme Scoring Method Site Speed Temperature Testing Time Tissue Harvesting Tissue Stains Tissues Toxic effect Toxicogenetics Toxicology Transgenic Organisms Validation age group base consumer product cost cytotoxicity design genotoxicity high throughput screening improved in vivo indexing male meetings method development micronucleus pre-clinical prototype public health relevance quinoline safety testing sex tissue processing tool treatment duration treatment group

项目摘要

DESCRIPTION (provided by applicant): Assessment of chemicals' potential to cause chromosomal damage is an established and important part of preclinical genotoxicity safety testing for many consumer products, industrial chemicals, and all pharmaceutical agents. Currently the mammalian erythrocyte micronucleus test is the most commonly employed assay for in vivo assessment of chromosomal damage, but this assay reports specifically on genotoxicity that occurs in the bone marrow. In order to obtain a more comprehensive understanding of potential genotoxicity, testing guidance's recommend evaluation of a second tissue. The liver, the site of metabolism and in many cases activation of genotoxicants, is usually regarded as the preferred second tissue. Even so, there is a lack of efficient and effective tools for studying liver genotoxicity. The Comet assay and transgenic rodent mutation models can be employed to study the liver, but these assays suffer from methodological and cost issues that limit their utility. Another important consideration is that these assays are not highly amenable to integration with on-going toxicology studies, meaning additional animals are required for the liver genotoxicity assessment. One alternative approach is to examine liver hepatocytes for the formation of micronuclei, an established indicator of chromosomal damage. However existing methods for examining liver micronuclei are still emerging and currently based on a multi-step sample processing scheme followed by manual scoring by microscopy. This approach is subjective and labor- intensive, and results in too few cells being scored for reliable enumeration of micronucleated hepatocytes, a situation that diminishes the ability of the test to detect weakly genotoxic agents. We will overcome these deficiencies by combining simple, rapid tissue processing and staining with high-speed flow cytometric analysis to greatly improve the execution of liver micronucleus scoring. Furthermore, we will multiplex several cytotoxicity measurements into the liver micronucleus assay, thereby providing information that we predict will be important for interpreting the genotoxicity results. The methodology will be reduced to practice in the form of commercially available kits, and will contribute to the reduction and refinement of animal testing, as it will make it feasible to integrate a liver genotoxicity assay ito ongoing toxicology studies. Overall, this project will meet a critical need in the practice of genetic toxicology by improving chemical safety assessments in several meaningful ways.