PUBLICATION
Use of fish liver PLHC-1 cells and zebrafish embryos in cytotoxicity assays
- Authors
- Krone, P.H., Blechinger, S.R., Evans, T.G., Ryan, J.A., Noonan, E.J., and Hightower, L.E.
- ID
- ZDB-PUB-050120-1
- Date
- 2005
- Source
- Methods (San Diego, Calif.) 35(2): 176-187 (Journal)
- Registered Authors
- Evans, Tyler, Krone, Patrick H.
- Keywords
- Heat shock proteins; Cytotoxicity assays; PLHC-1 cells; Zebrafish; Embryos; Cadmium; Cellular stress response
- MeSH Terms
-
- Animals
- Animals, Genetically Modified
- Cadmium/metabolism
- Cell Line
- Cell Line, Tumor
- Cell Survival
- Cytoskeleton/metabolism
- DNA/metabolism
- Dose-Response Relationship, Drug
- Electrophoresis, Polyacrylamide Gel
- Embryo, Nonmammalian/metabolism*
- Embryonic Development
- Fishes
- Gene Expression Regulation, Developmental
- Genes, Reporter
- Green Fluorescent Proteins/metabolism
- HSP70 Heat-Shock Proteins/genetics
- HSP70 Heat-Shock Proteins/metabolism
- Liver/metabolism*
- Signal Transduction
- Statistics as Topic
- Zebrafish
- PubMed
- 15649845 Full text @ Methods
Citation
Krone, P.H., Blechinger, S.R., Evans, T.G., Ryan, J.A., Noonan, E.J., and Hightower, L.E. (2005) Use of fish liver PLHC-1 cells and zebrafish embryos in cytotoxicity assays. Methods (San Diego, Calif.). 35(2):176-187.
Abstract
Heat shock proteins (HSPs) indicate exposure to cellular stress and adverse cellular effects, thus serving as biomarkers of these effects. The highly conserved Hsp70 proteins are expressed under proteotoxic conditions, whereas small HSPs are expressed in response to stressors acting on the cytoskeleton and cell signaling pathways. Poeciliopsis lucida hepatocellular carcinoma line 1 (PLHC-1) cells have been used extensively for studying effects of cytotoxicity. A number of assays have been developed to examine DNA levels, protein levels, growth rate, morphological changes, and viability. The boundary between sub-lethal and lethal effects of particular stressors has been determined. The methodology and analytical framework for these techniques along with sample assays using cadmium stressed PLHC-1 cells are described. A range of methodologies have been developed in the past decade that allow the analysis and interpretation of gene expression and function in vivo in zebrafish embryos, and many of these are now being applied to the development of embryotoxicity assays. Here we provide the theoretical background and methodology for utilizing Hsp70 expression as an indicator of toxicity in the zebrafish embryo. Hsp70 expression is activated in a tissue-specific manner in zebrafish larvae following exposure to a number of different toxicants, including cadmium. This has allowed the development of an hsp70/eGFP reporter gene system in stable transgenic zebrafish that serves as a reliable yet extremely quick indicator of cell-specific toxicity in the context of the multicellular, living embryo.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping