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Transfection of human mesothelial cells mediated by different asbestos fiber types
L Gan1, E F Savransky, T M Fasy
1Department of Pathology, Mount Sinai School of Medicine, New York, New York 10029.
Abstract:
Several different asbestos fiber types mediate transfection of human mesothelial cells by exogenous DNA. We have employed the human MeT-5A mesothelial cell line, which allows the use of DNA replication as an assay for entry of DNA when plasmids bearing the SV40 origin of replication are used for transfection. We find that Canadian chrysotile, Calidria chrysotile, amosite, and crocidolite are each capable of introducing plasmid pSVod DNA into MeT-5A cells followed by subsequent replication of a fraction of the plasmid DNA. A significant fraction of the input plasmid DNA associated with the cells in the presence of asbestos is fragmented, and this fragmentation is particularly evident with crocidolite. Each of the fiber types is highly cytotoxic for the MeT-5A cells, and these cells actively accumulate the added fibers from the surrounding environment as visualized by phase-contrast microscopy. MeT-5A cells were transfected at higher efficiency with calcium phosphate than were several other primate cell lines. Calcium phosphate, however, did not induce fragmentation of the input plasmid DNA. Compared with several different mineral agents, including glass fibers, kaolin, and talc, Calidria chrysotile fibers were most effective at mediating transfection of the MeT-5A cells. Results provide a mechanism by which transfection can contribute to mutagenicity of asbestos fibers and indicate that this mechanism can operate in human mesothelial cells.
Insights
Asbestos fibers like chrysotile and crocidolite can deliver DNA into human mesothelial cells, potentially causing mutations. This asbestos-mediated transfection mechanism highlights a pathway for asbestos-induced genotoxicity.
Area of Science:
- Cell Biology
- Toxicology
- Environmental Health
Background:
- Asbestos exposure is linked to mesothelioma and other cancers.
- The precise mechanisms by which asbestos induces cellular damage and mutagenicity are not fully understood.
Purpose of the Study:
- To investigate the ability of different asbestos fiber types to mediate the transfection of human mesothelial cells with exogenous DNA.
- To explore the potential role of asbestos-induced DNA fragmentation and cytotoxicity in this transfection process.
Main Methods:
- Utilized the human MeT-5A mesothelial cell line for DNA transfection assays.
- Employed plasmids with the SV40 origin of replication to assess DNA entry and replication.
- Visualized fiber uptake and cytotoxicity using phase-contrast microscopy.
- Compared asbestos fiber transfection efficiency with other mineral agents.
Main Results:
- Canadian chrysotile, Calidria chrysotile, amosite, and crocidolite effectively mediated DNA transfection in MeT-5A cells.
- A significant fraction of the introduced plasmid DNA was fragmented, particularly with crocidolite.
- All tested asbestos fiber types were cytotoxic to MeT-5A cells, which actively accumulated the fibers.
- Calidria chrysotile demonstrated the highest transfection efficiency among the tested mineral agents.
Conclusions:
- Asbestos fibers can facilitate the entry and replication of exogenous DNA in human mesothelial cells.
- DNA fragmentation induced by asbestos fibers may contribute to their mutagenic potential.
- These findings suggest a mechanism for asbestos-induced genotoxicity operating in human mesothelial cells.