Development of a mechanistically-based genetically engineered PC12 cell system to detect p53-mediated cytotoxicity

Erwin van Vliet, Chantra Eskes, Silvia Stingele, Joanne Gartlon, Anna Price, Massimo Farina, Jessica Ponti, Thomas Hartung, Enrico Sabbioni, Sandra Coecke

Research output: Contribution to journalArticle

Abstract

The human wild type p53 gene, key for apoptosis, was introduced into the pheochromocytoma (PC12) cell line, to create a mechanistically-based in vitro test model for the detection of p53-mediated toxicity. Expression of the wt p53 gene was regulated by a system, which allowed or blocked expression p53 by absence or presence of tetracycline in the culture media. Western blot analyses confirmed an inducible and tetracycline-dependent expression of the wt p53 protein. Functionality of the p53 protein was verified by camptothecin treatment, known to induce p53-dependent apoptosis. Results showed that p53-expressing cells were significantly more sensitive to camptothecin induced cytotoxicity compared to non-expressing cells, and presented a significantly higher incidence of apoptosis. A screening study on 31 metal compounds, showed that the classified human carcinogens (NaAsO2, CdSO4 · 8H2O, Na2CrO4 · 4H2O, MnCl2, (NH4)2PtCl6) significantly increased cytotoxicity in p53-expressing cells compared to non-expressing cells, suggesting that their cytotoxicity was p53-mediated. Finally, acute and subchronic treatment with methyl mercury showed no significant differences in cytotoxicity and the percentage of apoptosis or necrosis between p53-expressing and non-expressing differentiated cells, suggesting that methyl mercury cytotoxicity was p53-independent.

Original languageEnglish (US)
Pages (from-to)698-705
Number of pages8
JournalToxicology in Vitro
Volume21
Issue number4
DOIs
StatePublished - Jun 1 2007
Externally publishedYes

Keywords

  • Apoptosis
  • Arsenic
  • Camptothecin
  • In vitro
  • Mercury
  • Metals
  • p53

ASJC Scopus subject areas

  • Toxicology

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  • Cite this

    van Vliet, E., Eskes, C., Stingele, S., Gartlon, J., Price, A., Farina, M., Ponti, J., Hartung, T., Sabbioni, E., & Coecke, S. (2007). Development of a mechanistically-based genetically engineered PC12 cell system to detect p53-mediated cytotoxicity. Toxicology in Vitro, 21(4), 698-705. https://doi.org/10.1016/j.tiv.2006.12.004