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Open Access Research

Cytosolic Ca2+ shifts as early markers of cytotoxicity

Philippe Wyrsch, Christian Blenn, Theresa Pesch, Sascha Beneke and Felix R Althaus*

Author Affiliations

Institute of Pharmacology and Toxicology, University of Zurich-Vetsuisse, Winterthurerstrasse 260, Zurich, CH-8057, Switzerland

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Cell Communication and Signaling 2013, 11:11  doi:10.1186/1478-811X-11-11

Published: 6 February 2013

Abstract

The determination of the cytotoxic potential of new and so far unknown compounds as well as their metabolites is fundamental in risk assessment. A variety of strategic endpoints have been defined to describe toxin-cell interactions, leading to prediction of cell fate. They involve measurement of metabolic endpoints, bio-energetic parameters or morphological cell modifications. Here, we evaluated alterations of the free cytosolic Ca2+ homeostasis using the Fluo-4 dye and compared results with the metabolic cell viability assay Alamar Blue. We investigated a panel of toxins (As2O3, gossypol, H2O2, staurosporine, and titanium(IV)-salane complexes) in four different mammalian cell lines covering three different species (human, mouse, and African green monkey). All tested compounds induced an increase in free cytosolic Ca2+ within the first 5 s after toxin application. Cytosolic Ca2+ shifts occurred independently of the chemical structure in all tested cell systems and were persistent up to 3 h. The linear increase of free cytosolic Ca2+ within the first 5 s of drug treatment correlates with the EC25 and EC75 values obtained in Alamar Blue assays one day after toxin exposure. Moreover, a rise of cytosolic Ca2+ was detectable independent of induced cell death mode as assessed by caspase and poly(ADP-ribose) polymerase (PARP) activity in HeLa versus MCF-7 cells at very low concentrations. In conclusion, a cytotoxicity assay based on Ca2+ shifts has a low limit of detection (LOD), is less time consuming (at least 24 times faster) compared to the cell viability assay Alamar Blue and is suitable for high-troughput-screening (HTS).

Keywords:
Alamar blue; Arsenic trioxide; Fluo-4; Gossypol; H2O2; Staurosporine