Induction of apoptosis by intracellular potassium ion depletion: using the fluorescent dye PBFI in a 96-well plate method in cultured lung cancer cells.

Abstract

Depletion of intracellular potassium ions (K+) is necessary for cells to shrink, activate caspases and induce DNA fragmentation, events which are features of apoptosis. Here we describe a 96-well plate method using the cell permeable form of K+ binding benzofuran isophtalate (PBFI-AM) to measure intracellular K+ content in relation to untreated control. Cultured human pulmonary mesothelioma cells (P31) and small-cell lung cancer cells (U1690) were treated with K+ flux modulators in order to deprive the cells of intracellular K+. The combination of K+ influx inhibition with 10 micromol/L bumetanide plus 10 micromol/L ouabain and K+ efflux stimulation with 3 mg/L amphotericin B or 5 micromol/L nigericin efficiently reduced the intracellular K+ content after 3 h. Manipulation of K+ fluxes with subsequent intracellular K+ depletion induced apoptosis of lung cancer cells, as detected by caspase-3 activity after 3 h K+ depletion followed by 24 h proliferation and TUNEL positive staining after 48 h proliferation. We concluded that the PBFI-AM assay was a useful tool to determine intracellular K+ content in relation to untreated control, and that intracellular K+ depletion of lung cancer cells by clinically used drugs of relevant concentrations induced apoptosis. These findings may lead to novel therapeutic strategies in the treatment of lung cancer.

• Authors: B Andersson, V Janson, P Behnam-Motlagh, R Henriksson, K Grankvist
• Journal: Toxicology in vitro : an international journal published in association with BIBRA (Toxicol In Vitro) Vol. 20 Issue 6 Pg. 986-94 (Sep 2006) ISSN: 0887-2333 [Print] England
• PMID: 16483738 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Benzofurans
• Ethers, Cyclic
• potassium-binding benzofuran-isophthalate acetoxymethyl ester
• CASP3 protein, human
• Caspase 3
• Caspases
• Potassium

Topics

• Apoptosis (drug effects)
• Benzofurans (pharmacology)
• Caspase 3
• Caspases (metabolism)
• Cell Line, Tumor
• Ethers, Cyclic (pharmacology)
• Humans
• In Situ Nick-End Labeling
• Lung Neoplasms (pathology)
• Potassium (analysis, metabolism)