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Contains fulltext : 117662.pdf (Publisher’s version ) (Open Access) Radboud Universiteit Nijmegen, 15 mei 2013 Promotor : Dekhuijzen, P.N.R. Co-promotores : Elshout, F.J.J. van den, Vos, P.J.E., Heijdra, Y.F.

The aim of this study was to explore the possibility that heat-induced alterations in calcium homeostasis are the cause of hyperthermic cell killing. Therefore, the intracellular free calcium concentration ([Ca2+](i)) was determined spectrofluorometrically, using the fluorescent calcium probe fura-2/acetoxy methylester (AM), at both physiological and hyperthermic temperatures in cell suspensions from six different tumor cell lines. For all cell lines fura-2 leakage appears to contribute to a change in the fluorescence signal and hence leads to a false indication of an increase in [Ca2+](i), especially at the hyperthermic temperature. Two methods were introduced that circumvent this problem and results in true values of [Ca2+](i). Also, measurements of [Ca2+](i) in single cells using a fluorescent microscopical technique (not affected by dye leakage) were used for comparison. All three ap preaches show that a hyperthermic treatment that kills >90% of the cells does not lead to changes in the [Ca2+](i) in most cell lines. Therefore, heat-induced alterations of calcium homeostasis cannot be considered the general cause for hyperthermic cell killing.