¹Johns Hopkins Oncology Center, Johns Hopkins School of Medicine, Baltimore, MD 21231, USA

²Department of Physiology, Université catholique de Louvain, Brussels B-1200, Belgium

³Department of Urology, James Buchanan Brady Urological Institute, Johns Hopkins School of Medicine, Baltimore, MD 21231, USA

Correspondence to: B Tombal, Department of Physiology, UCL St-Luc, UCL/Harvey/FYMU 5540, Avenue Hippocrate, 55, B-1200 Brussels, Belgium. Tel: 32 2 764 5540; Fax: 32 2 764 5580; E-mail: bertrand.tombal@fymu.ucl.ac.be

Edited by J Cidlowsky

Many agents, such as the endoplasmic reticulum Ca²⁺ ATPase inhibitor, thapsigargin, or the ionophore, ionomycin, induce apoptosis by transiently elevating [Ca²⁺]_i. The role of [Ca²⁺]_i in apoptosis induced by agents that do not immediately increase [Ca²⁺]_i, such as 5-FdUr, TGF-1, doxorubicin, or radiation, is far more controversial. In the present paper, [Ca²⁺]_i was measured continuously for 120 h. in prostate and bladder cancer cell lines exposed to these four agents: 5-FdUR, TGF-1, doxorubicin, or radiation. Each of them consistently induced a delayed [Ca²⁺]_i rise associated with the morphological changes that characterize the execution phase of apoptosis (i.e. rounding, blebbing). This [Ca²⁺]_i rise occurred in two consecutive steps (10 M and >10 M) and resulted from a Ca²⁺ influx from the extracellular medium. This delayed supramicromolar [Ca²⁺]_i rise was also observed previously in breast, prostate and bladder cancer cell lines exposed to thapsigargin. This influx regulated transcriptional reprogramming of Gadd153 and is required to activate cytochrome c release, caspase-3 activation, loss of clonal survival and DNA fragmentation. When cells were maintained in low extracellular Ca²⁺ media, these phenomena were temporarily delayed but occurred on return to normal Ca²⁺ medium. Similarly, apoptosis could be delayed by overexpressing the Ca²⁺-binding proteins, Calbindin-D_28K and parvalbumin. As this delayed 10 M [Ca²⁺]_i elevation was observed in a number of cell lines exposed to a variety of different agents, we conclude that such elevation constitutes a key and general event of apoptosis in these malignant cells.

Cell Death and Differentiation (2002) 9, 561-573. DOI: 10.1038/sj/cdd/4400999

apoptosis; calcium; Fura Dextran; Prostate; DNA fragmentation

5-FdUr, 5-fluorodeoxyuridine; TGF-1, transforming growth factor b1