title: A supramicromolar elevation of intracellular free calcium ([Ca(2+)](i)) is consistently required to induce the execution phase of apoptosis.
abstract: Many agents, such as the endoplasmic reticulum Ca(2+) ATPase inhibitor, thapsigargin, or the ionophore, ionomycin, induce apoptosis by transiently elevating [Ca(2+)](i). The role of [Ca(2+)](i) in apoptosis induced by agents that do not immediately increase [Ca(2+)](i), such as 5-FdUr, TGF beta-1, doxorubicin, or radiation, is far more controversial. In the present paper, [Ca(2+)](i) was measured continuously for 120 h. in prostate and bladder cancer cell lines exposed to these four agents: 5-FdUR, TGF beta-1, doxorubicin, or radiation. Each of them consistently induced a delayed [Ca(2+)](i) rise associated with the morphological changes that characterize the execution phase of apoptosis (i.e. rounding, blebbing). This [Ca(2+)](i) rise occurred in two consecutive steps (< or = 10 microM and >10 microM) and resulted from a Ca(2+) influx from the extracellular medium. This delayed supramicromolar [Ca(2+)](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 Ca(2+) media, these phenomena were temporarily delayed but occurred on return to normal Ca(2+) medium. Similarly, apoptosis could be delayed by overexpressing the Ca(2+)-binding proteins, Calbindin-D(28K) and parvalbumin. As this delayed >or = 10 microM [Ca(2+)](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.
authors.affiliation: Johns Hopkins Oncology Center, Johns Hopkins School of Medicine, Baltimore, MD 21231, USA. bertrand.tombal@fymu.ucl.ac.be
authors.forename: B
authors.initials: B
authors.lastname: Tombal
authors.forename: S R
authors.initials: SR
authors.lastname: Denmeade
authors.forename: J-M
authors.initials: JM
authors.lastname: Gillis
authors.forename: J T
authors.initials: JT
authors.lastname: Isaacs
mesh_terms: D017209:Apoptosis / Q000502:physiology*; D064026:Calbindins; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000235:genetics / Q000378:metabolism; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism; D003574:Cytochrome c Group / Q000378:metabolism; D053938:DNA Fragmentation / Q000502:physiology; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D004317:Doxorubicin / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000473:pathology / Q000502:physiology / Q000528:radiation effects; D006801:Humans; D007424:Intracellular Fluid / Q000737:chemistry; D010320:Parvalbumins / Q000235:genetics / Q000378:metabolism*; D011839:Radiation, Ionizing; D064030:S100 Calcium Binding Protein G / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000502:physiology; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D053773:Transforming Growth Factor beta1; D014407:Tumor Cells, Cultured
chemical_list: D064026:Calbindins; D002135:Calcium-Binding Proteins; D003574:Cytochrome c Group; D010320:Parvalbumins; D064030:S100 Calcium Binding Protein G; C508960:TGFB1 protein, human; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; D003841:Deoxycytidine; D004317:Doxorubicin; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; C007746:5-fluoro-2'-deoxycytidine; D002118:Calcium