Arsenic trioxide (As2O3) induced calcium signals and cytotoxicity in two human cell lines: SY-5Y neuroblastoma and 293 embryonic kidney (HEK).
Arsenic trioxide (As(2)O(3)) has anticancer properties; however, its use also leads to neuro-, hepato- or nephro-toxicity, and therefore, it is important to understand the mechanism of As(2)O(3) toxicity. We studied As(2)O(3) influence on intracellular calcium ([Ca(2+)](i)) homeostasis of human neuroblastoma SY-5Y and embryonic kidney cells (HEK 293). We also relate the As(2)O(3) induced [Ca(2+)](i) modifications with cytotoxicity. We used Ca(2+) sensitive dyes (fluo-4 and rhod-2) combined with laser scanning microscopy or fluorescence activated cell sorting to measure Ca(2+) changes during the application of As(2)O(3) and we approach evaluation of cytotoxicity. As(2)O(3) (1 microM) increased [Ca(2+)](i) in SY-5Y and HEK 293 cells. Three forms of [Ca(2+)](i)-elevations were found: (1) steady-state increases, (2) transient [Ca(2+)](i)-elevations and (3) Ca(2+)-spikes. [Ca(2+)](i) modifications were independent from extracellular Ca(2+) but dependent on internal calcium stores. The effect was not reversible. Inositol triphosphate (IP(3)) and ryanodine (Ry) receptors are involved in regulation of signals induced by As(2)O(3). 2-APB and dantrolene significantly reduced the [Ca(2+)](i)-rise (p<0.001, t-test) but did not completely abolish [Ca(2+)](i)-elevation or spiking. This indicates that other Ca(2+) regulating mechanisms are involved. In cytotoxicity tests As(2)O(3) significantly reduced cell viability in both cell types. Staining with Hoechst 33342 showed occurrence of apoptosis and DNA damage. Our data suggest that [Ca(2+)](i) is an important messenger in As(2)O(3) induced cell death.
Florea
Ana-Maria
AM
Institut für Physiologie, Universitätsklinikum, Universität Duisburg Essen, Hufelandstrasse 55, 45122 Essen, Germany.
Splettstoesser
Frank
F
Büsselberg
Dietrich
D
D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D000077237:Arsenic Trioxide; D001152:Arsenicals / Q000494:pharmacology*; D001896:Boron Compounds / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects*; D002460:Cell Line; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004249:DNA Damage; D003620:Dantrolene / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D006801:Humans; D053496:Inositol 1,4,5-Trisphosphate Receptors / Q000378:metabolism; D018613:Microscopy, Confocal; D008938:Mitosis / Q000187:drug effects; D010087:Oxides / Q000494:pharmacology*; D019837:Ryanodine Receptor Calcium Release Channel / Q000378:metabolism; D013997:Time Factors
D000970:Antineoplastic Agents; D001152:Arsenicals; D001896:Boron Compounds; D053496:Inositol 1,4,5-Trisphosphate Receptors; D010087:Oxides; D019837:Ryanodine Receptor Calcium Release Channel; C109986:2-aminoethoxydiphenyl borate; D003620:Dantrolene; D000077237:Arsenic Trioxide; D002118:Calcium