Abstract
The present study is aimed to determine the role of Ca2+ signaling evoked by hydrogen peroxide (H2O2) on caspase activation in human leukemia cell line HL-60. We have analysed cytosolic free Ca2+ concentration ([Ca2+]c) determination, mitochondrial membrane potential and caspase-3 and -9 activity by fluorimetric methods, using the fluorescent ratiometric Ca2+ indicator Fura-2, the dye JC-1, and specific fluorogenic substrate, respectively. Our results indicated that treatment of HL-60 cells with H2O2 induced a transient increase in [Ca2+]c due to Ca2+ release from internal stores. The stimulatory effect of H2O2 on Ca2+ signal was followed by a mitochondrial membrane depolarization. Our results also indicated that H2O2 was able to increase the caspase-3 and -9 activities. The effect of H2O2 on caspase activation was time dependent, reaching a maximal caspase activity after 120 min of stimulation. Loading of cells with dimethyl BAPTA, an intracellular Ca2+ chelator, significantly reduced H2O2-induced mitochondrial depolarization and caspase activation. Similar results were obtained when the cells were pretreated with Ru360, a specific blocker of Ca2+ uptake into mitochondria. The findings suggest that H2O2-induced caspase-3 and -9 activation and mitochondrial membrane depolarization is dependent on rises in [Ca2+]c in human myeloid HL-60 cells.
Keywords: Apoptosis, caspases, Ca2+ signal, hydrogen peroxide, HL-60 cells
Current Signal Transduction Therapy
Title: Oxidative Stress-Induced Caspases are Regulated in Human Myeloid HL-60 Cells by Calcium Signal
Volume: 5 Issue: 2
Author(s): D. Gonzalez, I. Bejarano, C. Barriga, A.B. Rodriguez and J.A. Pariente
Affiliation:
Keywords: Apoptosis, caspases, Ca2+ signal, hydrogen peroxide, HL-60 cells
Abstract: The present study is aimed to determine the role of Ca2+ signaling evoked by hydrogen peroxide (H2O2) on caspase activation in human leukemia cell line HL-60. We have analysed cytosolic free Ca2+ concentration ([Ca2+]c) determination, mitochondrial membrane potential and caspase-3 and -9 activity by fluorimetric methods, using the fluorescent ratiometric Ca2+ indicator Fura-2, the dye JC-1, and specific fluorogenic substrate, respectively. Our results indicated that treatment of HL-60 cells with H2O2 induced a transient increase in [Ca2+]c due to Ca2+ release from internal stores. The stimulatory effect of H2O2 on Ca2+ signal was followed by a mitochondrial membrane depolarization. Our results also indicated that H2O2 was able to increase the caspase-3 and -9 activities. The effect of H2O2 on caspase activation was time dependent, reaching a maximal caspase activity after 120 min of stimulation. Loading of cells with dimethyl BAPTA, an intracellular Ca2+ chelator, significantly reduced H2O2-induced mitochondrial depolarization and caspase activation. Similar results were obtained when the cells were pretreated with Ru360, a specific blocker of Ca2+ uptake into mitochondria. The findings suggest that H2O2-induced caspase-3 and -9 activation and mitochondrial membrane depolarization is dependent on rises in [Ca2+]c in human myeloid HL-60 cells.
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Gonzalez D., Bejarano I., Barriga C., Rodriguez A.B. and Pariente J.A., Oxidative Stress-Induced Caspases are Regulated in Human Myeloid HL-60 Cells by Calcium Signal, Current Signal Transduction Therapy 2010; 5 (2) . https://dx.doi.org/10.2174/157436210791112172
DOI https://dx.doi.org/10.2174/157436210791112172 |
Print ISSN 1574-3624 |
Publisher Name Bentham Science Publisher |
Online ISSN 2212-389X |
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