Activation of caspase is independent to cyclin E expression in camptothecin-induced apoptosis in HL-60 cells

Abstract

Introduction: In response to the DNA topoisomerase I inhibitor camptothecin(CAM), leukemia HL-60 cells undergo characteristic morphological changes,caspase activation, and DNA fragmentation typical of apoptosis. The sequence ofapoptotic phenomena in an individual CAM-treated HL-60 cell is said to dependon the stage of proliferation of that cells when it encounters the CAM. This studydescribes further investigation on the dose and time effect of CAM on the apop-totic phenomena using the broad-spectrum caspase inhibitor Z-VAD-FMK. Therelationship between caspase and cyclin E in CAM-induced apoptosis was alsoinvestigated. Methods: The dose-response (10 nM, 50 nM and 1000 nM) of CAMon cell cycle progression and cell death on HL-60 cells in the presence andabsence of the caspase inhibitor was monitored by flow cytometry during 24-hcultures. Cell death was monitored by changes in cell light scatter and binding ofannexin V-fluorescein isothiocyanate to PS. Results: High doses (50, 1000 nM)of CAM executed all S-phase cells to undergo apoptosis after 4h whereas low (10nM) dose perturbed the whole cell cycle progression resulted with late apoptoticcell death after 12h. Caspase inhibition prevented PS exposure mainly from S-phase cells but had no effect on the cell death pathway of the non-S-phase cells.Cyclin E expression appeared earlier than activation of PS exposure and contin-ued to rise with treatment time; independent to concentrations of CAM and thepresence of general caspase inhibitor. Conclusion: The apoptotic phenomena inan individual HL-60 cell depend on the stage of proliferation of that cells, doseand time of CAM added. Our data show that caspases are only required for PSexposure-related cell death. The overexpression of cyclin E by CAM aims to sen-sitize the HL-60 cells to undergo apoptosis.