Roles Of Chloride Channels In Apoptosis Induced By Disulfiram/copper In Molt4 Cells

Objective:To assess the roles of chloride channels in apoptosis induced by disulfiram/copper in human acute lymphoblastic leukemia Molt4 cells.Methods:The whole-cell patch clamp technique was used to record volume-activated chloride currents. Volume changes of live cells were detected by the time-lapse imaging techniques. The permeability of the cell membrane and nuclear changes were evaluated by Hoechst 33258 staining. Apoptosis was monitored by flow cytometry.Results:1) Exposures of cells to 47%hypotonic challenges activated a volume-sensitive chloride. The current was outward-rectified and inhibited by the chloride channel blocker NPPB.2) 47%hypotonic bath solution swelled the cells and induced a regulatory volume decrease (RVD) which could be inhibited by NPPB.3) A volume-sensitive background chloride current was recorded in isotonic solutions. NPPB blocked the current and swelled the cells under isotonic conditions.4) Exracellular applications of disulfiram plus CuCl2 activated a volume-sensitive chloride current and induced apoptotic volume decrease (AVD) in Molt4 cells. These effects were depleted by NPPB.5) Hoechst 33258 staining indicated that the permeability of the cell membrane was increased significantly by disulfiram/copper treatments. The permeability increment was attenuated by NPPB.6) Incubation of cells in the medium containing disulfiram/copper induced apoptosis, demonstrated by flow cytometry. The chloride channel blocker NPPB prevented the cells from disulfiram/copper-induced apoptosis.Conclusion:1) Molt4 cells express volume-sensitive chloride channels.2) Volume-sensitive chloride channels are involved in cell volume regulation under hypotonic and isotonic conditions and play important role in maintenance of normal cell volume.3) The volume-sensitive chloride channels may be involved in disulfiram/copper-induced apoptosis via induction of AVD and permeabilization of the cell membrane.