Activation of cloned BKCa channels in nitric oxide-induced apoptosis of HEK293 cells

The large conductance Ca2+-activated K+ (BKCa) channels are highly expressed in vascular smooth muscle cells (VSMCs) and play an essential role in the regulation of various physiological functions. Besides its electrophysiological function in vascular relaxation, BKCa has also been reported to be implicated in nitric oxide (NO)-induced apoptosis of VSMCs. However, the molecular mechanism is not clear and has not been determined on cloned channels. The present study was designed to clarify whether activation of cloned BKCa channel was involved in NO-induced apoptosis in human embryonic kidney 293 (HEK293) cell. The cDNA encoding the α-subunit of BKCa channel, hSloα, was transiently transfected into HEK293 cells. The apoptotic death in HEK-hSloα cells was detected using immunocytochemistry, analysis of fragmented DNA by agarose gel electrophoresis, MTT test, and flow cytometry assays. Whole-cell and single-channel characteristics of HEK-hSloα cells exhibited functional features similar to native BKCa channel in VSMCs. Exposuring of HEK- hSloα cells to S-nitroso-N-acetyl-penicillamine increased the hSloα channel activities of whole-cell and single-channel, and then increased percentage of cells undergoing apoptosis. However, blocking hSloα channels with 1 mM tetraethylammonia or 100 nM iberiotoxin significantly decreased the NO-induced apoptosis, whereas 30 μM NS1619, the specific agonist of BKCa, independently increased hSloα currents and induced apoptosis. These results indicated that activation of cloned BKCa channel was involved in NO-induced apoptosis of HEK293 cells.