Inhibition of Hypertonicity-induced Cation Channels Sensitizes HeLa Cells to Shrinkage-induced Apoptosis

Hypertonicity-induced cation channels (HICCs) are an effective mechanism of regulatory volume increase (RVI), which is a restoration process of cell volume after osmotic cell shrinkage, in HeLa cells. Since a reduction of cell size is a hallmark of programmed cell death, we tested whether a blockage of HICCs sensitizes HeLa cells to shrinkage-induced apoptosis by using proliferation assays, apoptosis assays, and patch-clamp recordings. Under control conditions, increasing osmolality up to 600 mosmol/kg-H2O had no detectable effect on either cell proliferation or apoptosis. With HICCs blocked by flufenamate and Gd3+, however, a significant reduction of proliferation and a stimulation of apoptosis were observed. Both effects exhibited virtually identical sensitivity profiles to osmotic stress as well as to flufenamate and Gd3+. Moreover, the observed concentration dependency of flufenamate and Gd3+ on proliferation and apoptosis was in excellent accordance with that on HICC inhibition. These results suggest that persistent cell shrinkage may function as a specific signal in the induction of apoptosis. In addition, they provide further evidence for the interplay of proliferation vs. apoptosis and the actual role that mechanisms of cell volume regulation do play in these processes.