Hypoxia-induced increase in intracellular calcium concentration in endothelial cells: Role of the Na+-glucose cotransporter

Hypoxia is a common denominator of many vascular disorders, especially thos e associated with ischemia. To study the effect of oxygen depletion on endo thelium, we developed an in vitro model of hypoxia on human umbilical vein endothelial cells (HUVEC). Hypoxia strongly activates HUVEC, which then syn thesize large amounts of prostaglandins and platelet-activating factor. The first step of this activation is a decrease in ATP content of the cells, f ollowed by an increase in the cytosolic calcium concentration ([Ca2+](i)) w hich then activates the phospholipase A(2) (PLA(2)). The link between the d ecrease in ATP and the increase in [Ca2+](i) was not known and is investiga ted in this work. We first showed that the presence of extracellular Na+ wa s necessary to observe the hypoxia-induced increase in [Ca2+](i) and the ac tivation of PLA,. This increase was not due to the release of Ca2+ from int racellular stores, since thapsigargin did not inhibit this process. The Na/Ca2+ exchanger was involved since dichlorobenzamil inhibited the [Ca2+](i) and the PLA(2) activation. The glycolysis was activated, but the intracell ular pH (pH(i)) in hypoxic cells did not differ from control cells. Finally , the hypoxia-induced increase in [Ca2+](i) and PLA(2) activation were inhi bited by phlorizin, an inhibitor of the Na+-glucose cotransport. The propos ed biochemical mechanism occurring under hypoxia is the following: glycolys is is first activated due to a requirement for ATP, leading to an influx of Na+ through the activated Na+-glucose cotransport followed by the activati on of the Na+/Ca2+ exchanger, resulting in a net influx of Ca2+. (C) 2001 W iley-Liss, Inc.