Implication of Na+ kinetics in PGI2 generation of cultured human vascular endothelial cells

The regulatory mechanism of prostacyclin (PGI2) generation and its enhancement by ouabain and monensin in cultured human vascular endothelial cells was investigated with references to the kinetics of Ca++ and Na+. Na+-K+ ATPase activity was assayed as ouabain-sensitive 86Rb uptake. Cytosolic free calcium ion concentration ([Ca++]i) was measured using fura-2. The kinetics of 45Ca of 22Na, and [Ca++]i preceded PGI2 generation. Ouabain inhibited Na+-K+ ATPase activity (inhibition of 22Na release and 86Rb uptake) and increased 22Na uptake, while monensin enhanced 22Na uptake and Na+-K+ ATPase activity at 2.5min after starting incubation. Both of these reagents decreased 45Ca release and increased 45Ca uptake (inhibition of Na+-Ca++ exchange systems) at 2.5min. And all of these effects were attenuated at 10min. Ouabain and monensin increased [Ca++]i. Through these results it was concluded that the enhanced PGI2 generation by ouabain or monensin was speculated to be brought about by the increased [Ca++]i and Ca++ uptake which was based on the decreased Ca++ release via the inhibition of Na+-Ca++ exchange systems, and that enhanced PGI2 generation might be autoregulated by the attenuation of increased [Na+]i, which was derived from the attenuation of ouabain-induced inhibition of Na+-K+ ATPase activity, or from the enhancement of monensin-induced increase of its activity.