Human umbilical vein endothelial cells: specific binding of platelet-activating factor and cytosolic calcium flux

An interaction of the platelet-activating factor (Paf) with endothelial cells was investigated using human umbilical vein endothelial cells. Confluent endothelial cells bound [3H]Paf in the presence of 0.25% fatty acid-free serum albumin after culture in media containing either heat-inactivated foetal calf serum or serum substitute. The Scatchard analysis of the saturated specific [3H]Paf binding showed a Bmax of 2.5 fmol indicating 2800 binding sites per endothelial cell. [3H]Paf binding was partially reversible at 20 degrees and 4 degrees and endothelial cells partially metabolized [3H]Paf at 20 degrees but not at 4 degrees. [3H]Paf binding and Paf-mediated increase of cytosolic free calcium were inhibited by specific Paf receptor antagonists which do not interfere with Paf metabolism. Immortalized umbilical vein endothelial cells bound [3H]Paf specifically after culture in the presence of insulin (20 hr, 0.4 U/mL) with non-specific binding in the absence of insulin. The results show that specific Paf binding mediated calcium flux in human endothelial cells.