Possible involvement of a lipocortin in the initiation of DNA synthesis by human endothelial cells.

This work focused on three themes. First, evidence was obtained for the presence of proteins of 34, 35, 32, and 69 kDa immunologically related to lipocortins I, II, V, and VI, respectively, in human umbilical vein endothelial (HUVE) cells. The 69-kDa protein (p69), but not proteins related to lipocortins I, II, and V, exhibited an increased phosphorylation after exposure of cells to basic fibroblast growth factor (bFGF) and phorbol ester PMA. Second, treatment of HUVE cell particulate fractions with EGTA and hydrophobic affinity chromatography in combination with conventional techniques provided extracts rich in p69 and purified p69. p69 from control cells and extracts from control, bFGF-treated, and PMA-treated cells were found to possess anti-phospholipase A2 (PLA2) activity of lipocortin. In contrast, a striking reverse effect occurred when extracts were obtained from cells exposed to bFGF plus PMA. Third, the combination of bFGF and PMA induced a stimulated PLA2-catalyzed release of arachidonic acid in HUVE cells. This arachidonate production was shown to be involved in the decision of cells to enter into DNA synthesis. Taken together, the present results suggest that phosphorylation of p69 is causally involved in the control of commitment to growth in HUVE cells by acting as a coupling mechanism between surface stimuli and arachidonate pathways.