INTRACELLULAR POOL OF VASCULAR ENDOTHELIAL GROWTH-FACTOR IN HUMAN NEUTROPHILS

Vascular endothelial growth factor (VEGF), an endothelial cell mitogen , is a potent angiogenic factor produced by several cell types. Whethe r human neutrophils are potential producers of VEGF has not yet been d escribed. The present work shows that phorbol-12-myristate 13-acetate (PMA), fMet-Leu-Phe, and tumor necrosis factor-alpha (TNF-alpha) trigg ered a time-dependent secretion of VEGF by human neutrophils. Cells in cubated with 50 ng/mL of PMA released significant amounts of VEGF afte r 15 minutes. Because the extracellular content of VEGF in human neutr ophils supernatants remained constant over a period of 2 to 24 hours a nd because PMA is a potent inducer of human neutrophil degranulation, the PMA-induced secretion of VEGF may be due to a preexisting intracel lular pool of this molecule. This hypothesis was reinforced by the abs ence of cycloheximide effect on the PMA-induced secretion of VEGF. The existence of an intracellular pool of VEGF was confirmed by measuring the intracellular content of VEGF in resting neutrophils. A dose-depe ndent inhibition of PMA-induced VEGF secretion was observed when the c ells were incubated in the presence of pentoxifylline, a methylxanthin e known to inhibit neutrophil degranulation. To confirm the implicatio n of neutrophil degranulation in VEGF release, the effects of two indu cers of physiologic degranulation, fMet-Leu-Phe and TNF-alpha, were de termined. Both agonists induced a release of VEGF in the absence of cy tochalasin B, confirming the involvement of neutrophil degranulation a nd suggesting the intracellular localization of VEGF in the specific g ranule fraction.; In addition, the kinetics of fMet-Leu-Phe- and TNF-a lpha-induced secretion of lactoferrin were similar to those of VEGF re lease induced by these two both agonists. The subcellular fractionatio n of human neutrophils showed a granule-specific distribution of the i ntracellular pool of VEGF in resting neutrophils. The finding that hum an neutrophils contain an intracellular pool of VEGF, secreted in the extracellular space under PMA-, fMet-Leu-Phe-, and TNF-alpha-induced d egranulation, suggests a role for human neutrophils as cellular effect ers of physiologic as well as pathologic angiogenesis. (C) 1997 by The American Society of Hematology.