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Enhancement of thrombin‐ and lonomycin‐stimulated prostacyclin and platelet‐activating factor production in cultured endothelial cells by a tumor‐promoting phorbol ester
Author(s) -
Zavoico George B.,
Hrbolich Janet K.,
Gimbrone Michael A.,
Schafer Andrew I.
Publication year - 1990
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.1041430326
Subject(s) - ionomycin , phorbol , arachidonic acid , prostacyclin , umbilical vein , platelet activating factor , thrombin , platelet , chemistry , stimulation , incubation , biochemistry , medicine , endocrinology , biology , protein kinase c , in vitro , immunology , enzyme
Tumor‐promoting phorbol esters such as 4β‐phorbol 12‐myristate 13‐acetate (PMA) have been shown to act synergistically with Ca 2+ ionophores in cell activation, including stimulation of arachidonic acid metabolism. The effects of PMA on unstimulated and Ca 2+ ionophore‐ or thrombin‐stimulated PGI 2 and platelet‐activating factor (PAF) production in cultured bovine aortic endothelial cells (BAEC) and human umbilical vein endothelial cells (HUVEC) were investigated. Incubation of BAEC or HUVEC for 5–10 min with 100 nM PMA alone slightly increased basal PGI 2 production. PGI 2 production was rapidly stimulated in BAEC and HUVEC treated with the Ca 2+ ionophore ionomycin. Preincubation of BAEC of HUVEC with 100 nM PMA for 5–10 min followed by ionomycin for up to 60 min enhanced PGI 2 production up to 2.5‐fold. Pretreatment with 100 nM PMA for 5 min also caused a 2‐fold enhancement of thrombin‐stimulated (1 U/ml) PGI 2 production in HUVEC. The production of other prostaglandins, PGF 2α , PGE 2 , and PGD 2 , was also enhanced. In contrast, PMA had no effect on PGI 2 synthesized directly from exogenous arachidonic acid or PGH 2 . The inactive phorbol ester 4α‐phorbol 12,13‐didecanoate was without effect. Since the biosyntheses of both PGI 2 and PAF share a common first step, the hydrolysis of their respective phospholipid precursors by phospholipase A 2 , we investigated whether PMA preincubation could also enhance PAF biosynthesis. Incubation of HUVEC with 100 nM PMA alone had a negligible effect on PAF production. However, thrombin‐stimulated (1 U/ml) PAF production was enhanced 2.6‐fold by preincubation with 100 nM PMA. The protein kinase C inhibitors H‐7 and staurosporine ablated the enhancing effect of PMA on thrombin‐stimulated PGI 2 and PAF biosynthesis. These results demonstrate that PMA can significantly alter the production of PGI 2 and PAF in vascular endothelial cells, and suggest that protein kinase C activation modulates phospholipase A 2 activity in this cell type.