Cell-cell interaction between platelets and IL-1 beta-stimulated vascular smooth muscle cells in synthesis of thromboxane A2.

Transcellular biosynthesis of thromboxane (Tx) A2 between vascular smooth muscle cells (SMC) and platelets has been investigated by using 14C-arachidonic acid (AA) radiolabeled rat SMC (or platelets) and the fate of the label in phospholipids and eicosanoid fractions was studied using radioimmunoassay (RIA) and thin-layer chromatography (TLC). Stimulation of SMC with interleukin-1 beta (IL-1 beta) resulted in production of cyclooxygenase metabolites (e.g. 6-keto-PGF1 alpha, PGE2, PGF2 alpha, PGD2), 15-, 11-, 5-HETE, and free AA1 with a coincident decline of phosphatidylcholine (PC) in SMC. IL-1 beta did not induce TXB2 production, a stable metabolite of TXA2 measured by TLC and radioimmunoassay, either in human platelets from 0.01-100 U/ml for 1 h or in SMC for 24 h. However, human platelets converted exogenous PGH2 to TXA2 despite cyclooxygenase inhibition or PGH2 receptor blockade. Furthermore, TXB2 was produced in large quantities during co-incubation of IL-1 beta-stimulated SMC with human platelets for 30 min in concert with a significant decrease of 6-keto-PGF1 alpha and eicosanoids (PGE2, PGF2 alpha and PGD2) compared with control (P < 0.01). Pretreatment of SMC with cycloheximide and actinomycin not only inhibited IL-1 beta-induced eicosanoid synthesis and phospholipid breakdown but also diminished TXB2 production when co-incubated with platelets. These data suggest that a cell-cell interaction, i.e. platelet utilizing SMC-derived endoperoxides for its TXA2 production, might cause an excess thromboxane A2 synthesis.

Cell-cell interaction of macrophages and vascular smooth muscle cells in the synthesis of leukotriene b(4).

Biosynthesis of LTB(4) during cell-cell interaction between vascular smooth muscle cells (SMC) and alveolar macrophages (AM) has been investigated by use of both high-pressure Hquid chromatography (HPLC) and radtoimmunoassay (RIA). Both interleukin-beta (IL-beta) and tumour necrosis factor-alpha (TNFalpha) induced a time- and dose-dependent synthesis of 15-, and 5-hydroxyeicosatetraenoic acids (HETEs) from cultured SMC. However, neither TNFalpha nor IL-1beta induced a significant LTB(4) production in SMC alone or AM alone after 24 h of incubation. Addition of IL-1beta and TNFalpha simultaneously to SMC resulted in a dose-dependent synergistic increase of HETEs. Macrophages dose-dependently transformed extremely low concentrations of exogenous LTA(4) into LTB(4). Incubation of vascular SMC with various numbers of AM in the presence of IL-1beta (5 units/ml) and TNFalpha (10 units/ml) induced a great increase of LTB(4) synthesis in comparison with the detectable levels of LTB(4) produced by macrophages alone. Pretreatment of SMC with NDGA, cycloheximide, and actinomycin not only inhibited IL-1 and TNT induced HETEs synthesis but also abolished LTB(4) production when co-incubated with macrophages. These results suggest that LTB(4) in a mixture of SMC and macrophages could originate from a transcellular metabolism, i.e. macrophages transforming SMC-derived LTA(4) into LTB(4).