Endotoxin and mast cell granule proteases synergistically activate human coronary artery endothelial cells to generate interleukin-6 and interleukin-8

Mast cells (MC) are strategically located along blood vessels and, on activ ation, exocytose granules that contain many vasoactive mediators. Endotheli al cell (EC) activation, which includes the production of such cytokines as interleukin-6 (IL-6) and IL-8, is a key event in vascular inflammation. In this study, the effects of purified MC granules (MCG) on the production of IL-6 and IL-8 by human coronary artery EC (HCAEC) were examined. HCAEC wer e cocultured with MCG in the presence or absence of lipopolysaccharide (LPS ), and IL-6 and IL-8 levels in the culture medium were assayed by ELISA, Un activated HCAEC produced only low levels of IL-6 or IL-8, and the addition of MCG alone resulted in little or no increase in production of these cytok ines, LPS-activated HCAEC produced significant amounts of IL-6 and IL-8 in a dose-dependent and time-dependent fashion, which was amplified 23-fold by MCG at EC/MC ratios of 16:1-2:1, Scanning electron microscopy revealed dir ect communication between MCG and HCAEC. The enhancement of IL-6 and IL-8 p roduction by MCG was abrogated when MCG were pretreated with the serine pro tease inhibitor phenylmethylsulfonyl fluoride (PMSF), These results demonst rate that MCG interaction with HCAEC causes amplification of endotoxin-stim ulated cytokine production via serine proteases present in MCG, The synergi stic activation of EC by endotoxin and MCG proteases emphasizes the role of MC in amplifying vascular inflammation.