Characterization of endothelin secretion by vascular endothelial cells

The characterization of mechanisms that regulate ET-LP secretion from bovine adrenal cortical capillary endothelial cells (ACE) in culture was performed by developing radioimmunoassays that distinguish between ET 1–21 (AbET 1–21) and ET 1–39 (AbET 1–39). The conditioned media (DMEM) content of ET-like immunoreactivity (ET 1–21LI) increased from 50 to 350 pg/ml over a 24 h period. Addition of 10% calf serum or 0.1% BSA enhanced ET 1–21LI release 2–3 fold. Authenticity of ET 1–21LI was examined using reversed phase liquid chromatography. All ET 1–21LI co-eluted with authentic ET-1. Examination of ET 1–39IR by liquid chromatography revealed two peaks of immunoreactivity, one co-eluting with authentic ET 22–39 and a later running peak co-eluting with authentic ET 1–39. Neither ET 1–21LI nor ET 1–39LI was detected in the extracts of sonicated ACE cells. Treatment of cells with various forms of TGFβ significantly augmented ET 1–21LI release. These data suggest that ACE secretion of ET-LP in vitro occurs spontaneously and can be enhanced by TGFβ. Since neither ET 1–21LI nor ET 1–39LI was detectable in ACE cells it is unlikely that ET-LP are stored prior to their secretion.