Conversion of big endothelin‐1 to endothelin‐1 by two types of metalloproteinases derived from porcine aortic endothelial cells

Abstract

Incubation of big endothelin‐1 (big ET‐1_1–39) with either the cytosolic or membrane fraction obtained from cultured endothelial cells, resulted in an increase in immunoreactive‐endothelin (IR‐ET), which was markedly inhibited by metal chelators. Phosphoramidon, a metalloproteinase inhibitor, specifically suppressed the membrane fraction‐induced increase in IR‐ET, whereas the increase in IR‐ET observed with the cytosolic fraction was not influenced by phosphoramidon. Reverse‐phase (RP)‐HPLC of the incubation mixture of big ET‐1 with the cytosolic or membrane fraction revealed one major IR‐ET component corresponding to the elution position of synthetic ET‐1_1–21. Simultaneously, immunoreactivities like the C‐terminal fragment (CTF_22–39) of big ET‐1 were present, as deduced from the RP‐HPLC coupled with the radioimmunoassay for CTF. Our results indicate the presence of two types of metalloproteinases, which convert big ET‐1 to ET‐1 via a single cleavage between Trp²¹ and Val²², in vascular endothelial cells.