C. Plumpton et al., MEASUREMENT OF C-TERMINAL FRAGMENT OF BIG ENDOTHELIN-1 - A NOVEL METHOD FOR ASSESSING THE GENERATION OF ENDOTHELIN-1 IN HUMANS, Journal of cardiovascular pharmacology, 26, 1995, pp. 34-36
We developed a procedure for individual measurement of big endothelin-
1 (big ET-1) and the two products of big ET-1 conversion (ET-1 and C-t
erminal fragment (CTF) of big ET-1 (big ET-1(22-38), using selective s
olid-phase extraction and specific radioimmunoassays. These techniques
were used to measure the levels of these peptides in extracts of huma
n plasma after brachial artery infusions of big ET-1. Infusion of big
ET-1 (50 pmol/min, n = 6) caused a decrease in forearm blood flow from
3.03 to 1.55 ml/dl/min after 60 min (p < 0.05). In agreement, the lev
els of plasma immunoreactive (IR) big ET-1, ET, and CTF were significa
ntly increased from basal levels in the infused arm (from undetectable
to 386 pM, 2.2-7.5 pM, and 0.17-37 pM, respectively; p < 0.05). In th
e presence of the metalloprotease inhibitor phosphoramidon (30 nmol/mi
n), the increase in IR-ET and the associated vasoconstriction were abo
lished. However, IR-CTF was still detected, suggesting that either som
e conversion by phosphoramidon-insensitive endothelin-converting enzym
e (ECE) was occurring and/or that CTF was being preserved from further
proteolysis by phosphoramidon. These data confirm that exogenous big
ET-1 is locally converted to ET-1 and CTF in the human forearm, at lea
st in part by a phosphoramidon-sensitive ECE. Furthermore, because mea
surable levels of newly synthesized ET-1 are probably rapidly reduced
as a result of receptor binding, the assay of IR-CTF may be a more sen
sitive measure of the overexpression of ET-1 in disease.