BACKGROUND: Endothelin-1 is an endothelium-derived potent vasoconstrictor peptide of 21 amino acids. To establish reference values in different models of hypertension and in human subjects an assay for plasma immunoreactive endothelin-1 (ET-1) was optimized. METHODS: ET-1 is extracted by acetone from 1 mL of plasma and subjected to a sensitive enzyme-linked immunosorbent assay. RESULTS: The detection limit for plasma ET-1 is 0.05 fmol/mL. Mean recoveries of the 1, 2, 5, and 10 fmol of ET-1 added to 1 mL of plasma were 66%, 75%, 85%, and 92%, respectively. Within- and between-assay coefficients of variation were < or =12% and < or =10%, respectively. Assay accuracy was demonstrated by consistent recoveries of added ET-1 over the entire physiologic range of plasma concentrations and by the linearity of ET-1 concentrations measured in serially diluted plasma extracts (r = 0.99). No ET-1 was detected when albumin buffer was extracted instead of plasma. Using this method, we found increased ET-1 levels in plasma of three experimental rat models of hypertension: stroke prone spontaneously hypertensive rats (SP-SHR), deoxycorticosterone acetate-salt hypertensive rats, and one kidney-one clip hypertensive rats. In contrast, plasma ET-1 levels of SHR were half those of normotensive Wistar rats. In two kidney-one clip hypertensive rats, plasma ET-1 concentrations were not different from those found in sham-operated control rats. Plasma ET-1 concentrations of 37 healthy men were 0.85 +/- 0.26 fmol/ml (mean +/- SD). CONCLUSIONS: The present assay reliably measures ET-1 levels in rat and human plasma. It allows to discriminate between different forms of hypertension with high or low circulating levels of ET-1.
BACKGROUND:Endothelin-1 is an endothelium-derived potent vasoconstrictor peptide of 21 amino acids. To establish reference values in different models of hypertension and in human subjects an assay for plasma immunoreactive endothelin-1 (ET-1) was optimized. METHODS:ET-1 is extracted by acetone from 1 mL of plasma and subjected to a sensitive enzyme-linked immunosorbent assay. RESULTS: The detection limit for plasma ET-1 is 0.05 fmol/mL. Mean recoveries of the 1, 2, 5, and 10 fmol of ET-1 added to 1 mL of plasma were 66%, 75%, 85%, and 92%, respectively. Within- and between-assay coefficients of variation were < or =12% and < or =10%, respectively. Assay accuracy was demonstrated by consistent recoveries of added ET-1 over the entire physiologic range of plasma concentrations and by the linearity of ET-1 concentrations measured in serially diluted plasma extracts (r = 0.99). No ET-1 was detected when albumin buffer was extracted instead of plasma. Using this method, we found increased ET-1 levels in plasma of three experimental rat models of hypertension: stroke prone spontaneously hypertensiverats (SP-SHR), deoxycorticosterone acetate-salthypertensiverats, and one kidney-one clip hypertensiverats. In contrast, plasma ET-1 levels of SHR were half those of normotensive Wistar rats. In two kidney-one clip hypertensiverats, plasma ET-1 concentrations were not different from those found in sham-operated control rats. Plasma ET-1 concentrations of 37 healthy men were 0.85 +/- 0.26 fmol/ml (mean +/- SD). CONCLUSIONS: The present assay reliably measures ET-1 levels in rat and human plasma. It allows to discriminate between different forms of hypertension with high or low circulating levels of ET-1.
Authors: Philipp Wolf; Alexander Mohr; Georgina Gavins; Victoria Behr; Karin Mörl; Oliver Seitz; Annette G Beck-Sickinger Journal: Chembiochem Date: 2021-10-26 Impact factor: 3.461