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Digitalis-like factors in plasma of healthy human adults

1986; Linguagem: Inglês

10.1007/978-3-662-11292-2_21

Sudesh Vasdev, L. Longerich, E. Johnson, David A. Brent, M.H. Gault,

Mass Spectrometry Techniques and Applications

Digitalis-like factors (DLF) are defined as endogenous compounds which react positively in digoxin radioimmunoassays (RIA) in the absence of digoxin. Plasma from 11 healthy subjects was assayed for DLF and dehydroepiandrosterone sulfate (DHEA-S). DLF were extracted on C-18 Bond Elut columns with methanol and measured by the New England Nuclear (NEN) digoxin RIA and DHEA-S by RIA. Plasma values (means ± SD) were for DLF 44 ± 6 pmol/L digoxin equivalents, and for DHEA-S 11 ± 3 µmol/L. We studied DLF (NEN) and DHEA-S for the presence of 3 digitalis-like properties. Both plasma DLF, and DHEA-S (25–1600 µmol/L) displaced 125I-digoxin from antibody to digoxin, inhibited hog brain Na,K-ATPase, displaced (3H)-ouabain from ATPase in a concentration dependent manner and gave curves parallel to those found with digoxin. Increasing concentrations of DHEA-S were added to plasma and assayed for DLF by NEN and Kallestad digoxin RIAs and standard curves of DHEA-S vs. DLF concentration plotted. Using these curves to calculate the DLF equivalents for each plasma DHEA-S concentration, we found that 62–100% of the plasma DLF by NEN could be accounted for by the DHEA-S in the 11 subjects and similar proportions using Kallestad reagents. A plasma extract from one subject was fractionated by HPLC and showed several DLF peaks when the 20 fractions were assayed for digoxin (NEN). The largest peak had the same retention time as the DHEA-S standard and when the fractions under this peak were assayed for DHEA-S, it was found that DHEA-S could account for 80% of the DLF. FAB mass spectrometry and accurate mass measurement confirmed the presence of DHEA-S in the fraction forming the major DLF peak. DHEA-S has an apparent weak cross-reactivity (0.00024%) with NEN antidigoxin antibody and has other digitalis like properties only in high concentration. Yet its relatively high concentration in plasma appears to make it an important component of the total plasma digitalis-like factors in healthy subjects. Endogenous digitalis-like factors (DLF), measured by radioimmunoassay (RIA) of digoxin, have been reported in plasma of neonates (21, 22), pregnant women (6), patients with hepatic (19) and/or renal failure (11, 14) and in healthy and hypertensive subjects (1, 12). These digitalis-like factors are reported to have one or more of the following characteristics: interaction with antidigoxin antibody, inhibition of Na,K-ATPase, displacement of labelled ouabain from Na,K-ATPase and natriuretic activity (1, 3, 8, 11, 12, 17, 23, 30). DLF have been suggested to be steroid, lipid, or peptide (3, 8, 11, 17, 23, 30), but the exact chemical nature of these substances in healthy adults has not been established. We report that dehydroepiandrosterone sulfate (DHEAS), an important circulating androgen, forms a major portion of the DLF measured by digoxin radioimmunoassay (RIA) in plasma of healthy adults, inhibits Na,K-ATPase and displaces [3H]-ouabain from Na,K-ATPase.