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  • Title: Identification and quantification of diadenosine polyphosphate concentrations in human plasma.
    Author: Jankowski J, Jankowski V, Laufer U, van der Giet M, Henning L, Tepel M, Zidek W, Schlüter H.
    Journal: Arterioscler Thromb Vasc Biol; 2003 Jul 01; 23(7):1231-8. PubMed ID: 12738682.
    Abstract:
    OBJECTIVE: Diadenosine polyphosphates have been demonstrated to be involved in the control of vascular tone as well as the growth of vascular smooth muscle cells and hence, possibly, in atherogenesis. In this study we investigated the question of whether diadenosine polyphosphates are present in human plasma and whether a potential source can be identified that may release diadenosine polyphosphates into the circulation. METHODS AND RESULTS: Plasma diadenosine polyphosphates (ApnA with n=3 to 6) were purified to homogeneity by affinity-, anion exchange-, and reversed phase-chromatography from deproteinized human plasma. Analysis of the homogeneous fractions with matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) revealed molecular masses ([M+H]+) of 757, 837, 917, and 997 d. Comparison of the postsource decay matrix-assisted laser desorption/ionization mass spectrometry mass spectra of these fractions with those of authentic diadenosine polyphosphates revealed that these isolated substances were identical to Ap3A, Ap4A, Ap5A, and Ap6A. Enzymatic analysis showed an interconnection of the phosphate groups with the adenosines in the 5'-positions of the ribose moieties. The mean total plasma diadenosine polyphosphate concentrations (micromol/L; mean +/- SEM) in cubital veins of normotensive subjects amounted to 0.89+/-0.59 for Ap3A, 0.72+/-0.72 for Ap4A, 0.33+/-0.24 for Ap5A, and 0.18+/-0.18 for Ap6A. Cubital venous plasma diadenosine polyphosphate concentrations from normotensives did not differ significantly from those in the hypertensive patients studied. There was no significant difference between arterial and venous diadenosine polyphosphate plasma concentrations in 5 hemodialysis patients, making a significant degradation by capillary endothelial cells unlikely. Free plasma diadenosine polyphosphate concentrations are considerably lower than total plasma concentrations because approximately 95% of the plasma diadenosine polyphosphates were bound to proteins. There were no significant differences in the diadenosine polyphosphate plasma concentrations depending on the method of blood sampling and anticoagulation, suggesting that platelet aggregation does not artificially contribute to plasma diadenosine polyphosphate levels in significant amounts. The ApnA (with n=3 to 6) total plasma concentrations in adrenal veins were significantly higher than the plasma concentrations in both infrarenal and suprarenal vena cava: adrenal veins: Ap3A, 4.05+/-1.63; Ap4A, 6.18+/-2.08; Ap5A, 0.53+/-0.28; Ap6A, 0.59+/-0.31; infrarenal vena cava: Ap3A, 1.25+/-0.66; Ap4A, 0.91+/-0.54; Ap5A, 0.25+/-0.12; Ap6A, 0.11+/-0.06; suprarenal vena cava: Ap3A, 1.40+/-0.91; Ap4A, 1.84+/-1.20; Ap5A, 0.33+/-0.13; Ap6A, 0.11+/-0.07 (micromol/L; mean +/- SEM; each P<0.05 (concentration of adrenal veins versus infrarenal or suprarenal veins, respectively). CONCLUSIONS: The presence of diadenosine polyphosphates in physiologically relevant concentrations in human plasma was demonstrated. Because in adrenal venous plasma significantly higher diadenosine polyphosphate concentrations were measured than in plasma from the infrarenal and suprarenal vena cava, it can be assumed that, beside platelets, the adrenal medulla may be a source of plasma diadenosine polyphosphates in humans.
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