title: Adenosine transport in cultured human umbilical vein endothelial cells is reduced in diabetes.
abstract: Adenosine transport in cultured human umbilical vein endothelial cells (HUVEC) was characterized and shown to be mediated by a single facilitated diffusion mechanism. Initial rates of adenosine influx at 22 degrees C were saturable [apparent Michaelis constant, 69 +/- 10 microM; maximum velocity (Vmax), 600 +/- 70 pmol.10(6) cells-1.s-1] and inhibited by nitrobenzylthioinosine (NBMPR). Formycin B had an unusually high affinity [inhibitory constant (Ki), 18 +/- 4.3 microM], whereas inosine had a low affinity (Ki, 440 +/- 68 microM) and nucleobases were without effect on adenosine influx. The number of transporters (1.2 x 10(6) sites/cell) was estimated by NBMPR equilibrium binding (apparent dissociation constant, 0.11 +/- 0.01 nM; maximum binding, 2.0 +/- 0.15 pmol/10(6) cells). In addition, we compared these endothelial cells with those obtained from cords from pregnancies complicated by diabetes (HUVEC-D), since embriopathy may occur in these conditions. HUVEC-D exhibited a 2.3-fold reduction in both the Vmax for adenosine influx and the maximum number of NBMPR binding sites (260 +/- 40 pmol.10(6) cells-1.s-1 and 0.86 +/- 0.08 pmol/10(6) cells, respectively). However, the turnover number for each nucleoside transporter in normal and diabetic HUVEC was similar (approximately 300 adenosine molecules/s). Adenosine metabolism at 10 microM in HUVEC-D was modified compared with normal cells. Intracellular phosphorylation (> 90%) was the predominant pathway in normal HUVEC, whereas in HUVEC-D, substantial levels of adenine and adenosine were detected. The present results demonstrate therefore the downregulation of the NBMPR-sensitive nucleoside transporter and changes in adenosine metabolism in HUVEC from diabetic pregnancies.
authors.affiliation: Department of Physiology and Biophysics, University of Chile, Santiago.
authors.forename: L
authors.initials: L
authors.lastname: Sobrevía
authors.forename: S M
authors.initials: SM
authors.lastname: Jarvis
authors.forename: D L
authors.initials: DL
authors.lastname: Yudilevich
mesh_terms: D000241:Adenosine / Q000378:metabolism*; D001692:Biological Transport; D002455:Cell Division; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism* / Q000473:pathology; D005260:Female; D006801:Humans; D011247:Pregnancy; D011254:Pregnancy in Diabetics / Q000378:metabolism* / Q000473:pathology; D012016:Reference Values; D013868:Thioinosine / Q000031:analogs & derivatives / Q000378:metabolism; D013997:Time Factors; D014471:Umbilical Veins / Q000378:metabolism* / Q000473:pathology
chemical_list: D013868:Thioinosine; C001789:4-nitrobenzylthioinosine; D000241:Adenosine