Adenosine transport in cultured human umbilical vein endothelial cells is reduced in diabetes.
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.
Sobrevía
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Department of Physiology and Biophysics, University of Chile, Santiago.
Jarvis
S M
SM
Yudilevich
D L
DL
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
D013868:Thioinosine; C001789:4-nitrobenzylthioinosine; D000241:Adenosine