HYPOXANTHINE ENTERS HUMAN VASCULAR ENDOTHELIAL-CELLS (ECV-304) VIA THE NITROBENZYLTHIOINOSINE-INSENSITIVE EQUILIBRATIVE NUCLEOSIDE TRANSPORTER

The transport properties of the nucleobase hypoxanthine were examined in the human umbilical vein endothelial cell line ECV 304. Initial rat es of hypoxanthine influx were independent of extracellular cations: r eplacement of Na+ with Li+, Rb+, N-methyl-D-glucamine or choline had n o significant effect on hypoxanthine uptake by ECV 304 cells. Kinetic analysis demonstrated the presence of a single saturable system for th e transport of hypoxanthine in ECV 304 cells with an apparent K-m of 3 20+/-10 mu M and a V-max of 5.6+/-0.9 pmol/10(6) cells per s. Hypoxant hine uptake was inhibited by the nucleosides adenosine, uridine and th ymidine (apparent K-i 41+/-6, 240+/-27 and 59+/-8 mu M respectively) a nd the nucleoside transport inhibitors nitrobenzylthioinosine (NBMPR), dilazep and dipyridamole (apparent K-i 2.5+/-0.3, 11+/-3 and 0.16+/-0 .006 mu M respectively), whereas the nucleobases adenine, guanine and thymine had little effect (50% inhibition at >1 mM). ECV 304 cells wer e also shown to transport adenosine via both the NBMPR-sensitive and - insensitive nucleoside carriers. Hypoxanthine specifically inhibited a denosine transport via the NBMPR-insensitive system in a competitive m anner (apparent K-i 290+/-14 mu M). These results indicate that hypoxa nthine entry into ECV 304 endothelial cells is mediated by the NBMPR-i nsensitive nucleoside carrier present in these cells.