Characterization and regulation of adenosine transport in T84 intestinal epithelial cells.
Adenosine release from mucosal sources during inflammation and ischemia activates intestinal epithelial Cl- secretion. Previous data suggest that A2b receptor-mediated Cl- secretory responses may be dampened by epithelial cell nucleoside scavenging. The present study utilizes isotopic flux analysis and nucleoside analog binding assays to directly characterize the nucleoside transport system of cultured T84 human intestinal epithelial cells and to explore whether adenosine transport is regulated by secretory agonists, metabolic inhibition, or phorbol ester. Uptake of adenosine across the apical membrane displayed characteristics of simple diffusion. Kinetic analysis of basolateral uptake revealed a Na(+)-independent, nitrobenzylthioinosine (NBTI)-sensitive facilitated-diffusion system with low affinity but high capacity for adenosine. NBTI binding studies indicated a single population of high-affinity binding sites basolaterally. Neither forskolin, 5'-(N-ethylcarboxamido)-adenosine, nor metabolic inhibition significantly altered adenosine transport. However, phorbol 12-myristate 13-acetate significantly reduced both adenosine transport and the number of specific NBTI binding sites, suggesting that transporter number may be decreased through activation of protein kinase C. This basolateral facilitated adenosine transporter may serve a conventional function in nucleoside salvage and a novel function as a regulator of adenosine-dependent Cl- secretory responses and hence diarrheal disorders.
Mun
E C
EC
Department of Surgery, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts 02215, USA.
Tally
K J
KJ
Matthews
J B
JB
D000241:Adenosine / Q000493:pharmacokinetics*; D019830:Adenosine-5'-(N-ethylcarboxamide) / Q000494:pharmacology; D000345:Affinity Labels / Q000494:pharmacology; D001692:Biological Transport / Q000187:drug effects; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D015536:Down-Regulation; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D007700:Kinetics; D008565:Membrane Proteins / Q000378:metabolism*; D033703:Nucleoside Transport Proteins; D012964:Sodium / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013868:Thioinosine / Q000031:analogs & derivatives / Q000494:pharmacology; D014665:Vasodilator Agents / Q000494:pharmacology
D000345:Affinity Labels; D002352:Carrier Proteins; D008565:Membrane Proteins; D033703:Nucleoside Transport Proteins; D014665:Vasodilator Agents; D005576:Colforsin; D019830:Adenosine-5'-(N-ethylcarboxamide); D013868:Thioinosine; D012964:Sodium; C001789:4-nitrobenzylthioinosine; D000241:Adenosine; D013755:Tetradecanoylphorbol Acetate