1. Equilibrative nucleoside transporter 2 is expressed in human umbilical vein endothelium, but is not involved in the inhibition of adenosine transport induced by hyperglycaemia.
- Author
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Aguayo C, Casado J, González M, Pearson JD, Martín RS, Casanello P, Pastor-Anglada M, and Sobrevia L
- Subjects
- Cells, Cultured, Dose-Response Relationship, Drug, Down-Regulation, Drug Combinations, Endothelium, Vascular cytology, Endothelium, Vascular drug effects, Equilibrative Nucleoside Transporter 1 genetics, Equilibrative-Nucleoside Transporter 2 genetics, Glucose pharmacology, Humans, Hypoxanthine pharmacology, Nucleoside Transport Proteins metabolism, RNA, Messenger metabolism, Reverse Transcriptase Polymerase Chain Reaction, Thioinosine analogs & derivatives, Thioinosine pharmacology, Umbilical Veins, Adenosine metabolism, Endothelium, Vascular metabolism, Equilibrative Nucleoside Transporter 1 metabolism, Equilibrative-Nucleoside Transporter 2 metabolism, Hypoglycemia metabolism
- Abstract
Human equilibrative, Na(+)-independent nucleoside transport is mediated by membrane proteins sensitive (system es, hENT1) or insensitive (system ei, hENT2) to nitrobenzylthioinosine (NBMPR). Gestational diabetes and elevated extracellular concentrations of D-glucose reduce adenosine transport in human umbilical vein endothelium (HUVEC). We studied hENT2 and hENT1 expression in HUVEC, and the effect of D-glucose on their activity and expression in HUVEC preincubated with 25 mM D-glucose (24 h). hENT2 and hENT1 mRNA were quantified by real-time reverse transcription polymerase chain reaction, and their proteins were detected by Western blotting. hENT2 and hENT1 proteins are co-expressed in HUVEC and are located at the plasma membrane, however, hENT2 was mainly cytoplasmatic and perinuclear in location. D-Glucose reduced hENT1 and hENT2 mRNA expression, but only hENT1 protein abundance at the plasma membrane. Adenosine transport was inhibited by D-glucose and NMBPR (1 microM) in intact cells and membrane vesicles. Hypoxanthine inhibited adenosine transport in the absence or in the presence of 1 microM NBMPR. D-Glucose reduced NBMPR maximal binding in intact cells, membrane vesicles, and plasma membrane fractions. In conclusion, the present study demonstrates that hENT2 and hENT1 are co-expressed in HUVEC, and even when adenosine transport is also mediated by hENT2, the hENT2-mediated transport activity is not involved in the d-glucose-induced down-regulation of total adenosine transport.
- Published
- 2005
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