Nucleoside/nucleobase transport and metabolism by microvascular endothelial cells isolated from ENT1^-/- mice

Nucleoside and nucleobase uptake is integral to mammalian cell function, and its disruption has significant effects on the cardiovasculature. The predominant transporters in this regard are the equilibrative nucleoside transporter subtypes 1 (ENT1) and 2 (ENT2). To examine the role of ENT1 in more detail, we have assessed the mechanisms by which microvascular endothelial cells (MVECs) from ENT1^-/- mice transport and metabolize nucleosides and nucleobases. Wild-type murine MVECs express mainly the ENT1 subtype with only trace levels of ENT2. These cells also have a Na⁺-independent equilibrative nucleobase transport mechanism for hypoxanthine (ENBT1). In the ENT1^-/- cells, there is no change in ENT2 or ENBT1, resulting in a very low level of nucleoside uptake in these cells, but a high capacity for nucleobase accumulation. Whereas there were no significant changes in nucleoside transporter subtype expression, there was a dramatic increase in adenosine deaminase and adenosine A_2a receptors (both transcript and protein) in the ENT1^-/- tissues compared with WT. These changes in adenosine deaminase and A_2a receptors likely reflect adaptive cellular mechanisms in response to reduced adenosine flux across the membranes of ENT1^-/- cells. Our study also revealed that mouse MVECs have a nucleoside/nucleobase transport profile that is more similar to human MVECs than to rat MVECs. Thus mouse MVECs from transgenic animals may prove to be a useful preclinical model for studies of the effects of purine metabolite modifiers on vascular function.