Nucleoside and nucleobase transporters of primary human cardiac microvascular endothelial cells: characterization of a novel nucleobase transporter

• Published in: American Journal of Physiology: Cell Physiology 2007-12, Vol.293 (6), p.H3325-H3332
• Main Authors: Bone, Derek BJ, Hammond, James R
• Format: Article
• Language: English
• ISSN: 0363-6143; 1522-1563

Levels of cardiovascular active metabolites, like adenosine, are regulated by nucleoside transporters of endothelial cells. We characterized the nucleoside and nucleobase transport capabilities of primary human cardiac microvascular endothelial cells (hMVECs). hMVECs accumulated 2-[ super(3)H]chloroadenosine via the nitrobenzylmercaptopurine riboside-sensitive equilibrative nucleoside transporter 1 (ENT1) at a V sub(max) of 3.4 plus or minus 1 pmol. mu l super(-1).s super(-1), with no contribution from the nitrobenzylmercaptopurine riboside-insensitive ENT2. Inhibition of 2-chloroadenosine uptake by ENT1 blockers produced monophasic inhibition curves, which are also compatible with minimal ENT2 expression. The nucleobase [ super(3)H]hypoxanthine was accumulated within hMVECs (K sub(m) = 96 plus or minus 37 mu M; V sub(max) = 1.6 plus or minus 0.3 pmol. mu l super(-1).s super(-1)) despite the lack of a known nucleobase transport system. This novel transporter was dipyridamole-insensitive but could be inhibited by adenine (K sub(i) = 19 plus or minus 7 mu M) and other purine nucleobases, including chemotherapeutic analogs. A variety of other cell types also expressed the nucleobase transporter, including the nucleoside transporter-deficient PK(15) cell line (PK15NTD). Further characterization of [ super(3)H]hypoxanthine uptake in the PK15NTD cells showed no dependence on Na super(+) or H super(+). PK15NTD cells expressing human ENT2 accumulated 4.5-fold more [ super(3)H]hypoxanthine in the presence of the ENT2 inhibitor dipyridamole than did PK15NTD cells or hMVECs, suggesting trapping of ENT2-permeable metabolites. Understanding the nucleoside and nucleobase transporter profiles in the vasculature will allow for further study into their roles in pathophysiological conditions such as hypoxia or ischemia.