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Nucleoside transporter subtype expression and function in rat skeletal muscle microvascular endothelial cells.

ABSTRACT: 1. Microvascular endothelial cells (MVECs) form a barrier between circulating metabolites, such as adenosine, and the surrounding tissue. We hypothesize that MVECs have a high capacity for the accumulation of nucleosides, such that inhibition of the endothelial nucleoside transporters (NT) would profoundly affect the actions of adenosine in the microvasculature. 2. We assessed the binding of [(3)H]nitrobenzylmercaptopurine riboside (NBMPR), a specific probe for the inhibitor-sensitive subtype of equilibrative NT (es), and the uptake of [(3)H]formycin B (FB), by MVECs isolated from rat skeletal muscle. The cellular expression of equilibrative (ENT1, ENT2, ENT3) and concentrative (CNT1, CNT2, CNT3) NT subtypes was also determined using both qualitative and quantitative polymerase chain reaction techniques. 3. In the absence of Na(+), MVECs accumulated [(3)H]FB with a V(max) of 21+/-1 pmol microl(-1) s(-1). This uptake was mediated equally by es (K(m) 260+/-70 microm) and ei (equilibrative inhibitor-insensitive; K(m) 130+/-20 microm) NTs. 4. A minor component of Na(+)-dependent cif (concentrative inhibitor-insensitive FB transporter)/CNT2-mediated [(3)H]FB uptake (V(i) 0.008+/-0.005 pmol microl(-1) s(-1) at 10 microm) was also observed at room temperature upon inhibition of ENTs with dipyridamole (2,6-bis(diethanolamino)-4,8-dipiperidinopyrimido-[5,4-d]pyrimidine)/NBMPR. 5. MVECs had 122,000 high-affinity (K(d) 0.10 nm) [(3)H]NBMPR binding sites (representing es transporters) per cell. A lower-affinity [(3)H]NBMPR binding component (K(d) 4.8 nm) was also observed that may be related to intracellular es-like proteins. 6. Rat skeletal muscle MVECs express es/ENT1, ei/ENT2, and cif/CNT2 transporters with characteristics typical of rat tissues. This primary cell culture model will enable future studies on factors influencing NT subtype expression, and the consequent effect on adenosine bioactivity, in the microvasculature.

SUBMITTER: Archer RG

PROVIDER: S-EPMC1575281 | biostudies-literature | 2004 Sep

REPOSITORIES: biostudies-literature

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Nucleoside transporter subtype expression and function in rat skeletal muscle microvascular endothelial cells.

Archer Richard G E RG Pitelka Václav V Hammond James R JR

British journal of pharmacology 20040802 1

1. Microvascular endothelial cells (MVECs) form a barrier between circulating metabolites, such as adenosine, and the surrounding tissue. We hypothesize that MVECs have a high capacity for the accumulation of nucleosides, such that inhibition of the endothelial nucleoside transporters (NT) would profoundly affect the actions of adenosine in the microvasculature. 2. We assessed the binding of [(3)H]nitrobenzylmercaptopurine riboside (NBMPR), a specific probe for the inhibitor-sensitive subtype of ...[more]

PMID: 15289294

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Project description:Key pointsMultiple clinical studies report that acute hyperglycaemia (induced by mixed meal or oral glucose) decreases arterial vascular function in healthy humans. Feeding, however, impacts autonomic output, blood pressure, and insulin and incretin secretion, which may themselves alter vascular function. No prior studies have examined the effect of acute hyperglycaemia on both macro- and microvascular function while controlling plasma insulin concentrations. Macrovascular and microvascular functional responses to euglycaemia and hyperglycaemia were compared. Octreotide was infused throughout both protocols to prevent endogenous insulin release. Acute hyperglycaemia (induced by intravenous glucose) enhanced brachial artery flow-mediated dilatation, increased skeletal muscle microvascular blood volume and flow, and expanded cardiac muscle microvascular blood volume. Compared to other published findings, the results suggest that vascular responses to acute hyperglycaemia differ based on the study population (i.e. normal weight vs. overweight/obese) and/or glucose delivery method (i.e. intravenous vs. oral glucose).AbstractHigh glucose concentrations acutely provoke endothelial cell oxidative stress and are suggested to trigger diabetes-related macro- and microvascular injury in humans. Multiple clinical studies report that acute hyperglycaemia (induced by mixed meal or oral glucose) decreases arterial vascular function in healthy humans. Feeding, however, impacts autonomic output, blood pressure, and insulin and incretin secretion, which may each independently alter vascular function and obscure the effect of acute hyperglycaemia per se. Surprisingly, no studies have examined the acute effects of intravenous glucose-induced hyperglycaemia on both macro- and microvascular function while controlling plasma insulin concentrations. In this randomized study of healthy young adults, we compared macrovascular (i.e. brachial artery flow-mediated dilatation, carotid-femoral pulse wave velocity and post-ischaemic brachial artery flow velocity) and microvascular (heart and skeletal muscle perfusion by contrast-enhanced ultrasound) functional responses to euglycaemia and hyperglycaemia. Octreotide was infused throughout both protocols to prevent endogenous insulin release. Acute intravenous glucose-induced hyperglycaemia enhanced brachial artery flow-mediated dilatation (P = 0.004), increased skeletal muscle microvascular blood volume and flow (P = 0.001), and expanded cardiac muscle microvascular blood volume (P = 0.014). No measure of vascular function changed during octreotide-maintained euglycaemia. Our findings suggest that unlike meal-provoked acute hyperglycaemia, 4 h of intravenous glucose-induced hyperglycaemia enhances brachial artery flow-mediated dilatation, provokes cardiac and skeletal muscle microvascular function, and does not impair aortic stiffness. Previous findings of acute large artery vascular dysfunction during oral glucose or mixed meal ingestion may be due to differences in study populations and meal-induced humoral or neural factors beyond hyperglycaemia per se. (ClinicalTrials.gov number NCT03520569.).

Dataset Information

Nucleoside transporter subtype expression and function in rat skeletal muscle microvascular endothelial cells.

Publications

Nucleoside transporter subtype expression and function in rat skeletal muscle microvascular endothelial cells.

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