Insulin restores glucose inhibition of adenosine transport by increasing the expression and activity of the equilibrative nucleoside transporter 2 in human umbilical vein endothelium

Abstract

L-Arginine transport and nitric oxide (NO) synthesis (L-arginine/NO pathway) are stimulated by insulin, adenosine or elevated extracellular D-glucose in human umbilical vein endothelial cells (HUVEC). Adenosine uptake via the human equilibrative nucleoside transporters 1 (hENT1) and 2 (hENT2) has been proposed as a mechanism regulating adenosine plasma concentration, and therefore its vascular effects in human umbilical veins. Thus, altered expression and/or activity of hENT1 or hENT2 could lead to abnormal physiological plasma adenosine level. We have characterized insulin effect on adenosine transport in HUVEC cultured in normal (5 mM) or high (25 mM) D-glucose. Insulin (1 nM) increased overall adenosine transport associated with higher hENT2-, but lower hENT1-mediated transport in normal D-glucose. Insulin increased hENT2 protein abundance in normal or high D-glucose, but reduced hENT1 protein abundance in normal D-glucose. Insulin did not alter the reduced hENT1 protein abundance, but blocked the reduced hENT1 and hENT2 mRNA expression induced by high D-glucose. Insulin effect on hENT1 mRNA expression in normal D-glucose was blocked by N G -nitro-L-arginine methyl ester (L-NAME, NO synthase inhibitor) and mimicked by S-nitroso-Nacetyl-L,D-penicillamine (SNAP, NO donor). L-NAME did not block insulin effect on hENT2 expression. In conclusion, insulin stimulation of overall adenosine transport results from increased hENT2 expression and activity via a NO-independent mechanism. These findings could be important in hyperglycemia-associated pathological pregnancies, such as gestational diabetes, where plasma adenosine removal by the endothelium is reduced, a condition that could alter the blood flow from the placenta to the fetus affecting fetus growth and development.

Adenosine mediates transforming growth factor‐beta 1 release in kidney glomeruli of diabetic rats

“…The expression and activity of ENTs is also altered by the diabetic syndrome, glucose and insulin [21,26]. Experiments performed in rats revealed that on day 10 following streptozotocin administration, the mRNA level of rENT1 was slightly (10%) lowered whilst the level of rENT2 mRNA was reduced by 40% in total kidney [27].…”

Treatment with insulin inhibits poly(ADP-ribose)polymerase activation in a rat model of endotoxemia

“…In HUVECs insulin stimulates adenosine and L-arginine transport and nitric oxide (NO) synthesis (Munoz et al, 2006). Likewise, recent data indicate that PARP can be regulated by alternative pathways that do not involve free radical generation or DNA damage .…”

Adenosine signalling in diabetes mellitus—pathophysiology and therapeutic considerations

“…94 In the past few years, increasing attention has focused on the link between increased levels of adenosine and the onset of endothelial dysfunction in human umbilical veins during gestational diabetes mellitus. [95][96][97][98][99][100] Extracellular levels of adenosine in the culture media of primary cultures of endothelial cells of umbilical veins isolated from women with gestational diabetes mellitus are higher than those isolated from women with normal pregnancies. 95 Increased levels of adenosine upregulate expression of endothelial nitric oxide synthase via A 2A adenosine receptors, which leads to increased nitric oxide synthesis, a potential cause of vascular dysfunction in gestational diabetes mellitus.…”

“…95 Mechanistic studies investigating the cause of increased adenosine levels have shown that the expression and activity of ENT1, but not ENT2, is reduced in human umbilical vein endothelial cells by elevated levels of glucose, 97 which are offset by treatment with insulin. 98,101 Leukocyte expression of ADORA2B (encoding the A 2B adenosine receptor) is higher in women with gestational diabetes mellitus than in pregnant women with normal glucose tolerance. Furthermore, increased expression of ADORA2B positively correlated with altered expression of 19 genes involved in several aspects of insulin action, glucose and lipid metabolism, oxidative stress and inflammation, which indicates that ADORA2B is part of a complex gene network associated with gestational diabetes mellitus 102 and that leukocyte A 2B adenosine receptors could serve as potential biomarkers for gestational diabetes mellitus.…”