Modulation of adenosine transport by insulin in human umbilical artery smooth muscle cells from normal or gestational diabetic pregnancies

• Published in: The Journal of physiology 2001-07, Vol.534 (1), p.243-254
• Main Authors: Aguayo, Claudio, Flores, Carlos, Parodi, Jorge, Rojas, Romina, Mann, Giovanni E., Pearson, Jeremy D., Sobrevia, Luis
• Format: Article
• Language: English
• Subjects: Adenosine - metabolism; Biological Transport - drug effects; Biological Transport - physiology; Cells, Cultured; Cyclic AMP - physiology; Diabetes, Gestational - metabolism; Female; Humans; Insulin - pharmacology; Insulin - physiology; Muscle, Smooth, Vascular - cytology; Muscle, Smooth, Vascular - metabolism; Nitric Oxide - physiology; Original; Pregnancy - metabolism; Reference Values; Umbilical Arteries - cytology; Umbilical Arteries - metabolism
• ISSN: 0022-3751; 1469-7793
• DOI: 10.1111/j.1469-7793.2001.00243.x

Adenosine transport was measured in human cultured umbilical artery smooth muscle cells, isolated from non-diabetic or gestational diabetic pregnancies, under basal conditions and after pretreatment in vitro with insulin. Adenosine transport in non-diabetic smooth muscle cells was significantly increased by insulin (half-maximal stimulation at 0.33 ± 0.02 n m , 8 h) and characterized by a higher maximal rate (V max ) for nitrobenzylthioinosine (NBMPR)-sensitive ( es ) saturable nucleoside transport (17 ± 5 vs. 52 ± 12 pmol (μg protein) −1 min −1 , control vs. insulin, respectively) and maximal binding sites ( B max ) for [ 3 H]NBMPR (0.66 ± 0.07 vs. 1.1 ± 0.1 fmol (μg protein) −1 , control vs. insulin, respectively), with no significant changes in Michaelis-Menten ( K m ) and dissociation ( K d ) constants. In contrast, in smooth muscle cells from diabetic pregnancies, where the values of V max for adenosine transport (59 ± 4 pmol (μg protein) −1 min −1 ) and B max for [ 3 H]NBMPR binding (1.62 ± 0.16 fmol (μg protein) −1 ) were significantly elevated by comparison with non-diabetic cells, insulin treatment (1 n m , 8 h) reduced the V max for adenosine transport and B max for [ 3 H]NBMPR binding to levels detected in non-diabetic cells. In non-diabetic cells, the stimulatory effect of insulin on adenosine transport was mimicked by dibutyryl cGMP (100 n m ) and reduced by inhibitors of phosphatidylinositol 3-kinase (10 n m wortmannin), nitric oxide synthase (100 μ m N G -nitro- l -arginine methyl ester, l -NAME) or protein synthesis (1 μ m cycloheximide), whereas inhibition of adenylyl cyclase (100 μ m SQ-22536) had no effect. Wortmannin or SQ-22536, but not l -NAME or cycloheximide, attenuated the inhibitory action of insulin on the diabetes-induced stimulation of adenosine transport. Protein levels of inducible NO synthase (iNOS) were similar in non-diabetic and diabetic cells, but were increased by insulin (1 n m , 8 h) only in non-diabetic smooth muscle cells. Our results suggest that adenosine transport via the es nucleoside transporter is modulated differentially by insulin in either cell type. Insulin increased adenosine transport in non-diabetic cells via NO and cGMP, but inhibited the diabetes-elevated adenosine transport via activation of adenylyl cyclase, suggesting that the biological actions of adenosine may be altered under conditions of sustained hyperglycaemia in uncontrolled diabetes.