Nitric oxide, cGMP and cAMP modulate nitrobenzylthioinosine-sensitive adenosine transport in human umbilical artery smooth muscle cells from subjectswith gestational diabetes
C. Aguayo et L. Sobrevia, Nitric oxide, cGMP and cAMP modulate nitrobenzylthioinosine-sensitive adenosine transport in human umbilical artery smooth muscle cells from subjectswith gestational diabetes, EXP PHYSIOL, 85(4), 2000, pp. 399-409
Adenosine transport was characterized in human umbilical artery smooth musc
le cells isolated from nondiabetic and diabetic pregnant subjects. Transpor
t of adenosine was mediated by a Na+-independent transport system inhibited
by nanomolar concentrations of nitrobenzylthioinosine (NBMPR) in both cell
types. Diabetes increased adenosine transport, an effect that was associat
ed with a higher maximal velocity (V-max) for NBMPR-sensitive (es) saturabl
e nucleoside transport (18 +/- 2 vs. 61 +/- 3 pmol (mu g protein)(-1) min(-
1), P < 0.05) and the maximal number of binding sites (B-max) for specific
[H-3]NBMPR binding (74 +/- 4 vs. 156 +/- 10 pmol (mu g protein)(-1), P < 0.
05), with no significant changes in the Michaelis-Menten (K-m) and dissocia
tion (K-d) constants, respectively. Adenosine transport was unaltered by in
hibition of nitric oxide (NO) synthase (with 100 mu M N-G-nitro-L-arginine
methyl ester, L-NAME) or protein synthesis (with 1 mu M cycloheximide), but
was increased by inhibition of adenylyl cyclase activity (with 100 mu M, S
Q-22536) in nondiabetic cells. Diabetes-induced adenosine transport was blo
cked by L-NAME and associated with an increase in L-[H-3]citrulline formati
on from L-[H-3]arginine and intracellular cGMP, but with a decrease in intr
acellular cAMP compared with non-diabetic cells. Expression of inducible NO
synthase (iNOS) was unaltered by diabetes. Dibutyryl cGMP (dbcGMP) increas
ed, but dibutyryl cAMP (dbcAMP) decreased, adenosine transport in non-diabe
tic cells. dbcGMP or the NO donor S-nitrosoacetylpenicillamine (SNAP, 100 m
u M) did not alter the diabetes-elevated adenosine transport. However, acti
vation of adenylyl cyclase with forskolin (1 mu M), directly or after incub
ation of cells with dbcAMP, inhibited adenosine transport in both cell type
s. Our findings provide the first evidence that adenosine transport in huma
n umbilical artery smooth muscle cells is mediated by the NBMPR-sensitive t
ransport system es, and that its activity is upregulated by gestational dia
betes.