Fig. 2.

Adenosine-induced suppression of endothelial inflammation requires adenosine uptake. a Western blot detection and densitometric quantification of adhesion molecule expression in HUVECs. HUVECs, pretreated for 30 min with both 5 µM ZM 241385 and 5 µM MRS 1754 were incubated with 100 µM adenosine for 30 min and then stimulated with TNF-α at 10 ng/ml for 4 h (n = 4). b Western blot detection and densitometric quantification of adhesion molecule expression in HUVECs. HUVECs, transiently transfected with control or A_2AR siRNA, were pretreated for 30 min with 100 µM adenosine and then stimulated with 10 ng/ml TNF-α for 4 h (n = 4). c Western blot detection and densitometric quantification of adhesion molecule expression in HUVECs. HUVECs, transiently transfected with control or A_2BR siRNA, were pretreated for 30 min with 100 µM adenosine and then stimulated with 10 ng/ml TNF-α for 4 h (n = 4). d Quantification of intracellular adenosine by HPLC in adenosine (100 µM for 1 h)-treated HUVECs and in adenosine (100 µM for 1 h)-treated HUVECs preincubated with NBMPR at 10 µM for 30 min (n = 4). e Real-time-PCR (RT-PCR) analysis of mRNA levels of adhesion molecules in HUVECs. HUVECs, pretreated for 30 min with 10 µM NBMPR, were incubated with 100 µM adenosine for 30 min and then stimulated with TNF-α at 10 ng/ml for 2 h (n = 3). f Western blot detection and densitometric quantification of adhesion molecule expression in HUVECs. HUVECs, pretreated for 30 min with 10 µM NBMPR, were incubated with 100 µM adenosine for 30 min and then stimulated with TNF-α at 10 ng/ml for 4 h (n = 4). All images are representative. For all bar graphs, data are the mean ± SEM, *P < 0.05 and **P < 0.01 (one-way ANOVA with Tukey’s post hoc test)