Insulin Requires A Adenosine Receptors to Modulate the L-arginine/nitric Oxide Signalling in the Human Fetoplacental Vascular Endothelium from Late-onset Preeclampsia

Late-onset preeclampsia (LOPE) associates with reduced umbilical vein reactivity and endothelial nitric oxide synthase (eNOS) activity but increased human cationic amino acid (hCAT-1)-mediated L-arginine transport involving A adenosine receptor in the fetoplacental unit. This study addresses the A adenosine receptor (AAR)-mediated response to insulin in the fetoplacental vasculature from LOPE. Umbilical veins and HUVECs were obtained from women with normal (n = 37) or LOPE (n = 35) pregnancies. Umbilical vein rings reactivity to insulin was assayed in the absence or presence of adenosine and MRS-1754 (AAR antagonist) in a wire myograph. HUVECs were exposed to insulin, MRS-1754, BAY60-6583 (AAR agonist), NECA (general adenosine receptors agonist) or N-nitro-L-arginine methyl ester (NOS inhibitor). AAR, hCAT-1, total and phosphorylated eNOS, Akt and p44/42 protein abundance were determined by Western blotting. Insulin receptors A (IR-A) and B (IR-B), eNOS and hCAT-1 mRNA were determined by qPCR. Firefly/Renilla luciferase assay was used to determine -1606 bp SLC7A1 (hCAT-1) promoter activity. L-Citrulline content was measured by HPLC, L-[H]citrulline formation from L-[H]arginine by the Citrulline assay, and intracellular cGMP by radioimmunoassay. LOPE-reduced dilation of vein rings to insulin was restored by MRS-1754. HUVECs from LOPE showed higher AAR, hCAT-1, and IR-A expression, Akt and p44/42 activation, and lower NOS activity. MRS-1754 reversed the LOPE effect on AAR, hCAT-1, Akt, and eNOS inhibitory phosphorylation. Insulin reversed the LOPE effect on AAR, IR-A and eNOS, but increased hCAT-1-mediated transport. Thus, LOPE alters endothelial function, causing an imbalance in the L-arginine/NO signalling pathway to reduce the umbilical vein dilation to insulin requiring AAR activation in HUVECs.