Abstract 367: Eph-B4 Mediates Arteriovenous Fistula Maturation via Akt-1

Abstract

Objectives: Although the arteriovenous fistula (AVF) is the gold standard for dialysis access, low rates of fistula maturation prevent optimal fistula use. We have previously shown that stimulation of Eph-B4 in vein grafts prevents wall thickening during venous remodeling. Since Akt-1 is a downstream mediator of Eph-B4 signaling in endothelial cells, we hypothesized that Akt-1 mediates Eph-B4 function during AVF maturation. Methods: The infrarenal aorto-caval AVF model was created in wild type C57BL/6 mice and Akt-1 knockout mice as previously described. To stimulate Eph-B4 activity, mice were treated with either EphrinB2/Fc (20 μg IP) or a control vehicle at 48 hr intervals over a 21 day study period. Fistula maturation was monitored with weekly ultrasound measurements; AVF were harvested at day 21 and infrarenal IVC wall thickness was measured using computerized morphometry. Eph-B4, phospho-tyrosine, and Akt-1 immunoreactivity were measured with immunofluorescence. Results: In WT mice, Eph-B4 expression increased 4.6-fold after AVF compared to sham operation (p=0.007, t-test). IF revealed unmerged Eph-B4 and p-tyr signal at days 7 and 21. Stimulation of Eph-B4 in WT mice increased Eph-B4 and p-tyr merged signal 6-fold (p=0.005, ANOVA) at 21 days and resulted in a 49% decrease in phospho-Akt-1 signal (p=0.0013, t-test). Eph-B4 stimulation in WT mice was associated with reduced fistula diameter (45% vs. 98.6%; p<0.05, ANOVA) and reduced fistula wall thickness (8.9μm vs. 20.0μm; p=0.0261, t-test). However, in Akt-1 KO mice, Eph-B4 stimulation resulted in no change in fistula diameter (38% vs 22%; p=0.09, ANOVA) and no change in fistula wall thickness (15.3 vs 17μm; p=0.46, t-test). Conclusion: There is increased Eph-B4 expression in an AVF, although the Eph-B4 is not phosphorylated. Stimulation of Eph-B4 increases the percentage of phosphorylated Eph-B4 resulting in reduced Akt-1 phosphorylation with consequent reduced fistula diameter and wall thickness. Akt-1 knockout abolishes the effects of Eph-B4 stimulation on wall thickness and diameter. These findings suggest Eph-B4 is an inhibitor of Akt-1 activity and that Akt-1 mediates the effect of Eph-B4 on venous adaptation to the arterial environment.