Date of Award

January 2014

Document Type

Thesis

Degree Name

Medical Doctor (MD)

Department

Medicine

First Advisor

Alan Dardik

Subject Area(s)

Medicine, Surgery

Abstract

Despite the rise in percutaneous interventions, bypass surgery is an essential therapy for patients with advanced coronary and peripheral artery disease. Nearly half of the saphenous vein grafts used in these operations are eventually complicated by pathologic remodeling known as neointimal hyperplasia (NIH). Recent work in animal models has shown that the development of NIH can be reduced by stimulation of the receptor tryrosine kinase Eph-B4 by systemic injections of its ligand ephrin-b2. The purpose of this study was to determine whether the reduction of intimal hyperplasia observed with Eph-B4 stimulation in animal models translated into human tissues. Specifically we sought to determine whether the development of intimal hyperplasia in human saphenous vein (HSV) tissue could be reduced with ephrin-b2/fc treatments. We also investigated whether Eph-B4 stimulation in human endothelial cells caused similar downstream signaling changes observed in animal cells. Finally we wanted to assess endothelial Eph-B4 stimulation was achievable with adventitial delivery of ephrin-B2/fc. Excess saphenous vein from patients undergoing coronary artery bypass surgery was taken from the OR and put in organ culture for 14 days with or without treatment with ephrin-b2/fc. At the end of 14 days histologic measurements of the intima:media (I:M) were used to track the development of (NIH). The mean baseline I:M ratio was 0.456 ± 0.097 (n=19). Over 14 days in organ culture this ratio increased to 0.726 ± 0.142 (n=19) in untreated veins and 0.630 ± 0.132 (n=19) in treated veins which was significantly different (p=.017). Using quantitative reverse transcription polymerase chain reaction (qPCR) analysis gene expression changes in organ culture were measured. Eph-B4 and ephrin-b2 decreased while osteopontin expression increased in both treated and untreated groups. Using a human endothelial cell line, we observed increases in phosphorylated AKT and caveolin-1 with ephrin-b2/fc treatment. Finally using an arterial flow model bio-reactor we demonstrated that endothelial eph-B4 phosphorylation was achievable with adventitial application of ephrin-B2/fc in a pluronic hydrogel after 24 hours in arterial flow. In conclusion we show that Eph-B4 stimulation reduces the NIH development in vitro in HSV organ culture. Eph-B4 stimulation activates similar signaling cascades in human endothelial cells as in mice. Finally that local delivery of eprhin-b2/fc is possible through a pluronic hydrogel applied to the adventitia of a vein. Combined these results continue to point to Eph-B4 stimulation as a promising target for a therapy to reduce NIH in bypass recipients, however, longer term studies in large animals are necessary.

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