Chronic in vitro shear stress stimulates endothelial cell retention on prosthetic vascular grafts and reduces subsequent in vivo neointimal thickness,☆☆

Portions of this work were presented at the Surgical Forum, annual meeting of the American College of Surgeons, Chicago, Ill, Oct 1997.Supported by NIH NRSA fellowship HL09263-02 to A. Dardik, NIH grant DK47023, and American Heart Association grant-in-aid 95015650 to B. Ballermann. Corvita Corporation provided polyurethane grafts as a gift.
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Abstract

Objective: The absence of endothelial cells at the luminal surface of a prosthetic vascular graft potentiates thrombosis and neointimal hyperplasia, which are common causes of graft failure in humans. This study tested the hypothesis that pretreatment with chronic in vitro shear stress enhances subsequent endothelial cell retention on vascular grafts implanted in vivo. Methods: Cultured endothelial cells derived from Fischer 344 rat aorta were seeded onto the luminal surface of 1.5-mm internal diameter polyurethane vascular grafts. The seeded grafts were treated for 3 days with 1 dyne/cm2 shear stress and then for an additional 3 days with 1 or 25 dyne/cm2 shear stress in vitro. The grafts then were implanted as aortic interposition grafts into syngeneic rats in vivo. Grafts that were similarly seeded with endothelial cells but not treated with shear stress and grafts that were not seeded with endothelial cells served as controls. The surgical hemostasis time was monitored. Endothelial cell identity, density, and graft patency rate were evaluated 24 hours after implantation. Endothelial cell identity in vivo was confirmed with cells transduced in vitro with β-galactosidase complementary DNA in a replication-deficient adenoviral vector. Histologic, scanning electron microscopic, and immunohistochemical analyses were performed 1 week and 3 months after implantation to establish cell identity and to measure neointimal thickness. Results: The pretreatment with 25 dyne/cm2—but not with 0 or 1 dyne/cm2—shear stress resulted in the retention of fully confluent endothelial cell monolayers on the grafts 24 hours after implantation in vivo. Retention of seeded endothelial cells was confirmed by the observation that β-galactosidase transduced cells were retained as a monolayer 24 hours after implantation in vivo. In the grafts with adherent endothelial cells that were pretreated with shear stress, immediate graft thrombosis was inhibited and surgical hemostasis time was significantly prolonged. Confluent intimal endothelial cell monolayers also were present 1 week and 3 months after implantation. However, 1 week after implantation, macrophage infiltration was observed beneath the luminal cell monolayer. Three months after the implantation in vivo, subendothelial neointimal cells that contained α–smooth muscle actin were present. The thickness of this neointima averaged 41 ± 12 μm and 60 ± 23 μm in endothelial cell–seeded grafts that were pretreated with 25 dyne/cm2 shear stress and 1 dyne/cm2 shear stress, respectively, and 158 ± 46 μm in grafts that were not seeded with endothelial cells. Conclusion: The effect of chronic shear stress on the enhancement of endothelial cell retention in vitro can be exploited to fully endothelialize synthetic vascular grafts, which reduces immediate in vivo graft thrombosis and subsequent neointimal thickness. (J Vasc Surg 1999;29:157-67.)

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Reprint requests: Barbara J. Ballermann, MD, Johns Hopkins University School of Medicine, 954 Ross Research Bldg, 720 Rutland Ave, Baltimore, MD 21205.

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