Preformed confluent endothelial cell monolayers prevent early platelet deposition on vascular prostheses in baboons

Abstract

We assessed the capacity of preformed confluent endothelial cell (EC) monolayers on small-caliber prosthetic grafts to prevent early platelet deposition in a baboon model. Cultured human umbilical vein ECs were attached to expanded polytetrafluoroethylene (Gore-Tex, 4 mm inner diameter, 3 cm length) precoated with type I collagen and perfused in vitro for 2 hours at 15 ml/min with serum-containing culture medium to achieve cell spreading into confluent monolayers. Cell numbers were quantified by deoxyribonucleic acid assay or isotopic counting of indium 111-labeled ECs. Saturation density for cell attachment was 3.55 ± 0.29 × 105 cells per square centimeter of graft. After 1 hour of in vitro perfusion at 100 ml/min, 92.8% ± 1.8% of cells remained attached and the flow surface was morphologically confluent. When grafts were inserted as extension segments into arteriovenous silicone rubber (Silastic) shunts in baboons, thereby exposing the endothelialized grafts to native flowing blood (100 ml/min) for 1 hour, the EC monolayers remained confluent with 81.05% ± 5.88% of the cells attached. Indium 111—labeled platelet deposition onto grafts was quantified by dynamic scintillation camera imaging. Platelet deposition on 10 endothelialized grafts was markedly reduced (0.16 ± 0.04 × 109 platelets per graft) compared with 10 untreated control grafts (1.84 ± 0.59 × 109 platelets, p < 0.02), eight grafts with early attached unspread ECs (2.38 ± 0.66 × 109 platelets, p < 0.005), and 11 grafts treated with collagen alone (5.93 ± 0.72 × 109 platelets, p < 0.002). Thus, whereas early attached unspread ECs decreased platelet deposition onto collagen-coated grafts (p < 0.05), preformed confluent EC monolayers essentially abolished platelet deposition (p < 0.005 between spread and unspread ECs). We conclude that the use of in vitro preformed confluent EC monolayers may be a useful means for decreasing early thrombus formation on small-caliber vascular grafts.