The existence of a confluently covering endothelium that is free of any thrombotic appositions can be proved 30 days after clinical implantation of an in vitro endothelialized expanded polytetrafluoroethylene graft. The recipient of the mesosystemic H-graft was a 69-year-old man who had a thrombosed portal vein following pancreatitis. Autologous endothelial cells were obtained from the external jugular vein under local anesthesia, applying the in situ cannulation technique. After low-density plating, first-passage mass cultures of 1.22 × 106 endothelial cells were obtained 14 days after vein excision. After precoating was accomplished with fibrinolytically inhibited fibrin glue, a 10 mm expanded polytetrafluoroethylene graft was confluently lined with the autologous endothelial cells at a seeding density of 1.2 × 105 cells/cm2. After a maturation period of an additional 9 days and the microbiologic exclusion of a possible infection, an 11 cm graft segment was implanted between the superior mesenteric vein and the inferior vena cava. In spite of a patent shunt the patient had a repeat bleeding episode, needed parenteral nutrition, and died of sepsis on day 30. Immediately after the graft had been taken out, specimens were processed by scanning electron microscopy and light microscopy for the immunohistochemical proof of the endothelial nature of the surface-covering cell layer. The entire graft surface displayed a confluent cell lining that was free of any thrombotic appositions. A strongly positive stain result for both factor VIII – related antigen and the fixation-resistant CD34 molecule identified these cells as endothelial. No α-actin – positive cells could be detected. The underlying protein matrix was well preserved and unaltered in thickness and appearance, compared with preimplantation samples. None of the specimens showed any evidence of infection. This human demonstration of an intact endothelium on a patent venous prosthesis further establishes in vitro lining as a method that actually creates a persistent and functioning endothelium on a synthetic graft surface. (J VASC SURG 1994;19:549-54.)

Purpose: The creation of an endothelial coverage on prosthetic vascular surfaces may improve the performance of synthetic small diameter vascular grafts. In vitro lining with cultured autologous endothelial cells offers a confluent endothelium at the time of implantation.Methods: Between June 1989 and December 1991, 49 patients who had no saphenous vein available entered the study. Indication for operation was disabling claudication in 37 patients and critical ischemia in 12 patients. With a random 1:2 assignment, 33 patients were admitted to the endothelialized group and 16 control patients received an untreated polytetrafluoroethylene prosthesis. Cultured autologous endothelial cells from the external jugular vein were confluently lined onto polytetrafluoroethylene grafts precoated with fibrinolytically inhibited fibrin glue. The follow-up was based on angiography, platelet labeling studies with indium 111-labeled oxine, assessment of the ankle-brachial index, and duplex sonography.Results: first-passage mass cultures of 16 million endothelial cells-required for the confluent Lining of a 70 cm long 6 mm graft-were reached 25.1 +/- 11.2 days after vein excision. Growth failure occurred in 27.3%. After 32 months, the actuarial patency was 84.7% for endothelialized grafts and 55.4% for control grafts (p < 0.041 by Breslow test; p < 0.068 by Mantel-Cox test). The ankle-brachial index was continually diverging, reaching significantly lower values in the control group at 24 months (0.98 +/- 0.14 in the endothelialized group versus 0.70 +/- 0.12 in the control; p < 0.0023). The uptake of indium 111-labeled platelets-measured at 9 days, 3 months, 6 months, and 12 months - was significantly lower in the endothelialized group during the entire observation period.Conclusions: Our initial 3-year data demonstrate the early superiority of endothelialized synthetic grafts over commonly used untreated expanded polytetrafluoroethylene prostheses.

This study reports our results with vitro endothelialization of fresh nonpreserved homograft valve leaflets compared with mild alternatively preserved valves and valves treated by preservation procedures commonly used for commercially available tissue valves. In vitro lining of biological heart valves with cultured autologous endothelial cells might help prevent the detrimental effects of degeneration on valve durability. To investigate the growth characteristics of endothelial cells on valve bioprostheses, three different methods of storage and preservation were compared. After precoating with fibronectin and seeding of 4.4 x 10(4) endothelial cells/cm2 onto the different leaflet surfaces, primary adherence, growth kinetics, morphology, and maintenance of monolayer integrity were studied over a period of 10 days. On valve leaflet surfaces of group 1 (fresh nonpreserved homograft valve leaflets) and group 2 (mild alternatively preserved valves), endothelial cells grew to persistent monolayers between days 6 and 10. In contrast, endothelial cell proliferation with monolayer growth could not be achieved on the group 3 leaflets (preserved like commercially available biological valve prostheses). In that group, no viable endothelial cells could be found on the valve surfaces 2 days after seeding. These results demonstrate the theoretical feasibility of endothelializing biological heart valve leaflets in vitro if they are not preserved and stored according to commonly used procedures. Provided such an endothelium can withstand the mechanical forces after implantation in vivo, in vitro endothelialization might contribute either to the development of new biological heart valves for modern cardiac surgery or to the improvement of clinical results with homograft valve transplants.

The cultivation of autologous endothelial cells on the blood surface of artificial hearts might prevent their detrimental thromboembolic complications. To investigate the growth characteristics of endothelial cells on theoretically suitable biomaterials, we compared three polyurethanes (Pellethane, Biomer, Enka) and three silicone rubbers (Elastosil, 3145 RTV, Medical Adhesive). All synthetic surfaces were precoated with an extracellular matrix (group 1), fibronectin (group 2), or a glutaraldehyde-preserved cellular matrix (group 3). After the seeding of 2.5 X 10(4)/cm2 human endothelial cells onto the various surfaces, primary adherence, growth kinetics, and maintenance of monolayer integrity were studied for 13 days. On the three polyurethanes all precoating procedures resulted in endothelial cell proliferation and the formation of persistent monolayers. In contrast, on silicone rubbers a persistent coverage with a confluent endothelium could be achieved only on the glutaraldehyde-preserved cellular matrix. When endothelial cell growth was quantitatively assessed on all precoating substrates, the glutaraldehyde-preserved cellular matrix proved to be far superior on each of the synthetics (p < 0.001). These results demonstrate the theoretical feasibility of endothelialization of artificial hearts in vitro. Provided such an endothelium can withstand the mechanical forces within an artificial heart, in vitro endothelialization might contribute to a regained attractiveness of the elective long-term implantation of artificial hearts.