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The blood-surface interface of 12 mm ID x 5 cm ePTFE vena cava conduits, unseeded (n = 8) and seeded (n = 8) with enzymatically derived autologous endothelial cells, was studied in a canine model at 4 and 12 weeks after graft implantation. Acetylsalicylic acid (325 mg each day) and Coumadin (prothrombin time 1.4 to 1.7 times the control value) were administered preoperatively and continued 4 weeks postoperatively. Platelets labeled with 111In and 125I-labeled fibrinogen were administered 24 hours before graft removal. Luminal platelet adherence, expressed as 10(6) platelets/cm2 of graft surface, was 8.9 +/- 5.6 vs. 56.4 +/- 8.0 (p less than 0.008) and 4.0 +/- 0.9 vs. 12.4 +/- 2.3 (p less than 0.005) in seeded vs. unseeded grafts at 4 and 12 weeks, respectively. Luminal fibrinogen deposition, expressed in micrograms per square centimeter of graft surface, was 11.8 +/- 2.2 vs. 32.0 +/- 2.0 (p less than 0.06) and 6.1 +/- 2.4 vs. 12.4 +/- 6.3 (p less than 0.005) in seeded vs. unseeded grafts at 4 and 12 weeks, respectively. Cumulative 4- and 12-week luminal production of 6-keto-PGF1 alpha from seeded and unseeded grafts represented 11% and 5%, respectively, of that produced from the native iliac vein. Luminal endothelial cell coverage was 71% +/- 22% vs. 33% +/- 9% and 79% +/- 8% vs. 55% +/- 8% (p less than 0.05) in seeded and unseeded grafts at 4 and 12 weeks, respectively. Although endothelialization was not complete in seeded vena cava grafts, it is clear that seeded prostheses exhibited improved thromboresistance compared with unseeded conduits.

Endothelial cell-seeded bilateral carotid ePTFE grafts were studied in 11 dogs to determine the source of their eventual luminal linings. One graft was prepared with autologous cells and the other with nonautologous cells. Immunosuppression was accomplished with cyclosporin and prednisolone. At 2 and 3 weeks post implantation, grafts were removed, revealing 9 of 11 autologous and 8 of 11 nonautologous seeded conduits to be patent. Linings were harvested enzymatically and grown in tissue culture, with karyotype analysis limited to cultures that exhibited greater than or equal to 50% endothelium (n5 autologous, n7 nonautologous). Sex type was classified in 20 cells from each graft. All five autologous cell-seeded grafts revealed host cells. Among the seven nonautologous cell-seeded grafts, one harbored only non-autologous cells, three revealed only autologous cells, and three exhibited a predominance of autologous cells with lesser numbers of nonautologous endothelium. All 100 cells from the autologous-seeded grafts were the same sex as seeded cells, while only 28 of the 140 cells from the nonautologous seeded grafts were the same sex as seeded cells (P less than 0.001). Thus, nonautologous endothelial cell seeding with immunosuppression did not uniformly produce nonautologous cell linings, but was more often associated with autologous endothelial cell linings that may have been promoted by the nonautologous cell seeding.

Hemodynamic and hematologic responses to protamine sulfate reversal of heparin's anticoagulant effects were studied in 15 consecutive randomized patients undergoing aortic reconstructive surgery. In a double-blinded manner, patients were pretreated with either normal saline solution (n = 8) or protamine (0.75 mg/kg/3 min, n = 7) 5 minutes before heparinization (150 IU/kg). After aortic grafts were placed, protamine (1.5 mg/kg/3 min) was administered intravenously to reverse the heparin. Arterial blood pressure, heart rate, pulmonary artery and capillary wedge pressure, central venous pressure, and cardiac output were monitored, as were platelet count, white blood cell count, activated clotting time, total hemolytic complement levels, and C3a levels. Calculated parameters included systemic vascular resistance and pulmonary vascular resistance. Pretreatment with protamine compared with saline solution prevented the hypotension (+6 vs. -16 mm Hg, p less than 0.05) and declining pulmonary artery pressure (+1 vs. -7 mm Hg, p less than 0.01) observed with protamine reversal of heparin. Significant differences between the two groups in central venous pressure and pulmonary vascular resistance were of less clinical relevance. Protamine pretreatment lessened the thrombocytopenia found during reversal compared with saline-pretreated patients although the difference was not statistically significant. Minimal hypotension occurring after protamine pretreatment alone was not accompanied by hemodynamic or hematologic changes, other than decreased heart rate.(ABSTRACT TRUNCATED AT 250 WORDS)

Indium-111-labeled platelet adherence to ePTFE thoracoabdominal vascular prostheses in a canine model (n = 10) was quantitated by (1) an indium-111 excess technique, contrasting graft radioactivity to that in a reference region, and (2) a technetium-99m subtraction technique, with radioactivity of circulating platelets eliminated by discounting background blood activity. Variation in graft thrombogenicity was provided by seeding six prostheses with enzymatically derived autologous endothelial cells, and implanting four prostheses without seeding. Grafts were imaged at 1, 4, and 6 weeks postimplantation, with platelet labeling using indium-111-oxine and red blood cell labeling using technetium-99m. At 7 weeks grafts were excised and gamma activity was measured in proximal, middle, and distal segments. Luminal generation of TxB2 and 6-keto-PGF1 alpha from midportions of grafts was assayed. Indium-111 excess ratios at 6 weeks correlated with actual gamma activity of excised grafts (proximal r = 0.80, P less than 0.01; middle r = 0.73, P less than 0.05; distal r = 0.48, ns) but such a correlation did not exist for the technetium-99m subtraction technique (r = -0.05, -0.25, and 0.16, in the three segments, respectively, all ns). The ratio of graft to aortic TxB2 production revealed a positive correlation with graft gamma activity (r = 0.87, P less than 0.01), and the ratio of graft 6-keto-PGF1 alpha to TxB2 production also correlated with gamma counts (r = -0.64, P = 0.05). In this experimental setting technetium-99m subtraction analysis was an imprecise method of detecting graft platelet accumulation, whereas indium-111 excess ratios proved to be a more accurate method of quantitating vascular prosthetic thrombogenicity.

Platelet-related phenomena at the blood-surface interface of randomly placed knitted Dacron (n = 6) and polytetrafluoroethylene (ePTFE) (n = 6) interposition aortic grafts were studied in patients undergoing abdominal aortic aneurysmectomy. Luminal accumulation of platelets was assessed by infusing indium-111-oxine (400 microCi) labeled autologous platelets and imaging grafts at 1 week, 3 months, and 6 months after surgery. Image analysis included an indium ratio technique (comparing aortic graft radioactivity to that of an iliac artery) and a red blood cell technetium subtraction technique (excluding blood pool radioactivity from graft radioactivity, with the heart or iliac artery serving as reference regions). Plasma levels of beta-thromboglobulin and platelet factor 4 were correlated with platelet accumulations on the aortic prostheses. Differences in graft radioactivity or platelet-release products were not evident 1 week after surgery. Three months after implantation, Dacron and ePTFE conduits exhibited 87% and 47% (p less than 0.05) more radioactivity with the indium ratio technique than the iliac artery. Similarly, increased Dacron compared with ePTFE graft radioactivity was noted using technetium subtraction techniques: 71% vs 30% with a heart reference and 26% vs 11% with an iliac artery reference, respectively. Increases in graft radioactivity correlated with increases in both plasma beta-thromboglobulin and platelet factor 4 at 3 months (r = 0.6 to 0.9; p less than 0.05 to 0.001 depending on the imaging technique used). At 6 months, differences did not persist. In fact, technetium subtraction techniques suggested less Dacron conduit reactivity. It is speculated that differences in platelet accumulation and activation associated with different graft substrates may prove clinically important.

Initial adherence of 111Indium-oxine labeled, cultured canine venous endothelial cells to expanded polytetrafluorethylene (ePTFE) grafts was evaluated using different precoatings of the surface or different pretreated cells. The precoatings evaluated consisted of blood for 5 (Group I) or 15 min (Group II), fibronectin (Group IV), and cryoprecipitate (Group VI). In addition to endothelial cells subcultured immediately before labeling, cells subcultured 48 hours prior to labeling and kept in suspension were studied. Such cells were evaluated on grafts precoated with blood for 5 min (Group III) and fibronectin precoated surfaces (Group V). The amount of fibronectin that adhered to the graft surface with our technique was less den 1%. Seeding efficiency was higher with the blood precoated surfaces (5.3, 3.0, 2.2% in Groups I, II and III respectively) than fibronectin (1.7%, 1.7% in Groups IV, V respectively) or cryoprecipitate (1.9%, Group VI) precoated surfaces. No significant difference between cells immediately subcultured and these kept in suspension for 48 hours regarding adherence to blood (5.3 vs. 2.2%) or fibronectin precoated (1.7 vs. 1.7%) grafts was documented. The seeding efficiency of ePTFE grafts is low and further efforts to improve adherence must be made. From the present studies we recommend the use of blood precoating for about 5 min followed by cell incubation for approximately 10 min in order to achieve optimal seeding efficiency.

The efficacy of treating established vascular graft infections with rifampin and clindamycin (preferentially concentrated in leukocytes) and cefazolin (not concentrated in leukocytes) was studied in a canine model. Infrarenal aortic, 6 mm by 6 cm knitted Dacron double velour grafts were implanted and infected with 10(8) colony-forming units (CFU) of coagulase-positive Staphyloccus aureus organisms injected intravenously immediately after graft placement. Antibiotic therapy was instituted at 3 months postimplantation. Three groups were studied: (I) untreated controls (n = 3); (II) therapy with intravenous cefazolin 15 mg/kg/8 hr for 28 days (n = 7); and (III) combined therapy with intravenous rifampin 13 mg/kg/24 hr and intravenous clindamycin 13 mg/kg/8 hr for 28 days (n = 7). Grafts were removed for quantitative bacteriologic studies after the 28-day course of therapy. Two group I control grafts remained patent with 6.4 X 10(6) and 8.1 X 10(3) CFU S. aureus/gm of graft. The third control graft was thrombosed. Two group II animals demonstrated 1.6 X 10(7) and 2.3 X 10(5) CFU S. aureus organisms/gram of graft, respectively; the remaining five group II grafts were free of organisms. All group III grafts were sterile--a significant difference (p less than 0.05) from group I grafts. In this experimental model, established prosthetic graft infections were eradicated by intensive treatment with antibiotics preferentially concentrated in leukocytes.