Small-diameter Prosthetic Grafts Research Articles

Antibody-Conjugated Electrospun Vascular Scaffolds to Enhance In Situ Endothelialization.

Tissue-engineered vascular grafts (TEVGs) are promising alternatives to small-diameter prosthetic grafts. Previous methods of seeding tubular scaffolds with autologous vascular cells have been successful; however, these methods require significant preparation time. Endothelial cell (EC) growth on the luminal surface of vascular scaffolds may be critical for the integration of a TEVG to the host environment. An alternative approach for TEVGs includes the in situ endothelialization of acellular scaffolds by capturing circulating endothelial progenitor cells (EPCs) and ECs from the bloodstream through the biofunctionalization of the vascular scaffolds. In this study, fibrous scaffolds were electrospun with a 1:1 poly(ε-caprolactone) (PCL)/collagen blend solution. The electrospun fibrous scaffolds were surface-modified by immobilizing EC-specific antibodies: CD31, vascular endothelial cadherin (VE-CAD), vascular endothelial growth factor receptor 2 (VEGFR2), and von Willebrand factor (vWF). Antibodies most efficacious at capturing ECs were then paired to examine their potential synergistic cell-capturing capabilities. The study demonstrated that vascular scaffolds bioconjugated with dual antibodies demonstrated synergistic capture efficacy compared to bioconjugation with a single antibody. The capture of circulating EPCs and ECs can be optimized with bioconjugation of one or more antibodies on the luminal surface of TEVGs.

Short interposition with a small-diameter prosthetic graft for flow reduction of a high-flow arteriovenous fistula

ObjectiveThe objective of this study was to evaluate the outcome of a short interposition using a small-diameter prosthetic graft as a flow-limiting procedure to manage symptomatic high-flow arteriovenous fistula (AVF). MethodsA retrospective review of medical records on a case series was conducted. From June 2004 to April 2017, there were 25 patients with clinical symptoms of high output cardiac failure and progressive dilation of aneurysmal fistula vein due to high-flow AVF (≥1.5 L/min) who underwent short interposition with a 5-mm prosthetic graft at Saitama Medical Center. The primary outcome was the relief of clinical symptoms; other outcome measures included technical success, surgical complications, patency of vascular access, and postoperative changes in local and systemic hemodynamics as assessed by Doppler ultrasound. ResultsTwenty-five patients underwent short interposition for cardiac indications (n = 16) and aneurysmal dilation (n = 9). The technical success rate was 100%. The clinical symptoms were relieved in 24 patients (96.0%). Mean reduction in access blood flow was 52.4%. Cumulative primary unassisted patency rates (± standard error) at 1 year, 2 years, and 3 years were 76.2% ± 9.3%, 70.4% ± 10.3%, and 58.1% ± 11.6%, respectively. Secondary patency rates (± standard error) at 1 year, 2 years, and 3 years were 81.8% ± 8.2%, 71.5% ± 9.9%, and 71.5% ± 9.9%, respectively. Complications included access occlusion due to late thrombosis (n = 5 [21.7%]) and graft infection (n = 1 [4.3%]) in the median follow-up period of 3.9 years. ConclusionsShort interposition with a prosthetic graft is a simple, effective, and durable treatment option for end-stage renal disease patients with cardiac symptoms and progressive dilation of the fistula vein due to high-flow AVF, offering clinical symptom resolution while preserving the autologous behavior of the initial access.

Corrugated nanofiber tissue-engineered vascular graft to prevent kinking for arteriovenous shunts in an ovine model

ObjectiveProsthetic grafts are often needed in open vascular procedures. However, the smaller diameter prosthetic grafts (<6 mm) have low patency and often result in complications from infection. Tissue-engineered vascular grafts (TEVGs) are a promising replacement for small diameter prosthetic grafts. TEVGs start as a biodegradable scaffold to promote autologous cell proliferation and functional neotissue regeneration. Owing to the limitations of graft materials; however, most TEVGs are rigid and easily kinked when implanted in limited spaces, which precludes clinical application. We have developed a novel corrugated nanofiber graft to prevent kinking. MethodsTEVGs with corrugated walls (5-mm internal diameter by 10 cm length) were created by electrospinning a blend of poly-ε-caprolactone and poly(L-lactide-co-caprolactone). The biodegradable grafts were then implanted between the carotid artery and the external jugular vein in a U-shape using an ovine model. TEVGs were implanted on both the left and right side of a sheep (n = 4, grafts = 8). The grafts were explanted 1 month after implantation and inspected with mechanical and histologic analyses. Graft patency was confirmed by measuring graft diameter and blood flow velocity using ultrasound, which was performed on day 4 and every following week after implantation. ResultsAll sheep survived postoperatively except for one sheep that died of acute heart failure 2 weeks after implantation. The graft patency rate was 87.5% (seven grafts out of eight) with one graft becoming occluded in the early phase after implantation. There was no significant kinking of the grafts. Overall, endothelial cells were observed in the grafts 1 month after the surgeries without graft rupture, calcification, or aneurysmal change. ConclusionsOur novel corrugated nanofiber vascular graft displayed neotissue formation without kinking in large animal model.

Tissue-Engineered Small Diameter Arterial Vascular Grafts from Cell-Free Nanofiber PCL/Chitosan Scaffolds in a Sheep Model.

Tissue engineered vascular grafts (TEVGs) have the potential to overcome the issues faced by existing small diameter prosthetic grafts by providing a biodegradable scaffold where the patient’s own cells can engraft and form functional neotissue. However, applying classical approaches to create arterial TEVGs using slow degrading materials with supraphysiological mechanical properties, typically results in limited host cell infiltration, poor remodeling, stenosis, and calcification. The purpose of this study is to evaluate the feasibility of novel small diameter arterial TEVGs created using fast degrading material. A 1.0mm and 5.0mm diameter TEVGs were fabricated with electrospun polycaprolactone (PCL) and chitosan (CS) blend nanofibers. The 1.0mm TEVGs were implanted in mice (n = 3) as an unseeded infrarenal abdominal aorta interposition conduit., The 5.0mm TEVGs were implanted in sheep (n = 6) as an unseeded carotid artery (CA) interposition conduit. Mice were followed with ultrasound and sacrificed at 6 months. All 1.0mm TEVGs remained patent without evidence of thrombosis or aneurysm formation. Based on small animal outcomes, sheep were followed with ultrasound and sacrificed at 6 months for histological and mechanical analysis. There was no aneurysm formation or calcification in the TEVGs. 4 out of 6 grafts (67%) were patent. After 6 months in vivo, 9.1 ± 5.4% remained of the original scaffold. Histological analysis of patent grafts demonstrated deposition of extracellular matrix constituents including elastin and collagen production, as well as endothelialization and organized contractile smooth muscle cells, similar to that of native CA. The mechanical properties of TEVGs were comparable to native CA. There was a significant positive correlation between TEVG wall thickness and CD68+ macrophage infiltration into the scaffold (R2 = 0.95, p = 0.001). The fast degradation of CS in our novel TEVG promoted excellent cellular infiltration and neotissue formation without calcification or aneurysm. Modulating host macrophage infiltration into the scaffold is a key to reducing excessive neotissue formation and stenosis.

Enhancement of capillary and cellular ingrowth in ePTFE implants with a proangiogenic recombinant construct derived from fibronectin

Based on our discoveries of a unique, synergistic interplay between vascular endothelial growth factor (VEGF) and specific domains of the matrix protein fibronectin (FN), we used recombinant technology to create a new protein construct derived from the cell-binding and VEGF-binding domains of FN. We wished to test the hypothesis that this prototype recombinant FN (rFN) protein would enhance cellular and capillary ingrowth in vivo into expanded polytetrafluoroethylene (ePTFE) implants. ePTFE disks of high porosity (60 micron internodal distance) were embedded with fibrin gel and heparin, with/without mixtures of VEGF and rFN and were implanted subcutaneously in rats. Control implants embedded with fibrin glue and heparin alone showed an average of 8.5% (±0.51% standard error mean (SEM)) cellular ingrowth. The addition of either VEGF or rFN caused a modest but significant increase in cellular ingrowth (12.7 ± 1% and 11.8 ± 0.98%, respectively, p < 0.004). However, the combination of rFN/VEGF/heparin dramatically increased cellular ingrowth (27.6 ± 1.62%, p < 0.001), compared with all other treatments. Quantification of capillary ingrowth yielded the same pattern. These results suggest that the incorporation of such biological modulators into cardiovascular implants could offer new strategies for the design of a ready-made small diameter prosthetic graft with enhanced capacity for neovascularization and endothelialization.

Seeding HSVECs on small-caliber vascular grafts for the endothelialization

Objective To study the feasibility of seeding human saphenous vein endothelial cells (HSVECs) onto luminal surface of prosthetic grafts for the endotheliazation of expanded polytetrafluoroethylene (ePTFE) small diameter artificial blood vessel. Methods HSVECs were harvested and cultured in vitro for 13-15 days. The proliferated HSVECs were seeded onto the surface of ePTFE small diameter pros thetic grafts precoated with fibrin glue. After another 9-12 days of culture, morphological change was observed under scanning electron microscope. Results After seeding the cells, the adhesion, crawling and spreading of the HSVECs were observed onto the surface of ePTFE prosthetic graft under the scanning electron microscopy. On the 12th day, surface of ePTFE prosthetic graft was covered completely with endothelial monolayer. Conclusion Endothelial cells derived from human saphanous veins can be used as seed cells for the endothelialization of ePTFE prosthetic graft. Key words: Vascular endothelial cell; Proliferation; Vascular grafts

Optimal Prosthetic Graft Design for Small Diameter Vascular Grafts

Autogenous vein and arterial grafts, such as great saphenous veins and internal mammary and radial arteries, remain the gold standard conduits for vascular reconstruction. Expanded polytetrafluoroethylene (PTFE) grafts, which exhibit little inflammatory and thrombogenic reactivity, are the most commonly used material of choice for small diameter vascular grafts when autogenous grafts are not available. Several modifications of the basic graft have been attempted to enhance graft healing of expanded PTFE grafts, and little but definite experimental and clinical improvement has been achieved so far. The technique of vascular tissue engineering, in combination with stem cell research, may hold the key for the creation of a practical and successful small diameter prosthetic graft.

Coculture of endothelial and smooth muscle cells on a collagen membrane in the development of a small-diameter vascular graft

In this study, we have evaluated the feasibility of developing a biodegradable collagenous small diameter vascular graft of 2 mm diameter and 1 cm length. In brief, bi-layer type I collagen membrane was fabricated under vacuum suction and lyophilization methods. The smooth muscle cells were inoculated into the lower side of the porous membrane, while endothelial cells were seeded onto upper smooth side of the membrane. After cultured for 7 days, the vascular substitute was either harvested for in vitro examination or in vivo implanted in the subcutaneous layer for biocompatibility test. The tubular vascular prosthesis was then used as a temporary absorbable guide that served as an in vivo vascular graft to promote the complete regeneration of rat inferior vena cava. After implantation for 12 weeks, a thin continuous layer of endothelial cells and smooth muscle cells were lined with the vascular lumen and tunic media, respectively. Histology results showed that there were no signs of significant thrombogeneity and intima hyperplasia. This tissue engineered vascular substitute not only had enough tensile strength and good biocompatibility, but also advanced vascular regeneration. In the future, we suggest that this biodegradable vascular substitute will provide with the possibility in application on small diameter prosthetic grafts in artificial blood vessels.

Incorporation of nitric oxide-releasing crosslinked polyethyleneimine microspheres into vascular grafts.

Over the years, many attempts have been made to increase the patency of small- to medium-sized prosthetic vascular grafts. However, none of them has greatly affected long-term rates. Recently, nitric oxide (NO) has been shown to inhibit thrombus formation in such grafts, suggesting that local delivery of NO may help to increase graft patency. This study describes the site-specific delivery of NO by entrapping NO-releasing microspheres in the pores of a vascular graft. NO-releasing polyethyleneimine microspheres (PEIX) were developed using a novel water-in-oil emulsion technique involving chemical crosslinking with a bis-epoxide. The PEIX microspheres were then derivatized with NO forming the [N(O)NO]- moiety of the diazeniumdiolates formerly known as NONOates. These polymeric NO-releasing particles were found to spontaneously release 194 nmol NO/mg with a half-life of over 66 h under physiologic conditions. Fluorescein isothiocyanate-labeled microspheres were then embedded into the pores of a 60-micron nonreinforced Gore-tex vascular graft using a simple evacuation technique and evaluated for microsphere placement and NO release. Scanning electron microscopic analysis showed the microspheres entrapped in the pores of the vascular graft releasing 10 nmol NO/mg with a half-life of 51 h. The microspheres remained entrapped in the graft even after immersion and NO release, as confirmed by fluorescence of the medium. These results suggest that NO-releasing particles can be incorporated into the pores of a vascular graft to deliver therapeutic amounts of NO for the prevention of thrombosis in small-diameter prosthetic grafts.

Thrombogenicity of small-diameter prosthetic grafts: Relative contributions of graft-associated thrombin and factor Xa

We evaluated the contributions of coagulation factors IIa (thrombin) and Xa to small-diameter prosthetic graft thrombogenicity in vivo. Preclotted and nonpreclotted (collagen-coated) polyester grafts were studied before and 24 hours after implantation into pig femoral arteries. After incubation of explanted grafts was performed with plasma depleted of vitamin K-dependent coagulation factors by barium chloride adsorption (Ba-plasma), graft-associated thrombin activity was determined by radioimmunoassay for fibrinopeptide A. Fibrinopeptide A levels reflect thrombin-mediated fibrin formation. Factor Xa activity was characterized by measuring activation of prothrombin added to Ba-plasma. Thrombin and factor Xa were associated with the luminal surfaces of preclotted grafts before and 24 hours after implantation. Nonpreclotted grafts had negligible procoagulant activity before implantation. After 24 hours in vivo graft-associated factor Xa activity was similar in both nonpreclotted and preclotted grafts; however, more thrombin was bound to nonpreclotted coated grafts (p < 0.01). The procoagulant activity of small-diameter prosthetic grafts persists for 24 hours after implantation and is attributable not only to graft-associated thrombin but also to de novo thrombin elaboration induced by factor Xa. Moreover, graft-associated procoagulant activity is not dependent on preclotting because it develops on nonpreclotted, collagen-coated grafts as well. Treatment strategies to attenuate graft thrombosis may require the inhibition of both thrombin and factor Xa.

Optimization of Recombinant t-PA Secretion from Seeded Vascular Grafts

Seeding of vascular grafts with genetically engineered endothelial cells (EC) secreting anticoagulant or fibrinolytic agents offers a potential means of improving patency rates. Prior to the initiation of in vivo studies, we examined cell retention and tissue plasminogen activator (t-PA) secretion from small-diameter synthetic graft segments seeded with sheep venous EC genetically engineered to secrete human t-PA. Following retroviral-mediated gene transfer, EC were seeded at varying densities onto 4-mm-diameter synthetic graft segments of different composition, achieving confluent coverage of all materials. t-PA production from seeded grafts was evaluated under both static conditions and after flow exposure, for up to 3 days after seeding. t-PA secretion varied directly with increasing seeding density for all graft types, reaching a maximum of 20 ng/cm 2/24 hr. t-PA secretion correlated highly with the number of seeded cells as determined by measurement of DNA from lysates of seeded grafts ( r 2 = 0.90, P < 0.0001). For all graft types tested, approximately 50% of seeded cells were retained after exposure to flow in vitro. Retained EC remained viable as determined by t-PA secretion. The rate of t-PA secretion from collagen-impregnated Dacron grafts was higher than that obtained with other materials both under static conditions and after flow exposure. This higher rate was most likely due to the higher surface area presented by the Dacron grafts. These data demonstrate that small-diameter prosthetic graft materials can be coated with a layer of EC that (i) remains metabolically active and capable of secreting a fibrinolytic agent, and (ii) remains adherent to the graft surface after exposure to flow. These experiments provide a foundation for in vivo studies in which grafts are seeded with EC genetically engineered to increase local fibrinolysis.

Perioperative suppression of platelet adherence to small-diameter polytetrafluoroethylene (PTFE) grafts

The perioperative effect of platelet antagonists on small-diameter polytetrafluoroethylene (PTFE) grafts was evaluated in forty-six New Zealand white male rabbits receiving either dipyridamole (DPM) 100 mg/kg/day ( n = 10; group 1), aspirin (ASA) 10 mg/kg/day ( n = 10; group 2), a combination of ASA 10 mg/kg/day and DPM 10 mg/kg/day ( n = 9; group 3) or 100 mg/kg/day ( n = 10; group 4), or placebo ( n = 7) as single daily doses. All regimens began 72 hr prior to insertion of a 20 × 3-mm internal diameter PTFE interposition aortic graft. Autologous indium-111 labeled platelets were injected immediately after implantation. Graft and an equivalent segment of aorta were harvested after 48 hr. Graft platelet adherence index (GPAI) was calculated as the graft:reference aorta ratio of emissions. The GPAI in the control group was 238 ± 46 (mean ± SD). Single regimen antiplatelet agents in groups 1 and 2 reduced mean GPAI to 47 ± 38 and 40 ± 12, respectively. The combination regimen in group 3 lowered mean GPAI to 43 ± 8 and in group 4 to 21 ± 7. Platelet uptake in PTFE grafts at 48 hr is significantly lowered to 8.8 to 19.7% of control by perioperative antiplatelet agents given as a single daily oral dose ( P < 0.001). ASA alone and DPM alone provided similar suppression of platelet uptake, but combination ASA + low dose or high dose DPM gave significantly greater ( P < 0.001) suppression of early platelet deposition than the single agent regimens. These results support perioperative administration of combination oral antiplatelet agents as adjunctive therapy in small diameter prosthetic graft implantation.

Endothelial cell adhesion to vascular prosthetic surfaces

Improving the long term patency of small diameter prosthetic grafts remains an important but elusive objective in vascular surgery. In pursuit of a non-thrombogenic surface, we have cultured human-adult endothelial cells, examined their adhesive properties and their ability to colonize the inner surface of a Dacron® graft. To examine cell adhesion, endothelial cells were labelled with 111In-oxine and inoculated onto prosthetic wells previously prepared with either cold insoluble globulin (CIG), 1 % gelatin, alginate or left untreated as a control. At 100 min, mean percentage adhesion to CIG and gelatin precoated wells was 86.0 ± 9.9% (± sd) and 81.6 ± 2.9% respectively, whereas alginate at 57.5 ± 7.5% and the control well at 48.3 ± 9.9% showed significantly less adhesion. Further experiments examined the adhesion of Indiumoxine labelled endothelial cells to Dacron® graft (2 cm × 6 mm id) placed in an in vitro arterial circuit. An immediate loss of approximately 20% of the cells occurred within the first 30 s, whereafter, a stable population were adherent to the graft material. By 102 min, 73.4 ± 1.8% of cells remained attached when exposed to tissue culture medium, but only 64.1 ± 6.9% after exposure to blood. Cultured human adult endothelial cells adhere most effectively to prosthetic surfaces precoated with CIG or gelatin, and remain attached following exposure to shear forces.