Carotid Artery Interposition Grafts Research Articles

Long-Term Results of Autologous Bifurcated Carotid Artery Reconstructions

Carotid bifurcation revascularization using interposition grafts is rare. While internal carotid artery (ICA) revascularization is regarded as mandatory, the external carotid artery (ECA) is severed in most instances. Long-term results of an autologous bifurcated carotid artery reconstruction are discussed. Single centre, retrospective analysis of a consecutive series of patients treated at an academic vascular surgery center. From 12/2006 to 11/2019, fifty-one patients underwent reconstruction of the carotid artery using an autologous bifurcated interposition graft (38 male, 75%; median age: 68.7 years; IQR: 60.2-76.5). Thirty-eight patients were asymptomatic (74.5%). Indication for reconstruction was recurrent carotid stenosis unfavourable for endovascular treatment/redo patchplasty (n=32, symptomatic: n=7), carotid aneurysm (n=11, symptomatic: n=5), neck tumor with vascular involvement (n=7), trauma (n=1). Non-reversed valve depleted saphenous vein y-graft from the groin was used in 49 (94.2%) and reversed cubital vein bifurcation in three (5.8%) procedures. Median ICA cross-clamp time was 15 minutes (IQR: 13-20 minutes). In four procedures (7.7%), a shunt was inserted because of significant decrease of cerebral perfusion. In one patient, additional intraoperative stent placement of a proximal common carotid artery (CCA) stenosis was performed. Six patients suffered from ischemic neurological deficits (11.5%), all but 1 recovered with no or moderate symptoms. After a median follow-up of 5.2 years (IQR: 1.1-8.7 years), 7 significant asymptomatic stenoses (13.5%) and 2 occlusions (3.8%, one symptomatic) of the ICA, two significant asymptomatic CCA stenoses (3.8%), five significant stenoses (9.6%) and seven asymptomatic occlusions (13.5%) of the ECA were observed. This prompted 7 re-redo-interventions in 5 patients. Twenty-eight patients (54.9%) died after a median follow up of 3.3 years (IQR: .5-5.6 years). Two of these patients died due to conditions related to the performed carotid artery reconstruction. Autologous bifurcated carotid artery interposition graft provides results comparable to other complex ICA-revascularizations. Overall survival in this heterogeneous patient cohort is poor. Due to high risk of stroke and poor long-term outcome, this procedure should be reserved for symptomatic patients with no other option for revascularization.

Abstract 444: Apolipoprotein A-I (ApoAI) Vascular Gene Therapy Protects Against Vein-Graft Atherosclerosis

Introduction: Coronary artery venous bypass grafts typically fail because of accelerated atherosclerosis, driven by lipid infiltration and by accumulation of macrophages and other cells. We previously showed that transduction of carotid artery endothelium of fat-fed rabbits with a helper-dependent adenovirus expressing apolipoprotein AI (HDAdApoAI) reduced atherosclerotic lesion size and decreased intimal lipid and macrophage accumulation. We hypothesized that HDAdApoAI transduction of rabbit vein graft endothelium would reduce lipid and macrophage accumulation. Methods: Fat-fed rabbits received bilateral external jugular vein to carotid artery interposition grafts. Four wks later, one graft per rabbit was infused with HDAdApoAI and the other with HDAdNull (a non-expressing HDAd). Grafts were harvested 12 wks after HDAd infusion. Paired analysis was performed in all rabbits with bilateral patent grafts (n=22). Graft hemodynamics were measured at vector-infusion and harvest surgeries. In vivo graft diameters were measured before vector infusion and harvest with ultrasound. Grafts were cut into 8 segments, for histology and biochemical analyses. Results: Mean plasma cholesterol levels were 400-600 mg/dL. Graft patency rates were 89% after vein grafting and 100% after the vector-infusion surgery. Lumen diameter and hemodynamics did not differ between HDAdApoAI and HDAdNull-transduced grafts. HDAd genomes were detected in all vein grafts. High levels of apoAI mRNA were detected in 100% of HDAdApoAI-treated vein grafts (mean 155-fold higher than in HDAdNull grafts). HDAdApoAI-treated grafts had 25% lower cholesterol (by mass spectrometry; P=0.03) and 17% less median CD68 mRNA (by RT-qPCR; P=0.008; CD68 is expressed by macrophages and lipid-loaded SMC). No significant differences were detected in mRNA encoding several inflammatory cytokines and inflammation-associated proteins. There were nominal decreases in total intimal area (16%, P=0.9), percent stenosis (4%, P=0.08), and in intimal area positive for Oil Red O (11%, P=0.3), macrophages (11%, P=0.7), and VCAM-1 (8%, P=0.8). Conclusion: HDAdApoAI transduction of vein graft endothelium reduces some markers of atherosclerosis, potentially by stimulating cholesterol efflux.

Tissue-Engineered Carotid Artery Interposition Grafts Demonstrate High Primary Patency and Promote Vascular Tissue Regeneration in the Ovine Model.

Tissue-engineered vascular graft for the reconstruction of small arteries is still an unmet clinical need, despite the fact that a number of promising prototypes have entered preclinical development. Here we test Poly(3-hydroxybutyrate-co-3-hydroxyvalerate)Poly(ε-caprolactone) 4-mm-diameter vascular grafts equipped with vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and stromal cell-derived factor 1α (SDF-1α) and surface coated with heparin and iloprost (PHBV/PCL[VEGF-bFGF-SDF]Hep/Ilo, n = 8) in a sheep carotid artery interposition model, using biostable vascular prostheses of expanded poly(tetrafluoroethylene) (ePTFE, n = 5) as a control. Primary patency of PHBV/PCL[VEGF-bFGF-SDF]Hep/Ilo grafts was 62.5% (5/8) at 24 h postimplantation and 50% (4/8) at 18 months postimplantation, while all (5/5) ePTFE conduits were occluded within the 24 h after the surgery. At 18 months postimplantation, PHBV/PCL[VEGF-bFGF-SDF]Hep/Ilo grafts were completely resorbed and replaced by the vascular tissue. Regenerated arteries displayed a hierarchical three-layer structure similar to the native blood vessels, being fully endothelialised, highly vascularised and populated by vascular smooth muscle cells and macrophages. The most (4/5, 80%) of the regenerated arteries were free of calcifications but suffered from the aneurysmatic dilation. Therefore, biodegradable PHBV/PCL[VEGF-bFGF-SDF]Hep/Ilo grafts showed better short- and long-term results than bio-stable ePTFE analogues, although these scaffolds must be reinforced for the efficient prevention of aneurysms.

Improvement of a Novel Small-diameter Tissue-engineered Arterial Graft With Heparin Conjugation

Small diameter (<6 mm), bioabsorbable, arterial, tissue-engineered vascular grafts (TEVGs) remain limited by thromboembolism. The objective of this study was to test whether heparin-eluting (HE) TEVGs prevent early thrombosis in a large animal model. TEVGs were created with an outer poly-ε-caprolactone electrospun nanofiber layer, with a 15-μm average pore size and an inner layer composed of a 50:50 poly(L-lactide-co-ε-caprolactone) copolymer. Adult female sheep (n= 5) underwent bilateral carotid artery interposition grafting, with a control TEVG in 1 carotid artery and an HE TEVG in the contralateral position. Animals were followed for 8 weeks with weekly Duplex ultrasonography to monitor TEVG performance. All sheep survived to the designated endpoint. At 8 weeks all 5 HE TEVGs were patent. Three of 5 control TEVGs had early thrombotic occlusion at <1 week. More than 97% of heparin release occurred within the first 24 hours. Histologic evaluation of the HE TEVG displayed cellularity like a native carotid artery with no evidence of calcification. Significantly fewer platelets adhered to the HE TEVG than to the control TEVG (P < .001). This study suggests HE TEVGs prevent acute graft thrombosis. We hypothesize that the HE properties of the HE TEVG during vascular endothelialization is useful for maintaining TEVG patency. This technique may aid in the translation of small arterial TEVGs to the clinic.

The effect of pore diameter on neo-tissue formation in electrospun biodegradable tissue-engineered arterial grafts in a large animal model

To date, there has been little investigation of biodegradable tissue engineered arterial grafts (TEAG) using clinically relevant large animal models. The purpose of this study is to explore how pore size of electrospun scaffolds can be used to balance neoarterial tissue formation with graft structural integrity under arterial environmental conditions throughout the remodeling process. TEAGs were created with an outer poly-ε-caprolactone (PCL) electrospun layer and an inner sponge layer composed of heparin conjugated 50:50 poly (l-lactide-co-ε-caprolactone) copolymer (PLCL). Outer electrospun layers were created with four different pore diameters (4, 7, 10, and 15 µm). Fourteen adult female sheep underwent bilateral carotid artery interposition grafting (n = 3–4 /group). Our heparin-eluting TEAG was implanted on one side (n = 14) and ePTFE graft (n = 3) or non-heparin-eluting TEAG (n = 5) on the other side. Twelve of the fourteen animals survived to the designated endpoint at 8 weeks, and one animal with 4 µm pore diameter graft was followed to 1 year. All heparin-eluting TEAGs were patent, but those with pore diameters larger than 4 µm began to dilate at week 4. Only scaffolds with a pore diameter of 4 µm resisted dilation and could do so for up to 1 year. At 8 weeks, the 10 µm pore graft had the highest density of cells in the electrospun layer and macrophages were the primary cell type present. This study highlights challenges in designing bioabsorbable TEAGs for the arterial environment in a large animal model. While larger pore diameter TEAGs promoted cell infiltration, neotissue could not regenerate rapidly enough to provide sufficient mechanical strength required to resist dilation. Future studies will be focused on evaluating a smaller pore design to better understand long-term remodeling and determine feasibility for clinical use. Statement of SignificanceIn situ vascular tissue engineering relies on a biodegradable scaffold that encourages tissue regeneration and maintains mechanical integrity until the neotissue can bear the load. Species-specific differences in tissue regeneration and larger mechanical forces often result in graft failure when scaling up from small to large animal models. This study utilizes a slow-degrading electrospun PCL sheath to reinforce a tissue engineered arterials graft. Pore size, a property critical to tissue regeneration, was controlled by changing PCL fiber diameter and the resulting effects of these properties on neotissue formation and graft durability was evaluated. This study is among few to report the effect of pore size on vascular neotissue formation in a large animal arterial model and also demonstrate robust neotissue formation.

Vitrified Human Umbilical Arteries as Potential Grafts for Vascular Tissue Engineering.

The development of a biological based small diameter vascular graft (d < 6mm), that can be properly stored over a long time period at - 196°C, in order to directly be used to the patients, still remains a challenge. In this study the decellularized umbilical arteries (UAs) where vitrified, evaluated their composition and implanted to a porcine model, thus serving as vascular graft. Human UAs were decellularized using 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS) and sodium dodecyl sulfate (SDS) detergents. Then, vitrified with vitrification solution 55 (VS55) solution, remained for 6months in liquid nitrogen and their extracellular matrix composition was compared to conventionally cryopreserved UAs. Additionally, total hydroxyproline, sulphated glycosaminoglycan and DNA content were quantified in all samples. Finally, the vitrified umbilical arteries implanted as common carotid artery interposition graft to a porcine animal model. Decellularized and vitrified UAs characterized by proper preservation of extracellular matrix proteins and tissue architecture, whereas conventionally cryopreserved samples exhibited a disorganized structure. Total hydroxyproline content was preserved, although sulphated glycosaminoglycan and DNA contents presented significantly alterations in all samples. Implanted UAs successfully recellularized and remodeled as indicated by the histological analysis. Decellularized and vitrified UAs retained their structure function properties and can be possible used as an alternative source for readily accessible small diameter vascular grafts.

A Rabbit Model of Durable Transgene Expression in Jugular Vein to Common Carotid Artery Interposition Grafts.

Vein graft bypass surgery is a common treatment for occlusive arterial disease; however, long-term success is limited by graft failure due to thrombosis, intimal hyperplasia, and atherosclerosis. The goal of this article is to demonstrate a method for placing bilateral venous interposition grafts in a rabbit, then transducing the grafts with a gene transfer vector that achieves durable transgene expression. The method allows the investigation of the biological roles of genes and their protein products in normal vein graft homeostasis. It also allows the testing of transgenes for the activities that could prevent vein graft failure, e.g., whether the expression of a transgene prevents the neointimal growth, reduces the vascular inflammation, or reduces atherosclerosis in rabbits fed with a high-fat diet. During an initial survival surgery, the segments of right and left external jugular vein are excised and placed bilaterally as reversed end-to-side common carotid artery interposition grafts. During a second survival surgery, performed 28 days later, each of the grafts is isolated from the circulation with vascular clips and the lumens are filled (via an arteriotomy) with a solution containing a helper-dependent adenoviral (HDAd) vector. After a 20-min incubation, the vector solution is aspirated, the arteriotomy is repaired, and flow is restored. The veins are harvested at time points dictated by individual experimental protocols. The 28-day delay between the graft placement and the transduction is necessary to ensure the adaptation of the vein graft to the arterial circulation. This adaptation avoids rapid loss of transgene expression that occurs in vein grafts transduced before or immediately after grafting. The method is unique in its ability to achieve durable, stable transgene expression in grafted veins. Compared to other large animal vein graft models, rabbits have advantages of low cost and easy handling. Compared to rodent vein graft models, rabbits have larger and easier-to-manipulate blood vessels that provide abundant tissue for analysis.

A Rabbit Model of Durable Transgene Expression in Jugular Vein to Common Carotid Artery Interposition Grafts

A Rabbit Model of Durable Transgene Expression in Jugular Vein to Common Carotid Artery Interposition Grafts

Autologous Bifurcated Carotid Artery Reconstruction: Technique and 2 Year Follow Up

Carotid bifurcation resection with subsequent reconstruction is performed infrequently. While internal carotid artery (ICA) revascularisation is regarded as mandatory, the external carotid artery (ECA) is severed in most instances. Indications for and results of an autologous bifurcated carotid artery reconstruction are discussed. This was a single centre retrospective analysis of a consecutive series of patients treated at an academic vascular surgery centre. From December 2006 to August 2015, 47 patients underwent reconstruction of the carotid artery using an autologous bifurcated interposition graft (36 men; median age 68.3 years [range 41–87 years]; 33 asymptomatic [70%]). The indication for reconstruction was recurrent carotid stenosis unfavorable for endovascular treatment/redo patch plasty (n = 29; symptomatic: n = 12), carotid aneurysm (n = 10), neck tumour with vascular involvement (n = 7), and trauma (n = 1). Groups were not different with regard to vascular risk factors. Non-reversed valve depleted saphenous/accessory saphenous vein from the groin was used in 44 (94%), and reversed valve depleted cubital vein bifurcation graft in three patients (6%). Median ICA cross clamp time was 17 min (range 8–30 min). Five patients suffered from intra-operative ischaemic neurological deficits (11%), of which all were temporary except one. One tumour patient died on post-operative day 1 due to bleeding. Median follow up was 28.3 months (range 0.1–97.4 months). Tumour erosion required graft ligation in one patient 7 months after reconstruction. Five significant stenoses of the ICA (11%), nine (19%) stenoses of the ECA, and two (4%) common carotid artery stenoses, with four (8%) re-redo interventions, were observed. Thirteen patients (28%) died after a median follow up of 6.7 months (range 1.7–56.7 months), all unrelated to the vascular procedure. Autologous bifurcated carotid artery interposition graft avoids the use of prosthetic graft material, the ECA remains patent in 80%, and it provides mid-term results comparable with other redo carotid artery procedures. Therefore, this procedure should be reserved for symptomatic patients with no other revascularisation option.

Autologous Bifurcated Carotid Artery Reconstruction: Technique and 2 Year Follow Up

Carotid bifurcation resection with subsequent reconstruction is performed infrequently. While internal carotid artery (ICA) revascularisation is regarded as mandatory, the external carotid artery (ECA) is severed in most instances. Indications for and results of an autologous bifurcated carotid artery reconstruction are discussed. This was a single centre retrospective analysis of a consecutive series of patients treated at an academic vascular surgery centre. From December 2006 to August 2015, 47 patients underwent reconstruction of the carotid artery using an autologous bifurcated interposition graft (36 men; median age 68.3 years [range 41-87 years]; 33 asymptomatic [70%]). The indication for reconstruction was recurrent carotid stenosis unfavorable for endovascular treatment/redo patch plasty (n=29; symptomatic: n=12), carotid aneurysm (n=10), neck tumour with vascular involvement (n=7), and trauma (n=1). Groups were not different with regard to vascular risk factors. Non-reversed valve depleted saphenous/accessory saphenous vein from the groin was used in 44 (94%), and reversed valve depleted cubital vein bifurcation graft in three patients (6%). Median ICA cross clamp time was 17min (range 8-30min). Five patients suffered from intra-operative ischaemic neurological deficits (11%), of which all were temporary except one. One tumour patient died on post-operative day 1 due to bleeding. Median follow up was 28.3 months (range 0.1-97.4 months). Tumour erosion required graft ligation in one patient 7 months after reconstruction. Five significant stenoses of the ICA (11%), nine (19%) stenoses of the ECA, and two (4%) common carotid artery stenoses, with four (8%) re-redo interventions, were observed. Thirteen patients (28%) died after a median follow up of 6.7 months (range 1.7-56.7 months), all unrelated to the vascular procedure. Autologous bifurcated carotid artery interposition graft avoids the use of prosthetic graft material, the ECA remains patent in 80%, and it provides mid-term results comparable with other redo carotid artery procedures. Therefore, this procedure should be reserved for symptomatic patients with no other revascularisation option.

Abstract 254: The Bromo and Extraterminal Domain Protein (BET) Family Drives Endothelial-To-Mesenchymal Transition and Contributes to Vein Graft Stenosis

Background: Vein graft bypass remains the most commonly used open procedure for flow-limiting cardiovascular diseases. However, vein grafts are prone to stenosis, which ultimately leads to graft failure (4-year failure rate at ~43%). Previous studies have provided strong evidence supporting that dysfunctional endothelial cells (ECs) of the vein grafts contribute significantly to neoinitma formation (and hence stenosis) by undergoing EndoMT. Our group has recently shown that BRD4, a member of the BET epigenetic reader protein family, is up-regulated in the neointima lesion of stenotic failed vein grafts. Here we have determined whether BRD4, and possibly other BET proteins, would be involved in EndoMT in vein grafts, and ultimately vein graft stenosis/failure. Method: For in vitro experiments, we treated rat primary vascular ECs with 100ng/ml TGFβ-1 (to induce EndoMT), and/or BET inhibitors (Pan inhibitor: JQ1; Domain-selective inhibitors: Olinone and RVX208), and/or siRNAs for BRD2, 3, and 4. For in vivo experiments, we used a cuff-based technique to establish jugular vein to carotid artery interposition graft in rats, and lentiviral shRNA delivery to achieve specific knockdown of BRD4 in the graft. Results: BET pan-inhibition significantly reduced EndoMT of rat primary vascular ECs in vitro. Similar effects were observed via selective inhibition of bromodomain 2, but not bromodomain 1. Knockdown of BRD4 protein expression in vitro fully recapitulated the inhibitory effect of BET pan-inhibition on EndoMT, whereas knockdown of BRD2 and BRD3 could only lead to partial and marginal abrogation of EndoMT, respectively. In the rat vein graft model, local lentiviral delivery of BRD4 shRNA led to decrease of neointima formation. Conclusion: BET proteins, especially BRD4, play an important role in EndoMT of vascular ECs and also neointima formation in vein grafts. Blockage of BRD4 protein could potentially serve as a novel strategy to achieve prolonged patency of vein graft conduits.

A20 Haploinsufficiency Aggravates Transplant Arteriosclerosis in Mouse Vascular Allografts: Implications for Clinical Transplantation.

Inflammation is central to the pathogenesis of transplant arteriosclerosis (TA). We questioned whether physiologic levels of anti-inflammatory A20 influence TA severity. We performed major histocompatibility complex mismatched aorta to carotid artery interposition grafts, using wild type (WT) or A20 heterozygote (HET) C57BL/6 (H-2) donors and BALB/c (H-2) recipients, and conversely BALB/c donors and WT/HET recipients. We analyzed aortic allografts by histology, immunohistochemistry, immunofluorescence, and gene profiling (quantitative real-time reverse-transcriptase polymerase chain reaction). We validated select in vivo A20 targets in human and mouse smooth muscle cell (SMC) cultures. We noted significantly greater intimal hyperplasia in HET versus WT allografts, indicating aggravated TA. Inadequate upregulation of A20 in HET allografts after transplantation was associated with excessive NF-кB activation, gauged by higher levels of IkBα, p65, VCAM-1, ICAM-1, CXCL10, CCL2, TNF, and IL-6 (mostly localized to SMC). Correspondingly, cytokine-induced upregulation of TNF and IL-6 in human and mouse SMC cultures inversely correlated with A20 expression. Aggravated TA in HET versus WT allografts correlated with increased intimal SMC proliferation, and a higher number of infiltrating IFNγ and Granzyme B CD4 T cells and natural killer cells, and lower number of FoxP3 regulatory T cells. A20 haploinsufficiency in allograft recipients did not influence TA. A20 haploinsufficiency in vascular allografts aggravates lesions of TA by exacerbating inflammation, SMC proliferation, and infiltration of pathogenic T cells. A20 single nucleotide polymorphisms associating with lower A20 expression or function in donors of vascularized allografts may inform risk and severity of TA, highlighting the clinical implications of our findings.

Anti-inflammatory and anti-proliferative effects of CBS3830 in arterialized vein grafts in rats

Objective: To investigated whether CBS3830, a highly selectively inhibitor of p38MAPK, could ameliorate inflammation and intimal hyperplasia in arterialized vein grafts (AVGs).Methods: Sixty male Sprague-Dawley rats underwent a reversed right jugular vein to common carotid artery interposition graft and were randomly treatment with vehicle (control) or single-dose (3 mg/kg, preoperative) or double-dose (3 mg/kg, preoperative and 4 d postoperative) CBS3830. Twenty rats underwent sham operation. The levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) were determined by ELISA. Vein grafts were analyzed by intimal/medial morphometry, proliferating cell nuclear antigen (PCNA) expression, and p38MAPK phosphorylation.Results: TNF-α, IL-1β, and IL-6 gradually increased then slowly decreased in AVG rats. However, at 4 d and 7 d, TNF-α levels decreased by 37.5% and 29.5% (p = 0.003, 0.05, respectively) in the single-dose CBS3830 group, and by 37.6% and 32.5%, respectively (both p = 0.003) in the double-dose group compared with those of control. IL-1β levels significantly reduced at 4 d and 14 d in both dosage groups. IL-6 levels significantly reduced at 7 d in both groups. Intima and medial thickening were significantly reduced in both dosage treated groups at 7, 14, and 28 d (all p = 0.000) compared to the controls. Further study showed CBS3830 inhibited p38MAPK phosphorylation and decreased PCNA expression.Conclusions: CBS3830 significantly decreases inflammation and intimal hyperplasia in AVGs.

Time and flow-dependent changes in the p27kip1 gene network drive maladaptive vascular remodeling

Although clinical studies have identified that a single nucleotide polymorphism in the p27(kip1) gene is associated with success or failure after vein bypass grafting, the underlying mechanisms for this difference are not well defined. Using a high-throughput approach in a flow-dependent vein graft model, we explored the differences in p27(kip1)-related genes that drive the enhanced hyperplastic response under low-flow conditions. Bilateral rabbit carotid artery interposition grafts with jugular vein were placed with a unilateral distal outflow branch ligation to create differential flow states. Grafts were harvested at 2 hours and at 1, 3, 7, 14, and 28 days after implantation, measured for neointimal area, and assayed for cell proliferation. Whole-vessel messenger RNA was isolated and analyzed using an Affymetrix (Santa Clara, Calif) gene array platform. Ingenuity Pathway Analysis (Ingenuity, Redwood City, Calif) was used to identify the gene networks surrounding p27(kip1). This gene set was then analyzed for temporal expression changes after graft placement and for differential expression in the alternate flow conditions. Outflow branch ligation resulted in an eightfold difference in mean flow rates throughout the 28-day perfusion period (P < .001). Flow reduction led to a robust hyperplastic response, resulting in a significant increase in intimal area by 7 days (0.13 ± 0.04 mm(2) vs 0.014 ± 0.006 mm(2); P < .005) and progressive growth to 28 days (1.37 ± 0.05 mm(2) vs 0.39 ± 0.06 mm(2); P < .001). At 7 days, low-flow grafts demonstrated a burst of actively dividing intimal cells (36.4 ± 9.4 cells/mm(2) vs 11.5 ± 1.9 cells/mm(2); P = .04). Sixty-five unique genes within the microarray were identified as components of the p27(kip1) network. At a false discovery rate of 0.05, 26 genes demonstrated significant temporal changes, and two dominant patterns of expression were identified. Class comparison analysis identified differential expression of 11 genes at 2 hours and seven genes and 14 days between the high-flow and low-flow grafts (P < .05). At 2 hours, oncostatin M and cadherin 1 were the most differentially expressed. Cadherin 1 and protein kinase B exhibited the greatest differential expression at 14 days. Alterations in flow and shear stress result in divergent patterns of vein graft remodeling. Associated with the dramatic increase in neointimal expansion observed in low-flow vs high-flow grafts is a subset of differentially expressed p27(kip1)-associated genes that correlate with critical stages in the adaptive response. These represent potential biologic targets whose activity may be altered to augment maladaptive vascular remodeling.

Superiorin vivoCompatibility of Hydrophilic Polymer Coated Prosthetic Vascular Grafts

Protein adsorption, cell adhesion and graft patency was compared in hydrophilic versus hydrophobic polymer-coated prosthetic vascular grafts. We hypothesize that in vivo compatibility of hydrophilic polymer-coated prosthetic vascular grafts is superior to in vivo compatibility of hydrophobic grafts. A pairwise side-to-side common carotid artery interposition graft was placed eight female landrace goats (mean weight 55 kg). Protein adsorption was assessed using Western Blot in two hydrophilic and two hydrophobic grafts harvested after three days. Graft patency was monitored for 28 days in six goats with continuous wave Doppler ultrasonography. Adherence of endothelial cells, leukocytes and platelets was determined with ELISA and compared between the two graft types after 28 days. After three days, more ApoA-I, albumin and VEGF and less fibrin adsorbed to hydrophilic grafts. After 28 days, compared to hydrophobic grafts, higher numbers of endothelial cells were present on hydrophilic grafts (P=0.016), and less thrombocytes and leukocytes (P=0.012 and 0.024, respectively). Two out of eight hydrophobic grafts lost patency, while none of the hydrophilic grafts failed (P=0.157). Hydrophilic polymer-coated vascular grafts have superior in vivo compatibility when compared to hydrophobic grafts as characterized by reduced platelet and leukocyte adherence as well as higher endothelialization.

P38 MAPK Inhibitor, CBS3830 Limits Vascular Remodelling in Arterialised Vein Grafts

Following bypass surgery vein grafts undergo a remodelling process that can lead to restenosis and ultimately vein graft failure. Signalling through mitogen activated protein kinases (MAPKs) is a key mechanism involved in vein graft failure. Here, we investigated whether CBS3830 (c-a-i-r biosciences GmbH, Tübingen, Germany), a new highly selectively inhibitor of p38 MAPK, has a significant effect on inhibiting intimal, medial and adventitial hyperplasia. Sixty specific pathogen free Sprague Dawley male rats were randomly divided into three groups. The control group with a reversed right jugular vein, which is common to carotid artery interposition graft, was compared with sham-operated, and CBS3830 treated animals. Intimal, medial and adventitia morphometric examinations and expression of proliferating cell nuclear antigen (PCNA) were analysed after one, two and four weeks for vein grafts. Intimal, medial and adventitia thickening in CBS3830 group were significantly lower than in the control group at each time point. Moreover, CBS3830 significantly reduced the phosphorylation of p38 MAPK and PCNA expression compared to the control. On the basis of the present work, intima, media and adventitia of saphenous vein grafts undergo vascular remodelling after surgery. The new, highly selective p38 MAPK inhibitor, CBS3830, ameliorates intimal, medial, and adventitial remodelling by varying degrees.

Statin therapy prevents expansive remodeling in venous bypass grafts

BackgroundVenous grafts (VG) have high failure rates by 10 years in aortocoronary bypass surgery. We have previously shown that expansive remodeling followed by increased LDL retention are early atherosclerotic changes in experimental VG placed in the arterial circulation. The objective of this study was to determine whether statin therapy prevents these expansive remodeling changes. Methods and resultsReversed jugular vein-to-common carotid artery interposition graft was constructed in 27 cholesterol-fed (0.5%) rabbits. Rabbits were randomized either to control or atorvastatin (5mg/kg/day) groups, starting two weeks prior to vein graft implantation and continuing until sacrifice at 1 or 12 weeks post-surgery. Ultrasound measurements of arterial luminal cross-sectional area (CSA) were done at day 3 and at 4, 8 and 12 weeks post-surgery. Histomorphometric measurements were performed following sacrifice at 12 weeks. Atorvastatin treatment significantly decreased total plasma cholesterol levels at 4, 8 and 12 weeks (12 weeks: 6.7±4.2mmol/L versus control 38.7±10.6mmol/L, p<0.0002). Atorvastatin significantly reduced expansive remodeling at 4, 8 and 12 weeks (lumen CSA: 44.6±6.6mm2 versus control 77.6±10.7mm2, p<0.0001). Intimal CSA by histomorphometry was also significantly reduced by atorvastatin at 12 weeks (5.59±2.19mm2 versus control 9.57±2.43mm2, p<0.01). VG macrophage infiltration, MMP-2 activity and metalloelastase activity were reduced in the atorvastatin treated group. ConclusionAtorvastatin inhibits both expansive remodeling and intimal hyperplasia in arterialized VG, likely through inhibition of macrophage infiltration and reduction of tissue proteolytic activity. The mechanism proposed above may be important for preventing VG atherosclerosis and late VG failure.

Abstract P351: Statins Prevent Expansive Remodeling in Arterialized Vein Grafts

Vein grafts (VG) account for more than 70% of conduits used in aortocoronary bypass surgery, but have high failure rates by 10 years. We have previously shown in an experimental model that VG undergo marked expansion by 4 weeks, which was followed by increased total LDL retention at 12 weeks. The objective of this study was to determine whether statin therapy would prevent these expansive remodelling changes. Methods and Results Reversed jugular vein-to-common carotid artery interposition graft were constructed in 24 cholesterol-fed (0.5%) rabbits. Rabbits were randomized to either control or atorvastatin (5 mg/kg/day) groups, starting two weeks prior to vein graft implantation and continuing until sacrifice at 1 or 12 weeks post-surgery. Ultrasound measurements of arterial luminal cross-sectional area (LCSA) were done at day 3 and at 4, 8 and 12 weeks post-surgery. Histomorphometric measurements were performed following sacrifice at 12 weeks. Atorvastatin treatment significantly decreased total plasma cholesterol levels at 12 weeks (6.70±4.19mmol/L vs placebo 38.65±10.55mmol/L, p&lt;0.001). Atorvastatin significantly reduced expansive remodelling at all time points, including at 12 weeks (44.6±6.62mm 2 vs placebo 77.6±10.68mm 2 , p&lt;0.001). Intimal CSA by histomorphometry was significantly reduced by atorvastatin at 12 weeks (5.59±2.19mm 2 vs placebo 9.57±2.43mm 2 , p&lt;0.05). At 1 week, macrophage infiltration was significantly reduced (Ram11 positive intimal staining 0.018±0.019 vs 0.189±0.049, p&lt;0.05). Atorvastatin treated samples at 1 week showed reduction in both MMP 2 activity (gelatin zymography) and metalloelastase activity (casein zymography). Conclusion: Atorvastatin prevents expansive remodelling in arterialized VG, likely through inhibition of macrophage infiltration and reduction of tissue proteolytic activity. This may be an important mechanism preventing VG atherosclerosis and late VG failure.

Update on Lung Transplantation using Donation after Cardiac Death in NSW

P< 0.05) with hydraulic permeability and burst pressure equal to autologous conduits (HP: IMA 0ml/min/cm2, EC 0 P=ns; BP: IMA 2267 mmHg, EC 1922 P=ns). Fluorescence microscopy confirmed endothelial proliferation on electrospun fibres, and thrombogenicity studies demonstrated reducedplatelet attachment on rHE scaffolds. Discussion: Synthetic coronary conduits constructed using rHE/PCL fibres match the mechanical properties of the IMA. In addition, these scaffolds exhibit reduced thrombogenicity and the ability to support endothelialisation. The elastic conduits will be implanted as carotid artery interposition grafts in rabbits for assessment in vivo before ultimate study as coronary bypass graft candidates. These novel conduits show great potential as a synthetic internal mammary artery, the “Holy Grail” of coronary revascularisation surgery. doi:10.1016/j.hlc.2010.10.034 Update on Lung Transplantation using Donation after Cardiac Death in NSW E. Granger ∗, P. Spratt, M. Harkess, P. Jansz, K. Dhital, P. Macdonald, M. Malouf, A. Glanville The Lung TransplantUnit, St. Vincent’s Hospital, Darlinghurst, ever it will probably augment transplant numbers by only 15%. Current results are excellent despite “warm ischaemia”. Donors with longer warm ischaemic times and more “marginal” gas exchange may now be considered. doi:10.1016/j.hlc.2010.10.035 Beating Heart Preservation of Donor Hearts for Transplantation—A Working Heart Perfusion System for Pig Hearts to Extend Organ Preservation Time P. Pritzwald-Stegmann1,∗, J. Passage1,2, T. Muller 2, A. Muhle2, S. Krabbes1,2, S. Dhein2, F.-W. Mohr2 1Monash Medical Centre, Clayton, Australia Q4 2 Heart Centre Leipzig, Leipzig, Germany Introduction:Donor heart ischaemic time places significant geographic constraints on organ retrieval. Constant pressure perfusion systems can double current time limits in a pig model of heart transplantation from 4 to 8h, but the yield of successfully preserved hearts is low. The aim of this studywas to develop aworking heart perfusion system that would extend donor heart preservation time and allow the subsequent assessment of organ function post reperfusion. Methods: Beating hearts were harvested from 61± 3 kg pigs (n= 6) andperfused via a double lumen cannula using Sydney, NSW, Australia Introduction: Donation after cardiac death (DCD) from Maastrict category 3 donors consists of approximately 12% of lung transplant activity in New South Wales. We review the results of DCD lung transplants and discuss the future direction of the program at St. Vincent’s Hospital. Methodology: Since commencing in November 2007 there have been 25 referrals for DCD, leading to 11 attempted retrievals. Only 9 retrievals resulted in lung donation. Elective withdrawal of life-sustaining support and certification of death (limit of 60min fromwithdrawal) occurred in the Intensive Care Unit or Operating Theatre. Results: Average recipient age was 35 years (range 15–61) and aetiology of respiratory failure included cystic fibrosis (2), bronchiectasis (1), bronchiolitis obliterans (1), emphysema (3), interstitial lung disease (1) and pulmonary hypertension (1). The donor group average age was 37 years (range 14–56) and average PaO2/FiO2 ratio 412 (±51). Causes of death included subarachnoid haemorrhage (2), Traumatic head injury (2), hypoxic brain injury (3) and embolic CVA (2). Mean time to asystole after withdrawal of treatment was 27min (range 9–57), mean warm ischaemic time 19min (range 10–31) and mean cold ischaemic time (left lung) 279min (range 241–320). Early gas exchange was excellent: an average PaO2/FiO2 of 324 (141–503). Average intensive care stay was 5 days (3–10) and length of stay 23 days (12–36). All patients are still alive. Conclusion: Organ donation remains at a challenging low level in Australia. Donation after cardiac death may increase the “donor organ pool”, howaLangendorff-basedperfusioncircuit.Workingheart conditionswere created, at physiological perfusion pressures. After 7 hours, the hearts were arrested with cold HTK cardioplegia solution and underwent 1 hour of warm ischaemia. The hearts were subsequently reperfused in a non-working state at constant pressure for 1 hour and then weaned to again perform under the original working heart conditions. Pmax, max dp/dt, min dp/dt were monitored throughout. Tissue adenosine triphosphate (ATP) and malondialdehyde (MDA) were measured to assess myocardial energy content and oxidative stress respectively. Induction of apoptosis was assessed by staining for AIF. Results: All 6 hearts were successfully harvested, preserved and weaned following reperfusion. Pmax achieved during the working heart mode was higher than in the donor animal (142 vs 86mmHg, p= 0.002). There was no significant overall deterioration in systolic (max dp/dt 2030 vs 1812mmHg/s, p= 0.521) or diastolic (min dp/dt 1200 vs 1225mmHg/s, p= 0.902) function throughout preservation. ATP stores recovered to pre harvest levels (23.6 vs 22.1 pmol/ g, p= 0.57) and oxidative stress was well controlled (1.6 vs 1.8 mol/g, p= 0.926). Apoptosis levels were comparable to hearts that underwent conventional 4 h hypothermic arrest prior to transplantation (15.7 vs 12.7/hpf, p= 0.741). Discussion: A working heart perfusion system can be utilised to successfully increase donor heart preservation times. The system allows the real-time monitoring of donor organ function and canmimic the ischaemia and reperfusion events that occur during transplantation. It permits the specific modification of preservation parame-

The Role of ex-vivo Gene Therapy of Vein Grafts with Egr-1 Decoy in the Suppression of Intimal Hyperplasia

Objectives To test the hypothesis that vein graft intimal hyperplasia can be significantly suppressed by a single intra-operative transfection of the graft with a decoy oligonucleotide (ODN) binding the transcription factor Egr-1. Design Experimental study. Materials and methods Jugular vein to carotid artery interposition grafts in rabbits were treated with Egr-1 decoy, mutant decoy ODN, vehicle alone, using a non-distending pressure of 300 mmHg for 20 min, or were left untreated. All animals were fed a 2% cholesterol diet. The animals were sacrificed after 48 h, 6 weeks and 12 weeks. Paraffin-embedded vein sections were subjected to angiometric analysis. Results Successful delivery of the ODN was confirmed by DAPI staining. Quantitative real-time PCR revealed a 60% decrease of the Egr-1 gene expression in the animals in which the Egr-1 decoy ODN was delivered. Cellular proliferation was also significantly decreased as indicated by the Ki-67 labelling index. An increase in intimal and medial thickness was found in all vein grafts. However, intimal thickness was significantly reduced in the grafts treated with Egr-1 decoy ODN, whereas luminal area was significantly increased. Conclusion A single intra-operative pressure-mediated transfection of vein grafts with Egr-1 decoy ODN significantly suppresses intimal hyperplasia in a rabbit hypercholesterolaemic model.