Local Adenovirus-Mediated Delivery of Hirudin in a Rabbit Arterial Injury Model

Abstract

Intravascular delivery of an E1/E3 deleted adenovirus encoding the hirudin protein reduces neointimal formation in the rat arterial injury model. Given the interspecies variability in response to adenoviral vectors, we tested this same construct in the hirudin-sensitive cholesterol-fed rabbit arterial balloon injury model. We hypothesized that local delivery of an E1/E3-deleted adenovirus encoding hirudin (Ad-Hir) in addition to early hirudin infusion would limit neointimal formation compared to early hirudin alone. Methods and Results: Local delivery of Ad-Hir, 2.5 × 10¹⁰ PFU/ml, using a double balloon catheter [n = 6 vessels (v)] produced a 79% reduction in vessel wall thrombin activity at 48 h after balloon angioplasty (BA) compared with vehicle (Veh, n = 6v; p = 0.05). In chronic experiments, hypercholesterolemic rabbits underwent femoral BA, and received either early hirudin alone (n = 9v) or early hirudin plus locally delivered Ad-Hir (early hirudin + Ad-Hir; n = 9v), an E1/E3-deleted adenovirus encoding β-galactosidase (early hirudin + AdGal; n = 7v), or Veh (early hirudin + Veh; n = 10v). Early hirudin + Ad-Hir did not limit the arterial response to injury versus the other groups at 4 weeks after BA. Plaque area, cross-sectional luminal area narrowing by plaque, and T cell infiltration were significantly increased in the adenovirus- versus non-adenovirus-treated arteries. Plaque area correlated with T cell density. Conclusion: Following BA in cholesterol-fed rabbits, local transduction with A-Hir produced a marked reduction in vessel wall-associated thrombin activity. However, this strategy increased rather than decreased the arterial response to BA injury. Our results suggest that the lack of therapeutic effect resulted from adenovirus-stimulated plaque formation, possibly resulting from a T cell-mediated inflammatory response.

Selective α v β 3 -Receptor Blockade Reduces Macrophage Infiltration and Restenosis After Balloon Angioplasty in the Atherosclerotic Rabbit

“…14 The injury score of each artery was assessed with the following scale: 0, intact internal elastic lamina (IEL); 1, lacerated IEL, compression of media; 2, lacerated IEL and media; and 3, large transluminal laceration including disruption of the EEL. 14 …”

“…Paraffin-embedded sections were stained with the following antibodies: HHF-35, a mouse monoclonal anti-human ␣-actin, 0.3 g/mL (Enzo Diagnostics); RAM-11, mouse monoclonal anti-rabbit macrophage, 1.2 g/mL (Dako Corp) 14 ; polyclonal goat anti-rabbit T cell, 4 g/mL (Accurat Chem) 14 ; and polyclonal goat anti-mouse PECAM-1 (platelet and endothelial cell adhesion molecule-1), 1 g/mL (Santa Cruz Biotechnology, Inc). 15 Before labeling with T-cell or PECAM-1 antibodies, arterial tissue underwent microwave antigen retrieval (Vector Labs).…”

Gene Transfer Therapy in Vascular Diseases

“…However, this study demonstrated that adenoviral-mediated secretion of hirudin from ECs lining implanted femoral AVGs not only significantly reduced the proliferation of VSMCs but also the aggregation of platelets, formation of thrombus, and platelet deposition. Similar experiments testing the efficacy of hirudin to inhibit neointimal formation have been performed in the rat arterial injury model and cholesterol-fed rabbit arterial balloon-injury model (8). E1/E3-deleted adenoviral-mediated delivery of hirudin into the rabbit model resulted in a 79% reduction in vessel wall thrombin activity at 48 h after balloon angioplasty.…”

Cardiovascular gene delivery: The good road is awaiting☆

“…Genetic approaches to increase thromboresistance have been achieved by multiple groups through the overexpression of thrombomodulin [115], hydrolysis of adenosine nucleotides [116], addition of an ADPase [117], plasmin induction [118] or activators [119], and hirudin [120,121]. Of course, ECs themselves secrete many of these substances endogenously [122,123], so these approaches may be no more effective than a functioning endothelium, and thus, they may be best applied to SMCs or non-cellular prosthetic systems that are not expected to endothelialize over time.…”