Gene therapy is emerging as a potential treatment option in patients suffering from a wide spectrum of cardiovascular diseases including coronary artery disease, peripheral vascular disease, vein graft failure and in‐stent restenosis. Thus far preclinical studies have shown promise for a wide variety of genes, in particular the delivery of genes encoding growth factors such as vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF) to treat ischaemic vascular disease both peripherally and in coronary artery disease. VEGF as well as other genes such as TIMPs have been used to target the development of neointimal hyperplasia to successfully prevent vein graft failure and in‐stent restenosis in animal models. Subsequent phase I trials to examine safety of these therapies have been successful with low levels of serious adverse effects, and albeit in the absence of a placebo group some suggestion of efficacy. Phase 2 studies, which have incorporated a placebo group, have not confirmed this early promise of efficacy. In the next generation of clinical gene therapy trials for cardiovascular disease, many parameters will need to be adjusted in the search for an effective therapy, including the identification of a suitable vector, appropriate gene or genes and an effective vector delivery system for a specific disease target. Here we review the current status of cardiovascular gene therapy and the potential for this approach to become a viable treatment option. British Journal of Pharmacology (2007) 152, 175–188; doi:

Although successful, drug-eluting stents require significant periods of dual anti-platelet therapy with a persistent risk of late stent thrombosis due to inhibition of re-endothelialization. Endothelial regeneration is desirable to protect against in-stent thrombosis. Gene-eluting stents may be an alternative allowing inhibition of neointima and regenerating endothelium. We have shown that adenoviral endothelial nitric oxide synthase (eNOS) delivery can result in significantly decreased neointimal formation and enhanced re-endothelialization. Here, we examined non-viral reporter and therapeutic gene delivery from a stent. We coated lipoplexes directly onto the surface of stents. These lipostents were then deployed in the injured external iliac artery of either normal or hypercholesterolemic New Zealand White rabbits and recovered after 28 days. Lipoplexes composed of lipofectin and a reporter lacZ gene or therapeutic eNOS gene were used. We demonstrated efficient gene delivery at 28 days post-deployment in the media (21.3 ± 7.5%) and neointima (26.8 ± 11.2%). Liposomal delivery resulted in expression in macrophages between the stent struts. This resulted in improved re-endothelialization as detected by two independent measures compared with vector and stent controls (Po0.05 for both). However, in contrast to viral delivery of eNOS, liposomal eNOS does not reduce restenosis rates. The differing cell populations targeted by lipoplexes compared with adenoviral vectors may explain their ability to enhance re-endothelialization without affecting restenosis. Liposome-mediated gene delivery can result in prolonged and localized transgene expression in the blood vessel wall in vivo. Furthermore, lipoeNOS delivery to the blood vessel wall results in accelerated re-endothelialization; however, it does not reduce neointimal formation. Gene Therapy (2012) 19, 321-328; doi:10.1038/gt.2011.92; published online 30 June 2011Keywords: endothelialization; coronary artery stents; restenosis; late stent thrombosis; liposome INTRODUCTION Drug-eluting stents (DES) are now routinely used for occlusive atherosclerotic coronary lesions to reduce the problem of restenosis. However, DES have been associated with a higher frequency of very late stenosis and re-infarction (more than 1 year) compared with bare metal stents, 1.9% versus 0.6% per year, respectively. 1,2 In addition, animal studies have shown that DES can cause local toxicity to the vessel wall in the form of medial necrosis, intimal proliferation, chronic inflammation and delayed re-endothelialization of the stents. 1-3 Thus, although DES reduce the risk of restenosis, they are associated with delayed re-endothelialization. 4,5 An optimal therapeutic approach would involve a strategy, which inhibits intimal hyperplasia while promoting re-endothelialization and suppressing stent-or vector-related inflammatory side effects.Gene-eluting stents have the potential to provide an alternative treatment strategy for the prevention of restenosis. The safety and feasibility of viral-medi...

Tissue necrosis resulting from critical limb ischemia (CLI) leads to amputation in a significant number of patients. Autologous cell therapy using angiogenic cells such as endothelial progenitor cells (EPCs) holds promise as a treatment for CLI but a limitation of this treatment is that the underlying disease etiology that resulted in CLI may also contribute to dysfunction of the therapeutic EPCs. This study aimed to elucidate the mechanism of EPC dysfunction using diabetes mellitus as a model and to determine whether correction of this defect in dysfunctional EPCs ex vivo would improve the outcome after cell transplantation in the murine hind limb ischemia model. EPC dysfunction was confirmed in a homogenous population of patients with type 1 diabetes mellitus and a microarray study was preformed to identify dysregulated genes. Notably, the secreted proangiogenic protein osteopontin (OPN) was significantly downregulated in diabetic EPCs. Furthermore, OPN-deficient mice showed impaired recovery following hind limb ischemia, suggesting a critical role for OPN in postnatal neovascularization. EPCs isolated from OPN KO mice showed decreased ability to adhere to endothelial cells as well as impaired angiogenic potential. However, this dysfunction was reversed upon exposure to recombinant OPN, suggesting that OPN may act in an autocrine manner on EPCs. Indeed, exposure of OPN knockout (KO) EPCs to OPN was sufficient to induce the secretion of angiogenic proteins (IL-6, TGF-α, and FGF-α). We also demonstrated that vascular regeneration following hind limb ischemia in OPN KO mice was significantly improved upon injection of EPCs preexposed to OPN. We concluded that OPN acts in an autocrine manner on EPCs to induce the secretion of angiogenic proteins, thereby playing a critical role in EPC-mediated neovascularization. Modification of cells by exposure to OPN may improve the efficacy of autologous EPC transplantation via the enhanced secretion of angiogenic proteins.

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IntroductionPericytes, although traditionally considered as supporting cells, have recently been proposed to have a more active role in the repair and pathogenesis of various vascular diseases. In this study, we hypothesised that a pericyte-like stem cell population, termed vessel derived stem cells (VSCs), with chondrogenic and osteogenic potential exists in the vessel wall and in the presence of the inflammatory cytokines seen in atherosclerotic environment contributes, along with the circulating mesenchymal stem cells, to the calcification of atherosclerotic plaque which occurs through the endochondral pathway.MethodsVSCs from aortae of ApoE−/− mice and control C57BL/6 mice were isolated and characterised for cell surface markers by flow cytometry and immunocytochemistry. MSCs from the bone marrow of these mice were also isolated and characterised. Chondrogenic potential of these cells was investigated in presence or absence of inflammatory cytokines such as IL-6 and IFN-γ.Results and DiscussionIsolated VSCs were strongly positive for Sca-1, CD44 and negative for CD31 and CD34. A sub-population of VSCs also expressed 3G5, a specific pericyte marker. Differentiation assays demonstrated the ability of the cells to undergo osteogenesis and chondrogenesis. VSCs had significantly higher GAG/DNA ratio than MSCs indicating comparatively increased chondrogenesis. That both MSCs and VSCs from the ApoE−/− atherosclerotic mice generate a more mature hypertrophic chondrocyte than cells from the C57BL/6 mice is interesting and suggests that the atherosclerotic environment may modulate the stem cell phenotype. Collagen-type II and aggrecan expression and effect of oxidised LDL on these cells in vitro will be investigated to further test this hypothesis.