Systemic human Netrin-1 gene delivery by adeno-associated virus type 8 alters leukocyte accumulation and atherogenesis in vivo

Abstract

Atherosclerosis is an inflammatory disorder of arteries. Atherosclerotic plaque, in its early to intermediate stages, is composed largely of lipid-engorged foam cells. These foam cells are derived from the trafficking of monocytes (Mo) into the arterial intima, attracted to the site by chemoattractants. Given that foam cells are derived from the trafficking of Mo, the use of Netrin-1, an Mo chemorepellent, may be useful in limiting Mo accumulation and subsequent plaque formation. To investigate the potential of Netrin-1 for limiting atherosclerosis, we systemically delivered its human (h) cDNA by adeno-associated virus type 8 (AAV8, single-stranded structure) delivery into low-density lipoprotein receptor knockout (LDLRÀ/À) mice and placed the animals on a high cholesterol diet (HCD). Compared with control neomycin resistance (Neo) gene delivery/HCD, hNetrin-1 delivery resulted in a significant reduction in plaque formation, as determined by larger aortic lumen size, thinner intima-media thickness and lower blood velocity than the Neo/HCD control (all statistically significant). Indices of monocyte/macrophage (Mo/MF) accumulation, CD68, integrin, alpha M (ITGAM) and egf-like module containing, mucin-like, hormone receptor-like 1 (EMR-1), were reduced in hNetrin-1/HCD-treated animal's aortas and spleens compared with Neo/HCD-treated animals. Unexpectedly, CD25 and foxp3 (regulatory T cells (Tregs)) in the aorta were strongly upregulated. This is the first time the Mo/MF chemorepellent approach, and specific Netrin-1 gene delivery, has been performed for the reduction of Mo/MF burden and atherosclerosis. In addition, Netrin-1 has never before been linked to altered Treg levels. These data strongly suggest that hNetrin-1 gene delivery can reduce Mo/MF accumulation, inflammation and subsequent plaque formation.

Association of Neuroimmune Guidance Cue Netrin-1 and Its Chemorepulsive Receptor UNC5B With Atherosclerotic Plaque Expression Signatures and Stability in Human(s)

“…8 In another study, intravenous delivery of NTN1 to hypercholesterolemic mice using viral vectors increased NTN1 expression, presumably on endothelial cells, and resulted in prevention of MΦ infiltration of the nascent plaque. 13 Recent studies have shown that coronary endothelial cells express NTN1, which inhibits migration of human monocytes in vitro.…”

Systemic delivery of adeno-associated viral vectors

“…Today, impressive results have been reported in neonatal, adult and even aged animals. Some of these examples include AAV-1 mediated gene therapy for Pompe disease, limb-girdle muscular dystrophy (LGMD) and myotonic dystrophy [33-35], AAV-6 mediated gene therapy for DMD and facioscapulohumeral muscular dystrophy [25,36-38], AAV-8-mediated gene therapy for DMD, LGMD and atherosclerosis [26,39-41], and AAV-9 mediated gene therapy for cardiomyopathy, lysosomal storage disorders and neuronal diseases [42-51]. …”

Immunomodulatory Functions of Neuronal Guidance Proteins

“…As a result, ongoing inflammation within adipose tissue resulted in decreased glucose tolerance in a glucose tolerance test, with the development of insulin resistance [16]. Importantly, using an adenovirus vector to deliver hNetrin-1 in vivo resulted in a significant reduction in plaque formation, as determined by larger aortic lumen size, thinner intima-media thickness, and reduced blood velocity, thus demonstrating that netrin-1 is a viable target in atherosclerosis [81].…”