Adeno-associated virus serotype 9-mediated overexpression of extracellular superoxide dismutase improves recovery from surgical hind-limb ischemia in BALB/c mice

Abstract

Objective Neovascularization is a physiological repair process that is partly dependent on nitric oxide. Extracellular superoxide dismutase (EcSOD) is the major scavenger of superoxide and thus is an important regulator of nitric oxide bioavailability and thus protects against vascular dysfunction. We hypothesized that overexpression of EcSOD in skeletal muscle would improve recovery from hind-limb ischemia. Methods Adeno associated virus (AAV) vectors expressing EcSOD or luciferase (control) from the Cytomegalovirus (CMV) promoter were cross-packaged into AAV9 capsids and injected IM into hind-limb muscles (1×1011 viral genomes(vg)/limb) of 12 wk-old mice. Ischemia was then induced after IM injections. Limb perfusion was serially measured by laser Doppler on days 0, 7 & 14 post-injection and values were expressed as a ratio relative to the non-ischemic limb. EcSOD expression was measured by Western blotting. Capillary density was documented by immunohistochemical staining for platelet endothelial cell adhesion molecule (PECAM). Apoptosis was assessed by Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay and necrosis was visually evaluated daily. Results EcSOD expression was 2-fold up-regulated in EcSOD treated vs. control ischemic muscles at day 14. Capillary density was 1.9-fold higher in treated (1.65±0.02 capillaries/fiber) vs. control muscle (0.78±0.17 capillaries/fiber, p<0.05). Recovery of perfusion ratio at day 14 post-ischemia was 1.5-fold greater in EcSOD vs. control mice (p<0.05). The percentage of apoptotic nuclei was 1.3 ± 0.4% in EcSOD treated mice as compared to 4.2± 0.2% in controls (p<0.001). Limb necrosis was also significantly lower in EcSOD vs. control mice. Conclusion AAV9-mediated overexpression of EcSOD in skeletal muscle significantly improves recovery from hind-limb ischemia in mice, consistent with improved capillary density and perfusion ratios in treated mice.

Enhanced Skeletal Muscle Expression of Extracellular Superoxide Dismutase Mitigates Streptozotocin-Induced Diabetic Cardiomyopathy by Reducing Oxidative Stress and Aberrant Cell Signaling

“…27,28 To ascertain the potential protective function of EcSOD in the heart and its origin under the condition of exercise training, we first tested the extent to which EcSOD protein level in the heart and serum was promoted by endurance exercise training in mice. Four weeks of voluntary wheel running significantly increased EcSOD protein level in the heart and serum in reference to the sedentary control mice.…”

Unbalanced Oxidant‐Antioxidant Status: A Potential Therapeutic Target for Coronary Chronic Total Occlusion in Very Old Patients

“…Cyclic GMP levels were significantly elevated in PECGGp-treated group, thus providing evidence that PECGGp markedly increased levels of SOD3 and SOD3 expression elevated cyclic GMP levels in cardiac tissue of infarcted myocardium. The involvement of NO/cyclic GMP pathway in infarcted myocardium was confirmed [47, 48]. …”

Cardiac myocyte-specific transgenic ecSOD targets mitochondria to protect against Ca2+ induced permeability transition

“…In contrast to MnSOD or CuZnSOD that may be constrained by localization to the mitochondrial matrix, ecSOD can expand its antioxidant capabilities by boosting intracellular expression. These insights are important in understanding ecSOD's contributions to protecting the myocardium and its potential as a therapeutic such as that in gene therapy (Li et al, 1998, 2001; Qi et al, 2007a,b; Deng et al, 2010; Saqib et al, 2011). …”