Role of coronin 1B in PDGF-induced migration of vascular smooth muscle cells.

RATIONALE: The type I subclass of coronins, a family of actin-binding proteins, regulates various actin-dependent cellular processes, including migration. However, the existence and role of coronins in vascular smooth muscle cell (VSMC) migration has yet to be determined. OBJECTIVE: The goal of the present study was to define the mechanism by which coronins regulate platelet-derived growth factor (PDGF)-induced VSMC migration. METHODS AND RESULTS: Coronin 1B (Coro1B) and 1C (Coro1C) were both found to be expressed in VSMCs at the mRNA and protein levels. Downregulation of Coro1B by siRNA increases PDGF-induced migration, while downregulation of Coro1C has no effect. We confirmed through kymograph analysis that the Coro1B-downregulation-mediated increase in migration is directly linked to increased lamellipodial protraction rate and protrusion distance in VSMC. In other cell types, coronins exert their effects on lamellipodia dynamics by an inhibitory interaction with the ARP2/3 complex, which is disrupted by the phosphorylation of Coro1B. We found that PDGF induces phosphorylation of Coro1B on serine-2 via PKCε, leading to a decrease in the interaction of Coro1B with the ARP2/3 complex. VSMCs transfected with a phosphodeficient S2A Coro1B mutant showed decreased migration in response to PDGF, suggesting that the phosphorylation of Coro1B is required for the promotion of migration by PDGF. In both the rat and mouse, Coro1B phosphorylation was increased in response to vessel injury in vivo. CONCLUSIONS: Our data suggest that phosphorylation of Coro1B and the subsequent reduced interaction with ARP2/3 complex participate in PDGF-induced VSMC migration, an important step in vascular lesion formation.

Sildenafil reduces respiratory muscle weakness and fibrosis in the mdx mouse model of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is the most common form of muscular dystrophy caused by mutations in the dystrophin gene. Loss of dystrophin initiates a progressive decline in skeletal muscle integrity and contractile capacity which weakens respiratory muscles including the diaphragm, culminating in respiratory failure, the leading cause of morbidity and mortality in DMD patients. At present, corticosteroid treatment is the primary pharmacological intervention in DMD, but has limited efficacy and adverse side effects. Thus, there is an urgent need for new safe, cost-effective, and rapidly implementable treatments that slow disease progression. One promising new approach is the amplification of nitric oxide-cyclic guanosine monophosphate (NO-cGMP) signalling pathways with phosphodiesterase 5 (PDE5) inhibitors. PDE5 inhibitors serve to amplify NO signalling that is attenuated in many neuromuscular diseases including DMD. We report here that a 14-week treatment of the mdx mouse model of DMD with the PDE5 inhibitor sildenafil (Viagra(®), Revatio(®)) significantly reduced mdx diaphragm muscle weakness without impacting fatigue resistance. In addition to enhancing respiratory muscle contractility, sildenafil also promoted normal extracellular matrix organization. PDE5 inhibition slowed the establishment of mdx diaphragm fibrosis and reduced matrix metalloproteinase-13 (MMP-13) expression. Sildenafil also normalized the expression of the pro-fibrotic (and pro-inflammatory) cytokine tumour necrosis factor α (TNFα). Sildenafil-treated mdx diaphragms accumulated significantly less Evans Blue tracer dye than untreated controls, which is also indicative of improved diaphragm muscle health. We conclude that sildenafil-mediated PDE5 inhibition significantly reduces diaphragm respiratory muscle dysfunction and pathology in the mdx mouse model of Duchenne muscular dystrophy. This study provides new insights into the therapeutic utility of targeting defects in NO-cGMP signalling with PDE5 inhibitors in dystrophin-deficient muscle.

Sildenafil reverses cardiac dysfunction in the mdx mouse model of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is a progressive and fatal genetic disorder of muscle degeneration. Patients with DMD lack expression of the protein dystrophin as a result of mutations in the X-linked dystrophin gene. The loss of dystrophin leads to severe skeletal muscle pathologies as well as cardiomyopathy, which manifests as congestive heart failure and arrhythmias. Like humans, dystrophin-deficient mice (mdx mice) show cardiac dysfunction as evidenced by a decrease in diastolic function followed by systolic dysfunction later in life. We have investigated whether sildenafil citrate (Viagra), a phosphodiesterase 5 (PDE5) inhibitor, can be used to ameliorate the age-related cardiac dysfunction present in the mdx mice. By using echocardiography, we show that chronic sildenafil treatment reduces functional deficits in the cardiac performance of aged mdx mice, with no effect on normal cardiac function in WT controls. More importantly, when sildenafil treatment was started after cardiomyopathy had developed, the established symptoms were rapidly reversed within a few days. It is recognized that PDE5 inhibitors can have cardioprotective effects in other models of cardiac damage, but the present study reports a prevention and reversal of pathological cardiac dysfunction as measured by functional analysis in a mouse model of DMD. Overall, the data suggest that PDE5 inhibitors may be a useful treatment for the cardiomyopathy affecting patients with DMD at early and late stages of the disease.

Evaluation of the therapeutic utility of phosphodiesterase 5A inhibition in the mdx mouse model of duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is a devastating and ultimately fatal disease characterized by progressive muscle wasting and weakness. DMD is caused by the absence of a functional dystrophin protein, which in turn leads to reduced expression and mislocalization of dystrophin-associated proteins including neuronal nitric oxide (NO) synthase mu (nNOSµ). Disruption of nNOSµ signaling results in muscle fatigue and unopposed sympathetic vasoconstriction during exercise, thereby increasing contraction-induced damage in dystrophin-deficient muscles. The loss of normal nNOSµ signaling during exercise is central to the vascular dysfunction proposed over 40 years ago to be an important pathogenic mechanism in DMD. Recent preclinical studies focused on circumventing defective nNOSµ signaling in dystrophic skeletal and cardiac muscle by inhibiting phosphodiesterase 5A (PDE5A) have shown promising results. This review addresses nNOS signaling in normal and dystrophin-deficient muscles and the potential of PDE5A inhibition as a therapeutic approach for the treatment of cardiovascular deficits in DMD.