Search

NO mediates a variety of physiologic processes and is considered an important intracellular messenger in different cellular systems. Because of its complex regulation and multiple molecular and cellular targets, NO provides both stimulatory and suppressive properties in the immune system. Dendritic cells (DCs) are considered the most potent APCs, whose regulation has important implications in the induction of an effective immune response. In this study, we analyzed the effect of the compound NCX 2057, a new class of NO-releasing derivatives of ferulic acid, on activation and functional properties of DCs. NCX 2057 was able to modulate the inflammatory program, the cytokines production, and the cellular life cycle but not the maturation markers and the T cells stimulatory capacity of DCs in the presence or absence of LPS. The results indicate that NCX 2057 may modulate different aspects of the activation of DCs and suggest novel applications of NO donors in the contest of inflammatory response modulation through the life cycle regulation of DCs.

Neutrophil elastase (NE) is a crucial marker of neutrophilic inflammation in bronchiectasis (BE) and cystic fibrosis (CF). We aimed at comparing different techniques to detect active NE in sputum samples of 50 BE and 50 CF adults recruited during stable state at the Bronchiectasis and CF Programs of the Policlinico Hospital in Milan, Italy. Three methods including a ProteaseTag® Active NE Immunoassay (ELISA) and two enzymatic digestion assays (chromogenic –CS- and fluorogenic –FS- substrate) were compared. Results of active NE were also correlated with different clinical data. The three methods provided statistically different values for NE activity in the same sputum samples in both cohorts. The three methods were comparable in terms of consistency and agreement. In the BE cohort, the highest correlation between NE activity and Bronchiectasis Severity Index (rho=0.40, P

The JAK/STAT signalling plays a key role in both innate and adaptive immunity and it has been reported that JAK inhibition can reduce allergen-induced airway inflammation in different animal models of asthma. Here we studied the effects of the JAK inhibitor tofacitinib, orally administered at 15 and 30 mg/kg b.i.d., in two HDM-induced mouse models of asthma, one sensitive (HDM/Alum) and the other insensitive to inhaled steroids (HDM/CFA): modulation of inflammatory cell recruitment and cytokines in BALF along with the lung STAT3 phosphorylation, as a target engagement marker of JAK inhibition, were evaluated. In the HDM/Alum model characterised by eosinophilic inflammation and Th2 cytokine profile, tofacitinib, administered only during the HDM challenge, significantly reduced eosinophil recruitment at 30 mg/kg (41 ± 8%, p Conversely, in the HDM/CFA model, characterised by a predominant neutrophilic component and mixed Th1 and Th2 cytokine profile, tofacitinib failed to modulate inflammatory cell recruitment in BALF, even when administered during both the sensitisation and the challenge phase. Overall, these data suggest that the JAK/STAT pathway may represent a relevant target for eosinophilic allergic forms of asthma.

Aims Cyclooxygenase (COX)-inhibiting nitric oxide donors (CINODs) are a new class of drugs that structurally combine a COX inhibitor with a nitric oxide (NO) donating moiety. This combination reduces potential toxicity of the non-steroidal anti-inflammatory drugs (NSAIDs) whilst maintaining the analgesic and anti-inflammatory effects. The present study was undertaken to investigate the anti-inflammatory effects of NCX 429, a naproxen-based CINOD, and to assess the additional properties of NO donation beyond those related to naproxen. Main methods We evaluated the in vitro effects of NCX 429 on oxy-radical production, phagocytosis, cytokine release, MMP-9, PPARγ expression and NF-κB activation in human monocytes/MDM and compared to naproxen. Moreover, we compared the in vivo efficacy of NCX 429 and naproxen in a murine model of peritonitis. Key findings In all the experiments performed in vitro, NCX 429 reduced the inflammatory responses with equal or higher efficacy compared to naproxen. Moreover, in in vivo experiments, NCX 429, at the lowest dose tested, was able to significantly inhibit cell influx in response to IL-1β administration although naproxen was found to be more potent than NCX 429 at reducing PGE 2 in inflammatory exudates. Significance These results demonstrate that both in vitro and in vivo – in a murine model of peritonitis – NCX 429 elicits significant anti-inflammatory activity, beyond the simple COX inhibition or pure NO release. Therefore, NO donation along with COX inhibition may represent a strategy for investigating inflammatory diseases in which pain and function are not fully resolved by analgesics/anti-inflammatory drugs.

The role of neutrophil elastase (NE) is poorly understood in bronchiectasis because of the lack of preclinical data and so most of the assumptions made about NE inhibitor potential benefit is based on data from CF. In this context, NE seems to be a predictor of long-term clinical outcomes and a possible target of treatment. In order to better evaluate the role of NE in bronchiectasis, a systematic search of scientific evidence was performed. Two investigators independently performed the search on PubMed and included studies published up to May 15, 2017 according to predefined criteria. A final pool of 31 studies was included in the systematic review, with a total of 2679 patients. For each paper data of interest were extracted and reported in table. In this review sputum NE has proved useful as an inflammatory marker both in stable state bronchiectasis and during exacerbations and local or systemic antibiotic treatment. NE has also been associated with risk of exacerbation, time to next exacerbation and all-cause mortality. This study reviews also the role of NE as a specific target of treatment in bronchiectasis. Inhibition of NE is at a very early stage and future interventional studies should evaluate safety and efficacy for new molecules and formulations.

Inhibition of p38 MAPK is considered a possible new therapeutic approach for the treatment of COPD and severe asthma for which inhaled corticosteroids (ICS) have limited efficacy. We report here the effect of CHF6297, a potent and selective inhaled p38MAPK inhibitor, in three murine models of steroid-resistant lung inflammation. In mice exposed to tobacco smoke (TS), CHF6297, administered intranasally (i.n.) b.i.d. for 4 days at 0.03 or 0.3mg/kg, dose-dependently inhibited the corticosteroid-resistant TS-induced neutrophil influx in bronchoalveolar lavage (BAL) reaching a % of inhibition of 29.7±15.3 (n.s.) and 62±8.4 (p

The bronchoprotective effect of CHF6366, a novel dual antimuscarinic and β2 receptor-agonist (MABA) compound, was studied in a model of bronchospasm in anaesthetised guinea pigs that allows discrimination of the two pharmacological activities by using the spasmogens acetylcholine (ACh, -/+ β2 blocker for the assessment of the MABA and antimuscarinic effect, respectively) and histamine (β2 effect). Batefenterol, a reference MABA, was included in the study as a comparator. CHF6366 (0.3-1nmol/kg), administered intratracheally, exerted a dose-dependent prevention of ACh-induced bronchospasm at 1 hour post-administration with maximal efficacy at 1nmol/kg (92.9±2.9% inhibition) and good balance of the two activities (% inhibition M3: 83.9±4.5; β2: 72.7±4.4). CHF6366 peak effect was observed at 5 minutes post-dose with both activities fully effective within the first 15 minutes. Batefenterol was less potent and less balanced than CHF6366, exerting maximal inhibition at 3nmol/kg with a predominance of β2 activity. CHF6366 1nmol/kg, administered 24 hours before spasmogens, retained a robust and balanced residual activity (% inhibition MABA: 70.9±7.4; M3: 45.3±9.2; β2: 51.4±6.0). On the contrary, with batefenterol a slight overdose (10nmol/kg) was necessary to achieve an appreciable residual effect at 24 hours. Both CHF6366 and batefenterol did not induce significant lowering of blood potassium levels at bronchoprotective doses. In conclusion, CHF6366 is a new inhaled MABA compound with a favourable in vivo pharmacological profile holding promise as a new effective dual-acting bronchodilator for COPD.

Neutrophil elastase (NE) is a key proteolytic enzyme implicated in the pathogenesis of chronic inflammatory lung diseases, including chronic obstructive pulmonary disease (COPD), bronchiectasis and cystic fibrosis (CF). Here we studied the pharmacological profile of CHI25243, a novel NE inhibitor designed for inhaled treatment of NE-driven lung diseases. In in vitro assays, CHI25243 shows a high potency in inhibiting human NE (IC 50 =0.2nM), conserved across species (rat, IC 50 =4.1nM; dog, IC 50 =7.9nM), with more than 100-fold selectivity against other proteases. The in vivo effects of CHI25243 were studied in two rat models: human NE-induced lung injury and LPS-fMLP induced NE release. When administered intratracheally (i.t.), CHI25243 prevents human NE-induced lung haemorrhage with an ED 50 of 74µg/kg, while in the LPS-fMLP model it shows a potent, dose-dependent and long-lasting (up to 24h) inhibition of elastase activity (with 30µg/kg being the minimum dose providing 24h efficacy). In addition, CHI25243 shows efficacy in relevant disease models. Specifically, it significantly reduces lung neutrophil recruitment induced by 4-day cigarette smoke exposure in mice with a minimum effective dose (MED) of 100µg/kg. CHI25243administered i.t. to P. aeruginosa lung infected rats for 7 days, significantly reduces both lung tissue infection and inflammation with a MED of 100 µg/kg. Our data support the development of CHI25243, a novel potent and selective inhaled NE inhibitor, which is capable of inhibiting both pulmonary inflammation and infection in various preclinical animal models, thus suggesting the potential for the treatment of chronic inflammatory lung diseases.

Introduction: Myopathy is the most common side effect of statins. Because nitric oxide (NO) has a key role in regulating skeletal muscle function, we studied whether the NO-donating atorvastatin NCX 6560 could show a better profile on skeletal muscle function and structure compared with ator- vastatin. Methods: C57BL/6 mice received atorvastatin 40 mg/ kg/day or an equivalent dose of NCX 6560 for 2 months. Mus- cle function assessed by treadmill test, serum creatine kinase (CK) activity, citrate synthase (CS) activity, and muscle histol- ogy were evaluated. Results: Atorvastatin significantly (P < 0.001) reduced muscle endurance, increased serum CK by 6- fold, and induced muscle fiber atrophy. Conversely, NCX 6560 preserved muscle function, prevented CK increase and did not modify muscle structure. Interestingly, atorvastatin reduced CS activity, a marker for mitochondrial function, in gastrocnemius, diaphragm, and heart, whereas NCX 6560 prevented such decrease. Conclusions: These findings suggest that NO may prevent statin-induced myopathy. Muscle Nerve 47: 72-80, 2013

Graphical abstract, A resolutive therapy for muscular dystrophies, a heterogeneous group of genetic diseases leading to muscular degeneration and in the severe forms to death, is still lacking. Since inflammation and defects in nitric oxide generation are recognized key pathogenic events in muscular dystrophy, we have analysed the effects of a derivative of ibuprofen, NCX 320, belonging to the class of cyclooxygenase inhibiting nitric oxide donator (CINOD), in the α-sarcoglycan null mice, a severe mouse model of dystrophy. NCX 320 was administered daily in the diet for 8 months starting 1 month from weaning. Muscle functional recovery was evaluated by free wheel and treadmill tests at 8 months. Serum creatine kinase activity, as well as the number of diaphragm inflammatory infiltrates and necrotic fibres, was measured as indexes of skeletal muscle damage. Muscle regeneration was evaluated in diaphragm and tibialis anterior muscles, measuring the numbers of centronucleated fibres and of myogenic precursor cells. NCX 320 mitigated muscle damage, reducing significantly serum creatine kinase activity, the number of necrotic fibres and inflammatory infiltrates. Moreover, NCX 320 stimulated muscle regeneration increasing significantly the number of myogenic precursor cells and regenerating fibres. All these effects concurred in inducing a significant improvement of muscle function, as assessed by both free wheel and treadmill tests. These results describe the properties of a new compound incorporating nitric oxide donation together with anti-inflammatory properties, showing that it is effective in slowing muscle dystrophy progression long term. Of importance, this new compound deserves specific attention for its potential in the therapy of muscular dystrophy given that ibuprofen is well tolerated in paediatric patients and with a profile of safety that makes it suitable for chronic treatment such as the one required in muscular dystrophies.

Hypoxia of the retina and optic nerve head (ONH) is believed to be pivotal in the development of ocular vascular disorders, including diabetic macular edema (DME). Glucocorticoids are among the most effective agents for the treatment of back of the eye diseases. However, this class of compounds is highly liable to increase intraocular pressure (IOP) and does not improve ocular perfusion or tissue oxygenation. Nitric oxide (NO) has vasodilating properties and lowers IOP in experimental models and humans, suggesting that its properties might complement those of glucocorticoids. NCX 434 is an NO-donating triamcinolone acetonide (TA) that is less likely to increase IOP while targeting both the vascular and inflammatory components of DME.NCX 434 was studied in vitro with respect to its NO-releasing properties in isolated methoxamine-precontracted rabbit aortic rings and glucocorticoid-like activity in recombinant human glucocorticoid receptors. IOP and oxygen saturation in the ONH and overlaying arteries and veins were studied in the anesthetized cynomolgus monkey. Measurements were taken using, respectively, an applanation tonometer and a hyperspectral imaging system before and 7, 14, 21, 31 and 41 days after the intravitreal injection of NCX 434 (5.8 mg/eye) or TA equimolar doses (4.0 mg/eye).NCX 434 inhibited (3)H-dexamethasone-specific binding (IC(50)=34±5 nM) on human glucocorticoid receptors and elicited NO-dependent aortic ring relaxation (EC(50) of 0.5±0.1 μM, E(max) 98.9%). In monkey eyes, NCX 434 enhanced, whereas TA did not, oxygen saturation in various ONH areas (*P0.05 vs. basal), decreased it in veins, and did not affect it in the overlaying arteries. Neither NCX 434 nor TA altered IOP significantly at all time points. However, at 31 days post-treatment TA appeared to start increasing IOP (Δ(IOP)=+3.31±0.51 mmHg, 30.8%, over baseline, NS).NCX 434 enhances ocular tissue oxygenation. This feature appears to depend on its NO-donating properties; thus, the compound deserves to be further investigated for the treatment of DME and other ocular disorders with impaired ocular perfusion.

Novel bi-functional compounds with a nitric oxide (NO)-releasing moiety bound to a dorzolamide scaffold were investigated. Several compounds were synthesized and their activity as selective carbonic anhydrase inhibitors (CAI) evaluated in vitro on recombinant hCA type I, II and IV enzyme isoforms where they showed different degrees of potency and selectivity to hCA II. A high resolution X-ray crystal structure for the CA II adduct with 8 confirmed the high affinity of this class of compounds for the enzyme. Compounds 4, 6, and 8 showed highly potent and efficacious NO-mediated properties as assessed by their vascular relaxant effect on methoxamine-precontracted rabbit aortic rings. Finally, compounds 4 and 6 exerted potent intraocular pressure (IOP) lowering effects in vivo in normotensive rabbits thereby anticipating their potential for the treatment of hypertensive glaucoma.

Background In dystrophin-deficient muscles of Duchenne Muscular Dystrophy (DMD) patients and the mdx mouse model, nitric oxide (NO) signalling is impaired. Previous studies have shown that NO-donating drugs are beneficial in dystrophic mouse models. Recently, a long-term treatment (9 months) of mdx mice with naproxcinod, an NO-donating naproxen, has shown a significant improvement of the dystrophic phenotype with beneficial effects present throughout the disease progression. It remains however to be clearly dissected out which specific effects are due to the NO component compared with the anti-inflammatory activity associated with naproxen. Understanding the contribution of NO vs the anti-inflammatory effect is important, in view of the potential therapeutic perspective, and this is the final aim of this study. Methods Five-week-old mdx mice received either naproxcinod (30 mg/kg) or the equimolar dose of naproxen (20 mg/kg) in the diet for 6 months. Control mdx mice were used as reference. Treatments (or vehicle for control groups) were administered daily in the diet. For the first 3 months the study was performed in sedentary animals, then all mice were subjected to exercise until the sixth month. Skeletal muscle force was assessed by measuring whole body tension in sedentary animals as well as in exercised mice and resistance to fatigue was measured after 3 months of running exercise. At the end of 6 months of treatment, animals were sacrificed for histological analysis and measurement of naproxen levels in blood and skeletal muscle. Results Naproxcinod significantly ameliorated skeletal muscle force and resistance to fatigue in sedentary as well as in exercised mice, reduced inflammatory infiltrates and fibrosis deposition in both cardiac and diaphragm muscles. Conversely, the equimolar dose of naproxen showed no effects on fibrosis and improved muscle function only in sedentary mice, while the beneficial effects in exercised mice were lost demonstrating a limited and short-term effect. Conclusion In conclusion, this study shows that NO donation may have an important role, in addition to anti-inflammatory activity, in slowing down the progression of the disease in the mdx mouse model therefore positioning naproxcinod as a promising candidate for treatment of DMD. Electronic supplementary material The online version of this article (doi:10.1186/s13023-015-0311-0) contains supplementary material, which is available to authorized users.

Impaired nitric oxide (NO) signaling in the dystrophin-deficient muscles of patients with Duchenne muscular dystrophy (DMD) and the mdx mouse model of DMD renders the diseased muscles susceptible t...

In Duchenne muscular dystrophy (DMD) patients and the mouse model of DMD, mdx, dystrophin deficiency causes a decrease and mislocalization of muscle-specific neuronal nitric oxide synthase (nNOSμ), leading to functional impairments. Previous studies have shown that nitric oxide (NO) donation associated with anti-inflammatory action has beneficial effects in dystrophic mouse models. In this study, we have systematically investigated the effects of naproxcinod, an NO-donating naproxen derivative, on the skeletal and cardiac disease phenotype in mdx mice. Four-week-old mdx and C57BL/10 mice were treated with four different concentrations (0, 10, 21 and 41 mg/kg) of naproxcinod and 0.9 mg/kg of prednisolone in their food for 9 months. All mice were subjected to twice-weekly treadmill sessions, and functional and behavioral parameters were measured at 3, 6 and 9 months of treatment. In addition, we evaluated in vitro force contraction, optical imaging of inflammation, echocardiography and blood pressure (BP) at the 9-month endpoint prior to sacrifice. We found that naproxcinod treatment at 21 mg/kg resulted in significant improvement in hindlimb grip strength and a 30% decrease in inflammation in the fore- and hindlimbs of mdx mice. Furthermore, we found significant improvement in heart function, as evidenced by improved fraction shortening, ejection fraction and systolic BP. In addition, the long-term detrimental effects of prednisolone typically seen in mdx skeletal and heart function were not observed at the effective dose of naproxcinod. In conclusion, our results indicate that naproxcinod has significant potential as a safe therapeutic option for the treatment of muscular dystrophies.

Polymorphonuclear neutrophils, the first leukocytes to infiltrate the inflamed tissue, can make important contributions to vascular inflammatory processes driving the development of atherosclerosis. We herein investigated the effects of atorvastatin and NCX 6560 (a nitric oxide (NO)-donating atorvastatin derivative that has completed a successful phase 1b study) on neutrophilic inflammation in carotid arteries of normocholesterolemic rabbits subjected to perivascular collar placement. Atorvastatin or NCX 6560 were administered orally (5 mg/kg/day or equimolar dose) to New Zealand White rabbits for 6 days, followed by collar implantation 1 h after the last dose. Twenty-four hours later carotids were harvested for neutrophil quantification by immunostaining. Treatment with NCX 6560 was associated with a lower neutrophil infiltration (−39.5 %), while atorvastatin did not affect neutrophil content. The result was independent of effects on plasma cholesterol or differences in atorvastatin bioavailability, which suggests an important role of NO-related mechanisms in mediating this effect. Consistent with these in vivo findings, in vitro studies showed that NCX 6560, as compared to atorvastatin, had greater inhibitory activity on processes involved in neutrophil recruitment, such as migration in response to IL-8 and IL-8 release by endothelial cells and by neutrophils themselves. Pretreatment with NCX 6560, but not with atorvastatin, reduced the ability of neutrophil supernatants to promote monocyte chemotaxis, a well-known pro-inflammatory activity of neutrophils. Experimental data suggest a potential role of NO-releasing statins in the control of the vascular inflammatory process mediated by polymorphonuclear neutrophils.

Myopathy is the most common side effect of statins. Because nitric oxide (NO) has a key role in regulating skeletal muscle function, we studied whether the NO-donating atorvastatin NCX 6560 could show a better profile on skeletal muscle function and structure compared with atorvastatin.C57BL/6 mice received atorvastatin 40 mg/kg/day or an equivalent dose of NCX 6560 for 2 months. Muscle function assessed by treadmill test, serum creatine kinase (CK) activity, citrate synthase (CS) activity, and muscle histology were evaluated.Atorvastatin significantly (P0.001) reduced muscle endurance, increased serum CK by 6-fold, and induced muscle fiber atrophy. Conversely, NCX 6560 preserved muscle function, prevented CK increase and did not modify muscle structure. Interestingly, atorvastatin reduced CS activity, a marker for mitochondrial function, in gastrocnemius, diaphragm, and heart, whereas NCX 6560 prevented such decrease.These findings suggest that NO may prevent statin-induced myopathy.

Aims The aim of the present study was to assess whether the addition of a nitric oxide (NO)-donating moiety to atorvastatin enhances anti-inflammatory and anti-atherogenic effects in an animal model of endothelial dysfunction, systemic peroxidation and inflammation, and accelerated atherosclerosis. Methods and results Low-density lipoprotein receptor (LDLR)−/− mice kept on a high-fat diet (HFD) for 16 weeks underwent photochemical injury to the femoral artery with the local production of oxygen radicals. HFD markedly enhanced cholesterol, inflammatory biomarkers in plasma and in the femoral arterial wall, and atherosclerotic lesions in the aortic arch; inflammation and atherosclerosis were further increased by photochemically generated oxygen radicals. Treatment with the NO-donating atorvastatin NCX 6560 (11.7 mg/kg) was significantly more effective than atorvastatin (10 mg/kg) in reducing the following parameters: lipid-rich lesions in the aortic arch (surface covered: atorvastatin = 24 ± 5%; NCX 6560 = 14.7 ± 3.9%; P < 0.05); the production of radical oxygen species in the aorta (dichlorofluorescein fluorescence intensity per milligram of protein: atorvastatin = 2419 ± 136.7; NCX 6560 = 1766 ± 161.2; P < 0.05); femoral artery intima/media thickness (atorvastatin = 1.2 ± 0.11; NCX 6560 = 0.3 ± 0.14; P < 0.05); circulating interleukin-6 (atorvastatin = 34.3 ± 6.8 pg/mL; NCX 6560 = 17.7 ± 14.4 pg/mL; P < 0.05); and matrix metalloproteinase 2 in the arterial wall (atorvastatin = 55.2 ± 1.9 ng/µg of proteins; NCX 6560 = 45.8 ± 2.6 ng/µg of proteins; P < 0.05). Conclusion In conditions of severe endothelial dysfunction, systemic peroxidation and inflammation, and accelerated atherosclerosis, atorvastatin, even at high doses, displays suboptimal anti-atherogenic and anti-inflammatory effects, while the addition of a NO-donating property confers enhanced anti-atherogenic and anti-inflammatory effects.

A novel class of timolol derivatives with nitric oxide (NO)-donating moieties achieved chemical stability yet under physiologically relevant conditions released timolol and NO. Hindered esters A were designed and synthesized, whose 'triggered' release relied on enzymatic hydrolysis of the nitrate ester in A to B, that in turn cyclized to liberate timolol.

Nitric oxide (NO) plays a critical role in skeletal muscle function, including control of blood flow and muscle repair. In muscular dystrophies, synthesis of NO in the skeletal muscle is known to be defective therefore contributing to damage progression. In the present study, we evaluated the effects of naproxcinod, an NO-donating anti-inflammatory compound, in two models of muscular dystrophy, the alpha-sarcoglycan (alpha-SG) null mice, a model for limb-girdle muscle dystrophy, and the mdx mouse model for Duchenne muscle dystrophy (DMD). Naproxcinod (10 and 30 mg/kg/day) was orally administered for 7 months to mdx mice and for 4 months to alpha-SG null mice starting at 4 weeks of age. Muscle function was assessed by treadmill test at 4 months (both mdx and alpha-SG null mice) and 7 months of treatment (mdx mice). Serum creatine kinase (CK) was measured as index of skeletal muscle damage. Inflammatory infiltrates, as well as muscle regeneration were studied in diaphragm and tibialis anterior muscles. In mdx mice 30 mg/kg/day of naproxcinod significantly improved muscle function in terms of resistance to exercise with a recovery score of 12%; significantly reduced skeletal muscle inflammation and serum CK activity; and increased muscle regeneration. Likewise in alpha-SG null mice, 4 months of naproxcinod treatment led to a significant improvement of resistance to fatigue (recovery score of 59%), and to reduction of muscle inflammation and damage. Furthermore, both diaphragm and tibialis anterior muscles appeared fully regenerated, thus supporting the marked beneficial effects observed for muscle function. The results demonstrate that naproxcinod, through NO donation together with anti-inflammatory activity, produces significant and persistent therapeutic effects improving muscle function, reducing muscle inflammation and maintaining regeneration capacity of the muscle in two models of muscular dystrophies.

There is evidence that nitric oxide (NO) plays a critical role in skeletal muscle. In Duchenne muscular dystrophy (DMD) patients and the mdx mouse model of DMD, dystrophin deficiency causes a decrease and mislocalization of muscle-specific neuronal nitric oxide synthase (nNOSμ), leading to a variety of functional impairments such as muscle ischemia and compromised myogenesis. Previous studies have shown that NO donation associated with anti-inflammatory action showed beneficial effects in dystrophic mouse models. In this study, we have investigated the effects of naproxcinod, an NO donating naproxen, on skeletal and cardiac muscle function in mdx mice. 4-week-old mdx mice were orally treated for 9 months with three different doses of naproxcinod (10, 21 and 41 mg/kg/day) compared with 0.9 mg/kg of prednisolone. Functional and behavioral parameters using a grip strength meter and open field digiscan were measured at 3, 6, and 9 months of treatment using SOPs developed by TREAT-NMD. Additionally, in vitro EDL force contraction, optical imaging of inflammation, echocardiography and blood pressure were evaluated at the 9 months prior to sacrifice. Naproxcinod treatment at 10 and 21 mg/kg resulted in significant improvements in hindlimb grip strength as well as approximately a 25–30% decrease in inflammation in fore and hind limbs measured by in vivo optical imaging in mdx mice. Naproxcinod induced significant improvements in heart function as evidenced by ameliorated fraction shortening and ejection fraction measured using echocardiography along with improvements in systolic blood pressure. Moreover, the long term detrimental effects of prednisolone typically observed in mdx skeletal and heart function were not observed at the effective doses of naproxcinod. In conclusion, naproxcinod seems to have significant potential as a safe therapeutic option for the treatment of muscular dystrophies.