Diapocynin, a Dimer of the NADPH Oxidase Inhibitor Apocynin, Reduces ROS Production and Prevents Force Loss in Eccentrically Contracting Dystrophic Muscle

Abstract

Elevation of intracellular Ca2+, excessive ROS production and increased phospholipase A2 activity contribute to the pathology in dystrophin-deficient muscle. Moreover, Ca2+, ROS and phospholipase A2, in particular iPLA2, are thought to potentiate each other in positive feedback loops. NADPH oxidases (NOX) have been considered as a major source of ROS in muscle and have been reported to be overexpressed in muscles of mdx mice. We report here on our investigations regarding the effect of diapocynin, a dimer of the commonly used NOX inhibitor apocynin, on the activity of iPLA2, Ca2+ handling and ROS generation in dystrophic myotubes. We also examined the effects of diapocynin on force production and recovery ability of isolated EDL muscles exposed to eccentric contractions in vitro, a damaging procedure to which dystrophic muscle is extremely sensitive. In dystrophic myotubes, diapocynin inhibited ROS production, abolished iPLA2 activity and reduced Ca2+ influx through stretch-activated and store-operated channels, two major pathways responsible for excessive Ca2+ entry in dystrophic muscle. Diapocynin also prevented force loss induced by eccentric contractions of mdx muscle close to the value of wild-type muscle and reduced membrane damage as seen by Procion orange dye uptake. These findings support the central role played by NOX-ROS in the pathogenic cascade leading to muscular dystrophy and suggest diapocynin as an effective NOX inhibitor that might be helpful for future therapeutic approaches.

Apocynin inhibits the upregulation of TGF-β1 expression and ROS production induced by TGF-β in skeletal muscle cells

“…In this context, our data show that the use of apocynin partially inhibits the effect and signalling dependent on TGF-β, decreasing the NOX-induced ROS production. These findings are in agreement with previous reports in which the use of apocynin has beneficial and protective effects on the diaphragm during prolonged mechanical ventilation (McClung et al, 2009), in plantaris muscle after myocardial infarction (Bechara et al, 2014) by reducing ROS, and also prevented force loss of dystrophic muscle and reduced membrane damage (Ismail et al, 2014). Although the plant species from which apocynin has been isolated have not been used to evaluate their effect on muscle diseases, a previous report indicates that the treatment of diabetic rats, characterized by the loss of body weight and the increase of muscle wasting, with P. kurroa showed an increase on body weight compared to control animals, suggesting a protective role of this natural compound on muscle wasting (Husain et al, 2009).…”

“…Diapocynin, a dimer of apocynin, also prevented force loss of dystrophic muscle and reduced membrane damage (Ismail et al, 2014). These data suggest the importance of NOX-ROS axis in the pathogenic cascade leading to muscular diseases.…”

Pharmacological therapeutics targeting the secondary defects and downstream pathology of Duchenne muscular dystrophy

“…In dystrophic myotubes, diapocynin inhibited ROS production and reduced intracellular Ca 2+ overload through stretch-activated and store-operated channels. Diapocynin also prevented force loss after eccentric contractions and reduced membrane damage in mdx muscle 188 . Given the role of NOX in promoting oxidative stress, it would be of interest to assess diapocynin or another potent, efficacious, and selective NOX inhibitor in DMD patients.…”

Role of dystroglycan in limiting contraction-induced injury to the sarcomeric cytoskeleton of mature skeletal muscle

“…However, more recent reports have raised the possibility that the extent of contraction-induced injury may be affected indirectly by the absence of the DGC. Specifically, they demonstrated that, during either development or cycles of degeneration/regeneration, lack of the DGC leads to deleterious outcomes, such as inflammatory signaling (35,36) or abnormal localization/function of caveolae (37), membrane-bound enzyme complexes (38,39), and ion channels (40)(41)(42). These features have the potential to alter calcium influx/handling, the levels of reactive oxygen species, and cellular signaling, thereby exacerbating contraction-induced injury by increasing permeability of the sarcolemma, intensifying reactive oxygen species/calcium-mediated damage, and reducing release of calcium from the sarcoplasmic reticulum calcium (43).…”