Halofuginone and muscular dystrophy

Muscular dystrophies (MDs) include different inherited diseases that all result in progressive muscle degeneration, impaired locomotion and often premature death. The major focus of MD research has been on alleviating the primary genetic deficit - using gene therapy and myoblast-transfer approaches to promote expression of the deficient or mutated genes in the muscle fibers. Although promising, these approaches have not yet entered into clinical practice and unfortunately for MD patients, there is currently no cure. Thus, the development of complementary and supportive therapies that slow disease progression and improve patients' quality of life is critically important. The main features of MDs are sarcolemmal instability and increased myofiber vulnerability to mechanical stress, resulting in myofiber degeneration. Fibrosis, with progressive replacement of muscle tissue, is a prominent feature in some MDs, preventing complete regeneration and hampering muscle functions. TGFβ is the leading candidate for activating fibroblasts and eliciting overproduction of extracellular matrix (ECM) proteins. Halofuginone, an inhibitor of Smad3 phosphorylation downstream of TGFβ signaling, inhibits the activation of fibroblasts and their ability to synthesize ECM, regardless of their origin or location. In animal models of MDs with prominent muscle fibrosis, halofuginone treatment has resulted in both prevention of collagen production in young animals and resolution of established fibrosis in older ones: the reduction in muscle collagen content was associated with improved muscle histopathology and major improvements in muscle function. Recently, these halofuginone-dependent improvements were also observed in MD with minor fibrosis involvement, probably due to a direct effect of halofuginone on muscle cells, resulting in myotube fusion that is dependent on Akt and MAPK pathway activation. In summary, halofuginone improves muscle histopathology and muscle functions in various MDs, via inhibition of muscle fibrosis on the one hand, and increased myotube fusion on the other.     

Connective tissue metabolism in muscular dystrophy. Levels of collagen and mucopolysaccharides in embryonic chickens with genetic muscular dystrophy

There were marked differences between the levels of collagen (measured as hydroxyproline) and mucopolysaccharides (measured as hexosamine) found in embryonic chicks with genetic muscular dystrophy and their normal controls. The chief differences were that the dystrophic tissues (gastrocnemius muscle and tendon, pectoralis major and skin) had: (a) greater amounts of hexosamine early in embryonic development; (b) hydroxyproline levels that rose at a faster rate, yielding different slopes than their normal controls; (c) relatively greater amounts of hydroxyproline than hexosamine later in embryonic life (day 20). Connective tissue systems in muscles were preferentially affected. The connective tissue system associated with dystrophic tissues appeared to lag behind the normal rhythm pattern of embryological development. The changes in connective tissue metabolism observed in dystrophic chicks suggested that the collagen from dystrophic embryonic chicks may be of a different structure or composition than that found in the normals.     

Dystroglycan glycosylation and muscular dystrophy

Dystroglycan is an integral member of the skeletal muscle dystrophin glycoprotein complex, which links dystrophin to proteins in the extracellular matrix. Recently, a group of human muscular dystrophy disorders have been demonstrated to result from defective glycosylation of the α-dystroglycan subunit. Genetic studies of these diseases have identified six genes that encode proteins required for the synthesis of essential carbohydrate structures on dystroglycan. Here we highlight their known or postulated functions. This glycosylation pathway appears to be highly specific (dystroglycan is the only substrate identified thus far) and to be highly conserved during evolution.     

Oxidative stress and muscular dystrophy

Oxidative stress may be the fundamental basis of many of the structural, functional and biochemical changes characteristic of the inherited muscular dystrophies in animals and humans. The presence of by-products of oxidative damage, and the compensatory increases in cellular antioxidants, both indicate oxidative stress may be occurring in dystrophic muscle. Changes in the proportions and metabolism of cellular lipids, abnormal functions of cellular membranes, altered activity of membrane-bound enzymes such as the SR Ca2+-ATPase, disturbances in cellular protein turnover and energy production and a variety of other changes all indicate that these inherited muscular dystrophies appear more like the results of oxidative stress to muscle than any other type of underlying muscle disturbance. Particular details of these altered characteristics of dystrophic muscle, in combination with current knowledge on the processes of oxidative damage to cells, may provide some insight into the underlying biochemical defect responsible for the disease, as well as direct research towards the ultimate goal of an effective treatment.     

Myoblast senescence in muscular dystrophy

The limited proliferative capacity of normal diploid cells predicts that the utilization of cell divisions in vivo should reduce the lifespan of cells in culture. Because of the continuing demands for muscle regeneration in muscular dystrophy, myoblasts isolated from affected muscles should thus show a decrease in the number of cell divisions they are capable of expressing in culture. This hypothesis was tested by examining the proliferative capacity of myoblasts from different muscles for normal line 412 and dystrophic line 413 chickens of various ages. Prior to approx. 2 months of age, dystrophic myoblasts exhibited a relatively normal proliferative lifespan. By 5 months of age, myoblasts from the severely affected pectoralis major showed a 40% reduction in their proliferative potential, while myoblasts from the less affected posterior latissimus dorsi muscle showed a 25% decrease in their cultured lifespan. The time course of the appearance of a decreased proliferative capacity only after the disease has been clinically manifested strongly supports it representing a secondary response rather than it being an intrinsic property of dystrophic myoblasts. A hypothesis for manipulating the pattern of stem cell division in order to increase the mass of muscle produced from a constant number of cell divisions is presented. If myoblast senescence and the consequent failure of muscle regeneration is a contributing factor in the progressive deterioration of muscle function in the disease, then this hypothesis might provide an important therapeutic strategy for ameliorating the course of muscular dystrophy.     

Caveolae and caveolin-3 in muscular dystrophy

Caveolae are vesicular invaginations of the plasma membrane, and function as 'message centers' for regulating signal transduction events. Caveolin-3, a muscle-specific caveolin-related protein, is the principal structural protein of caveolar membrane domains in skeletal muscle and in the heart. Several mutations within the coding sequence of the human caveolin-3 gene (located at 3p25) have been identified. Mutations that lead to a loss of approximately 95% of caveolin-3 protein expression are responsible for a novel autosomal dominant form of limb-girdle muscular dystrophy (LGMD-1C) in humans. By contrast, upregulation of the caveolin-3 protein is associated with Duchenne muscular dystrophy (DMD). Thus, tight regulation of caveolin-3 appears essential for maintaining normal muscle health and homeostasis.