No abstract

A preclinical screening for prompt-to-use drugs that are safer than steroids and beneficial in Duchenne muscular dystrophy was performed. Compounds able to reduce calcium-induced degeneration (taurine or creatine 10% in chow) or to stimulate regeneration [insulin-like growth factor-1 (IGF-1); 50 or 500 g/kg s.c.] were administered for 4 to 8 weeks to mdx mice undergoing chronic exercise on a treadmill, a protocol to worsen dystrophy progression. ␣-Methyl-prednisolone (PDN; 1 mg/kg) was used as positive control. The effects were evaluated in vivo on forelimb strength and in vitro electrophysiologically on the macroscopic chloride conductance (gCl), an index of degeneration-regeneration events in mdx muscles, and on the mechanical threshold, a calcium-sensitive index of excitation-contraction coupling. The exercise produced a significant weakness and an impairment of gCl, by further decreasing the already low value of degenerating diaphragm (DIA) and fully hampering the increase of gCl typical of regenerating extensor digitorum longus (EDL) mdx muscle. The already negative voltage threshold for contraction of mdx EDL was also slightly worsened. Taurine Ͼ creatine Ͼ IGF-1 counteracted the exercise-induced weakness. The amelioration of gCl was drug-and muscle-specific: taurine was effective in EDL, but not in DIA muscle; IGF-1 and PDN were fully restorative in both muscles, whereas creatine was ineffective. An acute effect of IGF-1 on gCl was observed in vitro in untreated, but not in IGF-1-treated exercised mdx muscles. Taurine Ͼ PDN Ͼ IGF-1, but not creatine, significantly ameliorated the negative threshold voltage values of the EDL fibers. The results predict a potential benefit of taurine and IGF-1 for treating human dystrophy.

The mechanisms that regulate the formation of multinucleated muscle fibers from mononucleated myoblasts are not well understood. We show here that extracellular matrix (ECM) receptors of the beta1 integrin family regulate myoblast fusion. beta1-deficient myoblasts adhere to each other, but plasma membrane breakdown is defective. The integrin-associated tetraspanin CD9 that regulates cell fusion is no longer expressed at the cell surface of beta1-deficient myoblasts, suggesting that beta1 integrins regulate the formation of a protein complex important for fusion. Subsequent to fusion, beta1 integrins are required for the assembly of sarcomeres. Other ECM receptors such as the dystrophin glycoprotein complex are still expressed but cannot compensate for the loss of beta1 integrins, providing evidence that different ECM receptors have nonredundant functions in skeletal muscle fibers.

Erythropoietin (Epo) is required for the production of mature red blood cells. The requirement for Epo and its receptor (EpoR) for normal heart development and the response of vascular endothelium and cells of neural origin to Epo provide evidence that the function of Epo as a growth factor or cytokine to protect cells from apoptosis extends beyond the hematopoietic lineage. We now report that the EpoR is expressed on myoblasts and can mediate a biological response of these cells to treatment with Epo. Primary murine satellite cells and myoblast C2C12 cells, both of which express endogenous EpoR, exhibit a proliferative response to Epo and a marked decrease in terminal differentiation to form myotubes. We also observed that Epo stimulation activates Jak2/Stat5 signal transduction and increases cytoplasmic calcium, which is dependent on tyrosine phosphorylation. In erythroid progenitor cells, Epo stimulates induction of transcription factor GATA-1 and EpoR; in C2C12 cells, GATA-3 and EpoR expression are induced. The decrease in differentiation of C2C12 cells is concomitant with an increase in Myf-5 and MyoD expression and inhibition of myogenin induction during differentiation, altering the pattern of expression of the MyoD family of transcription factors during muscle differentiation. These data suggest that, rather than acting in an instructive or specific mode for differentiation, Epo can stimulate proliferation of myoblasts to expand the progenitor population during differentiation and may have a potential role in muscle development or repair.Erythropoietin (Epo) 1 is required for the development and maturation of erythroid cells and acts to stimulate the proliferation and differentiation of erythroid progenitor cells. Mice lacking expression of erythropoietin or its receptor die in utero due to insufficient erythropoiesis in the fetal liver (1). Erythropoietin production can be induced by hypoxia and provides physiologic regulation of the red cell mass. Erythropoietin receptor is a member of the cytokine receptor superfamily characterized by a single transmembrane domain, homology in the extracellular domain that includes a WSXWS motif, and a cytoplasmic domain that does not contain a kinase motif. Binding of erythropoietin to its receptor results in receptor dimerization, increased affinity for Jak2 to the receptor's membrane proximal region, and subsequent phosphorylation of Jak2 and tyrosines on the cytoplasmic region of the receptor (2). As with other members of this superfamily such as thrombopoietin, interleukin-3, granulocyte-macrophage colony-stimulating factor, and prolactin, Jak2 is required for signaling (3, 4) and Jak2 phosphorylation activates Stat5 (5, 6) and other signal transduction pathways. A role for calcium has been implicated in erythropoietin activity. For example, in erythroid progenitor cells, erythropoietin activates an increase in intracellular calcium in a dose-dependent manner mediated via tyrosine phosphorylation of the erythropoietin receptor requiring the cytoplasmic tyrosine 4...

Skeletal muscle atrophy occurs in aging and pathological conditions, including cancer, diabetes and AIDS. Treatment of atrophy is based on either preventing protein-degradation pathways, which are activated during atrophy, or activating protein-synthesis pathways, which induce muscle hypertrophy. Here we show that neuronal nitric oxide synthase (nNOS) regulates load-induced hypertrophy by activating transient receptor potential cation channel, subfamily V, member 1 (TRPV1). The overload-induced hypertrophy was prevented in nNOS-null mice. nNOS was transiently activated within 3 min after overload. This activation promoted formation of peroxynitrite, a reaction product of nitric oxide with superoxide, which was derived from NADPH oxidase 4 (Nox4). Nitric oxide and peroxynitrite then activated Trpv1, resulting in an increase of intracellular Ca(2+) concentration ([Ca(2+)](i)) that subsequently triggered activation of mammalian target of rapamycin (mTOR). Notably, administration of the TRPV1 agonist capsaicin induced hypertrophy without overload and alleviated unloading- or denervation-induced atrophy. These findings identify nitric oxide, peroxynitrite and [Ca(2+)](i) as the crucial mediators that convert a mechanical load into an intracellular signaling pathway and lead us to suggest that TRPV1 could be a new therapeutic target for treating muscle atrophy.