Abstract
X-linked myotubular myopathy (XLMTM) is a severe congenital disorder in male infants that leads to generalized skeletal muscle weakness and is frequently associated with fatal respiratory ...failure. XLMTM is caused by loss-of-function mutations in the MTM1 gene, which encodes myotubularin, the founder member of a family of 15 homologous proteins in mammals. We recently demonstrated the therapeutic efficacy of intravenous delivery of rAAV vectors expressing MTM1 in animal models of myotubular myopathy. Here, we tested whether the closest homologues of MTM1, MTMR1, and MTMR2 (the latter being implicated in Charcot-Marie-Tooth neuropathy type 4B1) are functionally redundant and could represent a therapeutic target for XLMTM. Serotype 9 recombinant AAV vectors encoding either MTM1, MTMR1, or MTMR2 were injected into the tibialis anterior muscle of Mtm1-deficient knockout mice. Two weeks after vector delivery, a therapeutic effect was observed with Mtm1 and Mtmr2, but not Mtmr1; with Mtm1 being the most efficacious transgene. Furthermore, intravenous administration of a single dose of the rAAV9-Mtmr2 vector in XLMTM mice improved the motor activity and muscle strength and prolonged survival throughout a 3-month study. These results indicate that strategies aiming at increasing MTMR2 expression levels in skeletal muscle may be beneficial in the treatment of myotubular myopathy.
Duchenne muscular dystrophy (DMD) associated cardiomyopathy is a major cause of morbidity and mortality. In an in vitro DMD cardiomyocyte model, nicorandil reversed stress-induced cell injury through ...multiple pathways implicated in DMD. We aimed to test the efficacy of nicorandil on the progression of cardiomyopathy in mdx mice following a 10-day treatment protocol.
A subset of mdx mice was subjected to low-dose isoproterenol injections over 5 days to induce a cardiac phenotype and treated with vehicle or nicorandil for 10 days. Baseline and day 10 echocardiograms were obtained to assess cardiac function. At 10 days, cardiac tissue was harvested for further analysis, which included histologic analysis and assessment of oxidative stress. Paired student's t test was used for in group comparison, and ANOVA was used for multiple group comparisons.
Compared to vehicle treated mice, isoproterenol decreased ejection fraction and fractional shortening on echocardiogram. Nicorandil prevented isoproterenol induced cardiac dysfunction. Isoproterenol increased cardiac fibrosis, which nicorandil prevented. Isoproterenol increased gene expression of NADPH oxidase, which decreased to baseline with nicorandil treatment. Superoxide dismutase 2 protein expression increased in those treated with nicorandil, and xanthine oxidase activity decreased in mice treated with nicorandil during isoproterenol stress compared to all other groups.
In conclusion, nicorandil is cardioprotective in mdx mice and warrants continued investigation as a therapy for DMD associated cardiomyopathy.
The Dlk1-Gtl2 imprinting locus is located on mouse distal chromosome 12 and consists of multiple maternally expressed non-coding RNAs and several paternally expressed protein-coding genes. The ...imprinting of this locus plays a crucial role in embryonic development and postnatal growth. At least one cis-element, the intergenic differentially methylated region (IG-DMR) is required for expression of maternally expressed genes and repression of silenced paternally expressed genes. The mechanism by which the IG-DMR functions is largely unknown. However, it has been suggested that the unmethylated IG-DMR acts as a positive regulator activating expression of non-coding RNAs. Gtl2 is the first non-coding RNA gene downstream of the IG-DMR. Although its in vivo function in the mouse is largely unknown, its human ortholog MEG3 has been linked to tumor suppression in human tumor-derived cell lines. We generated a knockout mouse model, in which the first five exons and adjacent promoter region of the Gtl2 gene were deleted. Maternal deletion of Gtl2 resulted in perinatal death and skeletal muscle defects, indicating that Gtl2 plays an important role in embryonic development. The maternal deletion also completely abolished expression of downstream maternally expressed genes, activated expression of silenced paternally expressed genes and resulted in methylation of the IG-DMR. By contrast, the paternal inherited deletion did not have this effect. These data strongly indicate that activation of Gtl2 and its downstream maternal genes play an essential role in regulating Dlk1-Gtl2 imprinting, possibly by maintaining active status of the IG-DMR.
Nebulin is a giant filamentous protein that is coextensive with the actin filaments of the skeletal muscle sarcomere. Nebulin mutations are the main cause of nemaline myopathy (NEM), with typical ...adult patients having low expression of nebulin, yet the roles of nebulin in adult muscle remain poorly understood. To establish nebulin's functional roles in adult muscle, we studied a novel conditional nebulin KO (Neb cKO) mouse model in which nebulin deletion was driven by the muscle creatine kinase (MCK) promotor. Neb cKO mice are born with high nebulin levels in their skeletal muscles, but within weeks after birth nebulin expression rapidly falls to barely detectable levels Surprisingly, a large fraction of the mice survive to adulthood with low nebulin levels (<5% of control), contain nemaline rods and undergo fiber-type switching toward oxidative types. Nebulin deficiency causes a large deficit in specific force, and mechanistic studies provide evidence that a reduced fraction of force-generating cross-bridges and shortened thin filaments contribute to the force deficit. Muscles rich in glycolytic fibers upregulate proteolysis pathways (MuRF-1, Fbxo30/MUSA1, Gadd45a) and undergo hypotrophy with smaller cross-sectional areas (CSAs), worsening their force deficit. Muscles rich in oxidative fibers do not have smaller weights and can even have hypertrophy, offsetting their specific-force deficit. These studies reveal nebulin as critically important for force development and trophicity in adult muscle. The Neb cKO phenocopies important aspects of NEM (muscle weakness, oxidative fiber-type predominance, variable trophicity effects, nemaline rods) and will be highly useful to test therapeutic approaches to ameliorate muscle weakness.
Dystrophinopathy is a class of genetic skeletal muscle disease characterized by myofiber degeneration and regeneration due to insufficient levels or functioning of dystrophin. Pathological evaluation ...for dystrophinopathy includes the identification of dystrophic skeletal muscle pathology and the immunohistochemical evaluation of dystrophin epitopes, but biopsies have become rare in recent years. However, the evaluation of dystrophin expression in the research setting has become critically important due to recent advances in genetic therapies, including exon skipping and gene therapy. Given the number of these therapies under evaluation in patients, it is likely that the traditional methods of evaluating dystrophinopathy will need to evolve in the near future. This review discusses current muscle biopsy diagnostic practices in dystrophinopathy and further focuses on how these practices have evolved in the context of therapeutic interventions for dystrophinopathy.
Highlights • Cardiomyopathy in BAG3 myofibrillar myopathy prior to onset of weakness. • Neuropathy associated with BAG3 overshadowing myopathy on EMG. • Severe axonal disease with ...demyelinating/remyelinating features.
Abstract
Nemaline myopathy (NM) is a heterogeneous congenital skeletal muscle disease with cytoplasmic rod-like structures (nemaline bodies) in muscle tissue. While weakness in NM is related to ...contractile abnormalities, myofiber smallness is an additional abnormality in NM that may be treatable. We evaluated the effects of mRK35 (a myostatin inhibitor developed by Pfizer) treatment in the TgACTA1D286G mouse model of NM. mRK35 induced skeletal muscle growth that led to significant increases in animal bodyweight, forelimb grip strength and muscle fiber force, although it should be noted that animal weight and forelimb grip strength in untreated TgACTA1D286G mice was not different from controls. Treatment was also associated with an increase in the number of tubular aggregates found in skeletal muscle. These findings suggest that myostatin inhibition may be useful in promoting muscle growth and strength in Acta1-mutant muscle, while also further establishing the relationship between low levels of myostatin and tubular aggregate formation.
Tetraspanins are a family of proteins known to assemble protein complexes at the cell membrane. They are thought to play diverse cellular functions in tissues by modifying protein-binding partners, ...thus bringing complexity and diversity in their regulatory networks. Previously, we identified the tetraspanin KAI/CD82 as a prospective marker for human muscle stem cells. CD82 expression appeared decreased in human Duchenne muscular dystrophy (DMD) muscle, suggesting a functional link to muscular dystrophy, yet whether this decrease is a consequence of dystrophic pathology or a compensatory mechanism in an attempt to rescue muscle from degeneration is currently unknown.
We studied the consequences of loss of CD82 expression in normal and dystrophic skeletal muscle and examined the dysregulation of downstream functions in mice aged up to 1 year.
Expression of CD82 is important to sustain satellite cell activation, as in its absence there is decreased cell proliferation and less efficient repair of injured muscle. Loss of CD82 in dystrophic muscle leads to a worsened phenotype compared to control dystrophic mice, with decreased pulmonary function, myofiber size, and muscle strength. Mechanistically, decreased myofiber size in CD82
dystrophic mice is not due to altered PTEN/AKT signaling, although increased phosphorylation of mTOR at Ser2448 was observed.
Basal CD82 expression is important to dystrophic muscle, as its loss leads to significantly weakened myofibers and impaired muscle function, accompanied by decreased satellite cell activity that is unable to protect and repair myofiber damage.
ABSTRACT We present a novel approach to studying the diversity of galaxies. It is based on a novel spectral graph technique, that of locally-biased semi-supervised eigenvectors. Our method introduces ...new coordinates that summarize an entire spectrum, similar to but going well beyond the widely used Principal Component Analysis (PCA). Unlike PCA, however, this technique does not assume that the Euclidean distance between galaxy spectra is a good global measure of similarity. Instead, we relax that condition to only the most similar spectra, and we show that doing so yields more reliable results for many astronomical questions of interest. The global variant of our approach can identify very finely numerous astronomical phenomena of interest. The locally-biased variants of our basic approach enable us to explore subtle trends around a set of chosen objects. The power of the method is demonstrated in the Sloan Digital Sky Survey Main Galaxy Sample, by illustrating that the derived spectral coordinates carry an unprecedented amount of information.
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