Analysis of serum miRNA profiles of myasthenia gravis patients.

Myasthenia gravis (MG) is an autoimmune disease characterized by the presence of autoantibodies, mainly against the acetylcholine receptor (AChR). The mechanisms triggering and maintaining this chronic disease are unknown. MiRNAs are regulatory molecules that play a key role in the immune system and are altered in many autoimmune diseases. The aim of this study was to evaluate miRNA profiles in serum of 61 AChR MG patients. We studied serum from patients with early onset MG (n = 22), late onset MG (n = 27) and thymoma (n = 12), to identify alterations in the specific subgroups. In a discovery cohort, we analysed 381 miRNA arrays from 5 patients from each subgroup, and 5 healthy controls. The 15 patients had not received any treatment. We found 32 miRNAs in different levels in MG and analysed 8 of these in a validation cohort that included 46 of the MG patients. MiR15b, miR122, miR-140-3p, miR185, miR192, miR20b and miR-885-5p were in lower levels in MG patients than in controls. Our study suggests that different clinical phenotypes in MG share common altered mechanisms in circulating miRNAs, with no additional contribution of the thymoma. MG treatment intervention does not modify the profile of these miRNAs. Novel insights into the pathogenesis of MG can be reached by the analysis of circulating miRNAs since some of these miRNAs have also been found low in MG peripheral mononuclear cells, and have targets with important roles in B cell survival and antibody production.

Altered RIG-I/DDX58-mediated innate immunity in dermatomyositis.

We investigated the molecular mechanisms involved in the pathogenesis of three inflammatory myopathies, dermatomyositis (DM), polymyositis (PM) and inclusion body myositis (IBM). We performed microarray experiments(†) using microdissected pathological muscle fibres from 15 patients with these disorders and five controls. Differentially expressed candidate genes were validated by immunohistochemistry on muscle biopsies, and the altered pathways were analysed in human myotube cultures. Up-regulation of genes involved in viral and nucleic acid recognition were found in the three myopathies but not in controls. In DM, retinoic acid-inducible gene 1 (RIG-I, DDX58) and the novel antiviral factor DDX60, which promotes RIG-I-mediated signalling, were significantly up-regulated, followed by IFIH1 (MDA5) and TLR3. Immunohistochemistry confirmed over-expression of RIG-I in pathological muscle fibres in 5/5 DM, 0/5 PM and 0/5 IBM patients, and in 0/5 controls. Stimulation of human myotubes with a ligand of RIG-I produced a significant secretion of interferon-ß (IFNß; p < 0.05) and up-regulation of class I MHC, RIG-I and TLR3 (p < 0.05) by IFNß-dependent and TLR3-independent mechanisms. RIG-I-mediated innate immunity, triggered by a viral or damage signal, plays a significant role in the pathogenesis of DM, but not in that of PM or IBM.

Bone marrow transplantation in dysferlin-deficient mice results in a mild functional improvement.

Dysferlinopathies are caused by mutations in the DYSF gene. Dysferlin is a protein mainly expressed in the skeletal muscle and monocytes. Cell therapy constitutes a promising tool for the treatment of muscular dystrophies. The aim of our study was to evaluate the effect of bone marrow transplantation (BMT) using the A/J Dysf(prmd) mouse model of dysferlinopathy. For that purpose, we studied dysferlin expression by western blot and/or immunohistochemistry in transplanted mice and controls. Computerized analyses of locomotion and electrophysiological techniques were also performed to test the functional improvement. We observed dysferlin expression in splenocytes, but not in the skeletal muscle of the transplanted mice. However, the locomotion test, electromyography studies, and muscle histology showed an improvement in all transplanted mice that was more significant in the animals transplanted with dysferlin⁺/⁺ cells. In conclusion, although BMT restores dysferlin expression in monocytes, but not in skeletal muscle, muscle function was partially recovered. We propose that the slight improvement observed in the functional studies could be related with factors, such as the hepatocyte growth factor, released after BMT that prevented muscle degeneration.

The increase of pericyte population in human neuromuscular disorders supports their role in muscle regeneration in vivo.

Pericytes are periendothelial cells that have been involved in many different functions including a possible role as mesodermal stem/progenitor cells. In the present study we demonstrate that alkaline phosphatase (AP) expression is specific for human muscular pericytes and can be used as a marker to identify them in skeletal muscle biopsies. We studied the pericyte population in skeletal muscle biopsies from controls, myopathic and neuropathic patients. We observed a significant increase in the number of pericytes only in myopathies that correlated with the number of NCAM(+) fibres, suggesting that an active muscular degenerative/regenerative process is related to an increase in the pericyte population. AP(+) pericytes sorted from skeletal muscle samples were able to activate the myogenic programme and fuse with both mononucleate satellite cells and mature multinucleated myotubes in vitro, demonstrating that they could participate in muscle regeneration. In accordance, pericytes expressing the myogenic transcription factor MyoD were found in biopsies of myopathic biopsies. All these data support the hypothesis that, apart from satellite cells, pericytes may play an important role in muscle regeneration in adult human muscles in vivo.