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Autosomal recessive hereditary inclusion-body myopathy (h-IBM) is caused by mutations of the UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase gene, a rate-limiting enzyme in the sialic acid metabolic pathway. Previous studies have demonstrated an abnormal sialylation of glycoproteins in h-IBM. h-IBM muscle shows the abnormal accumulation of proteins including amyloid-beta (Abeta). Neprilysin (NEP), a metallopeptidase that cleaves Abeta, is characterized by the presence of several N-glycosylation sites, and changes in these sugar moieties affect its stability and enzymatic activity. In the present study, we found that NEP is hyposialylated and its expression and enzymatic activity reduced in all h-IBM muscles analyzed. In vitro, the experimental removal of sialic acid by Vibrio Cholerae neuraminidase in cultured myotubes resulted in reduced expression of NEP. This was most likely because of a post-translational modification consisting in an abnormal sialylation of the protein that leads to its reduced stability. Moreover, treatment with Vibrio Cholerae neuraminidase was associated with an increased immunoreactivity for Abeta mainly in the form of distinct cytoplasmic foci within myotubes. We hypothesize that, in h-IBM muscle, hyposialylated NEP has a role in hampering the cellular Abeta clearing system, thus contributing to its abnormal accumulation within vulnerable fibers and possibly promoting muscle degeneration.

Neprilysin (NEP, EP24.11), a metallopeptidase originally shown to modulate signalling events by degrading small regulatory peptides, is also an amyloid-beta- (Abeta) degrading enzyme. We investigated a possible role of NEP in inclusion body myositis (IBM) and other acquired and hereditary muscle disorders and found that in all myopathies NEP expression was directly associated with the degree of muscle fibre regeneration. In IBM muscle, NEP protein was also strongly accumulated in Abeta-bearing abnormal fibres. In vitro, during the experimental differentiation of myoblasts, NEP protein expression was regulated at the post-transcriptional level with a rapid increase in the early stage of myoblast differentiation followed by a gradual reduction thereafter, coincident with the progression of the myogenic programme. Treatment of differentiating muscle cells with the NEP inhibitor dl-3-mercapto-2-benzylpropanoylglycine resulted in impaired differentiation that was mainly associated with an abnormal regulation of Akt activation. Therefore, NEP may play an important role during muscle cell differentiation, possibly through the regulation, either directly or indirectly, of the insulin-like growth factor I-driven myogenic programme. In IBM muscle increased NEP may be instrumental in (i) reducing the Abeta accumulation in vulnerable fibres and (ii) promoting a repair/regenerative attempt of muscle fibres possibly through the modulation of insulin-like growth factor I-dependent pathways.