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Item does not contain fulltext OBJECTIVE: Sporadic inclusion body myositis (sIBM) is an inflammatory myopathy characterized by both degenerative and autoimmune features. In contrast to other inflammatory myopathies, myositis-specific autoantibodies had not been found in sIBM patients until recently. We used human skeletal muscle extracts as a source of antigens to detect autoantibodies in sIBM and to characterize the corresponding antigen. METHODS: Autoantibodies to skeletal muscle antigens were detected by immunoblotting. The target antigen was immunoaffinity-purified from skeletal muscle extracts and characterized by mass spectrometry. A cDNA encoding this protein was cloned and expressed in vitro, and its recognition by patient sera was analyzed in an immunoprecipitation assay. Epitopes were mapped using microarrays of overlapping peptides. RESULTS: An Mr 44,000 polypeptide (Mup44) was frequently targeted by sIBM autoantibodies. The target protein was purified, and subsequent mass spectrometry analysis revealed that Mup44 is the cytosolic 5'-nucleotidase 1A (cN1A). By immunoprecipitation of recombinant cN1A, high concentrations of anti-Mup44 autoantibodies were detected in 33% of sIBM patient sera, whereas their prevalence in dermatomyositis, polymyositis, and other neuromuscular disorders appeared to be rare (4.2%, 4.5%, and 3.2%, respectively). Low concentrations of anti-Mup44 antibodies were found in myositis as well as other neuromuscular disorders, but not in healthy controls. Three major autoepitope regions of cN1A were mapped by using microarrays containing a set of overlapping peptides covering the complete cN1A amino acid sequence. INTERPRETATION: Anti-Mup44 autoantibodies, which are targeted to cN1A, represent the first serological biomarker for sIBM and may facilitate the diagnosis of this type of myositis.

Contains fulltext : 80248.pdf (Publisher’s version ) (Open Access) BACKGROUND: The Warburg phenotype in cancer cells has been long recognized, but there is still limited insight in the consecutive metabolic alterations that characterize its establishment. We obtained better understanding of the coupling between metabolism and malignant transformation by studying mouse embryonic fibroblast-derived cells with loss-of-senescence or H-RasV12/E1A-transformed phenotypes at different stages of oncogenic progression. RESULTS: Spontaneous immortalization or induction of senescence-bypass had only marginal effects on metabolic profiles and viability. In contrast, H-RasV12/E1A transformation initially caused a steep increase in oxygen consumption and superoxide production, accompanied by massive cell death. During prolonged culture in vitro, cell growth rate increased gradually, along with tumor forming potential in in vitro anchorage-independent growth assays and in vivo tumor formation assays in immuno-deficient mice. Notably, glucose-to-lactic acid flux increased with passage number, while cellular oxygen consumption decreased. This conversion in metabolic properties was associated with a change in mitochondrial NAD+/NADH redox, indicative of decreased mitochondrial tricarboxic acid cycle and OXPHOS activity. CONCLUSION: The high rate of oxidative metabolism in newly transformed cells is in marked contrast with the high glycolytic rate in cells in the later tumor stage. In our experimental system, with cells growing under ambient oxygen conditions in nutrient-rich media, the shift towards this Warburg phenotype occurred as a step-wise adaptation process associated with augmented tumorigenic capacity and improved survival characteristics of the transformed cells. We hypothesize that early-transformed cells, which potentially serve as founders for new tumor masses may escape therapies aimed at metabolic inhibition of tumors with a fully developed Warburg phenotype.