Amyloid toxicity in skeletal myoblasts: Implications for inclusion-body myositis.
Skeletal muscle disorder, inclusion-body myositis (IBM) has been known for accumulation of amyloid characteristic proteins in muscle. To understand the biophysical basis of IBM, the interaction of amyloid fibrils with skeletal myoblast cells (SMC) has been studied in vitro. Synthetic insulin fibrils and Abeta(25-35) fibrils were used for this investigation. From the saturation binding analysis, the calculated dissociation constant (K(d)) for insulin fibril and Abeta(25-35) fibrils were 69.37+/-11.17nM and 115.60+/-12.17nM, respectively. The fibrillar insulin comparatively has higher affinity binding to SMC than Abeta fibrils. The competitive binding studies with native insulin showed that the amount of bound insulin fibril was significantly decreased due to displacement of native insulin. However, the presence of native insulin is not altered the binding of beta-amyloid fibril. The cytotoxicity of insulin amyloid intermediates was measured. The pre-fibrillar intermediates of insulin showed significant toxicity (35%) as compared to matured fibrils. Myoblast treated with beta-amyloid fibrils showed more oxidative damage than the insulin fibril. Cell differentiating action of amyloidic insulin was assayed by creatine kinase activity. The insulin fibril treated cells differentiated more slowly compared to native insulin. However, beta-amyloid fibrils do not show cell differentiation property. These findings reinforce the hypothesis that accumulation of amyloid related proteins is significant for the pathological events that could lead to muscle degeneration and weakness in IBM.
Jayaraman
Murali
M
Bioorganic and Neurochemistry Laboratory, Central Leather Research Institute, Adyar, Chennai 600 020, India.
Kannayiram
Gomathi
G
Rajadas
Jayakumar
J
D016229:Amyloid beta-Peptides / Q000378:metabolism / Q000502:physiology*; D000818:Animals; D002478:Cells, Cultured; D003402:Creatine Kinase / Q000378:metabolism; D005452:Fluoresceins / Q000737:chemistry; D018482:Muscle, Skeletal / Q000201:enzymology / Q000503:physiopathology*; D018979:Myositis, Inclusion Body / Q000503:physiopathology*; D018384:Oxidative Stress; D010446:Peptide Fragments / Q000378:metabolism / Q000502:physiology*; D011485:Protein Binding; D051381:Rats; D017208:Rats, Wistar; D013050:Spectrometry, Fluorescence
D016229:Amyloid beta-Peptides; D005452:Fluoresceins; D010446:Peptide Fragments; C074636:amyloid beta-protein (25-35); C037631:2',7'-dichlorofluorescein; D003402:Creatine Kinase