Synergistic effect of β-amyloid protein and interferon gamma on nitric oxide production by C2C12 muscle cells

Abstract

Nitric oxide (NO) is an important mediator of diverse physiological and pathological responses. NO-induced oxidative stress has been proposed in the pathogenesis of muscle tissue damage in inclusion-body myositis (IBM), which is characterized by deposition of β-amyloid protein (Aβ) in vacuolated muscle fibres. To determine whether Aβ can induce NO production in skeletal muscle, we stimulated C2C12 mouse skeletal muscle cells in vitro with Aβ[1-42] or Aβ[25-35] peptides in the presence or absence of interferon gamma (IFN-γ). Neither Aβ peptides nor IFN-γ were able to stimulate nitrite (NO2-) production by C2C12 cells when given alone. However, combination of IFN-γ with either Aβ[1-42] or Aβ[25-35] resulted in significant NO2- release into cell-free supernatants. Northern blot analysis of RNA obtained from Aβ/IFN-γ-stimulated C2C12 cells revealed increased mRNA accumulation of inducible nitric oxide synthase (iNOS). Moreover, ~4% of muscle cells incubated with Aβ peptides and IFN-γ showed ultrastructural features of DNA fragmentation. These findings, taken together, indicate that the association of Aβ with IFN-γ stimulates NO2- production via induction of iNOS gene expression in skeletal muscle cells, with occasional evidence for nuclear changes suggesting apoptotic morphology. These data further support a role for Aβ deposition in the pathogenesis of postulated oxidative damage in IBM.