Fig. 1

The IL-1β signaling pathway is contained and active in C2C12 myocytes. a C2C12 muscle cells were treated with human recombinant IL-1β (10 ng/ml) or vehicle for 30 min and 1 h. Immunocytochemistry with anti-IRAK1 antibody shows cytoplasmic-to-nuclear translocation of IRAK1 in response to IL-1β after 30 min. Nuclei were stained in blue (DAPI). Scale bar = 50 μm. b Synthetic luciferase reporters with multimerized NF-κB sites (NF-κB-Luc) were transfected into C2C12 (b, left panel) and HeLa (b, right panel) cells, together with LacZ as transfection control. Cells were treated with recombinant IL-1β (10 ng/ml) for 24 h. n = 3. c, d C2C12 cells were differentiated for 8 days and treated with human recombinant IL-1β (10 ng/ml) for different time points as indicated. qRT-PCR analysis of Il6 and Nlrp3. mRNA expression was normalized to Gapdh. All data are reported as fold change ± SEM. e–h IL-1β increases Nlrp3 expression and induces atrophy in differentiated C2C12 myocytes in vitro. C2C12 cells were differentiated for 8 days and treated with human recombinant IL-1β (10, 20, and 50 ng/ml) for 72 h. Dexamethasone (10 μM/ml) treatment was used as atrophy control. e Representative light microscopy pictures. Scale bar = 250 μm. f, g Frequency distribution histograms of cell width of IL-1β (10, 20, and 50 ng/ml) and dexamethasone-treated myotubes, as indicated, compared to vehicle-treated myotubes, n = 100 cells per condition. h Mean myotube width. Data are presented as mean ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001, ****p ≤ 0.0001