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. 2016 Apr 1;7(17):23019–23032. doi: 10.18632/oncotarget.8552

Figure 1. ATGL is decreased in skeletal muscle of old mice.

Figure 1

A. Skeletal muscle of three young and three old mice was homogenized and 20 μg of total proteins were subjected to Western blot analysis of ATGL. Twenty μg of total proteins were derivatized with DNP and carbonylation was detected by Western blot with DNP antibody (upper panel). Five-hundred μg of total proteins were subjected to S-NO derivatization with biotin. After Western blot, biotin adducts were identified by incubating nitrocellulose membrane with HRP-conjugate streptavidin (middle panel). Twenty μg of total proteins were used to detect ubiquitinated proteins by immunoblot with an anti-ubiquitin monoclonal antibody (bottom panel). Tubulin was used as the loading control. All the immunoblots reported are from one experiment representative of four that gave similar results. B., C., D. Total RNA was isolated from skeletal muscle of three young and three old mice and relative mRNA levels of ATGL, IL-6, TNFα, Atrogin-1 and Murf-1 were analyzed by RT-qPCR. Data are expressed as means ± S.D. (n = 3, **p < 0.001, ***p < 0.0001 vs. young mice).