Abnormal expression of nuclear proteins in aged mice. A, Schematic diagram of mouse diaphragm. NSR, non-SR. Green line indicates nerve terminals and red patches represent AChR clusters. B, Volcano plot for DE genes (2415 downregulated, in green; 2621 upregulated, in red) in 24 M diaphragm SR samples compared with samples from 3 M mice. Green and red dots indicated statistical DE genes. C, Heatmap of DE genes in 24 M SR samples compared with that from 3 M mice. Each row of the heatmap represents the z score transformed log₂ (1+FPKM) values of one DE gene across samples (blue, low expression; red, high expression). D, Venn diagrams of downregulated genes in this study and other aged muscle DE datasets, illustrating overlapped decreased genes (47 genes) among three datasets. E, GO enrichment analysis of overlapped genes in D. Downregulated genes in aged mice were enriched in cellular components including “nucleus,” “nucleoplasm,” and “nuclear membrane.” F, List of overlapped genes in D. G, Reduced mRNA level of Lmna determined by real-time PCR for TA muscle from 12 and 24 M mice compared with 3 M. Data were shown as mean ± SEM 3 versus 12 M, t₍₄₎ = 7.973, **p = 0.0013; 3 versus 24 M, t₍₄₎ = 5.724, **p = 0.0046. Unpaired t test, n = 3 mice per group. H, Decreased lamin A/C protein level in muscles from 12 and 24 M mice compared with 3 M. I, Quantitative analysis for data in H. Data were shown as mean ± SEM TA, t₍₂₎ = 7.974, *p = 0.0154 for 12 M, t₍₂₎ = 4.772, *p = 0.0412 for 24 M; EDL, t₍₂₎ = 7.435, *p = 0.0176 for 12 M, t₍₂₎ = 9.819, *p = 0.0102 for 24 M; solues, t₍₂₎ = 3.212, p = 0.0848 for 12 M, t₍₂₎ = 14.68, **p = 0.0046 for 24 M; GA, t₍₂₎ = 5.439, *p = 0.0322 for 12 M, t₍₂₎ = 5.986, *p = 0.0268 for 24 M; diaphragm, t₍₂₎ = 5.757, *p = 0.0289 for 12 M, t₍₂₎ = 6.224, *p = 0.0249 for 24 M. Paired t test, n = 3 mice per group. J, Decreased lamin A/C protein in SR and NSR from 24 M diaphragm versus 3 M. K, Quantification for data in J indicated lamin A/C declined more in SR (average of 47 ± 15%) than in NSR (average of 22 ± 7%) in aged muscles. Data were shown as mean ± SEM t₍₄₎ = 2.805, *p = 0.0486 for SR; t₍₄₎ = 2.818, *p = 0.0479 for NSR, unpaired t test, n = 3 mice per group. L, Compromised nuclear proteins in SR from 24 M diaphragm showed nuclear defects and increased DNA damage in subsynaptic nuclei. M, Quantification for data in L. Data were shown as mean ± SEM t₍₄₎ = 3.78, *p = 0.0194 for LAP2; t₍₄₎ = 4.006, *p = 0.0161 for HP1; t₍₄₎ = 9.823, ***p = 0.0006 for Tri-Me-H3K9; t₍₄₎ = 4.564, *p = 0.0103 for γH2AX; unpaired t test, n = 3 mice per group. ns, no significant difference.