Exercise attenuates polyglutamine‐mediated neuromuscular degeneration in a mouse model of spinal and bulbar muscular atrophy

Abstract Background Spinal and bulbar muscular atrophy (SBMA) is a hereditary neuromuscular disorder caused by the expansion of trinucleotide cytosine–adenine–guanine (CAG) repeats, which encodes a polyglutamine (polyQ) tract in the androgen receptor (AR) gene. Recent evidence suggests that, in addition to motor neuron degeneration, defective skeletal muscles are also the primary contributors to the pathogenesis in SBMA. While benefits of physical exercise have been suggested in SBMA, underlying mechanism remains elusive. Methods We investigated the effect of running exercise in a transgenic mouse model of SBMA carrying human AR with 97 expanded CAGs (AR97Q). We assigned AR97Q mice to exercise and sedentary control groups, and mice in the exercise group received 1‐h forced running wheel (5 m/min) 5 days a week for 4 weeks during the early stage of the disease. Motor function (grip strength and rotarod performance) and survival of each group were analysed, and histopathological and biological features in skeletal muscles and motor neurons were evaluated. Results AR97Q mice in the exercise group showed improvement in motor function (~40% and ~50% increase in grip strength and rotarod performance, respectively, P < 0.05) and survival (median survival 23.6 vs. 16.7 weeks, P < 0.05) with amelioration of neuronal and muscular histopathology (~1.4‐fold and ~2.8‐fold increase in motor neuron and muscle fibre size, respectively, P < 0.001) compared to those in the sedentary group. Nuclear accumulation of polyQ‐expanded AR in skeletal muscles and motor neurons was suppressed in the mice with exercise compared to the sedentary mice (~50% and ~30% reduction in 1C2‐positive cells in skeletal muscles and motor neurons, respectively, P < 0.05). We found that the exercise activated 5′‐adenosine monophosphate‐activated protein kinase (AMPK) signalling and inhibited mammalian target of rapamycin pathway that regulates protein synthesis in skeletal muscles of SBMA mice. Pharmacological activation of AMPK inhibited protein synthesis and reduced polyQ‐expanded AR proteins in C2C12 muscle cells. Conclusions Our findings suggest the therapeutic potential of exercise‐induced effect via AMPK activation in SBMA.


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Figure S1 Study design and exercise intensity.(A) Aerobic exercise (5 m/min) was performed from 5 to 9 weeks of age in wild-type (WT) and AR97Q mice, and the phenotype of the mice was analyzed throughout the entire period.(B) Expression levels of citrate synthase (CS) in the quadriceps of WT and AR97Q mice after 4 weeks of 5 m/min exercise.

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Figure S2 Effect of exercise on behavior of wild-type mice.Exercise was started from 5 weeks of age and maintained for 4 weeks in wild-type (WT) mice.(A) Body weight, (B) grip strength, (C) rotarod performance, and (D) survival rate of WT mice in the sedentary (Sed, n = 14) and exercise (Ex, n = 15) groups are shown.Error bars indicate the standard error of the mean.

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Figure S3 Analysis of microarray data from the quadriceps of AR97Q mice at 9 weeks of age between the sedentary and exercise groups.(A) Principal component analysis.(B) GO term enrichment analysis of differentially expressed genes (DEGs) between the sedentary and early exercise groups (n = 3 per group).Top five GO terms (Cellular Component) of DEGs (false discovery rate < 0.05, fold-change ≤ 0.5 or ≥ 2) are listed.All of the top five categories are upregulated in the exercise group.The underlined genes are specifically expressed in fast-twitch skeletal muscles.Sed, sedentary; Ex, exercise.

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Figure S4 Differentially expressed genes of microarray data from the quadriceps of AR97Q mice between the sedentary and exercise groups.The top 30 differentially expressed genes that are up-(A) or downregulated (B).

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Figure S5 KEGG AMPK signaling pathway in the quadriceps of AR97Q mice.KEGG pathway analysis is performed by iDEP using the microarray data.Red and green represent up-and downregulated genes in the exercise group compared to the sedentary group, respectively.Red and blue circles indicate mitochondrial biogenesis and protein synthesis pathway, respectively.

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Figure S6 Autophagy, Igf-1/Akt signaling, and BDNF expression are not elevated by exercise in skeletal muscles of AR97Q.(A) Log fold-change (FC) of expression levels (Ex/Sed) of HGNC (https://www.genenames.org/)autophagy-related genes from microarray data of the quadriceps of AR97Q mice at 9 weeks of age.Red and blue represent up-and downregulated genes in the exercise group, respectively.(B) Immunoblots for Becn1 and LC3 of the quadriceps of WT and AR97Q mice (n = 3 per group).(C) Log FC (Ex/Sed) and false discovery rate (FDR) of Igf1 and Bdnf expression levels from microarray data of the quadriceps of AR97Q mice.(D) Immunoblots for phosphorylated (Ser-473) and total Akt of the quadriceps of AR97Q mice (n = 5 per group).Sed, sedentary; Ex, exercise, *P < 0.05 by unpaired t-test.