Overexpression of wild-type androgen receptor in muscle recapitulates polyglutamine disease

Abstract

We created transgenic mice that overexpress WT androgen receptor (AR) exclusively in their skeletal muscle fibers. Unexpectedly, these mice display androgen-dependent muscle weakness and early death, show changes in muscle morphology and gene expression consistent with neurogenic atrophy, and exhibit a loss of motor axons. These features reproduce those seen in models of Kennedy disease, a polyglutamine expansion disorder caused by a CAG repeat expansion in the AR gene. These findings demonstrate that toxicity in skeletal muscles is sufficient to cause motoneuron disease and indicate that overexpression of the WT AR can exert toxicity comparable with the polyglutamine expanded protein. This model has two clear implications for Kennedy disease: (i) mechanisms affecting AR gene expression may cause neuromuscular symptoms similar to those of Kennedy disease and (ii) therapeutic approaches targeting skeletal muscle may provide effective treatments for this disease.

Fig. 1.

Phenotype of HSA-AR male mice. (A) Photograph of L141 Tg male and age matched WT brother (top). L141 Tg males have reduced body weight (A and B), marked kyphosis (A) and motor deficits as revealed by paw print records (C) and the hang test (D). Despite reduced body weight, motor deficits are not observed in L78 Tg males, which express the AR transgene at a lower level than affected L141 Tg males (see SI Fig 6). Castrating L141 Tg males reverses deficits in hang time (E). Graphs represent mean ± SEM, open bars represent Tg males, and filled bars represent WT controls. *, significantly different from WT controls. WT 78 (n = 13, mean age in days: 294, range: 179–528), 78 Tg (n = 14–15, mean age: 281, range: 106–528), WT 141 (n = 12–15, mean age: 131, range: 72–218), Tg 141 (n = 6–7, mean age: 119, range: 72–179). Castrates: WT (n = 3, intact age: 80, castrate age: 90), Tg (n = 3, intact mean age: 74, range: 63–80, castrate mean age: 123, range: 90–190).

Fig. 2.

T treatment of asymptomatic 141 Tg females rapidly induces disease symptoms. Percent change relative to initial body weight, stride length during gait analysis, time on the hang test (seconds), and latency to fall on a constant speed rotarod during 9 days of T or vehicle (VEH) treatment of Tg or WT females. Motor dysfunction is therefore androgen-dependent in HSA-AR Tg mice, as in polyQ AR mice. *, significantly lower than Day 0 within a treatment group. Note that only the T-treated Tg females show a significant drop on these measures over time. However, T has no effect on fiber or axon number in L141 Tg females (ANOVAs indicate no significant effects of genotype or treatment, or a significant interaction), despite its debilitating effects on motor function. T does not have these effects on body weight or motor function of Tg L78 females (data not shown), paralleling the apparently normal behavioral phenotype of L78 Tg males. Means ± SEM of n = 7 mice/group (mean age/group: 102–103 days old; overall age range: 70–114 days old).

Fig. 3.

Histopathology in HSA-AR Male Mice. (A) Photomicrographs illustrating histopathology in EDL muscle sections stained with H&E or NADH from a WT male, or L78 or L141 Tg males. Muscle pathology seen in L78 and L141 Tg males is typical of that seen in KD, including grouped atrophic fibers (arrowheads), centralized nuclei (arrows), and increased NADH staining. Muscle pathology is notably greater in the symptomatic L141 Tg males than the nonsymptomatic L78 Tg males. (B and C) Whereas the number of EDL muscle fibers (B) is reduced in Tg males of both lines, the number of L5 motor axons (C) was significantly reduced relative to WT controls only in L141 Tg males, which is the line with particularly poor motor function. Graphs represent mean ± SEM, open bars represent Tg males, and filled bars represent age-matched WT males. *, significantly different from WT males. (D) Toluidine blue-stained cross sections of L5 ventral roots are shown for L141 WT and Tg males. WT 78 (n = 10–13, mean age: 317 days old, range: 179–528 days old), 78 Tg (n = 13, mean age: 300 days old, range: 106–528 days old), WT 141 (n = 12–13, mean age: 129 days old, range: 72–218 days old), Tg 141 (n = 6, mean age: 116 days old, range: 72–179 days old).

Fig. 4.

Alterations in gene expression in HSA-AR Male Mice consistent with neurogenic atrophy. Quantitative RT-PCR (qPCR) estimates of mRNA abundance of VEGF isoform 164 and 188 (VEGF), AChR alpha subunit (AChR), MyoD, and myogenin in muscles from WT and Tg males of the behavioral asymptomatic L78 line and the severely affected L141 line. Down-regulation of VEGF and up-regulation of AChR alpha, MyoD, and myogenin mRNA are also observed in muscle of KD mouse models and/or after denervation. All estimates are differences in mRNA relative to WT brothers within the same line. Graphs represent mean ± SEM. *, significantly different from WT brothers. L78 mice (n = 3 WT, 3 Tg; mean age = 145 days old, range = 106–222 days old); L141 mice (n = 4 WT, 4 Tg, mean age = 115 days old, range = 72–162 days old).