Decreased Peak Expiratory Flow Associated with Muscle Fiber-Type Switching in Spinal and Bulbar Muscular Atrophy

Abstract

The aim of this study was to characterize the respiratory function profile of subjects with spinal and bulbar muscular atrophy (SBMA), and to explore the underlying pathological mechanism by comparing the clinical and biochemical indices of this disease with those of amyotrophic lateral sclerosis (ALS). We enrolled male subjects with SBMA (n = 40) and ALS (n = 25) along with 15 healthy control subjects, and assessed their respiratory function, motor function, and muscle strength. Predicted values of peak expiratory flow (%PEF) and forced vital capacity were decreased in subjects with SBMA compared with controls. In SBMA, both values were strongly correlated with the trunk subscores of the motor function tests and showed deterioration relative to disease duration. Compared with activities of daily living (ADL)-matched ALS subjects, %PEF, tongue pressure, and grip power were substantially decreased in subjects with SBMA. Both immunofluorescence and RT-PCR demonstrated a selective decrease in the expression levels of the genes encoding the myosin heavy chains specific to fast-twitch fibers in SBMA subjects. The mRNA levels of peroxisome proliferator-activated receptor gamma coactivator 1-alpha and peroxisome proliferator-activated receptor delta were up-regulated in SBMA compared with ALS and controls. In conclusion, %PEF is a disease-specific respiratory marker for the severity and progression of SBMA. Explosive muscle strength, including %PEF, was selectively affected in subjects with SBMA and was associated with activation of the mitochondrial biogenesis-related molecular pathway in skeletal muscles.

Fig 1. Respiratory function profile of subjects with SBMA.

The actual and predicted values of forced vital capacity (FVC) (A, B), forced expiratory volume in 1 s (FEV_1.0) (C), the ratio of FEV_1.0 to FVC (D), and actual and predicted values of peak expiratory flow (PEF) (E, F) were compared among SBMA subjects (n = 40), ALS subjects (n = 25), or healthy controls (n = 15). Compared with the healthy controls, patients with SBMA exhibited decreased values for %FVC, PEF, and %PEF. The actual values of PEF were also lower in SBMA than in controls. When comparing SBMA and ALS subjects, %PEF was significantly decreased in SBMA, but no differences were detected for the other indices. ** p < 0.01. * p < 0.05. Data are presented as the mean ± SE. SBMA, spinal and bulbar muscular atrophy; ALS, amyotrophic lateral sclerosis; HC, healthy controls.

Fig 2. Relationships between %PEF and ALSFRS-R or %FVC in subjects with SBMA and ALS.

Comparison of the relationships between %PEF and total score of ALSFRS-R (A) or %FVC (B) in SBMA and ALS. The difference in %PEF between SBMA and ALS was significant after adjustment for the ALSFRS-R and %FVC with ANCOVA. %PEF, predicted values of peak expiratory flow; ALSFRS-R, the revised Amyotrophic Lateral Sclerosis Functional Rating Scale; %FVC, predicted forced vital capacity.

Fig 3. Longitudinal changes in respiratory function in SBMA.

Longitudinal changes in %PEF (A), %VC (B), and FEV_1.0/FVC (C) as a function of disease duration were analyzed. The solid lines indicate representative disease progression over disease duration calculated using a marginal model under an unstructured covariance matrix. The broken curvilinear line demonstrates the 95% confidence interval of these models. We calculated the estimated values at clinical onset (intercepts) and the change values per year (D). In an analysis using the marginal model, a generalized estimating equation (shown by the solid lines) identified that %PEF (A) and %FVC (B) demonstrated a slowly but with steady progression. PEF, peak expiratory flow; FVC, forced vital capacity; FEV_1.0, forced expiratory volume in 1 s.

Fig 4. Disease-specific fiber type alterations in SBMA and ALS.

(A, B) Representative images of anti-MHC immunostaining in the biceps muscles of subjects with SBMA (A, n = 2) and ALS (B, n = 2). Type I (blue), type IIa (strong green), and type IIx (strong red and intermediate green) fibers are shown in single- and merged-channel images of serial cross-sections of a human bicep incubated with an antibody cocktail (BAF-8, SC-71, and 6H1). Type IIx fibers were substantially decreased in SBMA compared with ALS, whereas type I fibers were atrophied in ALS compared with SBMA. Scale bar, 100 μm. SBMA, spinal and bulbar muscular atrophy; ALS, amyotrophic lateral sclerosis; MHC, myosin heavy chain.

Fig 5. Expression levels of myosin heavy chain and AMPK-PGC-1α pathway in the skeletal muscles.

The mRNA expression levels of MYH1 (encoding MHC type IIx), MYH2 (encoding MHC type IIa), and MYH7 (encoding MHC type I) normalized to β2-microglobulin levels in the intercostal muscles of SBMA (n = 5) and ALS (n = 5) subjects (A–C). The expression levels of MYH1 (A) and MYH2 (B) were significantly decreased in subjects with SBMA compared with subjects with ALS. The mRNA expression levels of MYH1, MYH2, and MYH7 normalized to β2-microglobulin levels in the iliopsoas muscles of SBMA (n = 5), ALS (n = 6), and DC (n = 4) subjects (D–F). The mRNA levels of MYH1 and MYH2 in the iliopsoas muscle were significantly decreased in subjects with SBMA compared with ALS subjects, as observed in the intercostal muscles (D, E). The expression levels of MYH7 were significantly higher in subjects with SBMA than in subjects with ALS (F). Expression levels of the genes known to regulate muscle fiber type switching in SBMA (n = 5), ALS (n = 6), and DC (n = 4) samples. The mRNA levels of PGC-1α and PPAR-δ, which regulate the oxidative fiber type profile, were significantly increased in SBMA compared with ALS and DC. The Mann-Whitney U test was performed to assess significant differences for each target gene between SBMA and ALS. ANOVA with Tukey’s post-hoc test was performed to compare the significance of differences in each target gene among SBMA, ALS, and DC. **p < 0.01. *p < 0.05. Data are presented as the mean ± SE. SBMA, spinal and bulbar muscular atrophy; ALS, amyotrophic lateral sclerosis; DC, disease control; PGC-1α, proliferator-activated receptor gamma coactivator 1-alpha; PPAR, peroxisome proliferator-activated receptor.