ABSTRACT
Title
Toward the identification of drug targets for contrasting in vivo muscle fatigability in muscular dystrophy: potential role of PGC-1α
Authors
G.M. Camerino1, R.F. Capogrosso1, A. Cozzoli1, M.M. Dinardo1, V. Sblendorio1, A. Scaramuzzi1, A. De Luca1
1Unit of Pharmacology, Department of Pharmaco-biology, Faculty of Pharmacy, University of Bari, Italy;
1Unit of Pharmacology, Department of Pharmaco-biology, Faculty of Pharmacy, University of Bari, Italy;
Abstract
In vivo fatigability is a major symptom of patients affected by Duchenne muscular dystrophy (DMD). In fact the 6 min walk test is a primary end-point for evaluation of clinical efficacy of therapeutic strategies in human trials (Bushby et al. 2010).
We have recently shown that in vivo fatigability also occurs in the mdx mouse, the most widely used animal model of DMD. In fact an endurance test on treadmill (5 m/min for 5 min followed by incremental speed every minute until exhaustion) showed that mdx mice run significantly less with respect to wild-type counterparts, with an impairment that is related to the amount of muscle activity previously experienced by the animal (Burdi et al., 2009).
In vivo fatigue may result from a complex adaptation of vascular and metabolic functions to mechano-transduction signals that are likely to be failing or aberrant in dystrophic muscle. Interestingly, we have previously demonstrated that a chronic treatment with the phosphodiesterase inhibitor pentoxifylline (PTX) significantly ameliorates the performance of dystrophic animals to endurance tests (Burdi et al., 2009). This effect can be related to the ability of PTX to enhance NO/cGMP pathways, known to play a role in both vascular adaptation and metabolic muscle performance. In fact NO/cGMP is one of the pathways stimulating peroxisome proliferator coactivator γ -1α (PGC-1α), a well known exercise-sensitive transcriptional co-activator of genes involved in muscle metabolisms, anti-oxidant defense and mitochondrial biogenesis. Then, we hypothesized that the proper activation of PGC-1α pathways are failing in dystrophic muscle due to the impaired mechano-transduction (Handschin et al., 2008). The verification of this hypothesis may help to identify novel targets for pharmacological treatment of dystrophy-related fatigue. To this aim we measured the resistance to exercise and the PGC-1α expression in mdx vs. wildtype mice, either sedentary or treadmill-exercised (12 m/min twice a week for 4 weeks). As expected mdx mice run a shorter distance with respect to wild-type animals (274 ± 50 m n=6 vs. 390 ± 25 m n=5). Interestingly, fatigue was further enhanced in mdx mice that previously underwent the chronic 4 weeks standard protocol of training. In fact in these animals the resistance to endurance test was further and significantly reduced to 163 ± 30 m (n=6) (p < 0.001 vs. wt mice). No significant alterations were instead observed in exercise performance of wild-type animals.
The evaluation PGC-1α expression by Taq-Man real-time PCR, expressed as ratio toward the housekeeping GAPDH gene, showed in fact no differences between wildtype and mdx mice in gastrocnemious muscle in basal conditions. However, the in vivo exercise protocol leads to the expected increase in expression in PGC-1α in wildtype gastrocnemious muscle (by 70%; p < 0.05 by Bonferroni t-test after ANOVA), while no changes have been observed in mdx mice. This result does corroborate the hypothesis of an impaired activation in response to physical challenge pathways.
A longer exercise protocol (12 weeks) did not lead to a further worsening of in vivo fatigue in mdx mice (170 ± 15m; n=7), underlying a possible slow adaptation of the dystrophic mice to the endurance test. The evaluation of PGC-1α expression in animals underlying longer exercise training as well as in mdx animals treated with drugs able to enhance PGC-1α activity through different pathways may help to elucidate the potential role of this co-activator as drug target in muscular dystrophy.
Bushby et al. (2010), Lancet Neurol. 9(1):77-93.
Burdi et al. (2009), J Appl Physiol.106(4):1311-24.
Handschin et al. (2008), Nature 24;454(7203):463-9
We have recently shown that in vivo fatigability also occurs in the mdx mouse, the most widely used animal model of DMD. In fact an endurance test on treadmill (5 m/min for 5 min followed by incremental speed every minute until exhaustion) showed that mdx mice run significantly less with respect to wild-type counterparts, with an impairment that is related to the amount of muscle activity previously experienced by the animal (Burdi et al., 2009).
In vivo fatigue may result from a complex adaptation of vascular and metabolic functions to mechano-transduction signals that are likely to be failing or aberrant in dystrophic muscle. Interestingly, we have previously demonstrated that a chronic treatment with the phosphodiesterase inhibitor pentoxifylline (PTX) significantly ameliorates the performance of dystrophic animals to endurance tests (Burdi et al., 2009). This effect can be related to the ability of PTX to enhance NO/cGMP pathways, known to play a role in both vascular adaptation and metabolic muscle performance. In fact NO/cGMP is one of the pathways stimulating peroxisome proliferator coactivator γ -1α (PGC-1α), a well known exercise-sensitive transcriptional co-activator of genes involved in muscle metabolisms, anti-oxidant defense and mitochondrial biogenesis. Then, we hypothesized that the proper activation of PGC-1α pathways are failing in dystrophic muscle due to the impaired mechano-transduction (Handschin et al., 2008). The verification of this hypothesis may help to identify novel targets for pharmacological treatment of dystrophy-related fatigue. To this aim we measured the resistance to exercise and the PGC-1α expression in mdx vs. wildtype mice, either sedentary or treadmill-exercised (12 m/min twice a week for 4 weeks). As expected mdx mice run a shorter distance with respect to wild-type animals (274 ± 50 m n=6 vs. 390 ± 25 m n=5). Interestingly, fatigue was further enhanced in mdx mice that previously underwent the chronic 4 weeks standard protocol of training. In fact in these animals the resistance to endurance test was further and significantly reduced to 163 ± 30 m (n=6) (p < 0.001 vs. wt mice). No significant alterations were instead observed in exercise performance of wild-type animals.
The evaluation PGC-1α expression by Taq-Man real-time PCR, expressed as ratio toward the housekeeping GAPDH gene, showed in fact no differences between wildtype and mdx mice in gastrocnemious muscle in basal conditions. However, the in vivo exercise protocol leads to the expected increase in expression in PGC-1α in wildtype gastrocnemious muscle (by 70%; p < 0.05 by Bonferroni t-test after ANOVA), while no changes have been observed in mdx mice. This result does corroborate the hypothesis of an impaired activation in response to physical challenge pathways.
A longer exercise protocol (12 weeks) did not lead to a further worsening of in vivo fatigue in mdx mice (170 ± 15m; n=7), underlying a possible slow adaptation of the dystrophic mice to the endurance test. The evaluation of PGC-1α expression in animals underlying longer exercise training as well as in mdx animals treated with drugs able to enhance PGC-1α activity through different pathways may help to elucidate the potential role of this co-activator as drug target in muscular dystrophy.
Bushby et al. (2010), Lancet Neurol. 9(1):77-93.
Burdi et al. (2009), J Appl Physiol.106(4):1311-24.
Handschin et al. (2008), Nature 24;454(7203):463-9