Low Intensity Training Provokes Adaptations on Muscle Fibrosis of a Muscular Dystrophy Model

Abstract

SUMMARY:Duchenne muscular dystrophy (DMD) is a genetic neuromuscular disorder with progressive clinical signs until death, around the second decade of life. Mdx is the most used animal model to pre-clinical studies of DMD. Parameters of exercise on this muscular disease are still unknown. This research aimed to investigate if the low intensity treadmill training would exacerbate the markers of muscle injury, fibrosis, and the composition of the extracellular matrix by type I and III collagens of the mdx model. Dystrophic 11-week-old male mice were separated in exercised (mdxE, n=8) and sedentary (mdxC, n=8) groups. Wild-type mice were used as control (WT, n=8). Exercised group underwent a LIT protocol (9 m/min, 30min, 3days/week, 60 days) on a horizontal treadmill. Gastrocnemius muscle was collected at day 60 and processed to morphological and morphometric analyzes. Sedentary mdx animals presented inflammatory infiltrate and necrotic fibers. Histochemical analysis revealed that the perimysium of the mdxC group is organized into thick and clustered collagen fibers, which generates a larger area of intramuscular collagen fibers for these animals. Histomorphometry attested that fraction area of collagen fibers of mdxC group was higher than mdxE group (p=0.04) and mdxE group values similar to WT group (p=1.00). Centrally located nuclei fibers and the variance coefficient (VC) of minimal Feret's diameter was similar in mdxE and mdxC groups (p=1.00) and both groups presented higher mean values than WT group (p<0.00). Immunohistochemistry revealed the presence of type I collagen mainly in the mdxC group. LIT protocol had not exacerbated muscle injuries resulting from the dystrophindeficiency membrane fragility at the same time that had reduced the intramuscular collagen deposition. LIT had positively influenced these markers of dystrophic muscle injury on gastrocnemius muscle of mdx model.

Beneficial Role of Exercise in the Modulation of mdx Muscle Plastic Remodeling and Oxidative Stress

“…Several studies have shown that treadmill exercise can efficiently modulate this process. Indeed, a low intensity exercise, at a speed of 9 m/min for two months, on a horizontal platform reduces collagen deposition in limb muscles of mdx mice [ 73 , 74 , 75 ]. In addition, low intensity running induced a strong beneficial effect on the degeneration-regeneration process of mdx muscle and a shift in favor of regeneration, both in limb muscles and in diaphragm [ 76 , 77 ], and reduced the percentage of necrotic area in mdx soleus and gastrocnemius muscles, while it seemed to exert opposite effects on mdx plantaris muscle [ 78 ].…”

“…Based on the above reported experimental evidences, we can conclude that beneficial effects on mdx limb muscles induced by horizontal treadmill running exercise depend primarily on training intensity, being mainly associated to running speed ranging between 4 and 9 m/min. These positive effects, including fibrosis reduction, increased regeneration, improvement of muscle performances and structure, such as muscle hypertrophy and fiber type switch, seem to be not influenced by age, since they can be found in mdx mice 4–5 weeks old [ 108 , 109 ], 8 weeks old [ 74 , 76 , 77 , 110 ], and up to 20 weeks old [ 73 , 75 , 79 ], or by the duration of training, given the persistence of positive effects for exercise duration ranging between 1 and 6 months. On the contrary, horizontal treadmill exercise at higher intensity, with speed equal or greater than 12 m/min, or downhill/uphill running, generally exert detrimental effects on mdx muscles, and are often used to worsen the mdx phenotype before assessing the efficacy of drug treatment [ 8 , 81 , 82 , 84 , 87 , 92 , 99 ].…”

“… Morici, G., et al (2017) [ 76 ] Male 8 weeks Pre-training : 3.2 m/min, 15 to 30 min/day, 5 days/week, 2 weeks Training : workload progressive increase, 4 to 4.8 m/min, 30 to 60 min/day, 5 days/week, 4 weeks DIA GAST QUAD Induces a trend for regeneration areas to be larger than necrosis areas in diaphragm and modulates the inflammatory status of hindlimb muscle. Pinto, P.A.F., et al (2018) [ 75 ] Male 11 weeks 9 m/min, 30 min/day, 3 days/week, 8 weeks Lateral GAST Reduces intramuscular fibrosis deposition and does not exacerbates markers of muscle injury. Zelikovich, A.S., et al (2019) [ 79 ] 4–5 months 4 m/min or 8 m/min, 30 min/day, 3 days/week, 6 months (Each exercise session consisted of 7’30’’ warm up followed by a 22’30’’ training at target speed) DIA GAST Heart QUAD TA Improves tetanic and specific force in TA muscle, increases respiratory capacity, attenuates cardiac decline associated with disease progression, increases adiponectin and reduces adipocyte cross sectional area, and induces a modest increase in expression of the PGC-1α gene in the gastrocnemius muscle.…”

>b<Efeito da associação da terapia celular e do treino de baixa intensidade no músculo distrófico de camundongo >i<mdx>/i<>/b<

“…O saco vitelino (SV) é uma das membranas extraembrionárias presente em todas as espécies de vertebrados, importante para a sobrevivência inicial do embrião e representa uma promissora via de pesquisa em células tronco (MANÇANARES et al, 2015;PINTO et al, 2021). Sabe-se que células isoladas do SV humano durante o estágio embrionário foram capazes de expressar marcadores de CTM, além de conter hemangioblastos, células progenitoras do sistema vascular e hematopoiético (MANÇANARES et al, 2019).…”

“…Estudos investigando o treinamento físico de baixa intensidade (low intensity training -LIT) em camundongos mdx encontraram resultados benéficos, como de melhora nos parâmetros fisiológicos, onde camundongos mdx treinados apresentaram uma redução dos níveis dos marcadores de estresse oxidativo (KACZOR et al, 2007;FERNANDES et al, 2019), redução de citocinas inflamatórias (HYZEWICZ et al, 2017) e foi capaz de modular proteínas envolvidas na defesa do estresse oxidativo (FONTANA et al, 2015). Camundongos mdx treinados também apresentam uma diminuição de áreas necróticas e infiltrados inflamatórios após treinamento (FRINCHI et al, 2014), além de apresentar uma menor área de fibrose intramuscular (GAIAD et al, 2017;PINTO et al, 2018;MORAIS et al, 2020).…”