Duchenne muscular dystrophy progression induced by downhill running is accompanied by increased endomysial fibrosis and oxidative damage DNA in muscle of mdx mice.

Duchenne muscular dystrophy (DMD) is characterized by progressive muscle necrosis. One of the major challenges for prescribing physical rehabilitation exercises for DMD patients is associated with the lack of a thorough knowledge of dystrophic muscle responsiveness to exercise. This study aims to understand the relationship between myogenic regulation, inflammation and oxidative stress parameters, and disease progression induced by downhill running in the skeletal muscle of an experimental model of DMD. Six-month-old C57BL/10 and C57BL/10-DMDmdx male mice were distributed into three groups: Control (C), mdx, and mdx + Exercise (mdx + Ex). Animals were trained in a downhill running protocol for seven weeks. The gastrocnemius muscle was subjected to histopathology, muscle regeneration (myoD and myogenin), inflammation (COX-2), oxidative stress (8-OHdG) immunohistochemistry markers, and gene expression (qPCR) of NF-kB and NADP(H)Oxidase 2 (NOX-2) analysis. In the mdx + Ex group, the gastrocnemius muscle showed a higher incidence of endomysial fibrosis and a lower myonecrosis percentage area. Immunohistochemical analysis revealed decreased myogenin immunoexpression in the mdx group, as well as accentuated immunoexpression of nuclear 8-OHdG in both mdx groups and increase in cytoplasmic 8-OHdG only in the mdx + Ex. COX-2 immunoexpression was related to areas of regeneration process and inflammatory infiltrate in the mdx group, while associated with areas of muscle fibrosis in the mdx + Ex. Moreover, the NF-kB gene expression was not influenced by exercise; however, a NAD(P)HOxidase 2 increase was observed. Oxidative stress and oxidative DNA damage play a significant role in the DMD phenotype progression induced by exercise, compromising cellular patterns resulting in increased endomysial fibrosis.

Articular cartilage degeneration and bone adaptation due to lack of dystrophin in mice.

INTRODUCTION: Duchenne muscular dystrophy is caused by the absence of dystrophin. This study aimed to investigate femoral morphological characteristics of lack of dystrophin in MDX mice, considering that this model, different from DMD patient, is not influenced by corticosteroids administration and limited ambulation. MATERIALS AND METHODS: Proximal femur of male 16-week-old Control and MDX mice were submitted to histological, morphometric (volume density of articular cartilage, compact bone, trabecular bone and bone marrow; articular cartilage layers area; articular cartilage cell area), and immunohistochemistry analysis for RUNX-2, RANK-L, MMP-2, MMP-9, Caspase-3 and KI-67. RESULTS: MDX showed loss of linearity of articular cartilage with subchondral bone transition and elevation of this subchondral bone to the articular surface when compared with control. In addition, MDX presented morphological difference in the pantographic network of collagen fibers. Volume density of trabecular bone tissue was higher in the MDX than Control, but volume density of articular cartilage was lower in MDX (p < 0.05). The articular cartilage layers and chondrocytes area were significantly smaller in MDX than Control. These results associated to MMPs and osteogenic markers of proximal femur revealed an adaptation process as a consequence of lack of dystrophin. CONCLUSIONS: The morphological changes observed in the bone tissue of the MDX may be not only secondary to muscle weakness or chronic use of corticosteroids but also our results indicate connections between decrease of cartilage thickness, collagen network alteration and consequent subchondral changes that may lead to articular cartilage degeneration and bone adaptation mechanism in MDX mice.

Duchenne muscular dystrophy progression induced by downhill running is accompanied by increased endomysial fibrosis and oxidative damage DNA in muscle of mdx mice

Duchenne muscular dystrophy (DMD) is characterized by progressive muscle necrosis. One of the major challenges for prescribing physical rehabilitation exercises for DMD patients is associated with the lack of a thorough knowledge of dystrophic muscle responsiveness to exercise. This study aims to understand the relationship between myogenic regulation, inflammation and oxidative stress parameters, and disease progression induced by downhill running in the skeletal muscle of an experimental model of DMD. Six-month-old C57BL/10 and C57BL/10-DMDmdx male mice were distributed into three groups: Control (C), mdx, and mdx + Exercise (mdx + Ex). Animals were trained in a downhill running protocol for seven weeks. The gastrocnemius muscle was subjected to histopathology, muscle regeneration (myoD and myogenin), inflammation (COX-2), oxidative stress (8-OHdG) immunohistochemistry markers, and gene expression (qPCR) of NF-kB and NADP(H)Oxidase 2 (NOX-2) analysis. In the mdx + Ex group, the gastrocnemius muscle showed a higher incidence of endomysial fibrosis and a lower myonecrosis percentage area. Immunohistochemical analysis revealed decreased myogenin immunoexpression in the mdx group, as well as accentuated immunoexpression of nuclear 8-OHdG in both mdx groups and increase in cytoplasmic 8-OHdG only in the mdx + Ex. COX-2 immunoexpression was related to areas of regeneration process and inflammatory infiltrate in the mdx group, while associated with areas of muscle fibrosis in the mdx + Ex. Moreover, the NF-kB gene expression was not influenced by exercise; however, a NAD(P)HOxidase 2 increase was observed. Oxidative stress and oxidative DNA damage play a significant role in the DMD phenotype progression induced by exercise, compromising cellular patterns resulting in increased endomysial fibrosis.

Insulin Modulates Myogenesis and Muscle Atrophy Resulting From Skin Scald Burn in Young Male Rats.

BACKGROUND: Burn injuries (BIs) due to scalding are one of the most common accidents among children. BIs greater than 40% of total body surface area are considered extensive and result in local and systemic response. We sought to assess morphological and myogenic mechanisms through both short- and long-term intensive insulin therapies that affect the skeletal muscle after extensive skin BI in young rats. MATERIALS AND METHODS: Wistar rats aged 21 d were distributed into four groups: control (C), control with insulin (C + I), scald burn injury (SI), and SI with insulin (SI + I). The SI groups were submitted to a 45% total body surface area burn, and the C + I and SI + I groups received insulin (5 UI/Kg/d) for 4 or 14 d. Glucose tolerance and the homeostatic model assessment of insulin resistance index were determined. Gastrocnemius muscles were analyzed for histopathological, morphometric, and immunohistochemical myogenic parameters (Pax7, MyoD, and MyoG); in addition, the expression of genes related to muscle atrophy (MuRF1 and MAFbx) and its regulation (IGF-1) were also assessed. RESULTS: Short-term treatment with insulin favored muscle regeneration by primary myogenesis and decreased muscle atrophy in animals with BIs, whereas the long-term treatment modulated myogenesis by increasing the MyoD protein. Both treatments improved histopathological parameters and secondary myogenesis by increasing the MyoG protein. CONCLUSIONS: Treatment with insulin benefits myogenic parameters during regeneration and modulates MuRF1, an important mediator of muscle atrophy.

Presence of metalloproteinases 2 and 9 and 8-OHdG in the fibrotic process in skeletal muscle of Mdx mice.

Inflammation and oxidative stress occurs in muscle of Duchenne muscular dystrophy (DMD). The relationship between a panel of biomarkers and the DMD outcome is necessary to indicate of disease progression and response to rehabilitation programs. The aim was to analyze the connective tissue of muscle of Mdx mice and immunoexpression of MMP-2, MMP-9, and 8-OHdG, which signalizes oxidative stress related to DNA damage. Biceps brachii of male C57BL/10 and C57BL/10-Dmdmdx mice was submitted to Hematoxylin-Eosin, Sirius red and immunohistochemistry (MMP-2, MMP-9 and 8-OHdG) analysis. Mdx showed focal lesions with intense inflammation and fibrosis related to immunoexpression of MMP-2 and MMP-9, proving the hypothesis that these MMPs are linked to muscular tissue degeneration, which can be regenerated by their inhibition, improving the treatment of DMD carriers. Histopathological findings related to centralized nuclei increase were related to higher 8-OHdG immunomarked nuclei in Mdx, which signalizes oxidative stress associated with DNA damage provoked by DMD. Such result shows that the evaluation of 8-OHdG during the evolution of the disease could be a method to evaluate DMD disease progression.

Short time insulin treatment post burn improves elastic-collagen rearrangement and reepithelization.

Extensive burn may cause acute resistance to insulin, which accentuates hypermetabolism, impairs glucose metabolism, immune dysfunction and risks of sepsis. To minimize these effects, insulin is used as a treatment. The purpose was to analyze the collagen-elastic arrangement effects of insulin on the burned skin. Wistar rats were assigned in groups: control (C); control with insulin (C + I); scald burn injury (SBI); and SBI with insulin (SBI+ I). SBI were submitted to 45% total body surface area burn and the insulin-treated groups received insulin (5 UI/Kg/day) for 4 or 14 days (d). Insulin levels, glucose tolerance test and HOMA index were determined. The skin sections were analyzed for histophatological and morphoquantitative data. Histopathological findings showed increased reepithelization of SBI+ I and formation of a new muscle layer after 14 days. In the collagen-elastic arrangement, insulin for 4 days increased the volume fraction (Vv) of thin collagen and elastic fibers. After 14 days, independently of injury, insulin decreased the elastic fibers. Insulin was able to reverse damages in the collagen-elastic rearrangement and stimulate reepithelization after 4 days. Untreated scald-burned animals showed higher Vv of thick collagen after 4 days, while those treated had a higher Vv of thin collagen. The Vv of elastic fibers was increased in SBI+ I for 4 days. In conclusion, insulin treatment was able to stimulate reepithelization. It also reversed the damages to the collagen-elastic arrangement in the scald-burned group, improving the organization of thin collagen and increasing the Vv of elastic fibers in the injured group treated with insulin for a short time, that is, for 4 days.

Therapeutic effects of vaccine derived from amastigote surface protein-2 (ASP-2) against Chagas disease in mouse liver.

This study investigated the efficacy of the vaccine in liver of mice infected with the Trypanosoma cruzi (T. cruzi) and immunized with AdASP-2. For this purpose, histopathological analysis and gene expression of COX-2, TNF-alpha, TNFR, iNOS, cytochrome C, caspase-3, TLR4, IL-6 and IL10 were evaluated. The following groups were used in this study: Group 1 - Control Group (CTRL) animals received AdßGal vehicle; Group 2 - Infected Group (TC) animals were infected with T. cruzi; Group 3 - Immunized Group (AdASP-2): animals were immunized by AdASP-2 vaccine; Group 4 - Immunized and Infected Group (AdASP-2+TC) animals were infected with T. cruzi and immunized by AdSP-2 vaccine. A significant decrease of amastigote nests was noticed in the group of animals that were immunized with AdASP-2 and infected on the same day. COX-2 and TNF-alpha gene expressions increased in TC group, whereas TNF-alpha decreased in the TC+AdASP-2 group. TNFR expression was high in AdASP-2+TC group. iNOS expression was high for all experimental groups whereas cytochrome C decreased for all experimental groups. Caspase 3 increased in TC and TC+AdASP-2 groups. The gene expression of TLR4 and IL-10 showed an increase in AdASP-2+TC group. Finally, hepatic fibrosis was noticed to TC and AdASP-2 + TC groups. Taken together, our results demonstrated that vaccination with AdASP-2 was effective against the acute phase of experimental Chagas disease as a result of a more powerful and rapid immune response closely related to expression of some inflammatory genes, such as iNOS, TNF-alpha, TLR 4, and IL-10.

Spontaneous physical activity and mediators of energy homeostasis in the hypothalamus of mice from 4 to 10 months of age.

NEW FINDINGS: What is the central question of this study? Is the initial decline of spontaneous physical activity (SPA) in mice related to impaired insulin and leptin signalling or brain-derived neurotrophic factor expression in the hypothalamus? What is the main finding and its importance? We showed that SPA started to decline at an early stage, concomitantly with an impairment of hypothalamic leptin signalling. Consequently, energy expenditure decreased and glucose tolerance worsened. Our results demonstrate the need to counteract the initial decline in SPA to avoid metabolic impairments and indicate the possible involvement of central leptin in the reduction in SPA with age. The biological control of physical activity is poorly understood. Age decreases insulin, leptin and brain-derived neurotrophic factor (BDNF) signalling in the hypothalamus, and all have been shown to modulate spontaneous physical activity (SPA). We investigated the age at which SPA starts to decline and whether this is associated with the emergence of hypothalamic insulin and leptin resistance and reduced BDNF expression. Spontaneous physical activity (and other parameters of locomotion) and energy expenditure were determined monthly in mice from the 4th to the 10th month of age. Metabolic and hypothalamic analyses were performed in 4-, 6- and 10-month-old mice. Spontaneous physical activity, distance travelled and speed of locomotion started to decrease in 6-month-old mice. The reduction in SPA became more evident from 8 months of age. Energy expenditure decreased from the 8th month. Hypothalamic BDNF protein expression and insulin signalling did not change throughout the time span studied. Leptin signalling decreased at 6 and 10 months compared with 4 months. Also, compared with 4 months, 6- and 10-month-old mice were glucose intolerant. In conclusion, SPA begins to decline in parallel with reduced hypothalamic leptin signalling. Metabolic impairment also manifests as SPA decreases, highlighting the need to understand the regulation of SPA in order to combat its decline.