Estudos in vivo e in vitro dos efeitos do treinamento físico sobre alterações musculares decorrentes da associação entre obesidade e envelhecimento

Abstract

Sarcopenia is one of the main outcomes of the dangerous association between aging and obesity, and is a syndrome characterized by functional and morphological impairment of skeletal muscle. Among the repercussions of sarcopenia, destabilization, fragility and systemic metabolic changes that directly affect quality of life stand out. On the other hand, physical training appears to be an effective tool against the effects of aging and obesity by contributing to reducing adiposity and protecting skeletal muscle mass. However, no studies were found that evaluated the effects of aerobic training when carried out throughout life or therapeutically on the changes caused by obesity in the early stages of aging. Objective: To investigate the effects of aerobic physical training performed therapeutically or throughout life on histological changes, redox balance, inflammation, and metabolism in skeletal muscle and cardiac ventricle caused by the association between aging and obesity in vivo and in vitro. Methods: The experiment was divided into a pilot study and a main study. The pilot study aimed to identify the age at which changes in muscle and physical performance begin in response to natural aging. Thus, 41 male Wistar rats aged 4 months were randomly distributed into 6 groups defined by age at eutanásia. Throughout the experimental protocol, the animals were weighed weekly, and physical performance was assessed monthly using the maximal running test on a motorized treadmill for rats. After defining the age at which muscle decline began (14 months), the influence of physical training on the changes observed was evaluated. To this end, 32 4-month-old male Wistar rats were randomly distributed into two groups: sedentary group (n = 16) and trained group (n = 16). Upon reaching 8 months of age, 8 animals from each group were euthanized, forming the young sedentary and trained groups. The remaining animals continued until they reached 14 months of age, and constituted the sedentary and trained aged groups. Training was carried out on a motorized treadmill for rats, at moderate intensity, on alternate days and lasting 1 hour per session. The animals were euthanized 24 hours after the last training session. Blood, adipose tissue and muscle samples were collected, duly measured and stored in an ultra-freezer -80 °C, or fixed. The frozen samples were used to quantify the inflammatory and oxidative profiles, and the samples were fixed for histological studies. The main study consisted of an in vivo and an in vitro study. The in vivo protocol was carried out with 32 male Wistar rats that were randomly distributed into four groups (n = 8/group): aged sedentary group (ESed), aging sedentary group fed a high-fat diet (HFD) (ASed+HFD), aging trained therapeutically and fed with HFD (ATT+HFD) and aging lifelong trained and fed with HFD (ALT+DHL). 24 hours after the last physical training session, all animals were euthanized and then the gastrocnemius muscle and cardiac ventricle were collected for histological, immunohistochemical, redox balance, inflammation and metabolism studies. The gastrocnemius muscle was used to analyze mRNA and microRNA expression by qPCR. Abdominal adipose tissue deposits were collected and weighed to determine body composition. For in vitro studies, primary macrophages obtained from the peritoneal exudate of 6 4-month-old male BALB/c mice, elicited through the administration of thioglycolate (3%), and primary myoblasts isolated from the gastrocnemius muscle of a 4 month old male rat were used. Cultures of macrophages and myoblasts were exposed to serum from the animals from the in vivo study for 24h. The cultures were used to analyze cell viability by MTT, total nitrites and hydrogen peroxide and to analyze the expression of genes involved in the inflammatory response and muscle metabolism. All variables of interest in this study were subjected to the Shapiro-Wilk normality test. Comparisons between pilot test groups were made using the ANOVA test for repeated measures (temporal analysis) and one-way ANOVA, followed by the Tukey post-test. For analyzes between young and aged, sedentary and trained groups, a factorial design (2x2) was adopted using the two-way ANOVA test. Analyzes of in vivo and in vitro studies were performed using Student's T test and one-way ANOVA test. Results: The results of the pilot study indicate that the decline in muscle function begins at 14 months of age, evidenced by the reduction in physical performance, lean mass and gastrocnemius muscle weight and antioxidant capacity of the muscle. On the other hand, the physical training routine up to 14 months improved body composition, inflammatory profile, redox status and protected the muscle against reduction in the cross-sectional area of the fibers. The in vivo study showed an unfavorable outcome in response to the association between aging and diet-induced obesity on muscle and cardiac parameters, morphology, fibrosis, inflammation, oxidative stress and metabolism. Furthermore, an increase in the expression of miRNA-21 and genes involved in the pro-inflammatory and catabolic response, and a reduction in the expression of genes involved in the antioxidant and anabolic response were observed in the gastrocnemius muscle. Physical training, especially the lifelong training protocol, attenuated the findings resulting from the association between aging and obesity on muscular and cardiac parameters, reduced the expression of miR-21 and increased the expression of miR-486. In in vitro studies, macrophages and myoblasts exposed to serum from aging and obese animals showed an increase in the pro-inflammatory and catabolic profile, respectively, while serum from trained animals, especially those that had undergone lifelong training, determined reduction of the pro-inflammatory and catabolic profile of cell cultures. Conclusion: In the animal model studied, the association between sedentary lifestyle, aging and obesity induced by a high-fat diet determined muscle damage, especially in the inflammatory, oxidative and metabolic profiles. On the other hand, regular physical exercise, especially throughout life, attenuated the changes caused by the association between aging and obesity in all parameters studied, in vivo and in vitro. Our findings reinforce the importance of adopting exercise routines throughout life as an effective tool against the effects of age, especially when associated with unhealthy eating habits.