MicroRNAs em plasticidade muscular: efeitos da superexpressão do miR-29c na modulação da massa muscular esquelética.

Abstract

SILVA, W. J. MicroRNAs in muscle plasticity: overexpression of miR-29c in modulation of skeletal muscle mass. 168 p. Ph.D. Thesis (Doctor in Sciences). São Paulo: University of São Paulo; 2019. The skeletal muscle is the body most abundant tissue. It plays an important role in daily life activities, such as movement, posture, and breathing. In addition, this tissue is crucial at physiological processes like metabolic equilibrium and immune defense. The substantial adaptability in response to environmental change marks skeletal muscle as a plastic organ. This plasticity could be coordinated by microRNAs, those are small non-protein-coding RNAs that regulate post-transcriptional gene expression. This knowledge has fostered new approaches that aim to optimize the skeletal muscle health. Thus, in this work, we intended to identify and characterize microRNAs that modulate the muscle mass and the regeneration process. An in silico analyses has allowed the identification of microRNAs who possibly bind genes from pathways of skeletal muscle mass control and regeneration. After, we manipulated the expression of those microRNAs on C2C12 cells and C57BL/6 mice. Finally, we measured the transcriptional levels of target genes and the impact of these alterations on the cells and on muscle tissue. In the first section of this work, we hypothesized that microRNAs could regulate MURF1 and MURF2 expression, both E3-ligases important for the regeneration process. In our analysis, MURF1 was a predictable target of miR-29c and miR-101a while for MURF2 were identified miR-133a and 133b. During the regeneration process, MURF1 is up-regulated and its predicted target microRNAs are downregulated. In this context the hyperexpression of both miR-29c and miR-101a in C2C12 cells induced MURF1 down-regulation. Furthermore, miR-29c promotes myogenesis with an increase in myotubes diameter and fusion index. In contrast, miR-101a expression reduces myotubes diameter with no change at the fusion index. Lastly, luciferase assay validated only miR-29c directly target MURF1 3`UTR. In the second section of this work, we identified miR-29c as a potential regulator of skeletal muscle mass. Then through gene delivery by electroporation, we induce miR-29c overexpression in mice skeletal muscle. This procedure promoted an increase in muscle mass as well as a gain in strength, endurance and sarcomere number, furthermore the number of activated satellite cells. Taken together our results found that miR-29c has a hypertrophic effect with gain in muscle function. Thus, the overexpression of this microRNA could be a useful tool for future therapeutics that manipulate muscle mass.