Therapeutic potential of the inhibition of microRNA-34c-3p: unravelling molecular regulators through aerobic exercise training

Author(s)

Clara Nóbrega

• Document type: Master's Dissertation
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Escola de Educação Física e Esportes (EEFE/BT)
• Defense date: 2019-04-16

Examining board members

Edilamar Menezes de Oliveira; Marcelo Alves da Silva Mori; Paulo Rizzo Ramires; Alice Cristina Rodrigues

• Advisor: Edilamar Menezes de Oliveira

Abstract

Cardiovascular diseases are the leading cause of death in Brazil. In contrast to this scenario, aerobic exercise training (AET) is an important tool to combat this condition, acting, among several mechanisms, on the expression of microRNAs (miRNA) and cardiac hypertrophy (CH). In this sense, would AET be able to reduce expression of miR-34c-3p in cardiac pathologies, generating improvement to the damages caused by them? In an attempt to elucidate this knowledge gap, male Wistar rats (2 months) underwent coronary occlusion, causing myocardial infarction (IMS; IMT) or induction of type I diabetes by STZ (DS: DT: 45mg kg). After proof that cardiac damage was caused, they were submitted to AET (10 wk), and left ventricles (LV) were dissected post mortem. Analyzing gene expression by RT-PCR, AET decreased (39%; p <0.02) miRNA-34c-3p expression in the LV of IMT vs. IMS. In agreement, there was a decrease (27%; p <0.04) of the miRNA in DT vs. DS. Investigating the therapeutic potential of this inhibition, through in vitro analysis in primary culture of neonatal rat cardiomyocytes treated with miRNA inhibitor (amiR-34c-3p), expression of miRNA- 34c-3p was decreased in 71% (50nM; p <0.01) vs. (C), whereas there was an increase of 41000% (50nM; p <0.01) between the superexpressor (mmiR) and C. Analyzing the pathologic HC markers between amiR and C, there was a reduction in skeletal a-actin (P <0.05), an increase in ANF (84.4%, p <0.05) and reduction in b-MHC (66%; p <0.007), demonstrating physiological HC in cardiomyocytes treated with amiR. Searching for miRNA-34c-3p targets responsible for these changes, eIF4E, the last non-canonically activated translation initiator of PI3K-AKT-mTOR, was analyzed. Its direct activation may represent an important advance in the search for therapies that induce physiological CH. The amiR increased eIF4E expression (64%, p <0.003), with validation for decreased luciferase activity (48%; p <0.02) by assaying 3\'UTR vector of the gene and miR-34c-3p. Another predicted target analyzed, KLF11, has homology with insulin gene and may be related to LV metabolic improvement. The treatment with amiR increased KLF 11 gene expression (192%; p <0.006), with a 55% reduction in luciferase activity and an increase in glucose uptake (13%; p <0.04). With the data presented, AET was efficient in inhibiting the expression of miRNA-34c-3p in LV of cardiopathic animals that, acting on their now validated targets, was able to promote physiological CH and metabolic improvement in cardiomyocytes. Considering the cardioprotective effects of inhibition of this miRNA, we can speculate a therapeutic potential of the inhibition of miR-34c-3p in patients in prospect of mitigating the damage caused by cardiopathies (AU)

• FAPESP's process: 16/14848-0 - Effects of aerobic exercise training on microRNA-34c* expression in heart of streptozotocin-induced diabetic rats
• Grantee: Clara Nóbrega
• Support Opportunities: Scholarships in Brazil - Master