Influence of microRNAs on angiogenic and antiangiogenic factors in the pathway of vascular endothelial growth factor in coronary artery disease patients

Author(s)

Larissa Ferreira dos Santos

• Document type: Doctoral Thesis
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina (FM/SBD)
• Defense date: 2020-12-11

Examining board members

Maria Urbana Pinto Brandão Rondon; Lígia de Moraes Antunes Corrêa; Carlos Eduardo Negrão; Edilamar Menezes de Oliveira

• Advisor: Maria Urbana Pinto Brandão Rondon

Abstract

INTRODUCTION: Endothelial dysfunction and inflammation are hallmarkers of coronary artery disease (CAD). Some circulating microRNAs (miRNAs) expressed in the vascular endothelium and regulators of the angiogenesis pathway have been described in CAD. However, the local expression in skeletal muscle of these miRNAs is unknown in CAD patients, as well as the association of these miRNAs with peripheral blood flow and with angiogenic and antiangiogenic factors of the vascular endothelial growth factor (VEGF) pathway in these patients. Therefore, the aims of this study were to evaluate the tissue and circulating miRNAs-126, -16, -21 and 92a expressions in CAD patients, the tissue gene and protein expression of the angiogenic and antiangiogenic factors of the VEGF pathway, to evaluate the peripheral blood flow (PBF), and to evaluate if the miRNAs expressions were associated with the PBF and with the protein expressions in these patients. METHODS: Twenty-two CAD patients without ventricular dysfunction (55±1 years) and fourteen healthy control subjects (CS) were selected. A blood sample was collected to assess circulating miRNAs and the vastus lateralis biopsy was made to assess tissue miRNAs, genes and protein expressions. The miRNAs and genes were analyzed by RT-qPCR, proteins by western blotting, and peripheral blood flow by Doppler ultrasound (shear rate, SR) and venous occlusion plethysmography (muscle blood flow, MBF). Mean arterial pressure (MAP, oscillometric) and cardiac frequency (HR, electrocardiogram) were assessed. RESULTS: The tissue expressions of the miRNAs -126, -16 and -21 was reduced in the CAD group compared to the CS group, (72 ± 6 vs. 100 ± 10, p = 0.03; 72 ± 5 vs. 100 ± 11, p = 0.03; 66 ± 8 vs. 100 ± 11, p = 0.04; respectively). The same behavior was observed for circulating miRNAs. The tissue and circulating expression of miRNA-92a was similar between groups. The gene expression of PI3KR2 was increased (p= 0.02), while iNOS (p=0.02) and Bcl-2 (p=0.01) were decreased in CAD compared to the CS. The gene expression of the other evaluated factors was not different between groups. Regarding tissue protein expression, the factors Bad, Bcl-2 and VEGFR-2 were increased (p < 0.05), pERK1/2/ERK1/2 was decreased (p=0.047) and p-Akt/Akt tended to be decreased (p= 0.06) in CAD patients. The other factors evaluated were similar between groups. At rest, the forearm and leg MBF, the forearm and leg vascular conductance (FVC and LVC, respectively), MAP and HR were not different between groups. However, during the handgrip and leg exercise, the CAD group had a lower FVC and LVC response compared to the CS group. The flow pattern, antegrade and medium shear rate (SR) of the brachial and femoral arteries were lower in the CAD group compared to the CS group. The retrograde and oscillatory SR were similar between groups. MiRNA-126 was associated with PI3KR2 protein expression and miRNA-16 with VEGFR-2. We did not observed associations between circulating miRNAs and brachial and femoral flow patterns. However, tissue miRNA-92a was associated with antegrade SR. Interestingly, circulating - 126, -16 and -21 miRNAs were associated with FBF and FVC at rest and during exercise. On the other hand, in the leg exercise, we only observed an association between miRNA-16 and the FBF. CONCLUSIONS: Patients with CAD have reduced circulating and tissue miRNAs -126, -16, and -21 expressions. These angiomiRs involved in the VEGF pathway are altered in these patients, which could interfere in the expression of target genes, altering the process of angiogenesis, activation of the apoptotic pathway, and protein synthesis. In addition, a positive association was observed between c-miRNAs - 126, -16 and -21 with FVC at rest and during exercise, thus suggesting the influence of these angiomiRs on the modulation of vascular function (AU)

• FAPESP's process: 17/24605-0 - Influence of microRNAs in angiogenic and antiangiogenic factors in the pathway of vascular endothelial growth factor in patients with Coronary Artery Disease
• Grantee: Larissa Ferreira dos Santos
• Support Opportunities: Scholarships in Brazil - Doctorate