1LMU Klinikum der Universität München Medizinische Klinik und Poliklinik I München, Deutschland; 2Icahn School of Medicine at Mount Sinai New York, USA; 3Technische Universität München (TUM) Institut für Pharmakologie und Toxikologie München, Deutschland; 4Université Laval Pulmonary Hypertension Research Group Québec, Kanada; 5Gene and Cell Therapy Institute Massachusetts General Brigham Boston, USA
Pulmonary arterial hypertension (PAH) is a severe vascular disease that leads to right heart failure and death. PAH therapies primarily target pulmonary vasoconstriction, but only modestly affect pulmonary vascular remodeling. Currently there is no cure for PAH and new therapeutic options targeting the roots of the disease are desperately needed. MicroRNAs (miRs) have emerged in the last decades as key regulators in health and disease. We identified miR-224 as a lung enriched miR and in silico approaches for PAH predicted miR-224 among the miRs that target PAH related genes. In this study we aimed to investigate the role of miR-224 in pulmonary vascular remodeling and to define the mechanism of miR-224 action in PAH.
We found pulmonary miR-224 levels to be increased in PH-diseased lungs (in mice, rat, pig and humans) and in human pulmonary artery smooth muscle cells (hPASMCs) isolated from PAH patients. In vitro studies revealed that miR-224 is enriched in hPASMCs and its expression is necessary and sufficient to induce hPASMCs proliferation.
We next tested the therapeutic effect of miR-224 inhibition using three different PAH animal models in mice and rats. In a first approach, we intra-tracheally aerosolized an adeno-associated virus1-tough decoy-miR-224 (AAV1-TuD-224) to PH diseased mice (Sugen/hypoxia model). AAV-Ctrl-treated mice displayed all the hallmarks of PAH (i.e., increased Fulton index, RVSP, cardiomyocyte hypertrophy and pulmonary arterial medial thickness), whereas AAV1-TuD-224-treated mice displayed a marked and significant decrease in these parameters. We next intra-tracheally delivered a chemically modified antisense oligonucleotide specific for miR-224 (LNA-224) to PH diseased mice and rats (Sugen/hypoxia model). Consistent with our genetic approach, LNA-224 significantly protected mice and rats from cardiac hypertrophy and pulmonary vascular remodeling at the tissue and cellular levels. We finally delivered LNA-224 to monocrotaline-treated rats and found (by magnetic resonance imaging, hemodynamics, morphometric and histological measurements) that miR-224 inhibition improves survival and suppresses PAH. Mechanistically, we found that miR-224 represses BMP signaling by directly targeting four pathway factors.
Our data suggests that miR-224 plays a pivotal role in pulmonary vascular remodeling by targeting the BMP pathway and may have therapeutic value for PAH.