MicroRNA-519 orchestrates cardiac repair post myocardial infarction by driving cardiomyocyte proliferation through p21/p57 repression
Y Gong, M Generali, H Renikunta, C Balbi, Y Wang, Z Zhang, A Mongelli, L Opitz, V Masciovecchio, S Mohammed, U Landmesser, F Ruschitzka, S Costantino, F Paneni, P JakobAbstract
Background
Myocardial infarction (MI) causes irreversible cardiomyocyte (CM) loss, adverse ventricular remodeling, and heart failure due to the limited regenerative capacity of the adult human heart. Effective cardiac repair requires the coordinated induction of CM proliferation and angiogenesis without promoting fibrosis. Following an unbiased high-throughput functional screen of 2,019 microRNAs (miRs) in human induced pluripotent stem cell-derived CMs (hiPSC-CMs), miR-519e-3p (miR-519) emerged as a potent activator of CM cell-cycle activity.
Purpose
To characterize the regenerative potential and underlying mechanisms of miR-519 in human multicellular systems and evaluate its therapeutic efficacy after MI in vivo.
Methods
MiR-519-mimics or miR-scrambled (control) were transfected into hiPSC-CMs, human aortic endothelial cells (HAECs), and human cardiac fibroblasts (cFBs). CM proliferation was assessed via immunofluorescence (IF) for 5-ethynyl-2′-deoxyuridine (EdU), phospho-Histone H3 (H3P), and Aurora B kinase (Au-B). Cell-cycle regulators were analyzed via Western blot (WB) and quantitative PCR (qPCR). Anti-apoptotic effects were evaluated following doxorubicin (Dox) exposure. Angiogenesis and fibrosis were assessed via tube formation and α-smooth muscle actin (α-SMA) IF staining. MI was induced in adult mice followed by intramyocardial miRNA delivery. Proliferative response was assessed by IF and molecular methods. Direct targets were identified via Argonaute-2 immunoprecipitation (Ago2-IP) following RNA-sequencing.
Results
MiR-519 significantly increased EdU-incorporation in hiPSC-CM and H3P- and Au-B-positive hiPSC-CM (p<0.01, n=10), with higher expression of Cyclin A2 and suppression of p21 (p<0.01, n=4), indicating progression from cell-cycle re-entry through cytokinesis. Viability was preserved in miR-519-treated hiPSC-CMs after doxorubicin exposure (p<0.05, n=6). In addition, miR-519 enhanced angiogenic activity, increasing mesh formation (p<0.001, n=6) and tube length (p<0.05, n=6) but had no effect on proliferation and α-SMA expression in cFBs (p=ns, n=8). In vivo, miR-519 delivery after MI increased CM proliferation in the infarct border zone, as assessed through H3P and Au-B-staining, suppressed p21 expression (p<0.05, n=5), and decreased fibrotic scar size (p<0.01, n=5), resulting in lower rates of death (p<0.05, n=12). Ago2-IP RNA-sequencing identified key cell cycle inhibitors p21 and p57 as direct targets of miR-519.
Conclusion
This study suggests that MiR-519 promotes cardiac repair through induction of CM proliferation, preserved cell survival and enhanced angiogenic capacity, thereby improving outcomes after myocardial infarction. These findings identify miR-519 as a promising therapeutic candidate for cardiac injury.