Double overexpression of miR-19a and miR-20a in induced pluripotent stem cell-derived mesenchymal stem cells effectively preserves the left ventricular function in dilated cardiomyopathic rat

Abstract

Abstract Background This study tested the hypothesis that double overexpression of miR-19a and miR-20a (dOex-mIRs) in human induced pluripotent stem cell (iPS)-derived mesenchymal stem cells (MSCs) effectively preserved left ventricular ejection fraction (LVEF) in dilated cardiomyopathy (DCM) (i.e., induced by doxorubicin) rat. Methods and results In vitro study was categorized into groups G1 (iPS-MSC), G2 (iPS-MSCdOex-mIRs), G3 (iPS-MSC + H2O2/100uM), and G4 (iPS-MSCdOex-mIRs + H2O2/100uM). The in vitro results showed the cell viability was significantly lower in G3 than in G1 and G2, and that was reversed in G4 but it showed no difference between G1/G2 at time points of 6 h/24 h/48 h, whereas the flow cytometry of intra-cellular/mitochondrial oxidative stress (DCFA/mitoSOX) and protein expressions of mitochondrial-damaged (cytosolic-cytochrome-C/DRP1/Cyclophilin-D), oxidative-stress (NOX-1/NOX2), apoptotic (cleaved-caspase-3/PARP), fibrotic (p-Smad3/TGF-ß), and autophagic (ratio of LC3B-II/LC3BI) biomarkers exhibited an opposite pattern of cell-proliferation rate (all p< 0.001). Adult-male SD rats (n=32) were equally divided into groups 1 (sham-operated control), 2 (DCM), 3 (DCM + iPS-MSCs/1.2 × 106 cells/administered by post-28 day’s DCM induction), and 4 (DCM + iPS-MSCdOex-mIRs/1.2 × 106 cells/administered by post-28 day’s DCM induction) and euthanized by day 60 after DCM induction. LV myocardium protein expressions of oxidative-stress signaling (p22-phox/NOX-1/NOX-2/ASK1/p-MMK4,7/p-JNK1,2/p-cJUN), upstream (TLR-4/MAL/MyD88/TRIF/TRAM/ TFRA6/IKKα/ß/NF-κB) and downstream (TNF-α/IL-1ß/MMP-9) inflammatory signalings, apoptotic (cleaved-PARP/mitochondrial-Bax), fibrotic (Smad3/TGF-ß), mitochondrial-damaged (cytosolic-cytochrome-C/DRP1/cyclophilin-D), and autophagic (beclin1/Atg5) biomarkers were highest in group 2, lowest in group 1 and significantly lower in group 4 than in group 3, whereas the LVEF exhibited an opposite pattern of oxidative stress (all p< 0.0001). Conclusion iPS-MSCdOex-mIRs therapy was superior to iPS-MSC therapy for preserving LV function in DCM rat.

Keywords

• Dilated cardiomyopathy
• Double overexpression of microRNAs
• Oxidative stress
• Inflammation
• Mitochondrial damage