Abstract 15253: Post-Transcriptional Downregulation of Let-7 Increases Susceptibility of Cardiac Myocytes to Ischemia-Induced Cell Death
Background: To a large extent, the pathophysiology of the ischemic heart is governed by changes in gene expression in the ischemic myocyte. Recently, microRNAs (miRs) have been shown to regulate gene expression and function in numerous cellular processes. MicroRNA-let-7 has been shown to act as a tumor suppressor and to regulate glucose homeostasis; however, its role in cardiac ischemia is unknown.
Hypothesis: To investigate the role of miR-let-7 in myocardial ischemic injury and identify its targets.
Methods and results: Transcriptional analysis revealed that miR-let-7c is significantly downregulated in a 2 hour permanent left anterior descending (LAD) artery occlusion model (0.695x vs. sham operated controls, p≤0.05). In a cell culture model of ischemia, levels of mature let-7c transcripts were similarly downregulated in neonatal rat ventricular myocytes (NRVM) (0.7x vs normoxic cells, p≤0.01) but not in cardiac fibroblasts. The MEK1/2 inhibitor U0126 blocked this downregulation, suggesting a role for ERK signaling, but cobalt chloride, a HIF-1 stabilizer, did not. Levels of the primary let-7c transcript remained unchanged, demonstrating post-transcriptional regulation of the mature transcript. To study the functional significance of let-7 loss, lentiviral overexpression of let-7c and/or let-7g was performed in NRVMs and effectively prevented the downregulation of mature let-7. MiR-let-7 “rescue” resulted in decreased cell death as determined by LDH release (30% vs 59%, let-7 vs control, p≤0.05) and reduced caspase-3 cleavage (0.35 vs 0.55 n.d.u. let-7 vs control, p≤0.05). Rescue from let-7 downregulation also activated cardioprotective STAT3 phosphorylation and hemeoxygenase induction (each ~2.0x vs control, p≤0.05). Using PAR-CLIP, in which the RISC complex is crosslinked to its target transcripts, we directly demonstrated myocyte let-7 targeting of LIN28A and additional genes.
Conclusion: During ischemic stress, let-7c is repressed in a myocyte-specific manner, which increases its susceptibility to ischemia-induced cell death. Preventing this downregulation increases the survival of myocyte under ischemic stress. Manipulating levels of let-7 may have therapeutic value for myocardial ischemic injury.
- © 2013 by American Heart Association, Inc.