A novel system-level approach using RNA-sequencing data identifies miR-30-5p and miR-142a-5p as key regulators of apoptosis in myocardial infarction

Jin Ock Kim,Wan Seok Kang,Seong Eui Hong,Youngkeun Ahn,Kyoung Jin Nho,Do Han Kim,Jei Hyoung Park,Taeyong Kim,Yong Sook Kim,Ji Young Lee,Chunghee Cho

doi:10.1038/s41598-018-33020-x

Abstract

This study identified microRNAs involved in myocardial infarction (MI) through a novel system-level approach using RNA sequencing data in an MI mouse model. This approach involved the extraction of DEGs and DEmiRs from RNA-seq data in sham and MI samples and the subsequent selection of two miRNAs: miR-30-5p (family) and miR-142a-5p, which were downregulated and upregulated in MI, respectively. Gene Set Enrichment Analysis (GSEA) using the predicted targets of the two miRNAs suggested that apoptosis is an essential gene ontology (GO)-associated term. In vitro functional assays using neonatal rat ventricular myocytes (NRVMs) demonstrated that miR-30-5p is anti-apoptotic and miR-142a-5p is pro-apoptotic. Luciferase assays showed that the apoptotic genes, Picalm and Skil, and the anti-apoptotic genes, Ghr and Kitl, are direct targets of miR-30-5p and miR-142a-5p, respectively. siRNA studies verified the results of the luciferase assays for target validation. The results of the system-level high throughput approach identified a pair of functionally antagonistic miRNAs and their targets in MI. This study provides an in-depth analysis of the role of miRNAs in the pathogenesis of MI which could lead to the development of therapeutic tools. The system-level approach could be used to identify miRNAs involved in variety of other diseases.

Highlights

Myocardial infarction (MI), a leading cause of death worldwide, is associated with a sudden loss of oxygen and nutrient supply to cardiac cells[1]
We demonstrated that miR-30-5p is anti-apoptotic, while miR-142a-5p is pro-apoptotic, and the apoptotic genes, Picalm and ski-like proto-oncogene (Skil), are direct targets of miR-30-5p, and the anti-apoptotic genes, growth hormone receptor (Ghr) and kit ligand (Kitl) are direct targets of miR-142a-5p
Left ventricle (LV) fractional shortening (FS) and ejection fraction (EF) were significantly reduced in 1 W and 8 W after MI induction compared with sham group (P < 0.05) (Supplementary Fig. 1B,C)

Summary

Introduction

Myocardial infarction (MI), a leading cause of death worldwide, is associated with a sudden loss of oxygen and nutrient supply to cardiac cells[1]. The expression of individual genes can be co-targeted by multiple miRNAs. the identification of a group of miRNAs that exert synergistic or antagonistic effects on disease may have important therapeutic implications. Most high-throughput studies on heart disease have been carried out using single layer-omics data www.nature.com/scientificreports/. Through integrated analysis using miRNA and mRNA expression data, Wang et al demonstrated that increased levels of miR-146b-5p mediates atrial fibrosis in patients with nonvalvular paroxysmal atrial fibrillation by targeting TIMP-44. We performed RNA-sequencing for both mRNA and miRNA, a revolutionary alternative that overcomes the limitations of microarray methods[5], to simultaneously profile the transcriptomes and miRomes at the early, middle, and end stages of MI. By using a two-layer omics data integration with a logical top-down approach, the miRNA-target networks implicated in the progression of MI were identified. We demonstrated that miR-30-5p is anti-apoptotic, while miR-142a-5p is pro-apoptotic, and the apoptotic genes, Picalm and Skil, are direct targets of miR-30-5p, and the anti-apoptotic genes, Ghr and Kitl are direct targets of miR-142a-5p

Methods

Results

Conclusion