MicroRNA miR-24-3p Reduces Apoptosis and Regulates Keap1-Nrf2 Pathway in Mouse Cardiomyocytes Responding to Ischemia/Reperfusion Injury.

Xu Xiao,Daniel H Shaw,Peter X Shaw,Kyle Tran,Briana Che,Victor Lin,Zhigang Lu,Zhihao Wang,Hongjun Du,Adam May

doi:10.1155/2018/7042105

Abstract

In recent years, microRNAs (miRNAs) have received increasing attention for their role in ischemia/reperfusion injury (I/RI), and many miRNAs have been demonstrated to play a very important role in cardiac I/RI. The miRNA miR-24-3p is a tumor suppressor that regulates multiple tumors; however, it remains unclear whether the expression level of miR-24-3p is altered in cardiac cells under I/RI. In this study, we used mouse primary cardiomyocytes and the H9C2 cardiomyocyte cell line to perform in vitro stimulated ischemia/reperfusion (SI/R) and then detected miR-24-3p expression level using quantitative real-time PCR (qRT-PCR). We discovered that the expression of miR-24-3p was significantly increased in cardiomyocytes following SI/R, and that the miR-24-3p level was inversely correlated to the ischemia marker HIF-1a. Furthermore, we transfected cardiomyocytes with miR-24-3p mimic or inhibitor to explore the role of miR-24-3p in cardiomyocyte ischemia/reperfusion injury in vitro. We performed flow cytometry to detect the apoptotic rate of H9C2 cardiomyocytes and found that the transfection of miR-24-3p mimic resulted in the decrease of the apoptosis rate of cardiomyocytes after SI/R, whereas the transfection of miR-24-3p inhibitor increased the number of apoptotic cardiomyocytes. These data suggest that the overexpression of miR-24-3p could reduce in vitro myocardial cell apoptosis induced by I/R injury. Finally, we applied the dual luciferase reporter gene system to verify whether miR-24-3p targets the Keap1 gene, and found that the luciferase signal intensity from a vector carrying the Keap1 wild-type reporter gene was significantly reduced after transfection with miR-24-3p mimic. The Keap1 protein level was also reduced following the transfection of miR-24-3p. The results from this study suggest a novel function of miR-24-3p in protecting cardiomyocytes from ischemia/reperfusion injury by the activation of the Nrf2-Keap1 pathway.

Highlights

Cardiac arrest (CA) is one of the most serious acute clinical conditions, requiring immediate rescue to prevent patient death [1, 2]
In order to investigate the changes of miR-24-3p expression in primary C57BL/6 cardiomyocytes, we performed qRT-PCR to detect the expression of miR-24-3p in stimulated ischemia/reperfusion (SI/R) cardiomyocytes and the control group
These results suggest that the miR-243p expression level is associated with SI/R cardiomyocyte injury and that potentially increasing its expression may have a protective effect

Summary

Introduction

Cardiac arrest (CA) is one of the most serious acute clinical conditions, requiring immediate rescue to prevent patient death [1, 2]. While cardiopulmonary resuscitation (CPR) is an effective rescue technique, the resulting restoration of spontaneous circulation (ROSC) exposes the body to numerous assaults, including acidosis, electrolyte imbalance, hypoxia, ischemia/reperfusion injury (I/RI), and oxidative stress. Following ROSC, patients often experience severe damage to the heart, brain, and other major organs, leading to systemic inflammatory response syndrome and eventually to multiple organ dysfunction [3,4,5]. Patients rescued from CA may exhibit myocardial dysfunction. The mechanism accounting for the different levels of myocardial dysfunction observed in CA-rescued patients remains poorly understood. Recent studies suggest that the degree of myocardial dysfunction

Methods

Results

Conclusion