miR-223 regulates myocardial ischemia-reperfusion damage via targeting NLRP3 in vitro and in vivo

Inflammatory responses are considered one of the leading causes of myocardial injury caused by ischemia-reperfusion (I/R). microRNA (miRNA) has been reported to participate in pathophysiological processes associated with myocardial I/R damage. Herein, we aimed to investigate the effects of the NOD-like receptor family pyrin domain containing 3 (NLRP3) in inflammatory responses and to assess miR-223 on myocardial I/R. Our results showed that cell necrosis and LDH activity were increased in H2O2-treated H9c2 cells in a dose-and time-dependent manner by flow cytometry and LDH detection assay, respectively. NLRP3 mRNA and protein expression were gradually elevated in H2O2-incubated H9c2 cells with increasing exposure time and I/R rats with ischemia time as determined by qRT-PCR and Western blotting. Moreover, overexpression of NLRP3 in H2O2-exposed H9c2 cells up-regulated PI+ cells and LDH activity. Additionally, when pre-challenged with NLRP3 mimic in I/R rats, myosin(+) cells and FSLVD% were increased and decreased respectively as compared to I/R rats. In addition, it was found that miR-223 was significantly reduced in H2O2-treated H9c2 cells over time. Forced miR-223 expression could suppress NLRP3 mRNA and protein expression, thereby we hypothesized that NLRP3 might negatively regulate miR-223, which was demonstrated by dual-luciferase reporter assay. Finally, we found that overexpressed miR-223 inhibited PI+ cells and LDH activity in H2O2-incubated H9c2 cells. Furthermore, miR-223 down-regulated and up-regulated myosin(+) cells and FSLVD% respectively in I/R rats. Thus, our data indicate that miR-223 could exert a cardioprotective role by targeting NLRP3 in H2O2- and I/R-induced myocardial damage in vitro and in vivo, which implies that miR-223 might be considered as a therapeutic target for modulation of NLRP3 in myocardial I/R injury.