Inhibition of circDGKZ ameliorates myocardial ischemia/reperfusion injury by targeting miR-345-5p/TLR4.

Abstract

This study aims to explore the molecular mechanism of circular RNAs' (circRNAs) potential involvement in myocardial ischaemia-reperfusion injury (MIRI). Differently expressed genes in myocardial infarction (MI) were identified by screening the GEO database. Serum was collected from MI patients and healthy volunteers (n=5 for each group). AC16 cells were cultured and exposed to hypoxia/reperfusion (H/R) treatment for the cell experiments. Then candidate genes were validated in human serum and the H/R model. Quantitative real-time PCR and western blot were used to detect expression of key molecules such as circDGKZ, miR-345-5p, and Toll-like receptor 4 (TLR4), as well as pyroptosis markers such as NOD-like receptor thermal protein domain-associated protein 3 (NLRP3), ASC, C-caspase1, interleukin (IL)-1β, and IL-18. CircDGKZ was positively correlated in human serum (P<0.05) and in AC16 cells (P<0.01). Knockdown of circDGKZ inhibited cardiomyocyte pyroptosis and the TLR4/nuclear factor kappa B (NF-κB) signalling pathway (all P<0.05). A luciferase assay was used to detect the molecule interaction. MiR-345-5p was regulated by circDGKZ and regulated TLR4 in cardiomyocytes both through direct interaction (P<0.01). The stability and distribution of circRNA or linear RNA were examined by subcellular localization and RNA decay assays. CircDGKZ was stably expressed in cardiomyocytes and mainly distributed in the cytoplasm (P<0.01). Knockdown of circDGKZ also promoted the degradation of NLRP3 by inducing autophagy (P<0.05). MIRI rat models were constructed (n=5 for each group), and the cellular results were further confirmed in rat models (P<0.05). Knockdown of circDGKZ interrupted pyroptosis and induced autophagy of cardiomyocytes via regulating miR-345-5p/TLR4/NF-κB.