Silencing of lncRNA NEAT1 alleviates acute myocardial infarction by suppressing miR-450-5p/ACSL4-mediated ferroptosis.

Ferroptosis is principally initiated by dysregulation of iron metabolism and excessive accumulation of ROS, which exacerbates myocardial injury during acute myocardial infarction (AMI). Previous studies have indeed demonstrated the significant involvement of long non-coding RNA (lncRNA) nuclear paraspeckle assembly transcript 1 (NEAT1) exerts its pleiotropic effects in the pathophysiology of myocardial infarction, heart failure and atherosclerosis by modulating inflammation, apoptosis, and oxidative stress. However, whether and how NEAT1 mediates myocardial ferroptosis remain unknown. In this study, we found that NEAT1 expression was significantly elevated in hypoxic HL-1 cells and AMI mice, while silencing of NEAT1 alleviated lipid peroxidation and myocardial ferroptosis both in vitro and in vivo. Mechanistically, NEAT1 directly sponged miR-450b-5p and negatively regulated its expression. In addition, miR-450b-5p directly targeted Acyl-CoA synthase long-chain family member 4 (ACSL4). Notably, inhibition of miR-450b-5p reversed the role of NEAT1 in AMI mice. Collectively, these findings newly illustrated that NEAT1 acts as a competitive endogenous RNA (ceRNA) of miR-450-5p in AMI. Especially, silencing of NEAT1 effectively ameliorated myocardium ischemia by suppression of ferroptosis via miR-450-5p/ACSL4 pathway, which providing a brand-new therapeutic strategy for myocardial ischemia injury.

Yu Q ，Li Y ，Zhang N ，Lu J ，Gan X ，Chen L ，Liang R ，Jian J ... -  《-》

EGCG attenuated acute myocardial infarction by inhibiting ferroptosis via miR-450b-5p/ACSL4 axis.

Epigallocatechin gallate (EGCG) has multiple biological effects such as anti-tumor multiple drug resistance, antioxidation and anti-inflammatory properties. Ferroptosis is the main driving factor of ischemic heart injury, thus inhibiting ferroptosis may prove to be an effective treatment strategy for cardiovascular diseases. However, the role of EGCG on ferroptosis in ischemic myocardium and underlying mechanisms remain uncertain. This study was aimed to investigate the effects and potential mechanisms of EGCG on myocardial ischemic-induced ferroptosis both in vitro and in vivo. Cardiomyocyte hypoxia model and mouse acute myocardial infarction (AMI) model were established in vitro and in vivo. MiR-450b-5p and ACSL4 silencing or overexpression plasmids were transfected, with or without EGCG pretreatment. Cell viability was determined by the CCK-8 assay. Hematoxylin and eosin (HE) staining and transmission electron microscopy (TEM) were used to evaluate the morphologic alterations. TTC staining was used to observe the infarction area, and echocardiography was adopted to appraise the heart function. Using flow cytometry, the presence of reactive oxygen species (ROS) was assessed. The content of cardiac troponin I (cTn I), glutathione (GSH), malondialdehyde (MDA), divalent iron ions (Fe2+) and superoxide dismutase (SOD) were detected using reagent kits. A luciferase activity assay was performed to assess the binding ability of miR-450b-5p to ACSL4. Expressions of related genes and proteins were measured by RT-qPCR and western blotting respectively. EGCG attenuated AMI-induced ferroptosis and improved myocardial ischemia injury, which was associated with reducing iron deposition and cTn I, inhibition of lipid peroxidation, decreasing TFR1 and ACSL4, and upregulating SLC7A11, FTH1 and GPX4. Meanwhile, EGCG pretreatment increased miR-450b-5p expression in ischemic myocardium. Further researches discovered that knockdown of miR-450b-5p partially compromised EGCG-generated protective effect in hypoxia HL-1 cells, while combination with miR-450b-5p mimic could strengthen the potency of EGCG on ischemic myocardium. The dual-luciferase test demonstrated that miR-450b-5p has binding to ACSL4. Furthermore, silencing of ACSL4 synergistically increased the cardioprotective effect of EGCG. More significantly, EGCG treatment regulated the ferroptosis-related proteins expression via miR-450b-5p/ACSL4 axis. In summary, the present study evidently demonstrated that EGCG attenuates myocardial ischemia injury by targeting ferroptosis. Our work revealed the role of miR-450b-5p/ACSL4 axis in AMI for the first time. Further, it also elucidated the molecular mechanisms of EGCG on inhibiting ferroptosis greatly depend on the miR-450b-5p/ACSL4 axis, suggesting that EGCG may act as a novel anti-ferroptosis agent and exert a therapeutic role in AMI.

Yu Q ，Zhang N ，Gan X ，Chen L ，Wang R ，Liang R ，Jian J ... -  《-》

lncRNA NEAT1 Downregulation Ameliorates the Myocardial Infarction of Mice by Regulating the miR-582-5p/F2RL2 Axis.

This study is aimed at effectively investigating the role of coagulation factor II thrombin receptor like 2 (F2RL2) in myocardial infarction (MI) as well as the upstream regulatory miRNA and lncRNA. Regulatory genes of F2RL2 were analyzed using StarBase and verified by dual-luciferase reporter assay. The MI mouse model was established. The left ventricular ejection fraction (EF) and fractional shortening (FS) were examined by echocardiography. The infarct area, pathological changes, and cell apoptosis in mouse myocardial tissue were evaluated using triphenyltetrazolium chloride and Evans blue, hematoxylin-eosin, and TUNEL staining assays. Oxygen-glucose deprivation- (OGD-) induced human cardiac myocytes (HCMs) were cultured and transfected. The cell viability, proliferation, and apoptosis were determined by CCK-8, EdU staining, and flow cytometry assays. The expressions of F2RL2, miR-582-5p, and nuclear paraspeckle assembly transcript 1 (NEAT1) in myocardial tissues and HCMs were quantified by qRT-PCR or Western blot. NEAT1 sponged miR-582-5p which targeted F2RL2. NEAT1 and F2RL2 were highly expressed while miR-582-5p was lowly expressed in MI mice. F2RL2 downregulation prevented the reduction in EF and SF and the elevation in infarct area and cell apoptosis of MI mice. Both F2RL2 and NEAT1 downregulations reversely modulated the decreased viability and proliferation and the increased apoptosis of OGD-induced HCMs, while miR-582-5p inhibitor did oppositely. NEAT1 silencing upregulated miR-582-5p level but downregulated F2RL2 level. miR-582-5p inhibitor upregulated the F2RL2 level. The role of NEAT1 silencing in OGD-induced HCMs was reversed by miR-582-5p inhibitor whose effect was further offset by F2RL2 downregulation. NEAT1 downregulation ameliorates MI by regulating the miR-582-5p/F2RL2 axis, providing novel biomarkers for MI treatment.

Wu Z ，Bai Y ，Qi Y ，Chang C ，Jiao Y ，Bai Y ，Guo Z ... -  《-》

Suppression of long noncoding RNA NEAT1 attenuates hypoxia-induced cardiomyocytes injury by targeting miR-378a-3p.

lncRNA NEAT1 is involved in the development of many diseases. However, the function of lncRNA NEAT1 in myocardial infarction is unclear. Therefore, this experimental design based on lncRNA NEAT1 to explore the pathogenesis of myocardial infarction. RT-qPCR was used to detect the expression of lncRNA NEAT1 and miR-378a-3p in peripheral blood and mouse cardiomyocytes of patients with myocardial infarction. MTT assay, flow cytometry, Caspase-3 kit and transwell assay were used to detect the effects of lncRNA NEAT1 and miR-378a-3p on cardiomyocyte proliferation, apoptosis and migration. Target gene prediction and screening, luciferase reporter assays were used to verify downstream target genes for lncRNA NEAT1 and miR-378a-3p. Western blotting was used to detect the protein expression of Atg12 and related autophagy genes. lncRNA NEAT1 was highly expressed in peripheral blood and mouse cardiomyocytes of patients with myocardial infarction. Moreover, lncRNA NEAT1 significantly promoted cell proliferation and migration of cardiomyocytes. In addition, lncRNA NEAT1 inhibited miR-378a-3p expression, and miR-378a-3p inhibited Atg12 expression, while lncRNA NEAT1 regulated expression of Atg12 and related autophagic factors via miR-378a-3p. Knockout of microRNA-378-3p reversed the effects of NEAT1 silencing on cell damage. lncRNA NEAT1 can regulate the proliferation of cardiomyocytes by regulating miR-378-3p/Atg12 axis, thus accelerating the occurrence and development of cardiomyocytes.

Zhao J ，Chen F ，Ma W ，Zhang P ... -  《-》

[IDI2-AS1 influences the development of acute myocardial infarction by regulating NR4A2 through microRNA-33b-5p].

Wu S ，Pang Z ，Wang R ，Cui J ，Li W ，Yang X ，Yao Z ... -  《-》