lncRNA ANRIL protects H9c2 cells against hypoxia-induced injury through targeting the miR-7-5p/SIRT1 axis.

BACKGROUND: Acute myocardial infarction (AMI) occurred in the heart, which underwent long-term ischemia, and was mainly caused by hypoxia. Recently, studies have uncovered the participation of long noncoding RNAs (lncRNAs) in the pathogenesis of heart disease. Here, we planned to probe the role and molecular basis of ANRIL in hypoxia-induced H9c2 cell injury. METHODS: Trypan blue exclusion assay and Transwell and flow cytometry assays were conducted to assess hypoxia-induced injury by determining the viability, migration, invasion, and apoptosis of H9c2 cells in different conditions, respectively. Gene expressions were evaluated by quantitative real-time polymerase chain reaction or western blot analysis as needed. RNA immunoprecipitation and luciferase reporter assays were applied to confirm the associations among genes. RESULTS: ANRIL expression was dramatically enhanced in hypoxia-injured H9c2 cells, and silencing ANRIL aggravated hypoxia-induced H9c2 cell injury. ANRIL positively regulated sirtuin 1 (SIRT1) expression via competitively binding with miR-7-5p. Moreover, inhibition of miR-7-5p counteracted ANRIL depletion-exacerbated injury in hypoxic H9c2 cells, meanwhile, forced SIRT1 expression attenuated the injury-promoting effect of miR-7-5p upregulation on hypoxic H9c2 cells. CONCLUSION: Our findings disclosed that ANRIL plays a protective part in hypoxia-induced H9c2 cell injury via modulating the miR-7-5p/SIRT1 axis, suggesting the great potential of ANRIL as a protective target for AMI.

Troxerutin attenuates myocardial cell apoptosis following myocardial ischemia-reperfusion injury through inhibition of miR-146a-5p expression.

The aim of the current study was to investigate the effects and the underlying mechanisms of troxerutin on myocardial cell apoptosis during ischemia-reperfusion (I/R) injury. Hypoxia/reoxygenation (H/R) model in neonatal rat cardiomyocytes, and I/R model in rats, were established following troxerutin preconditioning. The quantitative real-time polymerase chain reaction analysis was performed to examine the messenger RNA miR-146a-5p expression in cardiomyocytes and myocardial tissues. Hemodynamic parameters and serum creatine kinase, lactate dehydrogenase, tumor necrosis factor-α, and interleukin-10 were evaluated. Infarct size was examined by 2,3,5-triphenyltetrazolium chloride staining. Besides, myocardial apoptosis was detected by terminal deoxynucleotidyl transferase (dUTP) nick end labeling (TUNEL) assay. Western blot analysis was performed to determine the protein levels of caspase-3, Bax, and Bcl-2. The results showed that, troxerutin decreased rat cardiomyocyte apoptosis during H/R injury. Furthermore, the antiapoptotic effect of troxerutin against I/R injury was mediated by miR-146a-5p downregulation. In vivo experiments suggested that troxerutin alleviated myocardial I/R injury in rats via inhibition of miR-146a-5p. In conclusion, troxerutin exerted cardioprotective effects during I/R injury by downregulating miR-146a-5p.

Troxerutin Protects Against Myocardial Ischemia/Reperfusion Injury Via Pi3k/Akt Pathway in Rats.

BACKGROUND/AIMS: Troxerutin, also known as vitamin P4, has been commonly used in the treatment of chronic venous insufficiency (CVI) disease. However, its effect on in vivo myocardial ischemia/reperfusion (I/R) injury, a model that closely mimics acute myocardial infarction in humans, is still unknown. METHODS: The myocardial I/R injury rat model was created with troxerutin preconditioning. Myocardial infarct size was evaluated by the Evans blue-TTC method. Hemodynamic parameters, including the heart rate (HR), left ventricular end-diastolic pressure (LVEDP), left ventricular systolic pressure (LVSP), maximal rate of rise in blood pressure in the ventricular chamber (+dp/dt max), and maximal rate of decline in blood pressure in the ventricular chamber (-dp/dt max) were monitored. Serum TNF-α and IL-10 were determined by ELISA kit. Cell apoptosis was detected by MTT method. RESULTS: Troxerutin preconditioning significantly reduced myocardial infarct size, improved cardiac function, and decreased the levels of creatine kinase (CK), aspartate aminotransferase (AST) and lactate dehydrogenase (LDH) in the I/R injury rat model. The serum and mRNA levels of TNF-α and IL-10 as well as some apoptosis markers (Bax, Caspase 3) also decreased. Moreover, troxerutin pretreatment markedly increased the phosphorylation of Akt, and blocking PI3K activity by LY294002 abolished the protective effect of troxerutin on I/R injury. CONCLUSION: Troxerutin preconditioning protected against myocardial I/R injury via the PI3K/Akt pathway.

Oxidized phospholipids facilitate calcific aortic valve disease by elevating ATF4 through the PERK/eIF2α axis.

In this study we sought to analyze the critical role of oxidized phospholipid (OxPL) in the progression of calcific aortic valve disease (CAVD) with the involvement of activating transcription factor 4 (ATF4). Differentially expressed genes related to CAVD were identified using bioinformatics analysis. Expression of ATF4 was examined in mouse models of aortic valve calcification (AVC) induced by the high cholesterol (HC) diet. Valvular interstitial cells (VICs) were then isolated from mouse non-calcified valve tissues, induced by osteogenic induction medium (OIM) and co-cultured with OxPAPC-stimulated macrophages. The effect of OxPLs regulating ATF4 on the macrophage polarization and osteogenic differentiation of VICs was examined with gain- and loss-of-function experiments in VICs and in vivo. In aortic valve tissues and OIM-induced VICs, ATF4 was highly expressed. ATF4 knockdown alleviated the osteogenic differentiation of VICs, as evidenced by reduced expression of bone morphogenetic protein-2 (BMP2), osteopontin (OPN), and osteocalcin. In addition, knockdown of ATF4 arrested the AVC in vivo. Meanwhile, OxPL promoted M1 polarization of macrophages and mediated osteogenic differentiation of VICs. Furthermore, OxPL up-regulated ATF4 expression through protein kinase R-like endoplasmic reticulum kinase (PERK)/eukaryotic translation initiation factor 2 subunit alpha (eIF2α) pathway. In conclusion, OxPL can potentially up-regulate the expression of ATF4, inducing macrophages polarized to M1 phenotype, osteogenic differentiation of VICs and AVC, thus accelerating the progression of CAVD.

Exosomal miR-129 and miR-342 derived from intermittent hypoxia-stimulated vascular smooth muscle cells inhibit the eIF2α/ATF4 axis from preventing calcified aortic valvular disease.

This study aims to elucidate the role of miR-129/miR-342 loaded in exosomes derived from vascular smooth muscle cells (VSMCs) stimulated by intermittent hypoxia in calcified aortic valvular disease (CAVD). Bioinformatics analysis was conducted to identify differentially expressed miRs in VSMCs-derived exosomes and CAVD samples, and their potential target genes were predicted. VSMCs were exposed to intermittent hypoxia to induce stimulation, followed by isolation of exosomes. Valvular interstitial cells (VICs) were cultured in vitro to investigate the impact of miR-129/miR-342 on VICs' osteogenic differentiation and aortic valve calcification with eIF2α. A CAVD mouse model was established using ApoE knockout mice for in vivo validation. In CAVD samples, miR-129 and miR-342 were downregulated, while eIF2α and ATF4 were upregulated. miR-129 and miR-342 exhibited inhibitory effects on eIF2α through targeted regulation. Exosomes released from intermittently hypoxia-stimulated VSMCs contained miR-129 and miR-342. Overexpression of miR-129 and miR-342, or silencing ATF4, suppressed VICs' osteogenic differentiation and aortic valve calcification, which could be rescued by overexpressed eIF2α. Collectively, intermittent hypoxia stimulation of VSMCs leads to the secretion of exosomes that activate the miR-129/miR-342 dual pathway, thereby inhibiting the eIF2α/ATF4 axis and attenuating VICs' osteogenic differentiation and CAVD progression.

Cardioprotective Effect of circ_SMG6 Knockdown against Myocardial Ischemia/Reperfusion Injury Correlates with miR-138-5p-Mediated EGR1/TLR4/TRIF Inactivation.

Increased neutrophil recruitment represents a hallmark event in myocardial ischemia/reperfusion (I/R) injury due to the ensuing inflammatory response. Circular RNAs (circRNAs) are important regulatory molecules involved in cell physiology and pathology. Herein, we analyzed the role of a novel circRNA circ_SMG6 in the regulation of neutrophil recruitment following I/R injury, which may associate with the miR-138-5p/EGR1/TLR4/TRIF axis. Myocardial I/R injury was modeled in vivo by ligation of the left anterior descending (LAD) artery followed by reperfusion in mice and in vitro by exposing a cardiomyocyte cell line (HL-1) to hypoxia/reoxygenation (H/R). Gain- and loss-of-function experiments were performed to evaluate the effect of the circ_SMG6/miR-138-5p/EGR1/TLR4/TRIF axis on cardiac functions, myocardial infarction, myocardial enzyme levels, cardiomyocyte activities, and neutrophil recruitment. We found that the EGR1 expression was increased in myocardial tissues of I/R mice. Knockdown of EGR1 was found to attenuate I/R-induced cardiac dysfunction and infarction area, pathological damage, and cardiomyocyte apoptosis. Mechanistic investigations showed that circ_SMG6 competitively bound to miR-138-5p and consequently led to upregulation of EGR1, thus facilitating myocardial I/R injury in mice and H/R-induced cell injury. Additionally, ectopic EGR1 expression augmented neutrophil recruitment and exacerbated the ensuing I/R injury, which was related to the activated TLR4/TRIF signaling pathway. Overall, our findings suggest that circ_SMG6 may deteriorate myocardial I/R injury by promoting neutrophil recruitment via the miR-138-5p/EGR1/TLR4/TRIF signaling. This pathway may represent a potential therapeutic target in the management of myocardial I/R injury.

Circ_ZNF512-Mediated miR-181d-5p Inhibition Limits Cardiomyocyte Autophagy and Promotes Myocardial Ischemia/Reperfusion Injury through an EGR1/mTORC1/TFEB-Based Mechanism.

Studies have shown that circRNAs are important regulatory molecules involved in cell physiology and pathology. Herein, we analyzed the role of circ_ZNF512 in cardiomyocyte autophagy of myocardial ischemia/reperfusion (I/R) injury. A mouse model was induced by ligation of the left anterior descending artery followed by reperfusion. An in vitro model was also developed in cultured cardiomyocytes following hypoxia/reoxygenation (H/R) injury. It was established that EGR1 expression was increased in myocardial tissues of I/R mice and H/R-induced cardiomyocytes. Silencing of circ_ZNF512 attenuated its binding to miR-181d-5p, which in turn impaired the EGR1 expression by targeting its 3'-UTR, thus promoting the autophagy of cardiomyocytes and suppressing cell apoptosis to alleviate myocardial tissue injury. Additionally, the circ_ZNF512/miR-181d-5p/EGR1 crosstalk activated the mTORC1/TFEB signaling pathway, increasing mTORC1 expression while suppressing TFEB expression. Together, circ_ZNF512 knockdown protects against myocardial I/R injury, which may be a potential therapeutic approach for preventing myocardial I/R injury.

Simvastatin Improves Myocardial Ischemia Reperfusion Injury through KLF-Regulated Alleviation of Inflammation.

OBJECTIVE: To clarify the protective effect of simvastatin on myocardial ischemia reperfusion injury (MIRI) and the underlying mechanism. MATERIALS AND METHODS: The MIRI model in rats was firstly constructed. Twenty-four male rats were randomly assigned into the sham group, ischemia-reperfusion (I/R) group, and simvastatin group, with 8 rats in each group. Contents of superoxide dismutase (SOD) and malondialdehyde (MDA), as well as serum levels of CK and inflammatory factors, in rats were determined by the enzyme-linked immunosorbent assay (ELISA). Lactate dehydrogenase (LDH) activity in the three groups was examined. Through flow cytometry and Cell Counting Kit-8 (CCK-8) assay, apoptosis and viability in each group were detected, respectively. Relative levels of HMGB1, Kruppel-like factor 2 (KLF2), eNOS, and thrombomodulin (TM) were finally determined. RESULTS: Simvastatin treatment markedly enhanced SOD activity and reduced contents of MDA, LDH, and creatine kinase (CK) in MIRI rats. The increased apoptosis and decreased viability following MIRI were partially reversed by simvastatin treatment. Besides, MIRI resulted in the upregulation of inflammatory factors and chemokines. Their elevations were abolished by simvastatin. In MIRI rats, simvastatin upregulated KLF2 and p-eNOS. CONCLUSIONS: Simvastatin protects inflammatory response at post-MIRI through upregulating KLF2, thus improving cardiac function.

LncRNA CRNDE inhibits cardiomyocytes apoptosis by YAP1 in myocardial ischaemia/reperfusion injury.

BACKGROUND: Cardiomyocytes apoptosis is the basic pathological process of myocardial ischaemia/reperfusion (MI/R) injury, so inhibiting apoptosis of cardiomyocytes can effectively improve MI/R injury. Long non-coding RNA colorectal neoplasia differentially expressed (lncRNA CRNDE) can inhibit cell apoptosis, but its specific role in MI/R injury has not been studied. The aim of this study is to explore the specific effect of lncRNA CRNDE on cardiomyocytes apoptosis. METHODS: MI/R model in vivo and hypoxia/re-oxygenation (H/R) model in vitro were constructed. Apoptotic levels were assessed by TUNEL staining assay. QRT-PCR was used to validate lncRNA CRNDE level in myocardial tissues and HL-1 cells. The protein expressions of YAP1, Bcl-2 and cleaved caspase-3 were detected by western blot analysis. Flow cytometry was used to determine the apoptosis rate of cardiomyocytes. RIP assay was used to detect the interaction between lncRNA CRNDE and YAP1. RESULTS: The extent of cardiomyocytes apoptosis was significantly increased, and the levels of lncRNA CRNDE, YAP1 and Bcl-2 were down-regulated, while cleaved caspase-3 expression was up-regulated in MI/R mice and H/R-treated HL-1 cells. The expressions of YAP1 and Bcl-2 were decreased, while the expression of cleaved caspase-3 was increased after the knockdown of lncRNA CRNDE. Furthermore, lncRNA CRNDE could bind to YAP1 and regulated the protein level of YAP1 by ubiquitination and proteasomal degradation pathway. After transfection of Si-YAP1 in the H/R-treated HL-1 cells transfected with pc-DNA CRNDE, the protein level of Bcl-2 was decreased, while cleaved caspase-3 expression and the apoptosis rate were increased. CONCLUSION: Our study suggested that lncRNA CRNDE could regulate YAP1 level by ubiquitination and proteasomal degradation pathway, thus inhibiting cardiomyocytes apoptosis in MI/R injury.

Heart Failure and Main Pulmonary Artery Obstruction Caused by Right Ventricular Metastatic Adult Hepatoblastoma: A Case Report.

We report a case of a 18-year-old boy who presented with dyspnea and right upper quadrant abdominal dull pain. According to urgent echocardiography, a dense sessile mass occupied the right ventricule. Tumor resection was performed, followed by further adjuvant therapy. The specimen was histopathologically investigated and eventually diagnosed as metastatic adult hepatoblastoma. We discuss its clinical features and treatment in the light of the current knowledge. It is important for us to be aware that adjuvant chemotherapy might be an effective alternative in the treatment of hepatoblastoma combined with ventricle invasion. Early cardiac surgery may be advised in patients with cardiac function impairment.

Pulmonary mucoepidermoid carcinoma in Chinese population: a clinicopathological and radiological analysis.

BACKGROUND: Pulmonary mucoepidermoid carcinoma (PMEC) is a rare malignant neoplasm with remarkable resemblance to mucoepidermoid carcinoma of the salivary glands. It constitutes a unique set of patient population. In this study we briefly discussed the current state of knowledge of PMEC and described the clinical presentation and management of 27 PMEC cases. This study aimed to discuss the utility of surgical treatment in the patients with pulmonary mucoepidermoid carcinoma. METHODS: We retrospectively studied 27 cases with the diagnosis of PMEC, divided into low grade and high grade based on histopathological characteristics. The clinical symptoms, radiological manifestations, pathological characteristics, treatment strategy and prognosis were systemically analyzed. RESULTS: The tumor could occur in any lobe of the lungs. The treatment included surgical intervention and/or adjuvant therapies. While the sex-age distribution and initial staging was not different between low- and high- grade PMEC, the disease control rate (95%) and 5 year survival (95%) were much higher in low-grade PMEC than the high-grade cases (57.1% and 42.9%, respectively). CONCLUSION: The clinical, radiographical and pathological features of PMECs were systemically analyzed and summarized, and the utility of pathological grading system as the independent prognostic factor in addition to clinical staging was confirmed.