Platelet-Derived Microvesicles Mediate Cardiomyocyte Ferroptosis by Transferring ACSL1 During Acute Myocardial Infarction.

Acute myocardial infarction (AMI) is one of the critical health conditions often caused by the rupture of unstable coronary artery plaque, triggering a series of events, such as platelet activation, thrombus formation, coronary artery blockage, lasted severe ischemia, and hypoxia in cardiomyocytes, and culminating in cell death. Platelet-derived microvesicles (PMVs) act as intermediates for cellular communication. Nevertheless, the role of PMVs in myocardial infarction remains unclear. Initially, AMI-related messenger ribose nucleic acid (mRNA) and micro RNA (miRNA) datasets from the Gene Expression Omnibus (GEO) database were analyzed, specifically focusing on the expressed genes associated with Ferroptosis. Further, a miRNA-mRNA regulatory network specific to AMI was constructed. Then, the effect of PMVs on cardiomyocyte survival was further confirmed through in vitro experiments. High ACSL1 expression was observed in the platelets of AMI patients. The gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed that ACSL1, located in the mitochondria, played a key role in the PPAR signaling pathway. The elevated ACSL1 expression in a co-culture model of PMVs and AC16 cardiomyocytes significantly increased the AC16 cell Ferroptosis. Further, we validated that the platelet ACSL1 expression could be regulated by hsa-miR-449a. Together, these findings suggested that platelet ACSL1 could trigger myocardial cell death via PMV transport. In addition, this research provided a theoretical framework for attenuating myocardial cell Ferroptosis in patients with acute myocardial infarction.

Exosomal circ_HIPK3 reduces apoptosis in H2O2-induced AC16 cardiomyocytes through miR-33a-5p/IRS1 axis.

BACKGROUND: Exosomal circular RNAs (circRNAs) has been revealed to participate in the processes of cellular angiogenesis, growth and metastasis. Herein, the goal of this work was to investigate the role of exosomal circ_HIPK3 in cardiomyocyte apoptosis. METHODS: Exosomes were isolated using ultracentrifugation method and observed by transmission electron microscopy (TEM). Western blot was used to detect exosomes markers. The experimental group AC16 cells were exposed to hydrogen peroxide (H2O2). Levels of genes and proteins was detected by qRT-PCR and Western blot. EdU assay, CCK8 assay, flow cytometry, and Western blot were utilized to detect the function of exosomal circ_HIPK3 in proliferation, and apoptosis. The target relationship between miR-33a-5p and circ_HIPK3 or IRS1 (insulin receptor substrate 1). RESULTS: Circ_HIPK3 was packaged into exosomes and derived from AC16 cells. The expression of circ_HIPK3 was decreased by H2O2 treatment in AC16 cells, which also led to the decrease of circ_HIPK3 in exosomes. Functional analysis showed exosomal circ_HIPK3 promoted AC16 cell proliferation and reduced cell apoptosis under H2O2 treatment. Mechanistically, circ_HIPK3 acted as a sponge of miR-33a-5p to up-regulate the expression of its target IRS1. Functionally, forced expression of miR-33a-5p reversed the reduction of exosomal circ_HIPK3 in apoptosis of H2O2-stimulated AC16 cells. Moreover, miR-33a-5p inhibition contributed to the proliferation of H2O2-stimulated AC16 cells, which was abolished by IRS1 silencing. CONCLUSION: Exosomal circ_HIPK3 reduced H2O2-induced AC16 cardiomyocyte apoptosis through miR-33a-5p/IRS1 axis, suggesting a novel insight into the pathology of myocardial infarction.

LncRNA CDKN2B-AS1 hinders the proliferation and facilitates apoptosis of ox-LDL-induced vascular smooth muscle cells via the ceRNA network of CDKN2B-AS1/miR-126-5p/PTPN7.

OBJECTIVE: The patterns of lncRNA CDKN2B-AS1 in coronary heart disease (CHD) have been extensively studied. This study investigated the competing endogenous RNA (ceRNA) network of CDKN2B-AS1 in coronary atherosclerosis (CAS). METHODS: Microarray analyses were performed to screen out the CHD-related lncRNAs (CDKN2B-AS1) and the downstream microRNAs (miR-126-5p). The expression of CDKN2B-AS1 in serum of patients with CHD and healthy volunteers was detected. Vascular smooth muscle cells (VSMCs) were treated with oxidized low density lipoprotein (ox-LDL) to establish the cell model. Then pcDNA-CDKN2B-AS1 and/or miR-126-5p mimic were transfected into ox-LDL-treated VSMCs to estimate cell proliferation, apoptosis and inflammation. The ceRNA network of CDKN2B-AS1 along with the possible pathway in CHD was testified. RESULTS: CDKN2B-AS1 expression was low in patients with CHD and ox-LDL-treated VSMCs. Upon CDKN2B-AS1 overexpression, TNF-α, NF-κB and IL-1ß levels in VSMCs were decreased, the proliferation of VSMCs was inhibited and the apoptosis rate was increased. Overexpression of miR-126-5p could reverse these trends. CDKN2B-AS1 as a ceRNA competitively bound to miR-126-5p to upregulate PTPN7. CDKN2B-AS1 inhibited VSMC proliferation and accelerated apoptosis by inhibiting the PI3K-Akt pathway. CONCLUSION: LncRNA CDKN2B-AS1 upregulates PTPN7 by absorbing miR-126-5p and inhibits the PI3K-Akt pathway, thus hindering the proliferation and accelerating apoptosis of VSMCs induced by ox-LDL, thus being a therapeutic approach for CAS.

Proatherogenic stimuli induce HuR in atherosclerosis through MAPK/ErK pathway.

Atherosclerosis is a chronic inflammatory disease inflicting the arterial wall, and endothelial activation and dysfunction play an important role in its pathogenesis. The RNA-binding protein HuR has been associated with events of inflammation and activation in endothelial cells, however, its connection with atherosclerosis remains unclear. Here, we show that the expression and RNA-binding activity of HuR are upregulated in human and mouse atherosclerotic lesions. In addition, proatherogenic stimuli, such as inflammatory lipids (Ox-PAPC) and cytokines (TNF-α and IL-1ß), induce HuR in human aortic endothelial cells (HAECs) in vitro. Moreover, HuR is also induced in mouse aorta ECs fed a high-fat diet, and the inducible degree is correlated with proatherogenic hyperlipidemia. We further show that the MAPK/ErK pathway in ECs is activated by proatherogenic stimuli in vitro and by high-fat diet in vivo. Finally, we demonstrate that the MAPK/ErK pathway is required for HuR induction by proatherogenic stimuli. Altogether, our study uncovers the inducible effect of proatherogenic stimuli on HuR in ECs, and connects this effect to the activated MAPK/ErK pathway.

Identifying key genes associated with acute myocardial infarction.

BACKGROUND: This study aimed to identify key genes associated with acute myocardial infarction (AMI) by reanalyzing microarray data. METHODS: Three gene expression profile datasets GSE66360, GSE34198, and GSE48060 were downloaded from GEO database. After data preprocessing, genes without heterogeneity across different platforms were subjected to differential expression analysis between the AMI group and the control group using metaDE package. P < .05 was used as the cutoff for a differentially expressed gene (DEG). The expression data matrices of DEGs were imported in ReactomeFIViz to construct a gene functional interaction (FI) network. Then, DEGs in each module were subjected to pathway enrichment analysis using DAVID. MiRNAs and transcription factors predicted to regulate target DEGs were identified. Quantitative real-time polymerase chain reaction (RT-PCR) was applied to verify the expression of genes. RESULT: A total of 913 upregulated genes and 1060 downregulated genes were identified in the AMI group. A FI network consists of 21 modules and DEGs in 12 modules were significantly enriched in pathways. The transcription factor-miRNA-gene network contains 2 transcription factors FOXO3 and MYBL2, and 2 miRNAs hsa-miR-21-5p and hsa-miR-30c-5p. RT-PCR validations showed that expression levels of FOXO3 and MYBL2 were significantly increased in AMI, and expression levels of hsa-miR-21-5p and hsa-miR-30c-5p were obviously decreased in AMI. CONCLUSION: A total of 41 DEGs, such as SOCS3, VAPA, and COL5A2, are speculated to have roles in the pathogenesis of AMI; 2 transcription factors FOXO3 and MYBL2, and 2 miRNAs hsa-miR-21-5p and hsa-miR-30c-5p may be involved in the regulation of the expression of these DEGs.

CDKN2B-AS may indirectly regulate coronary artery disease-associated genes via targeting miR-92a.

OBJECTIVE: Coronary artery disease (CAD) has a high mortality rate and consists of multiple condition, including stable/unstable angina, sudden cardiac death, and myocardial infarction. This study is aimed to explore the pathogenesis of CAD. METHODS: Datasets of GSE20680 (including 87 CAD samples and 52 normal samples) and GSE20681 (including 99 CAD samples and 99 normal samples) were obtained from Gene Expression Omnibus database. The differentially expressed genes (DEGs) were identified by MetaDE. Effect Sizes in MetaDE package, and then were hierarchical clustered using pheatmap package in R. Subsequently, CAD-associated microRNAs (miRNAs) and their targets were obtained separately by miR2Disease and miRTarBase databases, and then used to construct an associated-miRNA-DEG regulatory network based on BioGRID, HPRD and DIP databases. Enrichment analysis was conducted for the involved DEGs using Fisher's exact test, and a support vector machine (SVM) classifier was constructed to optimize the feature genes. After CAD-associated long non-coding RNAs (lncRNAs) were predicted by lncRNA Disease database and their target miRNAs were predicted using miRcode and starBase databases, lncRNA-miRNA-DEG regulatory network was constructed. RESULTS: Total 1208 DEGs were screened, and 5 CAD-associated miRNAs (including miR-92a) were predicted associated with CAD. The SVM classifier was constructed based on the 41 featured genes and had high recognition efficiency. Only one lncRNA CDKN2B-AS targeting miR-92a was obtained. Finally, GATA2, MAP1B and ARG1 were involved in the CDKN2B-AS-miR-92a-feature gene regulatory network. CONCLUSION: GATA2, MAP1B and ARG1 indirectly regulated by CDKN2B-AS through miR-92a might be involved in CAD.

Short-Term Effects and Safety Analysis of Retrograde Autologous Blood Priming for Cardiopulmonary Bypass in Patients with Cardiac Valve Replacement Surgery.

This randomized, double-blind study evaluated the short-term effects and safety of perioperative retrograde autologous priming (RAP) for cardiopulmonary bypass (CPB) in patients with cardiac replacement surgery to determine if this approach is a better substitute for crystal liquids priming in patients with valvular heart disease. We observed that RAP significantly decreased the actual priming volume, preserved the hematocrit and hemoglobin level during CPB to a certain degree, and decreased lactate accumulation in CPB period. Moreover, RAP lowered the volume of transfusion and dosage blood products. Thus, our results showed that RAP approach effectively improved tissue perfusion and lowered intraoperative Lac levels, by reducing the hemodilution, which safely and reliably improve the microcirculation perfusion.

[Retraction] Generation of induced pluripotent stem cells using skin fibroblasts from patients with myocardial infarction under feeder-free conditions.

After the publication of the article, the authors decided they wished to retract their manuscript for the following reasons. We wish to retract our research article entitled 'Generation of induced pluripotent stem cells using skin fibroblasts from patients with myocardial infarction under feeder-free conditions' published on the Molecular Medicine Reports 9: 837-842, 2014. In this article, we generated human iPSCs from skin fibroblasts from myocardial infarction patients in feeder-independent conditions. However, in subsequent researches, all of the cells generated and believed to be iPSCs showed negative expression of the pluripotent markers, Nanog and Rex1, and the cell surface marker, SSEA-1 and SSEA-4. Therefore we think the established iPS cells might not be real pluripotent stem cells. Based on the above mentioned, we ascertained that there must have some serious disadvantages in our design of experiment fundamentally. As a result, all authors involved unanimously agreed to retract this article and redesign our experiment. We deeply apologize to the readers for any inconvenience caused by this retraction. [the original article was published in the Molecular Medicine Reports 9: 837-842, 2014 DOI: 10.3892/mmr.2014.1885].

Generation of induced pluripotent stem cells using skin fibroblasts from patients with myocardial infarction under feeder-free conditions.

Myocardial infarction (MI) is an increasing medical problem; however, its pathogenesis has yet to be elucidated and more effective treatment strategies are required. Induced pluripotent stem cells (iPSCs) were recently successfully generated using human somatic cells transfected with four transcription factors. The present study aimed to generate iPSCs from cells from patients with myocardial infarction. Six patients who had been diagnosed with myocardial infarction were enrolled in this study. The fibroblast cells from the biopsied skin were reprogrammed using octamer-binding transcription factor 4 (Oct­4), SRY-related HMG-box gene 2 (Sox­2), Kruppel-like factor 4 (Klf­4) and cellular myelocytomatosis oncogene (c­Myc) transcription factors. The generated cells were identified by karyotyping, in vitro and in vivo differentiation ability and staining for specific markers. These human MI­iPSCs expressed pluripotent genes and cell surface markers, and exhibited normal proliferation. The iPSCs also showed in vivo and in vitro differentiation ability, as indicated by teratoma and embryoid body formation, respectively. Moreover, the iPSCs differentiated into cardiomyocytes and neuronal cells. In conclusion, human iPSCs were successfully generated from skin fibroblasts from patients with MI under feeder­independent conditions, which increases their potential suitability for clinical applications. These results may encourage further study of MI pathogenesis and facilitate the development of safe downstream clinical applications of iPSC­based cell therapies.

A comprehensive study on long-term injury to nigral dopaminergic neurons following intracerebroventricular injection of lipopolysaccharide in rats.

Parkinson's disease (PD) is characterized by selective and progressive degeneration of dopaminergic neurons in the substantia nigra (SN). Lipopolysaccharide (LPS) can induce chronic inflammation and has been widely used to study the pathogenesis of PD. In this study, a single intracerebroventricular injection of LPS was used to induce neurotoxic effects on dopaminergic neurons in Sprague-Dawley rats. The long-term neurotoxic effects of LPS were evaluated at different time points. Microglia were activated in the hippocampus and striatum at 4 weeks, and in the SN at 24 weeks. Astrocytes were activated in the hippocampus and nigrostriatal system at 2 and 24 weeks. The expression of brain-derived neurotrophic factor in the SN increased at 4 weeks and decreased after 12 weeks, and tyrosine hydroxylase-positive neurons in the SN were shown to have an atrophic appearance, with cell loss evident after 24 weeks. Phospho-α-synuclein expression, a reflection of parkinsonian pathogenesis, increased at 12 weeks, and peaked at 24 weeks. Abnormal motor behavior appeared at 16 weeks and lasted up to 48 weeks. These results indicate that microglia are activated for several months after a single, low dose injection of LPS, which eventually results in progressive and selective damage to dopaminergic neurons in the SN.

Case control study of gastrointestinal complications after cardiopulmonary bypass heart surgery.

BACKGROUND: Gastrointestinal complications (GIC) after cardiopulmonary bypass (CPB) surgery are rare, but, nevertheless, extremely dangerous.The identification of risks for GIC may be helpful in planning appropriate perioperative management strategies. The aim of the present study was to analyze perioperative factors of GIC in patients undergoing CPB surgery. METHODS: We retrospectively analysed 206 patients who underwent GIC after cardiopulmonary bypass surgery from 2000 to 2007 and compared them with 206 matched control patients (matched for surgery, temperature, hemodilution and date). Univariate analysis and multiple logistic regression analysis were performed on 12 risk factors. RESULT: Sex and types of cardioplegia perfusate did not significantly influence the GIC after CPB surgery. Multiple logistic regression revealed that CPB time, preoperative serum creatinine (PSC) > or = 179 mg/dL, emergency surgery, perfusion pressure < or =40 mmHg, low cardiac output syndrome (LCOS), age > or = 61, mechanical ventilation > or =96 h, New York Heart Association (NYHA) class III and IV were predictors of the occurrence of GIC after CPB surgery. Perfusion pressure and aprotinin administration were protective factors. CONCLUSION: Gastrointestinal complications after CPB surgery could be predictive in the presence of the above risk factors. This study suggests that GIC can be reduced by maintenance of higher perfusion pressure and shortening the time on CPB and ventilation.

Anti-rejection effect of ethanol extract of Poria cocos wolf in rats after cardiac allograft implantation.

BACKGROUND: A living fetus within the maternal uterus provides an example of allogene tolerance in mammals. Poria cocos Wolf is the main component of many Chinese medicinal combination drugs that have therapeutic effects on recurrent spontaneous abortion and that can maintain pregnancy until delivery. It was hypothesized that this herbal medicine can also prolong allograft survival after organ transplantation. Here, in an in vivo study, we report the anti-rejection effect of the ethanol extract of Poria cocos Wolf (EEPCW) in rats after cardiac allograft implantation. METHODS: Ten normal rats were healthy controls. Eighty rats receiving homologous heart transplants were divided into 4 groups of 20 rats each based on type of treatment: olive oil 8 ml.kg(-1).d(-1), EEPCW 25 mg.kg(-1).d(-1), EEPCW 50 mg.kg(-1).d(-1) or cyclosporin A 5 mg.kg(-1).d(-1). Allograft survival was observed in 10 rats from each group. On the seventh day post transplantation, pathological lesions and percentages of CD3+, CD4+, and CD8+ lymphocytes and the CD4+/CD8+ ratio in peripheral blood were assessed in another 10 rats from each group and in 10 normal rats. RESULTS: The survival time of donor hearts in the two EEPCW groups was significantly prolonged, to (15.9 +/- 2.4) days and (30.0 +/- 0.0) days, respectively, compared with (6.7 +/- 0.8) days in the control group. Pathological lesions in the two EEPCW groups were also less severe, and the percentages of CD3+, CD4+, and CD8+ lymphocytes and CD4+/CD8+ ratio were significantly lower in the EEPCW groups. CONCLUSIONS: Acute rejection of heart transplants and cellular immune reaction can be effectively suppressed using the EEPCW. Taking advantage of novel immunosuppressants derived from Chinese medicinal herbs used to treat abnormal pregnancy provides a hopeful road for future research and treatment in organ transplantation.