Resveratrol protects cardiomyocytes against ischemia/reperfusion-induced ferroptosis via inhibition of the VDAC1/GPX4 pathway.

The present study aimed to explore how resveratrol (Res) confers myocardial protection by attenuating ferroptosis. In vivo and in vitro myocardial ischemia/reperfusion injury (MIRI) models were established, with or without Res pretreatment. The results showed that Res pretreatment effectively attenuated MIRI, as evidenced by increased cell viability, reduced lactate dehydrogenase activity, decreased infarct size, and maintained cardiac function. Moreover, Res pretreatment inhibited MIRI-induced ferroptosis, as shown by improved mitochondrial integrity, increased glutathione level, decreased prostaglandin-endoperoxide synthase 2 level, inhibited iron overload, and abnormal lipid peroxidation. Of note, Res pretreatment decreased or increased voltage-dependent anion channel 1/glutathione peroxidase 4 (VDAC1/GPX4) expression, which was increased or decreased via anoxia/reoxygenation (A/R) treatment, respectively. However, the overexpression of VDAC1 via pAd/VDAC1 and knockdown of GPX4 through Si-GPX4 reversed the protective effect of Res in A/R-induced H9c2 cells, whereas the inhibition of GPX4 with RSL3 abolished the protective effect of Res on mice treated with ischemia/reperfusion.Interestingly, knockdown of VDAC1 by Si-VDAC1 promoted the protective effect of Res on A/R-induced H9c2 cells and the regulation of GPX4. Finally, the direct interaction between VDAC1 and GPX4 was determined using co-immunoprecipitation. In conclusion, Res pretreatment could protect the myocardium against MIRI-induced ferroptosis via the VDAC1/GPX4 signaling pathway.

Exploring the molecular biology of ischemic cardiomyopathy based on ferroptosis­related genes.

Ischemic cardiomyopathy (ICM) is a serious cardiac disease with a very high mortality rate worldwide, which causes myocardial ischemia and hypoxia as the main damage. Further understanding of the underlying pathological processes of cardiomyocyte injury is key to the development of cardioprotective strategies. Ferroptosis is an iron-dependent form of regulated cell death characterized by the accumulation of lipid hydroperoxides to lethal levels, resulting in oxidative damage to the cell membrane. The current understanding of the role and regulation of ferroptosis in ICM is still limited, especially in the absence of evidence from large-scale transcriptomic data. Through comprehensive bioinformatics analysis of human ICM transcriptome data obtained from the Gene Expression Omnibus database, the present study identified differentially expressed ferroptosis-related genes (DEFRGs) in ICM. Subsequently, their potential biological mechanisms and cross-talk were analyzed, and hub genes were identified by constructing protein-protein interaction networks. Ferroptosis features such as reactive oxygen species generation, changes in ferroptosis marker proteins, iron ion aggregation and lipid oxidation, were identified in the H9c2 anoxic reoxygenation injury model. Finally, the diagnostic ability of Gap junction alpha-1 (GJA1), Solute carrier family 40 member 1 (SLC40A1), Alpha-synuclein (SNCA) were identified through receiver operating characteristic curves and the expression of DEFRGs was verified in an in vitro model. Furthermore, potential drugs (retinoic acid) that could regulate ICM ferroptosis were predicted based on key DEFRGs. The present article presents new insights into the role of ferroptosis in ICM, investigating the regulatory role of ferroptosis in the pathological process of ICM and advocating for ferroptosis as a potential novel therapeutic target for ICM based on evidence from the ICM transcriptome.

Tanshinone IIA confers protection against myocardial ischemia/reperfusion injury by inhibiting ferroptosis and apoptosis via VDAC1.

Tanshinone IIA (TSN) extracted from danshen (Salvia miltiorrhiza) could protect cardiomyocytes against myocardial ischemia/reperfusion injury (IRI), however the underlying molecular mechanisms of action remain unclear. The aim of the present study was to identify the protective effects of TSN and its mechanisms of action through in vitro studies. An anoxia/reoxygenation (A/R) injury model was established using H9c2 cells to simulate myocardial IRI in vitro. Before A/R, H9c2 cardiomyoblasts were pretreated with 8 µM TSN or 10 µM ferrostatin­1 (Fer­1) or erastin. The cell counting kit 8 (CCK­8) and lactate dehydrogenase (LDH) assay kit were used to detect the cell viability and cytotoxicity. The levels of total iron, glutathione (GSH), glutathione disulfide (GSSG), malondialdehyde (MDA), ferrous iron, caspase­3 activity, and reactive oxygen species (ROS) were assessed using commercial kit. The levels of mitochondrial membrane potential (MMP), lipid ROS, cell apoptosis, and mitochondrial permeability transition pore (mPTP) opening were detected by flow cytometry. Transmission electron microscopy (TEM) was used to observed the mitochondrial damage. Protein levels were detected by western blot analysis. The interaction between TSN and voltage­dependent anion channel 1 (VDAC1) was evaluated by molecular docking simulation. The results showed that pretreatment with TSN and Fer­1 significantly decreased cell viability, glutathione peroxidase 4 (GPX4) protein and GSH expression and GSH/GSSG ratio and inhibited upregulation of LDH activity, prostaglandin endoperoxide synthase 2 and VDAC1 protein expression, ROS levels, mitochondrial injury and GSSG induced by A/R. TSN also effectively inhibited the damaging effects of erastin treatment. Additionally, TSN increased MMP and Bcl­2/Bax ratio, while decreasing levels of apoptotic cells, activating Caspase­3 and closing the mPTP. These effects were blocked by VDAC1 overexpression and the results of molecular docking simulation studies revealed a direct interaction between TSN and VDAC1. In conclusion, TSN pretreatment effectively attenuated H9c2 cardiomyocyte damage in an A/R injury model and VDAC1­mediated ferroptosis and apoptosis served a vital role in the protective effects of TSN.

Role of dysregulated ferroptosis­related genes in cardiomyocyte ischemia­reperfusion injury: Experimental verification and bioinformatics analysis.

Acute myocardial infarction is a life-threatening condition with high mortality and complication rates. Although myocardial reperfusion can preserve ischemic myocardial tissue, it frequently exacerbates tissue injury, a phenomenon known as ischemia-reperfusion injury (IRI). However, the underlying pathological mechanisms of IRI remain to be completely understood. Ferroptosis is a novel type of regulated cell death that is associated with various pathological conditions, including angiocardiopathy. The purpose of this article was to elucidate the possible mechanistic role of ferroptosis in IRI through bioinformatics analysis and experimental validation. Healthy and IRI heart samples were screened for differentially expressed ferroptosis-related genes and functional enrichment analysis was performed to determine the potential crosstalk and pathways involved. A protein-protein interaction network was established for IRI, and 10 hub genes that regulate ferroptosis, including HIF1A, EGFR, HMOX1, and ATF3 were identified. In vitro, an anoxia/reoxygenation (A/R) injury model was established using H9c2 cardiomyoblasts to validate the bioinformatics analysis results, and extensive ferroptosis was detected. A total of 4 key hub genes and 3 key miRNAs were also validated. It was found that IRI was related to the aberrant infiltration of immune cells and the small-molecule drugs that may protect against IRI by preventing ferroptosis were identified. These results provide novel insights into the role of ferroptosis in IRI, which can help identify novel therapeutic targets.

Exploring Dysregulated Ferroptosis-Related Genes in Septic Myocardial Injury Based on Human Heart Transcriptomes: Evidence and New Insights.

Introduction: Sepsis is currently a common condition in emergency and intensive care units, and is defined as life-threatening organ dysfunction caused by a dysregulated host response to infection. Cardiac dysfunction caused by septic myocardial injury (SMI) is associated with adverse prognosis and has significant economic and human costs. The pathophysiological mechanisms underlying SMI have long been a subject of interest. Recent studies have identified ferroptosis, a form of programmed cell death associated with iron accumulation and lipid peroxidation, as a pathological factor in the development of SMI. However, the current understanding of how ferroptosis functions and regulates in SMI remains limited, particularly in the absence of direct evidence from human heart. Methods: We performed a sequential comprehensive bioinformatics analysis of human sepsis cardiac transcriptome data obtained through the GEO database. The lipopolysaccharide-induced mouse SMI model was used to validate the ferroptosis features and transcriptional expression of key genes. Results: We identified widespread dysregulation of ferroptosis-related genes (FRGs) in SMI based on the human septic heart transcriptomes, deeply explored the underlying biological mechanisms and crosstalks, followed by the identification of key functional modules and hub genes through the construction of protein-protein interaction network. Eight key FRGs that regulate ferroptosis in SMI, including HIF1A, MAPK3, NOX4, PPARA, PTEN, RELA, STAT3 and TP53, were identified, as well as the ferroptosis features. All the key FRGs showed excellent diagnostic capability for SMI, part of them was associated with the prognosis of sepsis patients and the immune infiltration in the septic hearts, and potential ferroptosis-modulating drugs for SMI were predicted based on key FRGs. Conclusion: This study provides human septic heart transcriptome-based evidence and brings new insights into the role of ferroptosis in SMI, which is significant for expanding the understanding of the pathobiological mechanisms of SMI and exploring promising diagnostic and therapeutic targets for SMI.

Dysregulated autophagy-related genes in septic cardiomyopathy: Comprehensive bioinformatics analysis based on the human transcriptomes and experimental validation.

Septic cardiomyopathy (SCM) is severe organ dysfunction caused by sepsis that is associated with poor prognosis, and its pathobiological mechanisms remain unclear. Autophagy is a biological process that has recently been focused on SCM, yet the current understanding of the role of dysregulated autophagy in the pathogenesis of SCM remains limited and uncertain. Exploring the molecular mechanisms of disease based on the transcriptomes of human pathological samples may bring the closest insights. In this study, we analyzed the differential expression of autophagy-related genes in SCM based on the transcriptomes of human septic hearts, and further explored their potential crosstalk and functional pathways. Key functional module and hub genes were identified by constructing a protein-protein interaction network. Eight key genes (CCL2, MYC, TP53, SOD2, HIF1A, CTNNB1, CAT, and ADIPOQ) that regulate autophagy in SCM were identified after validation in a lipopolysaccharide (LPS)-induced H9c2 cardiomyoblast injury model, as well as the autophagic characteristic features. Furthermore, we found that key genes were associated with abnormal immune infiltration in septic hearts and have the potential to serve as biomarkers. Finally, we predicted drugs that may play a protective role in SCM by regulating autophagy based on our results. Our study provides evidence and new insights into the role of autophagy in SCM based on human septic heart transcriptomes, which would be of great benefit to reveal the molecular pathological mechanisms and explore the diagnostic and therapeutic targets for SCM.

Identification and Validation of Ferroptosis-Related Biomarkers in Septic Cardiomyopathy via Bioinformatics Analysis.

Septic cardiomyopathy (SCM) is a cardiac dysfunction caused by severe sepsis and septic shock that increases the risk of heart failure and death and its molecular mechanism remains unclear. Ferroptosis, a novel form of programmed cell death, has been reported to be present in the heart tissue of patients with sepsis, which demonstrated that ferroptosis may be a potential mechanism of myocardial injury in SCM. Therefore, we explored the role of ferroptosis-related genes (FRGs) in SCM and aimed to identify pivotal ferroptosis-related targets in SCM and potential therapeutic targets involved in the pathological process of SCM. To explore the regulatory mechanisms of ferroptosis in SCM, we identified differentially expressed genes (DEGs) in SCM and FRGs by bioinformatics analysis, and further identified hub genes. And the crucial microRNAs (miRNAs)-FRGs regulatory network was subsequently constructed. Finally, several candidate drugs associated with the hub genes were predicted, and Real-time quantitative reverse Transcription PCR (qRT-PCR) and western blotting analysis were performed to confirm the abnormal expression of hub genes. In this study, we identified several FRGs that may be involved in the pathogenesis of SCM, which helps us further clarify the role of ferroptosis in SCM and deeply understand the molecular mechanisms and potential therapeutic targets of SCM.

Identification and Validation of Dilated Cardiomyopathy-Related Genes via Bioinformatics Analysis.

Purpose: Dilated cardiomyopathy (DCM) is a type of cardiomyopathy that can easily cause heart failure and has a high mortality rate. Therefore, there is an urgent need to study the underlying mechanism of action of dilated cardiomyopathy. In the present study, we aimed to explore potential miRNA-mRNA pairs and drugs related to DCM. Methods: The Microarray data were collected from the Gene Expression Omnibus (GEO) database. Bioinformatics analysis differentially expressed miRNAs and mRNAs in each microarray were obtained. The target genes of miRNAs were obtained from the miRWalk 2.0 database, and the intersection of these two gene sets (miRNA target genes and differentially expressed mRNAs in the microarray) was obtained. Pathway and Gene Ontology (GO) enrichment analyses were performed in the KOBAS database. Cytoscape software was used to construct the miRNA-mRNA network, and the final hub genes were obtained. Furthermore, we predicted several candidate drugs related to hub genes using DSigDB database. To confirm the abnormal expression of hub genes, qRT-PCR was performed. Results: In total, eight differentially expressed miRNAs and 92 differentially expressed mRNAs were identified. In addition, 47 differentially expressed miRNA target genes were identified. According to the analysis results of the miRNA-mRNA network, we identified hsa-miR-551b-3p, hsa-miR-770-5p, hsa-miR-363-3p, PIK3R1, DDIT4, and CXCR4 as hub genes in DCM. Several candidate drugs, which are related to the hug genes, were identified. Conclusion: In conclusion, in our study, we identified several hub genes that may be involved in the pathogenesis of DCM. Several drugs related to these hub genes may be used as clinical therapeutic candidates.

ITGB1-DT/ARNTL2 axis may be a novel biomarker in lung adenocarcinoma: a bioinformatics analysis and experimental validation.

BACKGROUND: Lung cancer is one of the most lethal malignant tumors that endangers human health. Lung adenocarcinoma (LUAD) has increased dramatically in recent decades, accounting for nearly 40% of all lung cancer cases. Increasing evidence points to the importance of the competitive endogenous RNA (ceRNA) intrinsic mechanism in various human cancers. However, behavioral characteristics of the ceRNA network in lung adenocarcinoma need further study. METHODS: Groups based on SLC2A1 expression were used in this study to identify associated ceRNA networks and potential prognostic markers in lung adenocarcinoma. The Cancer Genome Atlas (TCGA) database was used to obtain the patients' lncRNA, miRNA, and mRNA expression profiles, as well as clinical data. Informatics techniques were used to investigate the effect of hub genes on prognosis. The Cox regression analyses were performed to evaluate the prognostic effect of hub genes. The methylation, GSEA, and immune infiltration analyses were utilized to explore the potential mechanisms of the hub gene. The CCK-8, transwell, and colony formation assays were performed to detect the proliferation and invasion of lung cancer cells. RESULTS: We eventually identified the ITGB1-DT/ARNTL2 axis as an independent fact may promote lung adenocarcinoma progression. Furthermore, methylation analysis revealed that hypo-methylation may cause the dysregulated ITGB1-DT/ARNTL2 axis, and immune infiltration analysis revealed that the ITGB1-DT/ARNTL2 axis may affect the immune microenvironment and the progression of lung adenocarcinoma. The CCK-8, transwell, and colonu formation assays suggested that ITGB1-DT/ARNTL2 promotes the progression of lung adenocarcinoma. And hsa-miR-30b-3p reversed the ITGB1/ARNTL2-mediated oncogenic processes. CONCLUSION: Our study identified the ITGB1-DT/ARNTL2 axis as a novel prognostic biomarker affects the prognosis of lung adenocarcinoma.

Plasma Lipidomics Identifies Unique Lipid Signatures and Potential Biomarkers for Patients With Aortic Dissection.

Aortic dissection (AD) is a catastrophic cardiovascular emergency with a poor prognosis, and little preceding symptoms. Abnormal lipid metabolism is closely related to the pathogenesis of AD. However, comprehensive lipid alterations related to AD pathogenesis remain unclear. Moreover, there is an urgent need for new or better biomarkers for improved risk assessment and surveillance of AD. Therefore, an untargeted lipidomic approach based on ultra-high-performance liquid chromatograph-mass spectrometry was employed to unveil plasma lipidomic alterations and potential biomarkers for AD patients in this study. We found that 278 of 439 identified lipid species were significantly altered in AD patients (n = 35) compared to normal controls (n = 32). Notably, most lipid species, including fatty acids, acylcarnitines, cholesteryl ester, ceramides, hexosylceramides, sphingomyelins, lysophosphatidylcholines, lysophosphatidylethanolamines, phosphatidylcholines, phosphatidylinositols, diacylglycerols, and triacylglycerols with total acyl chain carbon number ≥54 and/or total double bond number ≥4 were decreased, whereas phosphatidylethanolamines and triacylglycerols with total double bond number <4 accumulated in AD patients. Besides, the length and unsaturation of acyl chains in triacylglycerols and unsaturation of 1-acyl chain in phosphatidylethanolamines were decreased in AD patients. Moreover, lysophosphatidylcholines were the lipids with the largest alterations, at the center of correlation networks of lipid alterations, and had excellent performances in identifying AD patients. The area under the curve of 1.0 and accuracy rate of 100% could be easily obtained by lysophosphatidylcholine (20:0/0:0) or its combination with lysophosphatidylcholine (17:0/0:0) or lysophosphatidylcholine (20:1/0:0). This study provides novel and comprehensive plasma lipidomic signatures of AD patients, identifies lysophosphatidylcholines as excellent potential biomarkers, and would be beneficial to the pathogenetic study, risk assessment and timely diagnosis and treatment of AD.

Iron Metabolism and Idiopathic Pulmonary Arterial Hypertension: New Insights from Bioinformatic Analysis.

Idiopathic pulmonary arterial hypertension (IPAH) is a rare vascular disease with a poor prognosis, and the mechanism of its development remains unclear. Further molecular pathology studies may contribute to a comprehensive understanding of IPAH and provide new insights into diagnostic markers and potential therapeutic targets. Iron deficiency has been reported in 43-63% of patients with IPAH and is associated with reduced exercise capacity and higher mortality, suggesting that dysregulated iron metabolism may play an unrecognized role in influencing the development of IPAH. In this study, we explored the regulatory mechanisms of iron metabolism in IPAH by bioinformatic analysis. The molecular function of iron metabolism-related genes (IMRGs) is mainly enriched in active transmembrane transporter activity, and they mainly affect the biological process of response to oxidative stress. Ferroptosis and fluid shear stress and atherosclerosis pathways may be the critical pathways regulating iron metabolism in IPAH. We further identified 7 key genes (BCL2, GCLM, MSMO1, SLC7A11, SRXN1, TSPAN5, and TXNRD1) and 5 of the key genes (BCL2, MSMO1, SLC7A11, TSPAN5, and TXNRD1) as target genes may be regulated by 6 dysregulated miRNAs (miR-483-5p, miR-27a-3p, miR-27b-3p, miR-26b-5p, miR-199a-5p, and miR-23b-3p) in IPAH. In addition, we predicted potential IPAH drugs-celastrol and cinnamaldehyde-that target iron metabolism based on our results. These results provide insights for further definition of the role of dysregulated iron metabolism in IPAH and contribute to a deeper understanding of the molecular mechanisms and potential therapeutic targets of IPAH.

Role of ferroptosis-related genes in Stanford type a aortic dissection and identification of key genes: new insights from bioinformatic analysis.

Stanford type A aortic dissection (TAAD) is one of the most dangerous vascular diseases worldwide, and the mechanisms of its development remain unclear. Further molecular pathology studies may contribute to a comprehensive understanding of TAAD and provide new insights into diagnostic markers and potential therapeutic targets. Recent studies have identified that ferroptosis, a form of cell death, may play a previously unrecognized role in influencing the development of TAAD. In this study, we explored the pathological role of ferroptosis in TAAD by performing bioinformatics analyses. Gene set enrichment analysis (GSEA) showed that the ferroptosis-related gene (FRG) set was significantly different between normal and TAAD aortic samples at an overall level (p < 0.001). Further Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses explored the potential functions and pathways of FRG in TAAD. We further identified six key genes (CA9, HMOX1, IL6, CDKN1A, HIF1A, MYC) from differentially expressed FRGs in TAAD by constructing a protein-protein interaction (PPI) network, all key genes were upregulated in TAAD. Four of the key genes (CA9, IL6, CDKN1A, and HIF1A) were demonstrated to be correlated with cigarette smoke extract-induced ferroptosis in aortic vascular smooth muscle cells. These results suggest that ferroptosis is one of the essential pathological processes in the development of TAAD, and some FRGs affect TAAD development by mediating cellular ferroptosis, which provides deepening insights into the molecular mechanisms and potential therapeutic targets of TAAD.