ABSTRACT
The molecular mechanisms by which FoxO transcription factors mediate diametrically opposite cellular responses, namely death and survival, remain unknown. Here we show that Mst1 phosphorylates FoxO1 Ser209/Ser215/Ser218/Thr228/Ser232/Ser243, thereby inhibiting FoxO1-mediated transcription of proapoptotic genes. On the other hand, Mst1 increases FoxO1-C/EBP-ß interaction and activates C/EBP-ß by phosphorylating it at Thr299, thereby promoting transcription of prosurvival genes. Myocardial ischemia/reperfusion injury is larger in cardiac-specific FoxO1 knockout mice than in control mice. However, the concurrent presence of a C/EBP-ß T299E phospho-mimetic mutation reduces infarct size in cardiac-specific FoxO1 knockout mice. The C/EBP-ß phospho-mimetic mutant exhibits greater binding to the promoter of prosurvival genes than wild type C/EBP-ß. In conclusion, phosphorylation of FoxO1 by Mst1 inhibits binding of FoxO1 to pro-apoptotic gene promoters but enhances its binding to C/EBP-ß, phosphorylation of C/EBP-ß, and transcription of prosurvival genes, which stimulate protective mechanisms in the heart.
Subject(s)
CCAAT-Enhancer-Binding Protein-beta , Forkhead Box Protein O1 , Myocardial Reperfusion Injury , Myocytes, Cardiac , Animals , Humans , Male , Mice , Rats , Apoptosis , CCAAT-Enhancer-Binding Protein-beta/metabolism , CCAAT-Enhancer-Binding Protein-beta/genetics , Forkhead Box Protein O1/metabolism , Forkhead Box Protein O1/genetics , Hepatocyte Growth Factor/metabolism , Mice, Knockout , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/pathology , Myocardial Reperfusion Injury/prevention & control , Myocytes, Cardiac/metabolism , Phosphorylation , Promoter Regions, Genetic , Protein Serine-Threonine Kinases/metabolism , Protein Serine-Threonine Kinases/genetics , Proto-Oncogene ProteinsABSTRACT
The specific pathophysiological pathways through which diabetes exacerbates myocardial ischemia/reperfusion (I/R) injury remain unclear; however, dysregulation of immune and inflammatory cells, potentially driven by abnormalities in their number and function due to diabetes, may play a significant role. In the present investigation, we simulated myocardial I/R injury by inducing ischemia through ligation of the left anterior descending coronary artery in mice for 40 min, followed by reperfusion for 24 h. Previous studies have indicated that protein kinase Cß (PKCß) is upregulated under hyperglycemic conditions and is implicated in the development of various diabetic complications. The Y4 RNA fragment is identified as the predominant small RNA component present in the extracellular vesicles of cardio sphere-derived cells (CDCs), exhibiting notable anti-inflammatory properties in the contexts of myocardial infarction and cardiac hypertrophy. Our investigation revealed that the administration of Y4 RNA into the ventricular cavity of db/db mice following myocardial I/R injury markedly enhanced cardiac function. Furthermore, Y4 RNA was observed to facilitate M2 macrophage polarization and interleukin-10 secretion through the suppression of PKCß activation. The mechanism by which Y4 RNA affects PKCß by regulating macrophage activation within the inflammatory environment involves the inhibition of ERK1/2 phosphorylation In our study, the role of PKCß in regulating macrophage polarization during myocardial I/R injury was investigated through the use of PKCß knockout mice. Our findings indicate that PKCß plays a crucial role in modulating the inflammatory response associated with macrophage activation in db/db mice experiencing myocardial I/R, with a notable exacerbation of this response observed upon significant upregulation of PKCß expression. In vitro studies further elucidated the protective mechanism by which Y4 RNA modulates the PKCß/ERK1/2 signaling pathway to induce M2 macrophage activation. Overall, our findings suggest that Y4 RNA plays an anti-inflammatory role in diabetic I/R injury, suggesting a novel therapeutic approach for managing myocardial I/R injury in diabetic individuals.
Subject(s)
Disease Models, Animal , Macrophages , Mice, Inbred C57BL , Myocardial Reperfusion Injury , Protein Kinase C beta , Signal Transduction , Animals , Protein Kinase C beta/metabolism , Myocardial Reperfusion Injury/pathology , Myocardial Reperfusion Injury/enzymology , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/prevention & control , Myocardial Reperfusion Injury/genetics , Macrophages/metabolism , Macrophages/enzymology , Male , Interleukin-10/metabolism , Interleukin-10/genetics , Mice , Diabetic Cardiomyopathies/enzymology , Diabetic Cardiomyopathies/pathology , Diabetic Cardiomyopathies/metabolism , Diabetic Cardiomyopathies/etiology , Diabetic Cardiomyopathies/genetics , Diabetic Cardiomyopathies/physiopathology , Cells, Cultured , Phenotype , Myocytes, Cardiac/enzymology , Myocytes, Cardiac/pathology , Myocytes, Cardiac/metabolism , Mitogen-Activated Protein Kinase 3/metabolism , Macrophage Activation , Mitogen-Activated Protein Kinase 1/metabolism , Ventricular Function, Left , PhosphorylationABSTRACT
Type 2 diabetes predisposes patients to heart disease, which is the primary cause of death across the globe. Type 2 diabetes often accompanies obesity and is defined by insulin resistance and abnormal glucose handling. Insulin resistance impairs glucose uptake and results in hyperglycemia, which damages tissues such as kidneys, liver, and heart. 2-oxoglutarate (2-OG)- and iron-dependent oxygenases (2-OGDOs), a family of enzymes regulating various aspects of cellular physiology, have been studied for their role in obesity and diet-induced insulin resistance. However, nothing is known of the 2-OGDO family member 2-oxoglutarate and iron-dependent prolyl hydroxylase domain containing protein 1 (OGFOD1) in this setting. OGFOD1 deletion leads to protection in cardiac ischemia-reperfusion injury and cardiac hypertrophy, which are two cardiac events that can lead to heart failure. Considering the remarkable correlation between heart disease and diabetes, the cardioprotection observed in OGFOD1-knockout mice led us to challenge these knockouts with high-fat diet. Wildtype mice fed a high-fat diet developed diet-induced obesity, insulin resistance, and glucose intolerance, but OGFOD1 knockout mice fed this same diet were resistant to diet-induced obesity and insulin resistance. These results support OGFOD1 down-regulation as a strategy for preventing obesity and insulin handling defects.
Subject(s)
Diet, High-Fat , Insulin Resistance , Mice, Knockout , Obesity , Animals , Obesity/metabolism , Obesity/genetics , Mice , Diet, High-Fat/adverse effects , Male , Prolyl Hydroxylases/metabolism , Prolyl Hydroxylases/genetics , Glucose Intolerance/metabolism , Glucose Intolerance/genetics , Mice, Inbred C57BL , Gene Deletion , Cardiomegaly/metabolism , Cardiomegaly/prevention & control , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/prevention & control , Myocardial Reperfusion Injury/geneticsABSTRACT
Myocardial infarction/reperfusion (I/R) injury significantly impacts the health of older individuals. We confirmed that the level of lncRNA Peg13 was downregulated in I/R injury. However, the detailed function of Peg13 in myocardial I/R injury has not yet been explored.To detect the function of Peg13, in vivo model of I/R injury was constructed. RT-qPCR was employed to investigate RNA levels, and Western blotting was performed to assess levels of endoplasmic reticulum stress and apoptosis-associated proteins. EdU staining was confirmed to assess the cell proliferation.I/R therapy dramatically produced myocardial injury, increased the infarct area, and decreased the amount of Peg13 in myocardial tissues of mice. In addition, hypoxia/reoxygenation (H/R) notably induced the apoptosis and promoted the endoplasmic reticulum (ER) stress of HL-1 cells, while overexpression of Peg13 reversed these phenomena. Additionally, Peg13 may increase the level of Sirt1 through binding to miR-34a. Upregulation of Peg13 reversed H/R-induced ER stress via regulation of miR-34a/Sirt1 axis.LncRNA Peg13 reduces ER stress in myocardial infarction/reperfusion injury through mediation of miR-34a/Sirt1 axis. Hence, our research might shed new lights on developing new strategies for the treatment of myocardial I/R injury.
Subject(s)
Endoplasmic Reticulum Stress , MicroRNAs , Myocardial Reperfusion Injury , RNA, Long Noncoding , Sirtuin 1 , Animals , MicroRNAs/genetics , MicroRNAs/metabolism , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism , Sirtuin 1/metabolism , Sirtuin 1/genetics , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/genetics , Mice , Male , Myocardial Infarction/metabolism , Myocardial Infarction/genetics , Apoptosis/genetics , Disease Models, Animal , Mice, Inbred C57BLABSTRACT
Myocardial ischemia-reperfusion injury (MIRI) refers to the secondary damage to myocardial tissue that occurs when blood perfusion is rapidly restored following myocardial ischemia. This process often exacerbates the injury to myocardial fiber structure and function. The activation mechanism of angiogenesis is closely related to MIRI and plays a significant role in the occurrence and progression of ischemic injury. In this study, we utilized sequencing data from the GEO database and employed WGCNA, Mfuzz cluster analysis, and protein interaction network to identify Stat3, Rela, and Ubb as hub genes involved in MIRI-angiogenesis. Additionally, the GO and KEGG analysis of differentially expressed genes highlighted their broad participation in inflammatory responses and associated signaling pathways. Moreover, the analysis of sequencing data and hub genes revealed a notable increase in the infiltration ratio of monocytes and activated mast cells. By establishing key cell ROC curves, using independent datasets, and validating the expression of hub genes, we demonstrated their high diagnostic value. Moreover, by scrutinizing single-cell sequencing data alongside trajectory analysis, it has come to light that Stat3 and Rela exhibit predominant expression within Dendritic cells. In contrast, Ubb demonstrates expression across multiple cell types, with all three genes being expressed at distinct stages of cellular development. Lastly, leveraging the CMap database, we predicted potential small molecule compounds for the identified hub genes and validated their binding activity through molecular docking. Ultimately, our research provides valuable evidence and references for the early diagnosis and treatment of MIRI from the perspective of angiogenesis.
Subject(s)
Biomarkers , Myocardial Reperfusion Injury , STAT3 Transcription Factor , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/pathology , Humans , STAT3 Transcription Factor/metabolism , STAT3 Transcription Factor/genetics , Biomarkers/metabolism , Transcription Factor RelA/metabolism , Transcription Factor RelA/genetics , Protein Interaction Maps/genetics , Neovascularization, Pathologic/genetics , Gene Expression Profiling , AngiogenesisABSTRACT
Myocardial ischemia/reperfusion injury (MIRI) is a significant challenge in the management of myocardial ischemic disease. Extensive evidence suggests that the macrophagemediated inflammatory response may play a vital role in MIRI. Mesenchymal stem cells and, in particular, exosomes derived from these cells, may be key mediators of myocardial injury and repair. However, whether exosomes protect the heart by regulating the polarization of macrophages and the exact mechanisms involved are poorly understood. The present study aimed to determine whether exosomes secreted by bone marrow mesenchymal stem cells (BMSCExo) harboring miR253p can alter the phenotype of macrophages by affecting the JAK2/STAT3 signaling pathway, which reduces the inflammatory response and protects against MIRI. An in vivo MIRI model was established in rats by ligating the anterior descending region of the left coronary artery for 30 min followed by reperfusion for 120 min, and BMSCExo carrying miR253p (BMSCExo253p) were administered through tail vein injection. A hypoxiareoxygenation model of H9C2 cells was established, and the cells were cocultured with BMSCExo253p in vitro. The results of the present study demonstrated that BMSCExo or BMSCExo253p could be taken up by cardiomyocytes in vivo and H9C2 cells in vitro. BMSCExo253p demonstrated powerful cardioprotective effects by decreasing the cardiac infarct size, reducing the incidence of malignant arrhythmias and attenuating myocardial enzyme activity, as indicated by lactate dehydrogenase and creatine kinase levels. It induced M1like macrophage polarization after myocardial ischemia/reperfusion (I/R), as evidenced by the increase in iNOS expression through immunofluorescence staining and upregulation of proinflammatory cytokines through RTqPCR, such as interleukin1ß (IL1ß) and interleukin6 (IL6). As hypothesized, BMSCExo253p inhibited M1like macrophage polarization and proinflammatory cytokine expression while promoting M2like macrophage polarization. Mechanistically, the JAK2/STAT3 signaling pathway was activated after I/R in vivo and in LPSstimulated macrophages in vitro, and BMSCExo253p pretreatment inhibited this activation. The results of the present study indicate that the attenuation of MIRI by BMSCExo253p may be related to JAK2/STAT3 signaling pathway inactivation and subsequent inhibition of M1like macrophage polarization.
Subject(s)
Exosomes , Macrophages , Mesenchymal Stem Cells , MicroRNAs , Myocardial Reperfusion Injury , STAT3 Transcription Factor , Animals , MicroRNAs/genetics , MicroRNAs/metabolism , Exosomes/metabolism , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/pathology , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/prevention & control , Rats , Macrophages/metabolism , Male , Mesenchymal Stem Cells/metabolism , Mesenchymal Stem Cells/cytology , STAT3 Transcription Factor/metabolism , Janus Kinase 2/metabolism , Signal Transduction , Rats, Sprague-Dawley , Disease Models, Animal , Myocytes, Cardiac/metabolism , Cell LineABSTRACT
Ischemia/reperfusion (I/R) injury is one of the major causes of cardiovascular disease. Gypenoside A (GP), the main active component of Gynostemma pentaphyllum, alleviates myocardial I/R injury. Circular RNAs (circRNAs) and microRNAs (miRNAs) are involved in the I/R injury. We explored the protective effect of GP on human cardiomyocytes (HCMs) via the circ_0010729/miR-370-3p/RUNX1 axis. Overexpression of circ_0010729 abolished the effects of GP on HMC, such as suppression of apoptosis and increase in cell viability and proliferation. Overexpression of miR-370-3p reversed the effect of circ_0010729 overexpression, resulting in the stimulation of HMC viability and proliferation and inhibition of apoptosis. The knockdown of miR-370-3p suppressed the effects of GP in HCMs. RUNX1 silencing counteracted the effect of miR-370-3p knockdown and maintained GP-induced suppression of apoptosis and stimulation of HMC viability and proliferation. The levels of RUNX1 mRNA and protein were reduced in cells expressing miR-370-3p. In conclusion, this study confirmed that GP alleviated the I/R injury of myocardial cell via the circ_0010729/miR-370-3p/RUNX1 axis.
Subject(s)
Core Binding Factor Alpha 2 Subunit , Gynostemma , MicroRNAs , Myocardial Reperfusion Injury , Myocytes, Cardiac , RNA, Circular , Humans , MicroRNAs/metabolism , MicroRNAs/genetics , RNA, Circular/genetics , RNA, Circular/metabolism , Core Binding Factor Alpha 2 Subunit/metabolism , Core Binding Factor Alpha 2 Subunit/genetics , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/pathology , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/drug effects , Apoptosis/drug effects , Cell Survival/drug effects , Cell Proliferation/drug effects , Plant ExtractsABSTRACT
BACKGROUND: Cuproptosis is known to regulate diverse physiological functions in many diseases, but its role in regulating Myocardial Ischemia-Reperfusion Injury (MI/RI) remains unclear. METHODS: For this purpose, the MI/RI microarray datasets GSE61592 were downloaded from the Gene Expression Omnibus (GEO) database, and the Differently Expressed Genes (DEGs) in MI/RI were identified using R software. Moreover, the MI/RI mice model was established to confirm further the diagnostic value of Pyruvate Dehydrogenase B (Pdhb), Dihydrolipoamide S-acetyltransferase (Dlat), and Pyruvate dehydrogenase E1 subunit alpha 1 (Pdhα1). RESULTS: The analysis of microarray datasets GSE61592 revealed that 798 genes were upregulated and 768 were downregulated in the myocardial tissue of the ischemia-reperfusion injury mice. Furthermore, Dlat, Pdhb, Pdhα1, and cuproptosis-related genes belonged to the downregulated genes. The receiver operating characteristics curve analysis results indicated that the Dlat, Pdhb, and Pdhα1 levels were downregulated in MI/RI and were found to be potential biomarkers for MI/RI diagnosis and prognosis. Similarly, analysis of Dlat, Pdhb, and Pdhα1 levels in the MI/RI mice revealed Pdhb being the key diagnostic marker. CONCLUSIONS: This study demonstrated the prognostic value of cuproptosis-related genes (Dlat, Pdhb, and Pdhα1), especially Pdhb, MI/RI, providing new insight into the MI/RI treatment.
Subject(s)
Computational Biology , Myocardial Reperfusion Injury , Animals , Myocardial Reperfusion Injury/genetics , Mice , Down-Regulation/genetics , Male , Disease Models, Animal , Up-Regulation , Mice, Inbred C57BL , Gene Expression Profiling/methods , Pyruvate Dehydrogenase (Lipoamide)/genetics , Biomarkers/analysis , Acetyltransferases/geneticsABSTRACT
Myocardial ischemia-reperfusion (I/R) injury is a prevalent cause of myocardial injury, involving a series of interconnected pathophysiological processes. However, there is currently no clinical therapy for effectively mitigating myocardial I/R injury. Here, we show that p85α protein levels increase in response to I/R injury through a comprehensive analysis of cardiac proteomics, and confirm this in the I/R-injured murine heart and failing human myocardium. Genetic inhibition of p85α in mice activates the Akt-GSK3ß/Bcl-x(L) signaling pathway and ameliorates I/R-induced cardiac dysfunction, apoptosis, inflammation, and mitochondrial dysfunction. p85α silencing in cardiomyocytes alleviates hypoxia-reoxygenation (H/R) injury through activating the Akt-GSK3ß/Bcl-x(L) signaling pathway, while its overexpression exacerbates the damage. Mechanistically, the interaction between MG53 and p85α triggers the ubiquitination and degradation of p85α, consequently enhancing Akt phosphorylation and ultimately having cardioprotective effects. Collectively, our findings reveal that substantial reduction of p85α and subsequently activated Akt signaling have a protective effect against cardiac I/R injury, representing an important therapeutic strategy for mitigating myocardial damage.
Subject(s)
Myocardial Reperfusion Injury , Myocytes, Cardiac , Proto-Oncogene Proteins c-akt , Signal Transduction , Animals , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/pathology , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/pathology , Myocardial Reperfusion Injury/genetics , Mice , Humans , Proto-Oncogene Proteins c-akt/metabolism , Proto-Oncogene Proteins c-akt/genetics , Glycogen Synthase Kinase 3 beta/metabolism , Glycogen Synthase Kinase 3 beta/genetics , Male , bcl-X Protein/metabolism , bcl-X Protein/genetics , Cell Survival , Class Ia Phosphatidylinositol 3-Kinase/metabolism , Class Ia Phosphatidylinositol 3-Kinase/genetics , Apoptosis , Mice, Inbred C57BL , Mice, Knockout , Phosphorylation , Tumor Suppressor Protein p53ABSTRACT
Ischemic heart diseases are a major global cause of death, and despite timely revascularization, heart failure due to ischemia-hypoxia reperfusion (IH/R) injury remains a concern. The study focused on the role of Early Growth Response 1 (EGR1) in IH/R-induced apoptosis in human cardiomyocytes (CMs). Human induced pluripotent stem cell (hiPSC)-derived CMs were cultured under IH/R conditions, revealing higher EGR1 expression in the IH/R group through quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting (WB). Immunofluorescence analysis (IFA) showed an increased ratio of cleaved Caspase-3-positive apoptotic cells in the IH/R group. Using siRNA for EGR1 successfully downregulated EGR1, suppressing cleaved Caspase-3-positive apoptotic cell ratio. Bioinformatic analysis indicated that EGR1 is a plausible target of miR-124-3p under IH/R conditions. The miR-124-3p mimic, predicted to antagonize EGR1 mRNA, downregulated EGR1 under IH/R conditions in qRT-PCR and WB, as confirmed by IFA. The suppression of EGR1 by the miR-124-3p mimic subsequently reduced CM apoptosis. The study suggests that treatment with miR-124-3p targeting EGR1 could be a potential novel therapeutic approach for cardioprotection in ischemic heart diseases in the future.
Subject(s)
Apoptosis , Down-Regulation , Early Growth Response Protein 1 , Induced Pluripotent Stem Cells , MicroRNAs , Myocytes, Cardiac , MicroRNAs/genetics , MicroRNAs/metabolism , Early Growth Response Protein 1/metabolism , Early Growth Response Protein 1/genetics , Humans , Myocytes, Cardiac/metabolism , Induced Pluripotent Stem Cells/metabolism , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/pathologyABSTRACT
BACKGROUND: The regenerative capacity of the adult mammalian hearts is limited. Numerous studies have explored mechanisms of adult cardiomyocyte cell-cycle withdrawal. This translational study evaluated the effects and underlying mechanism of rhCHK1 (recombinant human checkpoint kinase 1) on the survival and proliferation of cardiomyocyte and myocardial repair after ischemia/reperfusion injury in swine. METHODS AND RESULTS: Intramyocardial injection of rhCHK1 protein (1 mg/kg) encapsulated in hydrogel stimulated cardiomyocyte proliferation and reduced cardiac inflammation response at 3 days after ischemia/reperfusion injury, improved cardiac function and attenuated ventricular remodeling, and reduced the infarct area at 28 days after ischemia/reperfusion injury. Mechanistically, multiomics sequencing analysis demonstrated enrichment of glycolysis and mTOR (mammalian target of rapamycin) pathways after rhCHK1 treatment. Co-Immunoprecipitation (Co-IP) experiments and protein docking prediction showed that CHK1 (checkpoint kinase 1) directly bound to and activated the Serine 37 (S37) and Tyrosine 105 (Y105) sites of PKM2 (pyruvate kinase isoform M2) to promote metabolic reprogramming. We further constructed plasmids that knocked out different CHK1 and PKM2 amino acid domains and transfected them into Human Embryonic Kidney 293T (HEK293T) cells for CO-IP experiments. Results showed that the 1-265 domain of CHK1 directly binds to the 157-400 amino acids of PKM2. Furthermore, hiPSC-CM (human iPS cell-derived cardiomyocyte) in vitro and in vivo experiments both demonstrated that CHK1 stimulated cardiomyocytes renewal and cardiac repair by activating PKM2 C-domain-mediated cardiac metabolic reprogramming. CONCLUSIONS: This study demonstrates that the 1-265 amino acid domain of CHK1 binds to the 157-400 domain of PKM2 and activates PKM2-mediated metabolic reprogramming to promote cardiomyocyte proliferation and myocardial repair after ischemia/reperfusion injury in adult pigs.
Subject(s)
Cell Proliferation , Checkpoint Kinase 1 , Disease Models, Animal , Myocardial Reperfusion Injury , Myocytes, Cardiac , Animals , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/pathology , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/pathology , Myocardial Reperfusion Injury/enzymology , Myocardial Reperfusion Injury/genetics , Checkpoint Kinase 1/metabolism , Checkpoint Kinase 1/genetics , Humans , Pyruvate Kinase/metabolism , Pyruvate Kinase/genetics , HEK293 Cells , Swine , Cellular Reprogramming , Thyroid Hormone-Binding Proteins , Regeneration , Protein Binding , Sus scrofa , Ventricular Remodeling/physiology , Recombinant Proteins/metabolism , Recombinant Proteins/pharmacology , Energy Metabolism/drug effects , Thyroid Hormones/metabolism , Metabolic ReprogrammingABSTRACT
BACKGROUND: Ischemic postconditioning (IPostC) has been reported as a promising method for protecting against myocardial ischemia-reperfusion (MI/R) injury. Our previous study found that the infarct-limiting effect of IPostC is abolished in the heart of diabetes whose cardiac expression of DJ-1 (also called PARK7, Parkinsonism associated deglycase) is reduced. However, the role and in particular the underlying mechanism of DJ-1 in the loss of sensitivity to IPostC-induced cardioprotection in diabetic hearts remains unclear. METHODS: Streptozotocin-induced type 1 diabetic rats were subjected to MI/R injury by occluding the left anterior descending artery (LAD) and followed by reperfusion. IPostC was induced by three cycles of 10s of reperfusion and ischemia at the onset of reperfusion. AAV9-CMV-DJ-1, AAV9-CMV-C106S-DJ-1 or AAV9-DJ-1 siRNA were injected via tail vein to either over-express or knock-down DJ-1 three weeks before inducing MI/R. RESULTS: Diabetic rats subjected to MI/R exhibited larger infarct area, more severe oxidative injury concomitant with significantly reduced cardiac DJ-1 expression and increased PTEN expression as compared to non-diabetic rats. AAV9-mediated cardiac DJ-1 overexpression, but not the cardiac overexpression of DJ-1 mutant C106S, restored IPostC-induced cardioprotection and this effect was accompanied by increased cytoplasmic DJ-1 translocation toward nuclear and mitochondrial, reduced PTEN expression, and increased Nrf-2/HO-1 transcription. Our further study showed that AAV9-mediated targeted DJ-1 gene knockdown aggravated MI/R injury in diabetic hearts, and this exacerbation of MI/R injury was partially reversed by IPostC in the presence of PTEN inhibition or Nrf-2 activation. CONCLUSIONS: These findings suggest that DJ-1 preserves the cardioprotective effect of IPostC against MI/R injury in diabetic rats through nuclear and mitochondrial DJ-1 translocation and that inhibition of cardiac PTEN and activation of Nrf-2/HO-1 may represent the major downstream mechanisms whereby DJ-1 preserves the cardioprotective effect of IPostC in diabetes.
Subject(s)
Diabetes Mellitus, Experimental , Ischemic Postconditioning , Myocardial Reperfusion Injury , PTEN Phosphohydrolase , Protein Deglycase DJ-1 , Rats, Sprague-Dawley , Animals , Protein Deglycase DJ-1/metabolism , Protein Deglycase DJ-1/genetics , PTEN Phosphohydrolase/metabolism , PTEN Phosphohydrolase/genetics , Diabetes Mellitus, Experimental/metabolism , Male , Rats , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/pathology , Myocardial Reperfusion Injury/genetics , NF-E2-Related Factor 2/metabolism , NF-E2-Related Factor 2/genetics , Diabetes Mellitus, Type 1/metabolism , Diabetes Mellitus, Type 1/complications , Protein Transport , Streptozocin , Myocardial Infarction/metabolism , Myocardial Infarction/pathologyABSTRACT
BACKGROUND: This study was designed to investigate the mechanism by which miR-30a-5p mediates cardiomyocyte apoptosis after acute myocardial infarction (AMI) induced by hypoxia/reoxygenation (H/R). METHODS: Differentially expressed miRNAs were analyzed by RNA high-throughput sequencing in acute myocardial infarction (ST-elevation myocardial infarction) patients versus healthy individuals (controls). The H/R model was used to assess the regulatory mechanism of miRNAs in AMI. Lentivirus-associated vectors were used to overexpress or knock down miR-30a-5p in cellular models. The pathological mechanisms of miR-30a-5p regulating the development of acute myocardial infarction were serially explored by qPCR, bioinformatics, target gene prediction, dual luciferase, enzyme-linked immunosorbent assays (ELISAs) and Western blotting. RESULTS: The results showed that the expression of miR-30a-5p was significantly increased in AMI patients and H9C2 cells. Hypoxia decreased cardiomyocyte survival over time, and reoxygenation further reduced cell survival. Bax and Phosphatase and tensin homolog (PTEN)were suppressed, while Bcl-2 was upregulated. Additionally, miR-30a-5p specifically targeted the PTEN gene. According to the GO and KEGG analyses, miR-30a-5p may participate in apoptosis by interacting with PTEN. The miR-30a-5p mimic decreased the expression of apoptosis-related proteins and the levels of the proinflammatory markers IL-1ß, IL-6, and TNF-α by activating the PTEN/PI3K/Akt signaling pathway. Conversely, anti-miR-30a-5p treatment attenuated these effects. Additionally, silencing PTEN and anti-miR-30a-5p had opposite effects on H/R-induced cell apoptosis. CONCLUSIONS: miR-30a-5p plays a crucial role in cardiomyocyte apoptosis after hypoxia-induced acute myocardial infarction. Our findings provide translational evidence that miR-30a-5p is a novel potential therapeutic target for AMI.
Subject(s)
Apoptosis , Cell Hypoxia , MicroRNAs , Myocytes, Cardiac , PTEN Phosphohydrolase , Signal Transduction , Animals , Female , Humans , Male , Middle Aged , Rats , Case-Control Studies , Cell Line , Gene Expression Regulation , MicroRNAs/genetics , MicroRNAs/metabolism , Myocardial Infarction/genetics , Myocardial Infarction/pathology , Myocardial Infarction/metabolism , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/pathology , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/enzymology , Myocytes, Cardiac/pathology , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/enzymology , Phosphatidylinositol 3-Kinase/metabolism , Proto-Oncogene Proteins c-akt/metabolism , PTEN Phosphohydrolase/metabolism , PTEN Phosphohydrolase/geneticsABSTRACT
Exploring and discovering novel circRNAs is one of the ways to develop innovative drugs for the diagnosis and treatment of myocardial ischemia-reperfusion injury (MI/RI). In the work, some dysregulated circRNAs were found by microarray screening analysis in AC16 cells, and hsa_circRNA_104852 named circMIRIAF was screened, which was up-regulated in AC16 cells damaged by hypoxia-reoxygenation injury (H/RI). The comprehensive analysis of ceRNA network revealed the potential relationship of circMIRIAF/miR-544/WDR12. Then, the results of interaction research confirmed that circMIRIAF acted as sponge of miR-544 to positively regulate WDR12 protein expression. Further, the validation results indicate that miR-544 silencing increased the expression of WDR12, and WDR12 activated Notch1 signal to aggravate H/RI of AC16 cells and MI/RI of mice via regulating oxidative stress and inflammation. Furthermore, silencing circMIRIAF caused the decreased circMIRIAF levels and the increased miR-544 levels in cardiomyocytes, while excessive miR-544 inhibited WDR12 expression to alleviate the disorder. On the contrary, excessive circMIRIAF increased WDR12 expression by adsorbing miR-544 to exacerbate H/RI in AC16 cells. In addition, circMIRIAF siRNA reversed the aggravation of H/RI in cells caused by WDR12 overexpression. Overall, circMIRIAF can serve as a drug target or treating MI/RI, and circMIRIAF could sponge miR-544 and enhance WDR12 expression to aggravate MI/RI, which may provide a novel therapeutic strategy for MI/RI treatment.
Subject(s)
MicroRNAs , Myocardial Reperfusion Injury , RNA, Circular , Animals , Humans , Male , Mice , Cell Line , Disease Models, Animal , Gene Expression Regulation , MicroRNAs/genetics , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/pathology , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/pathology , Oxidative Stress , RNA, Circular/genetics , Signal TransductionABSTRACT
BACKGROUND: Myocardial ischemia-reperfusion injury (I/RI) is a major cause of perioperative cardiac-related adverse events and death. Studies have shown that sevoflurane postconditioning (SpostC), which attenuates I/R injury and exerts cardioprotective effects, regulates mitochondrial dynamic balance via HIF-1α, but the exact mechanism is unknown. This study investigates whether the PI3K/AKT pathway in SpostC regulates mitochondrial dynamic balance by mediating HIF-1α, thereby exerting myocardial protective effects. METHODS: The H9C2 cardiomyocytes were cultured to establish the hypoxia-reoxygenation (H/R) model and randomly divided into 4 groups: Control group, H/R group, sevoflurane postconditioning (H/R + SpostC) group and PI3K/AKT blocker (H/R + SpostC + LY) group. Cell survival rate was determined by CCK-8; Apoptosis rate was determined by flow cytometry; mitochondrial membrane potential was evaluated by Mito Tracker™ Red; mRNA expression levels of AKT, HIF-1α, Opa1and Drp1 were detected by quantitative real-time polymerase chain reaction (qRT-PCR); Western Blot assay was used to detect the protein expression levels of AKT, phosphorylated AKT (p-AKT), HIF-1α, Opa1 and Drp1. RESULTS: Compared with the H/R group, the survival rate of cardiomyocytes in the H/R + SpostC group increased, the apoptosis rate decreased and the mitochondrial membrane potential increased. qRT-PCR showed that the mRNA expression of HIF-1α and Opa1 were higher in the H/R + SpostC group compared with the H/R group, whereas the transcription level of Drp1 was lower in the H/R + SpostC group. In the H/R + SpostC + LY group, the mRNA expression of HIF-1α was lower than the H/R + SpostC group. There was no difference in the expression of Opa1 mRNA between the H/R group and the H/R + SpostC + LY group. WB assay results showed that compared with the H/R group, the protein expression levels of HIF-1α, Opa1, P-AKT were increased and Drp1 protein expression levels were decreased in the H/R + SpostC group. HIF-1α, P-AKT protein expression levels were decreased in the H/R + SpostC + LY group compared to the H/R + SpostC group. CONCLUSION: SpostC mediates HIF-1α-regulated mitochondrial fission and fusion-related protein expression to maintain mitochondrial dynamic balance by activating the PI3K/AKT pathway and increasing AKT phosphorylation, thereby attenuating myocardial I/R injury.
Subject(s)
Apoptosis , Hypoxia-Inducible Factor 1, alpha Subunit , Membrane Potential, Mitochondrial , Mitochondria, Heart , Mitochondrial Dynamics , Myocardial Reperfusion Injury , Myocytes, Cardiac , Phosphatidylinositol 3-Kinase , Proto-Oncogene Proteins c-akt , Sevoflurane , Signal Transduction , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Proto-Oncogene Proteins c-akt/metabolism , Animals , Myocytes, Cardiac/drug effects , Myocytes, Cardiac/pathology , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/enzymology , Sevoflurane/pharmacology , Myocardial Reperfusion Injury/pathology , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/prevention & control , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/enzymology , Mitochondrial Dynamics/drug effects , Cell Line , Rats , Apoptosis/drug effects , Phosphatidylinositol 3-Kinase/metabolism , Mitochondria, Heart/drug effects , Mitochondria, Heart/metabolism , Mitochondria, Heart/pathology , Mitochondria, Heart/enzymology , Membrane Potential, Mitochondrial/drug effects , Cell Hypoxia , Dynamins/metabolism , Dynamins/genetics , GTP Phosphohydrolases/metabolism , GTP Phosphohydrolases/genetics , Phosphoinositide-3 Kinase Inhibitors/pharmacology , Cytoprotection , Ischemic Postconditioning , PhosphorylationABSTRACT
Objectives. The aim of this study was to investigate the expression of long non-coding RNA (lncRNA) brain and reproductive organ-expressed protein (BRE) antisense RNA 1 (BRE-AS1) in patients with acute myocardial infarction (AMI) and its effect on ischemia/reperfusion (I/R)-induced oxidative stress and apoptosis of cardiomyocytes. Methods. Serum BRE-AS1 levels in patients with AMI was detected using quantitative real-time polymerase chain reaction (qRT-PCR). The diagnostic and prognostic values of BRE-AS1 were evaluated. H9c2 cells were treated with hypoxia/reoxygenation to establish an in vitro myocardial infarction cell model. The levels of inflammatory cytokines such as tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and IL-6 were detected by enzyme-linked immunosorbent assay (ELISA). Levels of lactate dehydrogenase (LDH), malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px) were determined by commercial kits. Cell counting kit-8 (CCK-8) and flow cytometry were used to evaluate the cell viability and cell apoptosis. Results. The expression of BRE-AS1 in serum of patients with AMI is upregulated, which shows the clinical diagnostic value for AMI. In the I/R injury cell model, the knockout of BRE-AS1 can significantly alleviate the increase in TNF-α, IL-1ß, and IL-6 levels, inhibit the production of LDH and MDA, increase the activities of SOD and GSH-Px, promote the cell viability and suppress cell apoptosis. Conclusions. Abnormally elevated BRE-AS1 has a high diagnostic value for AMI as well as a prognostic value for major adverse cardiovascular events (MACEs). The elevation of BRE-AS1 promoted oxidative stress injury and cell apoptosis in vitro.
Subject(s)
Apoptosis , Inflammation Mediators , Myocardial Infarction , Myocytes, Cardiac , Oxidative Stress , RNA, Long Noncoding , Animals , Female , Humans , Male , Rats , Case-Control Studies , Cell Line , Cytokines/metabolism , Cytokines/blood , Inflammation Mediators/metabolism , Inflammation Mediators/blood , Myocardial Infarction/metabolism , Myocardial Infarction/pathology , Myocardial Infarction/blood , Myocardial Infarction/genetics , Myocardial Infarction/diagnosis , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/pathology , Myocardial Reperfusion Injury/blood , Myocardial Reperfusion Injury/diagnosis , Myocardial Reperfusion Injury/genetics , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/pathology , RNA, Long Noncoding/blood , RNA, Long Noncoding/metabolism , RNA, Long Noncoding/genetics , Signal Transduction , Up-RegulationABSTRACT
BACKGROUND: Myocardial ischemia-reperfusion injury (MIRI) is caused by reperfusion after ischemic heart disease. LncRNA Snhg1 regulates the progression of various diseases. N6-methyladenosine (m6A) is the frequent RNA modification and plays a critical role in MIRI. However, it is unclear whether lncRNA Snhg1 regulates MIRI progression and whether the lncRNA Snhg1 was modified by m6A methylation. METHODS: Mouse cardiomyocytes HL-1 cells were utilized to construct the hypoxia/reoxygenation (H/R) injury model. HL-1 cell viability was evaluated utilizing CCK-8 method. Cell apoptosis, mitochondrial reactive oxygen species (ROS), and mitochondrial membrane potential (MMP) were quantitated utilizing flow cytometry. RNA immunoprecipitation and dual-luciferase reporter assays were applied to measure the m6A methylation and the interactions between lncRNA Snhg1 and targeted miRNA or target miRNAs and its target gene. The I/R mouse model was constructed with adenovirus expressing lncRNA Snhg1. HE and TUNEL staining were used to evaluate myocardial tissue damage and apoptosis. RESULTS: LncRNA Snhg1 was down-regulated after H/R injury, and overexpressed lncRNA Snhg1 suppressed H/R-stimulated cell apoptosis, mitochondrial ROS level and polarization. Besides, lncRNA Snhg1 could target miR-361-5p, and miR-361-5p targeted OPA1. Overexpressed lncRNA Snhg1 suppressed H/R-stimulated cell apoptosis, mitochondrial ROS level and polarization though the miR-361-5p/OPA1 axis. Furthermore, WTAP induced lncRNA Snhg1 m6A modification in H/R-stimulated HL-1 cells. Moreover, enforced lncRNA Snhg1 repressed I/R-stimulated myocardial tissue damage and apoptosis and regulated the miR-361-5p and OPA1 levels. CONCLUSION: WTAP-mediated m6A modification of lncRNA Snhg1 regulated MIRI progression through modulating myocardial apoptosis, mitochondrial ROS production, and mitochondrial polarization via miR-361-5p/OPA1 axis, providing the evidence for lncRNA as the prospective target for alleviating MIRI progression.
Subject(s)
Apoptosis , MicroRNAs , Mitochondrial Dynamics , Myocardial Reperfusion Injury , Myocytes, Cardiac , RNA, Long Noncoding , Animals , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolism , MicroRNAs/metabolism , MicroRNAs/genetics , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/pathology , Mice , Apoptosis/genetics , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/pathology , Cell Line , Male , Mice, Inbred C57BL , GTP Phosphohydrolases/metabolism , GTP Phosphohydrolases/genetics , Reactive Oxygen Species/metabolism , Adenosine/analogs & derivatives , Adenosine/metabolism , Base Sequence , Methylation , Membrane Potential, MitochondrialABSTRACT
BACKGROUND: Aberrant mitochondrial fission, a critical pathological event underlying myocardial ischemia/reperfusion (MI/R) injury, has emerged as a potential therapeutic target. The long non-coding RNA (lncRNA) Oip5-as1 is increasingly recognized for its regulatory roles, particularly in MI/R injury. However, its precise mechanistic role in modulating mitochondrial dynamics remains elusive. This study aims to elucidate the mechanistic role of Oip5-as1 in regulating mitochondrial fission and evaluate its therapeutic potential against MI/R injury. METHODS: To simulate in vitro MI/R injury, HL-1 cardiomyocytes were subjected to hypoxia/reoxygenation (H/R). Lentiviral vectors were employed to achieve overexpression or knockdown of Oip5-as1 in HL-1 cells by expressing Oip5-as1 or shRNA targeting Oip5-as1, respectively. The impact of Oip5-as1 on mitochondrial dynamics in HL-1 cells was assessed using CCK-8 assay, flow cytometry, immunofluorescence staining, and biochemical assays. MI/R injury was induced in mice by ligating the left anterior descending coronary artery. Conditional knockout mice for Oip5-as1 were generated using the CRISPR/Cas9 genome editing technology, while overexpression of Oip5-as1 in mice was achieved via intramyocardial administration of AAV9 vectors. In mice, the role of Oip5-as1 was evaluated through echocardiographic assessment, histopathological staining, and transmission electron microscopy. Furthermore, Western blotting, RNA pull-down, RNA immunoprecipitation, and co-immunoprecipitation assays were conducted to investigate Oip5-as1's underlying mechanisms. RESULTS: The expression levels of Oip5-as1 are significantly decreased in MI/R-injured HL-1 cells and myocardium. In HL-1 cells undergoing H/R injury, overexpression of Oip5-as1 attenuated excessive mitochondrial fission, preserved mitochondrial functionality, and reduced cellular apoptosis, while knockdown of Oip5-as1 exhibited the opposite effects. Furthermore, in a mouse model of MI/R injury, overexpression of Oip5-as1 diminished mitochondrial fission, myocardial infarct size and improved cardiac function. However, knockout of Oip5-as1 exacerbated myocardial injury and cardiac dysfunction, which were significantly reversed by treatment with a mitochondrial division inhibitor-1 (Mdivi-1). Mechanistically, Oip5-as1 selectively interacts with AKAP1 and CaN proteins, inhibiting CaN activation and subsequent DRP1 dephosphorylation at Ser637, thereby constraining DRP1's translocation to the mitochondria and its involvement in mitochondrial fission. CONCLUSIONS: Our study underscores the pivotal role of Oip5-as1 in mitigating excessive mitochondrial fission during MI/R injury. The findings not only enhance our comprehension of the molecular mechanisms underlying MI/R injury but also identify Oip5-as1 as a potential therapeutic target for ameliorating MI/R injury.
Subject(s)
Dynamins , Mitochondrial Dynamics , Myocardial Reperfusion Injury , Myocytes, Cardiac , RNA, Long Noncoding , Animals , Mice , Cell Line , Dynamins/metabolism , Dynamins/genetics , Mice, Inbred C57BL , Mice, Knockout , Mitochondrial Dynamics/genetics , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/pathology , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/pathology , Phosphorylation , RNA, Long Noncoding/genetics , RNA, Long Noncoding/metabolismABSTRACT
BACKGROUND: Hypothermic ischemia-reperfusion arrhythmia is a common complication of cardiothoracic surgery under cardiopulmonary bypass, but few studies have focused on this type of arrhythmia. Our prior study discovered reduced myocardial Cx43 protein levels may be linked to hypothermic reperfusion arrhythmias. However, more detailed molecular mechanism research is required. METHOD: The microRNA and mRNA expression levels in myocardial tissues were detected by real-time quantitative PCR (RT-qPCR). Besides, the occurrence of hypothermic reperfusion arrhythmias and changes in myocardial electrical conduction were assessed by electrocardiography and ventricular epicardial activation mapping. Furthermore, bioinformatics analysis, applying antagonists of miRNA, western blotting, immunohistochemistry, a dual luciferase assay, and pearson correlation analysis were performed to investigate the underlying molecular mechanisms. RESULTS: The expression level of novel-miR-17 was up-regulated in hypothermic ischemia-reperfusion myocardial tissues. Inhibition of novel-miR-17 upregulation ameliorated cardiomyocyte edema, reduced apoptosis, increased myocardial electrical conduction velocity, and shortened the duration of reperfusion arrhythmias. Mechanistic studies showed that novel-miR-17 reduced the expression of Cx43 by directly targeting Gja1 while mediating the activation of the PKC/c-Jun signaling pathway. CONCLUSION: Up-regulated novel-miR-17 is a newly discovered pro-arrhythmic microRNA that may serve as a potential therapeutic target and biomarker for hypothermic reperfusion arrhythmias.
Subject(s)
Arrhythmias, Cardiac , Connexin 43 , MicroRNAs , Protein Kinase C , Signal Transduction , Animals , Humans , Male , Rats , Apoptosis/genetics , Arrhythmias, Cardiac/metabolism , Arrhythmias, Cardiac/genetics , Arrhythmias, Cardiac/etiology , Arrhythmias, Cardiac/pathology , Connexin 43/metabolism , Connexin 43/genetics , MicroRNAs/genetics , MicroRNAs/metabolism , Myocardial Reperfusion Injury/metabolism , Myocardial Reperfusion Injury/genetics , Myocardial Reperfusion Injury/pathology , Myocardial Reperfusion Injury/etiology , Myocardium/metabolism , Myocardium/pathology , Myocytes, Cardiac/metabolism , Myocytes, Cardiac/pathology , Protein Kinase C/metabolism , Protein Kinase C/genetics , Proto-Oncogene Proteins c-jun/metabolism , Up-RegulationABSTRACT
This research is concentrated on investigating the role and mechanism of miR-652-3p in the protective effects of isoflurane (ISO) against myocardial ischemia-reperfusion (I/R) injury. H9c2 cells underwent pretreatment with varying concentrations of ISO, and subsequently, a hypoxia/reoxygenation (H/R) model was constructed. The levels of miR-652-3p, ISL LIM homeobox 1 (ISL1), and inflammatory cytokines interleukin (IL)-6 and tumor necrosis factor-alpha (TNF-α) were evaluated through reverse transcription polymerase chain reaction (RT-qPCR). Enzyme-linked immunosorbent assay was employed to investigate concentrations of myocardial injury markers, such as creatine kinase-MB (CK-MB) and cardiac troponin I (cTnI). Cell counting kit-8 was used to evaluate cell viability, while flow cytometry was utilized to measure apoptosis. Additionally, a dual luciferase reporter assay was conducted to validate the targeting relationship between ISL1 and miR-652-3p. Herein, we confirmed that the level of miR-652-3p was gradually increased with prolonged hypoxia; nevertheless, this increase was suppressed by ISO pretreatment (P < 0.05). Additionally, ISO pretreatment prevented the decrease in cell viability, increase in apoptosis, and overproduction of IL-6, TNF-α, CK-MB, and cTnI induced by H/R (P < 0.05). However, the inhibitory effects of ISO were counteracted by the increased levels of miR-652-3p (P < 0.05). ISL1 is a potential target of miR-652-3p. H/R induction suppressed ISL1 levels compared to the control, but ISO treatment increased its expression (P < 0.05). Overexpression of ISL1 inhibited the elimination of the protective effect of ISO on myocardial damage induced by the elevation of miR-652-3p (P < 0.05). The findings of this research confirm that miR-652-3p attenuated the protective effect of ISO on cardiomyocytes in myocardial ischemia by targeting ISL1.