Myocardial Ischemia-reperfusion Injury Group Research Articles

Investigation of the mechanism of poly (ADP-ribose) polymerase (PARP) in elderly mouse myocardial ischemia-reperfusion injury.

This study investigated the role of Poly (ADP-ribose) Polymerase (PARP) in myocardial ischemia-reperfusion injury (MIRI) in elderly mice. It involves 30 elderly male KM mice divided into three groups: Sham, MIRI, and DPQ, where the MIRI and DPQ groups undergo myocardial ischemia-reperfusion with the DPQ group also receiving DPQ for PARP-1 inhibition. Over three weeks, assessments include histological analysis of myocardial lesions, left ventricular ejection fraction (LVEF) measurements, and evaluations of serum cardiac enzymes and inflammatory markers. This approach aims to understand the protective effects of DPQ in MIRI, focusing on its impact on cardiac health and inflammation via the JAK2/STAT3 pathway. The findings suggest that PARP activation exacerbates cardiac dysfunction and inflammation in MIRI by possibly modulating the JAK2/STAT3 signaling pathway. Inhibition of PARP-1 with DPQ mitigates these effects, as indicated by reduced myocardial lesions and inflammatory infiltration, improved LVEF, and altered levels of inflammatory markers and signaling molecules. However, the differences in STAT3 and p-STAT3 protein expression between the DPQ and MIRI groups were not statistically significant, suggesting that while PARP inhibition affects many aspects of MIRI pathology, its impact on the JAK2/STAT3 pathway may not fully explain the observed benefits. This study contributes to our understanding of the complex mechanisms underlying myocardial ischemia-reperfusion injury, particularly in the context of aging. It highlights the potential of PARP inhibition as a therapeutic strategy to attenuate cardiac dysfunction and inflammation in MIRI, though further research is necessary to fully elucidate the underlying molecular pathways and to explore the clinical relevance of these findings in humans.

Bioinformatics integration reveals key genes associated with mitophagy in myocardial ischemia-reperfusion injury

BackgroundMyocardial ischemia is a prevalent cardiovascular disorder associated with significant morbidity and mortality. While prompt restoration of blood flow is essential for improving patient outcomes, the subsequent reperfusion process can result in myocardial ischemia–reperfusion injury (MIRI). Mitophagy, a specialized autophagic mechanism, has consistently been implicated in various cardiovascular disorders. However, the specific connection between ischemia–reperfusion and mitophagy remains elusive. This study aims to elucidate and validate central mitophagy-related genes associated with MIRI through comprehensive bioinformatics analysis.MethodsWe acquired the microarray expression profile dataset (GSE108940) from the Gene Expression Omnibus (GEO) and identified differentially expressed genes (DEGs) using GEO2R. Subsequently, these DEGs were cross-referenced with the mitophagy database, and differential nucleotide sequence analysis was performed through enrichment analysis. Protein–protein interaction (PPI) network analysis was employed to identify hub genes, followed by clustering of these hub genes using cytoHubba and MCODE within Cytoscape software. Gene set enrichment analysis (GSEA) was conducted on central genes. Additionally, Western blotting, immunofluorescence, and quantitative polymerase chain reaction (qPCR) analyses were conducted to validate the expression patterns of pivotal genes in MIRI rat model and H9C2 cardiomyocytes.ResultsA total of 2719 DEGs and 61 mitophagy-DEGs were identified, followed by enrichment analyses and the construction of a PPI network. HSP90AA1, RPS27A, EEF2, EIF4A1, EIF2S1, HIF-1α, and BNIP3 emerged as the seven hub genes identified by cytoHubba and MCODE of Cytoscape software. Functional clustering analysis of HIF-1α and BNIP3 yielded a score of 9.647, as determined by Cytoscape (MCODE). In our MIRI rat model, Western blot and immunofluorescence analyses confirmed a significant elevation in the expression of HIF-1α and BNIP3, accompanied by a notable increase in the ratio of LC3II to LC3I. Subsequently, qPCR confirmed a significant upregulation of HIF-1α, BNIP3, and LC3 mRNA in the MIRI group. Activation of the HIF-1α/BNIP3 pathway mediates the regulation of the degree of Mitophagy, thereby effectively reducing apoptosis in rat H9C2 cardiomyocytes.ConclusionsThis study has identified seven central genes among mitophagy-related DEGs that may play a pivotal role in MIRI, suggesting a correlation between the HIF-1α/BNIP3 pathway of mitophagy and the pathogenesis of MIRI. The findings highlight the potential importance of mitophagy in MIRI and provide valuable insights into underlying mechanisms and potential therapeutic targets for further exploration in future studies.

Ticagrelor alleviates pyroptosis of myocardial ischemia reperfusion-induced acute lung injury in rats: a preliminary study.

Pulmonary infection is highly prevalent in patients with acute myocardial infarction undergoing percutaneous coronary intervention. However, the potential mechanism is not well characterized. Myocardial ischemia-reperfusion injury (MIRI) induces acute lung injury (ALI) related to pulmonary infection and inflammation. Recent studies have shown that pyroptosis mediates ALI in several human respiratory diseases. It is not known whether MIRI induces pyroptosis in the lungs. Furthermore, ticagrelor is a clinically approved anti-platelet drug that reduces ALI and inhibits the expression levels of several pyroptosis-associated proteins, but the effects of ticagrelor on MIRI-induced ALI have not been reported. Therefore, we investigated whether ticagrelor alleviated ALI in the rat MIRI model, and its effects on pyroptosis in the lungs. Sprague-Dawley rats were randomly divided into four groups: control, MIRI, MIRI plus low ticagrelor (30 mg/kg), and MIRI plus high ticagrelor (100 mg/kg). Hematoxylin and Eosin (HE) staining was performed on the lung sections, and the HE scores were calculated to determine the extent of lung pathology. The wet-to-dry ratio of the lung tissues were also determined. The expression levels of pyroptosis-related proteins such as NLRP3, ASC, and Cleaved caspase-1 were estimated in the lung tissues using the western blot. ELISA was used to estimate the IL-1β levels in the lungs. Immunohistochemistry was performed to determine the levels of MPO-positive neutrophils as well as the total NLRP3-positive and Cleaved caspase-1-positive areas in the lung tissues. The lung tissues from the MIRI group rats showed significantly higher HE score, wet-to-dry ratio, and the MPO-positive area compared to the control group, but these effects were attenuated by pre-treatment with ticagrelor. Furthermore, lung tissues of the MIRI group rats showed significantly higher expression levels of pyroptosis-associated proteins, including NLRP3 (2.1-fold, P < 0.05), ASC (3.0-fold, P < 0.01), and Cleaved caspase-1 (9.0-fold, P < 0.01). Pre-treatment with the high-dose of ticagrelor suppressed MIRI-induced upregulation of NLRP3 (0.46-fold, P < 0.05), ASC (0.64-fold, P < 0.01), and Cleaved caspase-1 (0.80-fold, P < 0.01). Immunohistochemistry results also confirmed that pre-treatment with ticagrelor suppressed MIRI-induced upregulation of pyroptosis in the lungs. In summary, our data demonstrated that MIRI induced ALI and upregulated pyroptosis in the rat lung tissues. Pre-treatment with ticagrelor attenuated these effects.

Single-cell landscape dissecting the transcription and heterogeneity of innate lymphoid cells in ischemic heart.

Until now, few articles have revealed the potential roles of innate lymphoid cells (ILCs) in cardiovascular diseases. However, the infiltration of ILC subsets in ischemic myocardium, the roles of ILC subsets in myocardial infarction (MI) and myocardial ischemia-reperfusion injury (MIRI) and the related cellular and molecular mechanisms have not been described with a sufficient level of detail. In the current study, 8-week-old male C57BL/6J mice were divided into three groups: MI, MIRI and sham group. Single-cell sequencing technology was used to perform dimensionality reduction clustering of ILC to analyze the ILC subset landscape at a single-cell resolution, and finally flow cytometry was used to confirm the existence of the new ILC subsets in different disease groups. Five ILC subsets were found, including ILC1, ILC2a, ILC2b, ILCdc and ILCt. It is worth noting that ILCdc, ILC2b and ILCt were identified as new ILC subclusters in the heart. The cellular landscapes of ILCs were revealed and signal pathways were predicted. Furthermore, pseudotime trajectory analysis exhibited different ILC statuses and traced related gene expression in normal and ischemic conditions. In addition, we established a ligand-receptor-transcription factor-target gene regulatory network to disclose cell communications among ILC clusters. Moreover, we further revealed the transcriptional features of the ILCdc and ILC2a subsets. Finally, the existence of ILCdc was confirmed by flow cytometry. Collectively, by characterizing the spectrums of ILC subclusters, our results provide a new blueprint for understanding ILC subclusters' roles in myocardial ischemia diseases and further potential treatment targets.

Emodin Regulates lncRNA XIST/miR-217 Axis to Protect Myocardial Ischemia-Reperfusion Injury.

This study is aimed at investigating the effect of emodin on myocardial ischemia-reperfusion injury (MIRI) and mechanism. Eighty healthy adult male SD rats (weighing 230-250 g) were utilized to establish I/R model, which were randomly divided into five groups (16 rats in each group): sham operation group, myocardial ischemia-reperfusion injury group (I/R group), emodin group, emodin +NC group, and emodin +XIST group. The contents of CK, CK-MB, LDH, and HBDH in serum were determined by ELISA kit. LDH was detected by ELISA assay, SOD was detected by the xanthine oxidase method, and MDA was detected by the thiobarbituric acid method. The relative expression of XIST and miR-217 was evaluated by RT-qPCR. Western blot was applied to detect the protein expression. Flow cytometry was applied to detect cardiomyocyte apoptosis. Myocardial infarction area was obviously increased in I/R model rats, while emodin decreased the myocardial infarction in I/R model rats. In addition, cardiac enzymes (CK, CK-MB, LDH, and HBDH) and apoptosis were obviously increased in MIRI model rats, while emodin obviously decreased cardiac enzymes and apoptosis. The ROS and MDA levels were raised while the activities of SOD were declined in the I/R model group. The ROS and MDA levels were decreased while the activities of SOD were raised in the emodin group. The XIST expression was markedly raised in the I/R model group while decreased in the emodin group, and the overexpression of XIST reversed the protective effect of emodin on myocardial infarction, oxidative stress, and cardiomyocyte apoptosis. In addition, XIST directly regulated the expression of miR-217, and si-XIST inhibited H/R-induced oxidative damage of cardiomyocytes via inhibiting miR-217. Emodin protected MIRI both in vitro and in vivo via inhibiting lncRNA XIST to upregulate miR-217.

Serum oxidative stress factors predict myocardial ischemia reperfusion injury after percutaneous coronary intervention in patients with acute myocardial infarction and type 2 diabetes mellitus.

Serum oxidative stress factors may be considered to be essential parameters for indicating cell oxidative damage. We designed this study to investigate the clinical diagnostic value of serum oxidative stress factors (superoxide dismutase (SOD), malondialdehyde (MDA), and myeloperoxidase (MPO)) combined with ischemia-modified albumin (IMA) and heat shock protein 70 (HSP70) for myocardial ischemia-reperfusion injury (MIRI) after percutaneous coronary intervention (PCI) in elderly patients with acute myocardial infarction (AMI) and type 2 diabetes mellitus (T2DM). From November 2020 to August 2021, 94 patients with AMI + T2DM and 86 patients with AMI were enrolled in the study; they were sub-grouped into the MIRI and non-MIRI groups following the occurrence of MIRI within 48 h after PCI. SOD, MDA, MPO, IMA, and HSP70 levels were determined. The clinical values of the combined serum oxidative stress factors, IMA, and HSP70 levels to predict MIRI events were analyzed. There was a higher probability of MIRI events in the AMI + T2DM group than the AMI group (p < 0.05). The ROC curve for the combined prediction of SOD, MDA, MPO, IMA, and HSP70 for the occurrence of MIRI events was higher in both the AMI and the AMI + T2DM groups than for predictive factors alone (all p < 0.05). Combined prediction of SOD, MDA, MPO, IMA, and HSP70 has the highest diagnostic value for predicting MIRI events after PCI in AMI patients, especially in patients with AMI combined with T2DM.

Phycocyanin relieves myocardial ischemia-reperfusion injury in rats by inhibiting oxidative stress

Purpose: To investigate the effect of phycocyanin on myocardial ischemia-reperfusion injury, and the possible mechanisms involved.&#x0D; Methods: Twenty-four Sprague-Dawley (SD) rats were randomly divided into Sham group (only threading without ligation), IRI group (myocardial ischemia-reperfusion injury group) and phycocyanin group (phycocyanin pretreatment + myocardial ischemia-reperfusion injury group). The heart was harvested and cardiomyocytes were isolated. Colorimetry was used to determine the contents of cardiomyocyte serum creatine phospho-MB (CK-MB), lactate dehydrogenase (LDH) and malondialdehyde (MDA), and the activities of total antioxidant capacity (T-AOC), catalase (CAT), glutathione (GSH), total superoxide dismutase (SOD) and other related oxidative stress indicators. Furthermore, apoptosis was evaluated using TUNEL staining. Protein levels of cardiac factor E2 related factor 2 (Nrf2), heme oxygenase-1 (HO-1), human NADPH dehydrogenase 1 (NQO1) and nuclear factor-κB (NF-κB) were evaluated by Western blot and immunohistochemistry.&#x0D; Results: Compared with the myocardial IRI group, the contents of CK-MB, LDH, MAD and ROS in the treated group were significantly decreased (p &lt; 0.05), but the activities of SOD, GSH, SOD, CAT, and T-AOC in the myocardial tissues were significantly enhanced (p &lt; 0.05). Moreover, the pathological changes in myocardial tissue were significantly reduced. In addition, the expression levels of Nrf2, HO-1 and NQO-1 were significantly up-regulated after phycocyanin pretreatment, while expression of NF-κB was significantly down-regulated (p &lt; 0.05).&#x0D; Conclusion: Phycocyanin improves myocardial anti-oxidative stress via activation of Nrf2 signaling pathway, and also protects rats from myocardial ischemia-reperfusion injury by reducing inflammatory response via inhibition of NF-κB signaling pathway.

Effects of Mongolian medicine Shaosha-7 on myocardial ischemia/ reperfusion injury of rats

Objective: To investigate the effects of Shaosha-7 in rats with myocardial ischemia-reperfusion injury and its mechanisms. Methods: male SD rats were divided into sham operation group (n=10), myocardial ischemia-reperfusion injury group (n=10), low, medium and high dose of Shaosha-7 groups (n=10), and positive drug group (n=10). The rats of Shaosha-7 (low, medium and high dose) groups were treated with Shaosha-7 at the doses of 0.4, 0.8 and 1.6 g/kg respectively, once a day for 15 days. The rats of positive drug group were treated with 0.3 g/kg Danshen, once a day for 15 days. The rats of the sham operation group and myocardial ischemia-reperfusion injury were treated with 2 ml/100 g distilled water, once a day for 15 days. After 15 days, the rats of the model group and the treatment group underwent thoracotomy and ligation of coronary artery for 30 minutes, then thoracic cavity was closed after reperfusion. Rats in six groups were executed electrocardiographic examination and their hearts were taken for Hematoxylin-Eosin (HE) staining and 2, 3, 5-triphenyltetrazolium chloride (TTC) staining to observe infarct area and myocardial pathological changes. The contents of cTnI, CK-MB, LDH, MDA, SOD, GSH-Px, TNF-α, IL-18, IL-6 and IL-1 β in serum were detected by ELISA. The expression of NF-кB was detected by immunohistochemistry. Results: Compared with the sham operation group, the infarct size, the levels of cTnI, CK-MB, CK-MB, LDH, MDA, GSH-Px, TNF-α, IL-18, IL-6, IL-1β and NF-кB were increased and the content of SOD were decreased in rats with myocardial ischemia-reperfusion injury. Compared with the rats with myocardial ischemia-reperfusion injury, Shaosha-7 improved the arrhythmia and pathological changes, reduced the infarct area, decreased the contents of cTnI, CK-MB, LDH, MDA, GSH-Px, TNF-α, IL-18, IL-6, IL-1 β, increased the content of SOD, decreased the expression of NF-кB. Conclusion: Mongolian medicine Shaosha-7 can effectively alleviate myocardial ischemia-reperfusion injury in rats. This study provides a theoretical basis for the treatment of myocardial ischemia-reperfusion (I/R) injury with Shaosha-7.

Systematic Pharmacology Reveals the Antioxidative Stress and Anti-Inflammatory Mechanisms of Resveratrol Intervention in Myocardial Ischemia-Reperfusion Injury.

Objective To explore the oxidative stress and inflammatory mechanisms of resveratrol intervention in myocardial ischemia-reperfusion injury (MIRI). Methods The potential targets of resveratrol were predicted by PharmMapper. The MIRI genes were collected by Online Mendelian Inheritance in Man (OMIM), GeneCards is used to collect related disease genes, and String is used for enrichment analysis. Animal experiments were then performed to verify the systematic pharmacological results. Hematoxylin-eosin (HE) staining was used to observe myocardial damage. The levels of serum interleukin-1β (IL-1β), IL-6, and tumor necrosis factor-α (TNF-α) in each experimental group were detected. The protein and mRNA expressions of Toll-like receptor 4 (TLR4), nuclear factor-kappa (NF-κB) p65, IL-1β, IL-6, and TNF-α in rat myocardial tissue were measured. Results The results of systematic pharmacology showed that insulin resistance, FoxO signaling pathway, adipocytokine signaling pathway, insulin signaling pathway, PI3K-Akt signaling pathway, ErbB signaling pathway, T-cell receptor signaling pathway, peroxisome proliferator-activated receptors (PPAR) signaling pathway, Ras signaling pathway, TNF signaling pathway, and so on were regulated to improve MIRI. The results of animal experiments showed that the myocardial cells of the sham operation group were arranged in fibrous form, and the myocardial ischemia-reperfusion injury group had obvious cell morphology disorder. Compared with the MIRI group, the resveratrol group had a certain degree of relief. Compared with the MIRI group, serum IL-1β, TNF-α, and IL-6 in the resveratrol group was significantly reduced (P < 0.05), and myocardial tissue TLR4, NF-κB p65, IL-1β, IL-6, and TNF-α mRNA and protein expressions were significantly reduced (P < 0.05). Conclusion Resveratrol can effectively improve MIRI, and its mechanism may be related to antioxidative stress and anti-inflammatory.

Propofol relieves inflammation in MIRI rats by inhibiting Rho/Rock signaling pathway.

The aim of this study was to investigate the effects of propofol on myocardial ischemia-reperfusion injury (MIRI) and its mechanism by establishing in vivo rat models. Sprague-Dawley rats were selected for the construction of MIRI models in vivo. All rats were divided into three groups, including sham operation group (Sham operation), MIRI group and MIRI + propofol group. At 2 h after reperfusion, myocardial tissues and blood samples were collected from rats. The expression levels of serum lactic dehydrogenase (LDH) and creatine kinase-MB (CK-MB), as well as serum interleukin-6 (IL-6), IL-10 and tumor necrosis factor-α (TNF-α), were measured in each group of rats, respectively. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) assay was employed to detect the apoptosis of myocardial cells. Additionally, the messenger ribonucleic acid (mRNA) and protein expressions of Ras homolog gene family, member A (RhoA) and Rho-associated coiled-coil-containing protein kinase 2 (Rock2) were determined via quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and Western blotting, respectively. (1) The expression levels of serum LDH and CK-MB were significantly lower in MIRI + propofol group than those in MIRI group (p<0.05). (2) In comparison with MIRI group, MIRI + propofol group exhibited significantly reduced serum IL-6 and TNF-α levels (p<0.01) and elevated serum IL-10 level (p<0.01). (3) Compared with MIRI group, the apoptosis of myocardial cells was remarkably reduced in MIRI + propofol group after IRI (p<0.05). (4) The mRNA and protein expressions of RhoA and Rock2 were significantly lower in MIRI + propofol group than those in MIRI group (p<0.05). Propofol relieves MIRI and inflammation, reduces the level of oxidative stress and represses I/R-induced myocardial cell apoptosis in MIRI rats by inhibiting the activity of the Rho/Rock signaling pathway.

Protective effect of MAPK signaling pathway mediated by ITGB3 gene silencing on myocardial ischemia-reperfusion injury in mice and its mechanism.

To explore the effect of integrin β3 (ITGB3) gene silencing mediated mitogen-activated protein kinase (MAPK) signaling pathway on myocardial ischemia-reperfusion injury (MIRI) in mice. MIRI mice model was established, and myocardial tissues of MIRI mice and sham operation group mice were extracted. Hematoxylin-Eosin (HE) staining was used to observe the pathological changes of myocardial tissue; terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) staining was used to detect the apoptosis of myocardial cells; ELISA method was used to detect the levels of interleukin (IL)-1, IL-6, and tumor necrosis factor (TNF)-α in the two groups. The infarct size was measured by TTC staining. Myocardial cells of MIRI model mice were isolated and cultured, and then grouped and transfected. The cells were transfected with the grouping of MIRI group, negative control (NC) group, MAPK signal pathway agonist Anisomycin group, MAPK signal pathway inhibitor SB203580 group, ITGB3-siRNA group, SB203580 + ITGB3-siRNA group. Real-time quantitative polymerase chain reaction (qRT-PCR) and Western blot were used to detect the mRNA and protein expressions of ITGB3, p38MAPK/p-p38MAPK, GSK-3β/p-GSK-3β, Cx43/p-Cx43, pro-apoptotic factor Bax and anti-apoptotic factor Bcl-2. 3-(4,5)-dimethylthiahiazo (-z-y1)-3,5-di-phenytetrazoliumromide (MTT) assay was used to detect cell proliferation and flow cytometry to detect cell apoptosis. The expression of ITGB3 in myocardial tissue of MIRI mice was significantly higher than that of sham operated mice (p<0.05). Compared with the sham operation group, the apoptosis rate of myocardial cells in MIRI group was significantly increased, the expression levels of IL-1, IL-6 and TNF-α were significantly increased, and the myocardial infarction area was significantly increased (all p<0.05). Compared with MIRI and NC groups, ITGB3 mRNA and protein expression levels in ITGB3-siRNA group and SB203580 + ITGB3-siRNA group were significantly decreased (all p<0.05), but no significant change was found in Anisomycin group and SB203580 group (p>0.05). Furthermore, ITGB3-siRNA group and Anisomycin group had markedly decreased mRNA and protein expressions of ITGB3 and Bax, increased mRNA and protein expressions of p38MAPK/p-p38MAPK, GSK-3β/p-GSK-3β, Cx43/p-Cx43 and Bcl-2, as well as increased cell proliferation and decreased cell apoptosis (all p<0.05); SB203580 group indicated an opposite result with Anisomycin group; while SB203580 + ITGB3-siRNA revealed none significant statistical difference. In addition, compared with ITGB3-siRNA group, SB203580 + ITGB3-siRNA group showed significantly upregulated mRNA and protein expressions of Bax, downregulated mRNA and protein expressions of p38MAPK/p-p38MAPK, GSK-3β/p-GSK-3β, Cx43/p-Cx43 and Bcl-2, as well as decreased cell proliferation and increased cell apoptosis (all p<0.05). Silencing ITGB3 gene expression can promote the activation of MAPK signaling pathway, elevate the phosphorylation of GSK-3β and Cx43 in the downstream, promote the proliferation of mouse myocardial cells, inhibit myocardial cell apoptosis and inflammatory reaction, and thus have protective effect on MIRI in mice.

Effect of electroacupuncture preconditioning on cell apoptosis mediated by mitochondrial reactive oxygen species in myocardial ischemia/reperfusion injury rats

To observe the effect of electroacupuncture (EA) preconditioning on cell apoptosis and the content of mitochondrial reactive oxygen species (ROS) in rats with myocardial ischemia/reperfusion injury (MIRI), so as to explore their mechanisms underlying improvement of MIRI. Eighty male Wistar rats were randomly divided into control, sham, ischemia reperfusion (IR) and EA groups, with 20 rats in each group. Rats of the control, sham and IR groups were just banded on the boards for 30 min, once daily for 7 days. Before modeling, EA (2 Hz, 1 mA) was applied to "Neiguan" (PC6), "Zusanli" (ST36), "Guanyuan"(CV4) for 20 min, once daily for 7 successive days in the EA group. The MIRI model was established by occlusion of the left anterior descending branch of the left coronary artery for 20 min, followed by reperfusion for 30 min. Ventricular arrhythmia (VA) score was used to evaluate arrhythmia. Enzyme-linked immunosorbent assay (ELISA) was used to detect the level of CK-MB. DHE staining was used to detect the content of reactive oxygen species (ROS). The gene expression levels of cytochrome C (Cyt-C), Caspase-9 and Caspase-3 were detected by real-time fluorescent quantitative PCR. Compared with the sham group, VA score, serum CK-MB content, ROS content in heart tissue and Cyt-C, Caspase-9 and Caspase-3 gene expression were significantly up-regulated in the MIRI group (all P<0.01). Following the intervention, the increased VA score, serum CK-MB content, ROS content in heart tissue, Cyt-C, Caspase-9 and Caspase-3 gene expression were all reversed in the EA group compared with the MIRI group (all P<0.01). The protective effect of EA preconditioning on MIRI may be based on the regulation of ROS mediated-apoptosis pathway.

Effect of Vaspin on Myocardial Ischemia-Reperfusion Injury Rats and Expression of NLR Family Pyrin Domain Containing 3 (NLRP3)

Myocardial ischemia-reperfusion injury (MIRI) can cause myocardial damage. Vaspin can protect against myocardial damage. However, the effect of vaspin on MIRI rats and the expression of NLRP3 remains unclear. Sprague-Dawley rats were separated into sham group; MIRI group and Vaspin group, in which 100 ng/ml vaspin was administrated before model preparation followed by analysis of cardiac function by M-mode ultrasound, level of NLRP3, of type I collagen, IL-6 and TNF-α by ELISA, SOD activity and ROS by spectrophotometry and Bcl-2 and PI3K/AKT signaling protein expression by Western Blot. In MIRI group, left ventricular end-systolic diameter (LVESD), left ventricular mass index (LVMI), left ventricular end-diastolic diameter (LVEDD), NLRP3 expression, contents of type I collagen, IL-6, TNF-α as well as ROS were significantly increased and SOD activity was significantly decreased with decreased Bcl-2 expression and upregulated pAKT and pPI3K (P &lt; 0.05). In Vaspin group, LVESD, LVMI and LVEDD and NLRP3 expression as well as type I collagen, IL-6, TNF-α and ROS was decreased, SOD activity and Bcl-2 expression was significantly increased with downregulated pAKT and pPI3K (P &lt; 0.05). Vaspin can regulate PI3K/AKT signaling pathway, inhibit NLRP3 expression, regulate oxidative stress, inhibit inflammation, reduce apoptosis, improve and improve cardiac function of myocardial ischemia-reperfusion injury in rats.

Effects of electroacupuncture pretreatment on mitochondrial energy metabolism in the rats with myocardial ischemia reperfusion: 电针预处理对心肌缺血再灌注大鼠线粒体能量代谢的影响

ObjectiveTo explore the effects of electroacupuncture pretreatment on mitochondrial energy metabolism in the rats with myocardial ischemia reperfusion injury (MIRI). MethodsA total of 60 SPF Wistar rats were randomly divided into a sham-operation group (sham group), a myocardial ischemia reperfusion injury group (MIRI group) and an electroacupuncture pretreatment group (EA group), 20 rats in each one. The rats in the sham group and the MIRI group were binded for 7 days, once a day, 20 min each time. On the 8th day, the sample was collected after the heart exposed for 50 min in thoractomy in the sham group and the sample was collected after ischemia for 20 min and reperfusion for 30 min in thoractomy in the MIRI group. In the EA group, the pretreatment intervention with electroacupuncture was applied at "Nèiguān (内关PC6)", "Guānyuán (关元CV4)" and "Zúsānlĭ (足三里ST36)" in the rats for 7 days, once a day, 20 min each time. On the 8th day, after ischemia for 20 min and reperfusion for 30 min in thoractomy, the sample was collected in the EA group. The changes in STⅡ segment of electroacardiogram (ECG) were observed and measured. Using enzyme-linked immunosorbent assay (ELISA), the concentrations of cardiac troponin T (cTnT) and cardiac troponin I (cTnI)were detected. Using nitro blue tetrazolium chloride monohydrate (NBT) staining, the myocardial infarction weight percentage was measured. Using ELISA, the concentrations of mitochondrial adenosine monophosphate (AMP), adenosine diphosphate (ADP)and adenosine triphosphate (ATP)were detected. Results(1) STⅡ changes: in 20 min of ligation, compared with the sham group, the STⅡ segment of electrocardiograph (ECG) was elevated significantly in the MIRI group and EA group (both P < 0.01), but the elevation range in the EA group was lower than that of the MIRI group (P < 0.01). After reperfusion for 30 min, the STⅡ segment was fallen by over 50% in the MIRI group and the EA group. Simultaneously, the STⅡ segment in the EA group was lower than that of the MIRI group (P < 0.01). (2) Regarding myocardial infarction weight percentage, compared with the sham group, the infarction weight was larger in the MIRI group and the EA group (both P < 0.05) and the infarction weight in the EA group was lower than that of the MIRI group (P < 0.05). (3) Regarding the levels of serum cTnt and cTnI, compared with the sham group, the levels of serum cTnT and cTnI were higher in the MIRI group and the EA group (all P < 0.01) and the levels of cTnT and cTnI in the EA group were lower than that of the MIRI group (both P < 0.01). (4) Regarding the concentrations of AMP, ADP and ATP, compared with the sham group, ATP concentration was lower in the MIRI group and the EA group (both P < 0.01) and the concentrations of AMP and ADP were higher (P < 0.05, P < 0.01). Compared with the MIRI group, ATP concentration was higher in the EA group (P < 0.05) and the concentrations of AMP and ADP were lower (both P < 0.01). ConclusionsElectroacupuncture pretreatment reduces the elevation of ECG STⅡ segment, decreases the concentrations of myocardial injury markers, cTnT and cTnI and regulates the transfer among AMP, ATP and ADP. The protective effect of electroacupuncture pretreatment may result from the regulation of mitochondrial energy metabolism.

A study on protective effect of morphine against myocardial ischemia-reperfusion injury in rats via CAMP/PKA signaling pathway.

The purpose of this study was to explore the mitigating effect of morphine on the myocardial ischemia-reperfusion injury (MIRI) in rats through the cyclic adenosine monophosphate/protein kinase A (cAMP/PKA) pathway. A total of 30 male Wistar rats were assigned into sham group, MIRI group and morphine group using a random number table. The model of MIRI was routinely established. Then, the pathological changes in the morphology of myocardial tissues were observed via hematoxylin-eosin (HE) staining. The levels of the oxidative stress indicators superoxide dismutase (SOD) and malondialdehyde (MDA), the content of the inflammatory cytokine tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β) and IL-6 and the quantity of glutathione peroxidase (GSH-Px), lactate dehydrogenase (LDH), creatine kinase (CK), CK-MB and cardiac troponin I (cTnI) in the myocardial enzyme spectrum were determined and analyzed through enzyme-linked immunosorbent assay (ELISA). Moreover, the messenger ribonucleic acid (mRNA) and protein expressions of cAMP, PKA, cAMP-response element binding protein (CREB) and phosphorylated CREB (p-CREB) in the cAMP/PKA signaling pathway in the myocardial tissues were measured using real-time polymerase chain reaction (PCR) and Western blotting, respectively. The results manifested that compared with those in MIRI group, the levels of myocardial infarct size, LDH, CK, CK-MB, cTnI, MDA, TNF-α, IL-1β, IL-6 and p-CREB were decreased, while the levels of GSH-Px, SOD, PKA and CREB were increased in the morphine group. In conclusion, morphine may mitigate MIRI in rats through the cAMP/PKA signaling pathway.

Inhibition of miR-217 Protects Against Myocardial Ischemia–Reperfusion Injury Through Inactivating NF-κB and MAPK Pathways

Recent studies have demonstrated that miRNAs play a vital role in regulating myocardial ischemia/reperfusion injury (MIRI). MiR-217 has been proven to be implicated in cardiac diseases such as chronic heart failure and cardiac myxoma. However, the role of miR-217 in MIRI is not clear. A mouse MIRI model was established and the myocardial infarct size was evaluated by TTC staining. The expression level of miR-217 in I/R group was determined by real-time polymerase chain reaction. Subsequently, MIRI mice and H9C2 cells were administrated with miR-217 inhibitor in vivo and in vitro, respectively. The levels of TNF-α and IL-6 were measured by commercially available ELISA kits. Blood and cell samples were collected for the measurement of lactate dehydrogenase (LDH) level and caspase-3 activity. Cell viability was assessed with the CCK-8 assay. We then explored the detailed molecular mechanisms by TargetScan 7.1 database and further studies were performed to prove the prediction by dual-luciferase reporter assay. Larger stainless infarct areas were observed in the MIRI group, accompanied by inceased serum LDH activity, indicating the mouse MIRI model was successfully established. MiR-217 was up-regulated in MIRI mice and hypoxia/reoxygenation-treated H9C2 cells. MiR-217 knockdown alleviated the MIRI in MIRI mouse model, and also attenuated the myocardial hypoxia/reoxygenation injury in H9C2 cells. Moreover, dual specificity protein phosphatase 14 (DUSP14) was proved to be a target of miR-217. Besides, further study indicated that inhibition of miR-217 protected against MIRI through inactivating NF-κB and MAPK pathways via targeting DUSP14. MiR-217 inhibition protected against MIRI through inactivating NF-κB and MAPK pathways by targeting DUSP14. This study may provide valuable diagnostic and factors and therapeutic agents for MIRI.

Glutamine protects myocardial ischemia-reperfusion injury in rats through the PI3K/Akt signaling pathway.

To study the protective mechanism of glutamine (Gln) on myocardial ischemia-reperfusion (IR) injury in rats through the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway. A total of 30 healthy SD rats weighing 200-300 g were used in this experiment. They were randomly divided into 3 groups: sham group (n=10), myocardial IR injury group (IR group, n=10), IR+Gln group (n=10). The protein expression levels of phosphorylated Akt (p-Akt), total Akt (t-Akt), phosphorylated mammalian target of rapamycin (p-mTOR), mTOR, proliferating cell nuclear antigen (PCNA), P21, and Tubulin were determined by Western blotting (WB). Quantitative Polymerase Chain Reaction (qPCR) was applied to detect the messenger ribonucleic acid (mRNA) levels of Akt and mTOR. 3-(4,5)-dimethylthiazol(-z-y1)-3,5-diphenyl tetrazolium bromide (MTT) test was utilized to examine the proliferation ability of cardiomyocytes in vitro. Besides, the contents of the inflammatory cytokines, tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6) were measured via enzyme-linked immunosorbent assay (ELISA). Cell apoptosis in each group was examined through Hoechst staining. Compared with those in the sham group, ratios of p-AKT/AKT, p-mTOR/mTOR, and the level of PCNA extremely significantly decreased, but the level of P21 notably increased in IR group (p<0.01). In comparison with those in the IR group, ratios of p-AKT/AKT, p-mTOR/mTOR, and the level of PCNA were remarkably raised, while the level of P21 was remarkably reduced in IR+Gln group (p<0.05). QRT-PCR results manifested that there were no significant differences in the mRNA levels of Akt and mTOR among the three groups [no significant difference (NS)]. Moreover, the cell proliferation ability in IR group was remarkably lower than that in the sham group (p<0.01), while it was enhanced in the IR+Gln group compared with that in the IR group (p<0.05). Additionally, IR group had significantly elevated expression levels of TNF-α and IL-6 compared with the sham group (p<0.01), whereas the IR+Gln group had notably decreased expression levels of TNF-α and IL-6 compared with IR group (p<0.05). In comparison with that in the sham group, the apoptosis in IR group was significantly raised (p<0.01), and compared with that in the IR group, the apoptosis in the IR+Gln group prominently decreased (p<0.05). The contents of the inflammatory cytokines, TNF-α, and IL-6 presented the same trends among the three groups. Gln activates the PI3K/Akt signaling pathway by increasing the levels of p-AKT and p-mTOR. Gln can increase the PCNA level and reduce the P21 level, so as to enhance the proliferation ability of cardiomyocytes. Besides, Gln reduces the levels of inflammatory cytokines, TNF-α, and IL-6, and inhibits cell apoptosis. Finally, Gln can protect cells from myocardial IR injury by activating the PI3K/Akt signaling pathway.

Effect of ulinastatin on myocardial ischemia-reperfusion injury through JNK and P38 MAPK signaling pathways.

The aim of this study was to investigate the effect of ulinastatin (UTI) on myocardial ischemia-reperfusion injury (MIRI) through the c-Jun N-terminal kinase (JNK) signaling pathway and p38 mitogen-activated protein kinase (MAPK) signaling pathway. Healthy adult male Sprague-Dawley (SD) rats were randomly divided into 5 groups, including the sham group (n=10), MIRI group (model group, n=10), UTI group (n=10), UTI + JNK inhibitor SP600125 (UTI+SP600125 group, n=10) and UTI + p38 MAPK inhibitor SB203580 (UTI+SB203580 group, n=10). Hemodynamics, myocardial infarction and the messenger ribonucleic acid (mRNA) expressions of p38 MAPK, JNK, superoxide dismutase (SOD), malondialdehyde (MDA), nitric oxide (NO), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and high-sensitivity C-reactive protein (hs-CRP) were compared among groups. The protein levels of p38 MAPK and JNK in serum were detected via Western blotting. Furthermore, the correlations of serum p38 MAPK and JNK with TNF-α were analyzed. In the UTI group, the left ventricular systolic pressure (LVSP), the left ventricular end-diastolic pressure (LVEDP) and maximal rate of increase of the left ventricular pressure (+dp/dtmax) were significantly higher than those of the model group. However, the maximal rate of the decrease of the left ventricular pressure (-dp/dtmax), infarction area and ischemia area were significantly smaller than those of the model group. LVSP, LVEDP and +dp/dtmax in UTI+SP600125 group and UTI+SB203580 group were markedly higher than those of the UTI group. Meanwhile, the mRNA expressions of p38 MAPK and JNK in the UTI group were significantly lower than those of the model group. However, the mRNA expression levels of p38 MAPK and JNK in UTI+SP600125 group and UTI+SB203580 group were remarkably higher than the UTI group. Besides, the UTI group showed a markedly higher level of SOD and significantly lower levels of MDA, NO, TNF-α, IL-6 and hs-CRP than the model group. Furthermore, UTI+SP600125 group and UTI+SB203580 group showed significantly higher levels of MDA, NO, TNF-α, IL-6 and hs-CRP than the UTI group. The protein levels p38 MAPK and JNK were remarkably lower in the UTI group than those of the model group. However, they were remarkably higher in UTI+SP600125 group and UTI+SB203580 group than the UTI group. In addition, both p38 MAPK and JNK proteins were positively correlated with TNF-α (r=0.983 and 0.892, p=0.043 and p=0.033). UTI may inhibit MIRI caused by p38 MAPK signaling pathway and JNK signaling pathway by down-regulating TNF-α expression, thereby protecting and improving MIR.

Effects of propofol on myocardial ischemia reperfusion injury through inhibiting the JAK/STAT pathway.

The aim of this study was to investigate the effect of propofol (PPF) on myocardial ischemia-reperfusion injury (MIRI) by inhibiting the Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway, and to explore the possible underlying mechanism. A total of 60 Sprague-Dawley (SD) rats were randomly divided into 5 groups, including the Sham group (n=12), the MIRI model group (n=12), the PPF pretreatment group (n=12), the RG81640-CH (RG) pretreatment group (n=12) and the PPF+RG pretreatment group (n=12). The hemodynamic parameters of rats in each group were measured. Serum samples were collected from rats in each group. Meanwhile, the levels of lactate dehydrogenase (LDH), creatine kinase-muscle/brain (CK-MB), nicotinamide adenine dinucleotide+ (NAD+) and inflammatory factors, including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and monocyte chemotactic protein (MCP), were detected by enzyme-linked immunosorbent assay (ELISA). Myocardial infarction area of rats in each group was detected via 2,3,5-triphenyl tetrazolium chloride (TTC) staining. Moreover, the JAK/STAT pathway, as well as apoptosis indexes in myocardial cells of rats, were detected via Western blotting. Compared with the Sham group, the contents of LDH, CK-MB, NAD+ and inflammatory factors, as well as the area of myocardial infarction were significantly increased in the MIRI group (p<0.05). In terms of hemodynamic parameters, the left ventricular end-diastolic pressure (LVEDP) was significantly increased in the MIRI group. However, heart rate (HR), left ventricular developed pressure (LVDP) and maximal rate of the increase/decrease of left ventricular pressure (±dp/dtmax) were significantly decreased in the MIRI group when compared with those of the Sham group (p<0.05). Compared with the MIRI group, the contents of LDH, CK-MB, NAD+ and inflammatory factors, as well as the area of myocardial infarction and LVEDP were significantly declined in the PPF group. Meanwhile, HR, LVDP and ±dp/dtmax were remarkably increased (p<0.05). No significant differences in each index were found between the PPF + RG group and the MIRI group (p>0.05). Western blotting revealed that the protein level of B-cell lymphoma-2 (Bcl-2) was remarkably increased, while the activity of Caspase-3 was decreased in the PPF group when compared with the MIRI group (p<0.05). In addition, the protein expression levels of JAK1, STAT1 and STAT3 in the PPF group were significantly decreased than those of the MIRI group (p<0.05). However, completely opposite trends were found in the RG group. PPF reduces the release of inflammatory factors and alleviates tissue damage caused by myocardial apoptosis in MIRI rats by inhibiting the activation of the JAK/STAT pathway. Our findings indicate that PPF has a certain myocardial protective effect on MIRI.

Tongxinluo Ameliorates Myocardial Ischemia-Reperfusion Injury Mainly via Activating Parkin-Mediated Mitophagy and Downregulating Ubiquitin-Proteasome System.

To investigate the protective effects and mechanism of Chinese herbal compound Tongxinluo Capsule (, TXL) on the Parkin-mediated mitophagy and the ubiquitin-proteasome system in a rat model of myocardial ischemia-reperfusion injury (MIRI). Seventy adult male Sprague-Dawley rats were randomly divided into 7 groups: sham group, MIRI group, low- and high-dose TXL (0.5 and 1 g·kg-1·d-1, respectively) groups, atorvastatin (ATV) group (7.2 g·kg-1·d-1), chloroquine (CQ) group (10 g·kg-1·d-1), and highdose TXL + CQ group. After pharmacological administration for 7 days, rats underwent left anterior descending artery ligation surgery to establish the MIRI models with 50 min ischemia followed by 4 h reperfusion. Blood was taken for cardiac troponin I (cTnI) detection and hearts were harvested for infarct staining and apoptosis detection. The autophagy or mitophagy proteins and ubiquitinated proteins were detected by Western blotting. Compared with the sham group, the MIRI group exhibited a larger infarcted area (27.13%±0.01%, P<0.01), a higher apoptotic index (34.33%±2.03% vs.1.81%±0.03%, P<0.01), and higher cTnI expression (14.18±1.01 vs. 7.96±0.32, P<0.01). The mitochondrial integrity was damaged in the MIRI group, while TXL and ATV alleviated the damage of MIRI. More autophagosomes were observed in the high-dose TXL group than in the MIRI group (7.00±0.58 vs. 4.33±1.15, P<0.05). More amounts of PTEN-induced putative kinase protein 1 (PINK1) and Parkin translocated onto the mitochondria were detected in the high-dose TXL group than in the MIRI group (P<0.05). The ubiquitin response was signifificantly downregulated in the high-dose TXL group relative to the MIRI group (P<0.05). CQ administration abolished the activation of autophagy flux and the PINK1/ Parkin pathway induced by high-dose of TXL. TXL ameliorates MIRI via activating Parkin-mediated mitophagy in rats. The downregulation of the ubiquitin-proteasome system is also involved.