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This study aims to investigate the effect of salvianolic acid B (Sal B), the active ingredient of Salvia miltiorrhiza, on H9C2 cardiomyocytes injured by oxygen and glucose deprivation/reperfusion (OGD/R) through regulating mitochondrial fission and fusion. The process of myocardial ischemia-reperfusion injury was simulated by establishing OGD/R model. The cell proliferation and cytotoxicity detection kit (cell counting kit-8, CCK-8) was used to detect cell viability; the kit method was used to detect intracellular reactive oxygen species (ROS), total glutathione (t-GSH), nitric oxide (NO) content, protein expression levels of mitochondrial fission and fusion, apoptosis-related detection by Western blot. Mitochondrial permeability transition pore (MPTP) detection kit and Hoechst 33342 fluorescence was used to observe the opening level of MPTP, and molecular docking technology was used to determine the molecular target of Sal B. The results showed that relative to control group, OGD/R injury reduced cell viability, increased the content of ROS, decreased the content of t-GSH and NO. Furthermore, OGD/R injury increased the protein expression levels of dynamin-related protein 1 (Drp1), mitofusions 2 (Mfn2), Bcl-2 associated X protein (Bax) and cysteinyl aspartate specific proteinase 3 (caspase 3), and decreased the protein expression levels of Mfn1, increased MPTP opening level. Compared with the OGD/R group, it was observed that Sal B had a protective effect at concentrations ranging from 6.25 to 100 μmol·L-1. Sal B decreased the content of ROS, increased the content of t-GSH and NO, and Western blot showed that Sal B decreased the protein expression levels of Drp1, Mfn2, Bax and caspase 3, increased the protein expression level of Mfn1, and decreased the opening level of MPTP. In summary, Sal B may inhibit the opening of MPTP, reduce cell apoptosis and reduce OGD/R damage in H9C2 cells by regulating the balance of oxidation and anti-oxidation, mitochondrial fission and fusion, thereby providing a scientific basis for the use of Sal B in the treatment of myocardial ischemia reperfusion injury.
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This study explored the effects of Buyang Huanwu Decoction(BYHWD) on platelet activation and differential gene expression after acute myocardial infarction(AMI). SD rats were randomly divided into a sham-operated group, a model group, a positive drug(aspirin) group, and a BYHWD group. Pre-treatment was conducted for 14 days with a daily oral dose of 1.6 g·kg~(-1) BYHWD and 0.1 g·kg~(-1) aspirin. The AMI model was established using the high ligation of the left anterior descending coronary artery method. The detection indicators included myocardial infarct size, heart function, myocardial tissue pathology, peripheral blood flow perfusion, platelet aggregation rate, platelet membrane glycoprotein CD62p expression, platelet transcriptomics, and differential gene expression. The results showed that compared with the sham-operated group, the model group showed reduced ejection fraction and cardiac output, decreased peripheral blood flow, and increased platelet aggregation rate and CD62p expression, and activated platelets. At the same time, TXB_2 content increased and 6-keto-PGF1α content decreased in serum. Compared with the model group, BYHWD increased ejection fraction and cardiac output, improved blood circulation in the foot and tail regions and cardiomyocytes arrangement, reduced myocardial infarct size and inflammatory infiltration, down-regulated platelet aggregation rate and CD62p expression, reduced serum TXB_2 content, and increased 6-keto-PGF1α content. Platelet transcriptome sequencing results revealed that BYHWD regulated mTOR-autophagy pathway-related genes in platelets. The differential gene expression levels were detected using real-time quantitative PCR. BYHWD up-regulated mTOR, down-regulated autophagy-related FUNDC1 and PINK genes, and up-regulated p62 gene expression. The results demonstrated that BYHWD could regulate platelet activation, improve blood circulation, and protect ischemic myocardium in AMI rats, and its mechanism is related to the regulation of the mTOR-autophagy pathway in platelets.
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Ratas , Animales , Ratas Sprague-Dawley , Medicamentos Herbarios Chinos/uso terapéutico , Infarto del Miocardio/genética , Miocardio/metabolismo , Aspirina/uso terapéutico , Serina-Treonina Quinasas TOR/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas MitocondrialesRESUMEN
Objective:To investigate the effect of Shuangshen Ningxin capsules (SSNX) on cardiac hemodynamics and cardiac function in rats with coronary microvascular dysfunction. Method:Rats were randomly divided into a sham operation group, a model group, a nicorandil group (5 mg·kg<sup>-1</sup>), and high- (180 mg·kg<sup>-1</sup>), medium- (90 mg·kg<sup>-1</sup>), and low-dose (45 mg·kg<sup>-1</sup>) SSNX groups. Rats received corresponding drugs for 7 days. Two hours after the last administration, the model of coronary microvascular dysfunction was induced by left ventricular injection of embolic microspheres (40-120 μm, about 1 000 microspheres). Twenty-four hours after modeling, left ventricular internal dimension in diastole (LVIDd), left ventricular internal dimension in systole (LVIDs) left ventricular end-diastolic volume (LVEDV), left ventricular end-systolic volume (LVESV), stroke volume (SV), cardiac output (CO), left ventricular ejection fraction (EF), and left ventricular shortening rate (FS) were detected by echocardiography. Cardiac catheterization was used to observe the arterial systolic blood pressure (SBP), diastolic blood pressure (DBP), left ventricular systolic pressure (LVSP), left ventricular end-diastolic pressure (LVEDP), maximum rate of increase in left ventricular pressure (LV+dp/dt<sub>max</sub>), and maximum rate of decrease in left ventricular pressure (LV-dp/dt<sub>max</sub>), and the mean arterial pressure (MAP) was calculated. Heart rate (HR) was calculated according to Ⅱ lead ECG. Biochemical analysis was carried out to detect the activities of creatine kinase (CK), creatine kinase-MB (CK-MB), and lactate dehydrogenase (LDH). Enzyme-linked immunosorbent assay (ELISA) was used to detect serum cardiac troponin T (cTnT). Western blot was used to detect the protein expression of Caspase-3, Bcl-2, and Bax, and 2,3,5-triphenyltetrazolium chloride (TTC) staining to observe the area of myocardial infarction. Hematoxylin-eosin (HE) staining was used to observe the morphological changes of the myocardium. Result:As revealed by echocardiography, compared with the sham operation group, the model group showed reduced SV, CO, EF, and FS (<italic>P</italic><0.01), and increased LVIDs and LVEDV (<italic>P</italic><0.01). Compared with the model group, the SSNX groups showed increased EF (<italic>P</italic><0.05, <italic>P</italic><0.01) and FS (<italic>P</italic><0.01), and the high- and medium-dose SSNX groups displayed reduced LVIDs and LVESV, and increased LVEDV, SV, and CO (<italic>P</italic><0.05, <italic>P</italic><0.01). SBP, DBP, MAP, LVSP, LV+dp/dt<sub>max</sub>, and LV-dp/dt<sub>max</sub> in the model group were lower than those in the sham operation group (<italic>P</italic><0.01), while there was no significant difference in HR. SSNX improved hemodynamics of rats, and increased SBP, DBP, MAP, LVSP, LV+dp/dt<sub>max</sub>, LV-dp/dt<sub>max</sub>, and HR as compared with the model group (<italic>P</italic><0.05, <italic>P</italic><0.01). The serum CK, LDH, CK-MB, and cTnT levels in the model group were higher than those in the sham operation group (<italic>P</italic><0.01). Compared with the model group, SSNX groups reduced serum CK, LDH, CK-MB, and cTnT (<italic>P</italic><0.05,<italic> P</italic><0.01). Compared with the sham operation group, the model group displayed increased expression of Caspase-3 protein in the myocardium (<italic>P</italic><0.01) and reduced expression of Bcl-2 protein (<italic>P</italic><0.05). The expression of Caspase-3 protein in the myocardium of SSNX groups was lower than that in the model group, and statistical difference was observed between the low-dose SSNX group and the model group (<italic>P</italic><0.05). Compared with the model group, the SSNX groups exhibited increased expression of Bcl-2 in the rat myocardium, and the statistical difference was observed in the high-dose SSNX group <italic>(P</italic><0.01). As demonstrated by the TTC staining, compared with the model group, SSNX groups showed reduced areas of myocardial infarction (<italic>P</italic><0.01). The HE staining indicated that the pathological injury in myocardial tissues of the SSNX groups was relieved as compared with that in the model group. Conclusion:SSNX can significantly enhance the cardiac function after coronary microvascular dysfunction caused by embolic microspheres, improve cardiac hemodynamics, reduce the area of myocardial infarction, and decrease CK, LDH, CK-MB, and cTnT levels. The mechanism may be related to the inhibition of cardiomyocyte apoptosis to protect the myocardium.
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The present study evaluated the curative efficacy of Chinese herbal injection on unstable angina pectoris( UAP) by network Meta-analysis. The databases,including Pub Med,Cochrane Library,Web of Science,CNKI,CBM,VIP and Wanfang were searched for randomized controlled trial( RCT) of Chinese herbal injection in the treatment of UAP. All researchers independently screened the articles,extracted the data and evaluated the quality. Open BUGS and Stata were employed for the analysis of the trials that met the quality standards. Fifty-eight studies were finally included in this study,involving 20 intervention measures. In terms of the effective rate,16 injections such as Dengzhan Xixin Injection,Xuesaitong Injection and Danshen Injection combined with western medicine exhibited significant efficacy. In terms of ECG,Puerarin Injection,Ginkgo Leaf Extract and Dipyridamole Injection( GDI),Breviscapine Injection combined with western medicine were superior to western medicine. In terms of the reduction of the angina attack times,Sodium Tanshinone ⅡASulfonate Injection,GDI and Dazhu Hongjingtian Injection combined with western medicine showed better effects than western medicine. In terms of shortening the angina duration,Shenmai Injection combined with western medicine was superior to western medicine. As revealed by the results,Dengzhan Xixin Injection,Xuesaitong Injection,Danshen Injection,Breviscapine Injection,Danshen Ligustrazine Injection combined with western medicine displayed prominent curative efficacy,which were recommended for clinical application. Meanwhile,appropriate intervention measures should be selected according to individual conditions. Limited by the quality of the included trials,the conclusions still need to be further verified.
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Humanos , Angina de Pecho , Angina Inestable/tratamiento farmacológico , China , Medicamentos Herbarios Chinos , Metaanálisis en Red , Resultado del TratamientoRESUMEN
OBJECTIVE@#Using network pharmacology to explore the mechanism of the 'invigorating qi and promoting blood circulation' drug pair Ginseng-Danshen (Salvia miltiorrhiza) on treatment of ischemic heart disease (IHD).@*METHODS@#The chemical constituents of ginseng and Danshen drug pair were identified by searching the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), and the potential targets of the pair were identified. The pharmacodynamics of the pair was analyzed using network pharmacology. The targets of IHD were identified by database screening. Using protein-protein interaction network, the interaction targets of Ginseng-Danshen on IHD were constructed. A "constituent-target-disease" interaction network was constructed using Cytoscape software, Gene Ontology (GO) term enrichment analysis and biological pathway enrichment analysis were carried out, and the mechanism of improving myocardial ischemia by the Ginseng-Danshen drug pair was investigated.@*RESULTS@#Seventeen active constituents and 53 targets were identified from ginseng, 53 active constituents and 61 targets were identified from Danshen, and 32 protein targets were shared by ginseng and Danshen. Twenty GO terms were analyzed, including cytokine receptor binding, cytokine activity, heme binding, and antioxidant activity. Sixty Kyoto Encyclopedia of Genes and Genomes (KEGG) signaling pathways were analyzed, including phosphatidylinositol 3-kinase-serine-threonine kinase (PI3K-AKT) signaling pathway, p53 signaling pathway, interleukin 17 signaling pathway, tumor necrosis factor signaling pathway, and the advanced glycation end product (AGE)-the receptor for AGE (RAGE) signaling pathway in diabetic complications.@*CONCLUSION@#The specific mechanism of Ginseng-Danshen drug pair in treating IHD may be associated with improving the changes of metabolites inbody, inhibiting the production of peroxides, removing the endogenous oxygen free radicals, regulating the expression of inflammatory factors, reducing myocardial cell apoptosis and promoting vascular regeneration.
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Objective:To observe the effect of Shuangshen Ningxin capsule in alleviating myocardial ischemia/reperfusion injury in rats by regulating mitochondrial adenosine triphosphate(ATP)-sensitive potassium channels.Method:A total of 56 adult male Sprague-Dawley rats were randomly divided into sham-operated control group (sham), model group (model), Shuangshen Ningxin group (SSNX, 90 mg·kg-1).Shuangshen Ningxin and mitochondrial ATP-sensitive potassium channel (MitoKATP) channel inhibitor group 5-hydroxyl-acid group (SSNX+5-HD, 5 mg·kg-1), with 14 rats in each group. Except the sham operation group, the other three groups received occlusion of left anterior descending coronary artery (LAD) for 45 min, and were sacrificed 3 h after reperfusion. Myocardial ischemia and infarct size were observed by TSC Evans blue staining, and myocardial tissue damage degree was observed by hematoxylin-eosin(HE) staining. The kit was used to measure serum lactate dehydrogenase (LDH), creatine kinase (CK) and creatine kinase isoenzyme (CK-MB). The ultrastructural changes of mitochondria and mitochondrial autophagy were observed under transmission electron microscope. The changes of mitochondrial membrane potential in cardiomyocytes were detected by fluorescent probe.Result:Compared with the sham group, myocardial infarct size and myocardial ischemic area percentage in the model group were significantly increased, myocardial tissue arrangement was disordered and loose, individual myocardial fibers were broken, cardiomyocytes were necrotic, and serum CK, CK-MB, LDH activities were significantly increased (P<0.01). Mitochondrial membrane potential was significantly decreased (P<0.01), and mitochondrial structure was destroyed by transmission electron microscopy. Compared with the model group, the myocardial tissue of the SSNX group was arranged orderly, and a small amount of cell edema was mildly degenerated. The percentage of myocardial infarct size and myocardial ischemic area was significantly decreased, serum CK, CK-MB, and LDH activities were significantly decreased (P<0.01), while mitochondrial membrane potential increased (P<0.01). Compared with the model group, the SSNX+5-HD group had mild myocardial tissue disorder and mild degeneration of cell edema in some areas, the percentage of myocardial infarct size and myocardial ischemic area was significantly reduced, serum CK, CK-MB, and LDH activities were significantly decreased (P<0.01), and mitochondrial membrane potential increased (P<0.01). Compared with SSNX group, SSNX+5-HD group had significant increase in serum CK, CK-MB and LDH activities (P<0.01), significant increase in the percentage of myocardial infarct size and myocardial ischemic area, and mitochondrial membrane potential Reduced (P<0.05).Conclusion:SSNX protects rat myocardial ischemia-reperfusion injury by opening mitochondrial ATP-sensitive potassium channel.
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The aim of this paper was to investigate whether the mechanism of salvianolic acid B in protecting H9 c2 cardiomyocytes from hypoxia/reoxygenation injury is related to the regulation of mitochondrial autophagy mediated by NIX. H9 c2 cardiomyocytes were cultured in vitro and divided into normal group, model group and salvianolic acid B group(50 μmol·L~(-1)). Hypoxia/reoxygenation injury model was established by hypoxia for 4 h and reoxygenation for 2 h. In normal group, high glucose DMEM medium was used for culture. Those in model group were cultured with DMEM medium without glucose and oxygen, and no drugs for hypoxia and reoxyge-nation. In salvianolic acid B group, salvianolic acid B prepared by glucose-free DMEM medium was added during hypoxia, and the other process was as same as the model group. The cell viability was evaluated by CCK-8 assay. The leakage of lactate dehydrogenase(LDH) was detected by microplate method. The levels of intracellular reactive oxygen species(ROS) and mitochondrial membrane potential(ΔΨm) were measured by chemical fluorescence method. The level of intracellular adenosine triphosphate(ATP) was mea-sured by fluorescein enzyme method. The autophagy related proteins LC3-Ⅰ, LC3-Ⅱ, apoptosis related protein cleaved caspase-3 and mitochondrial autophagy receptor protein NIX were detected by Western blot. As compared with the normal group, the activity of H9 c2 cardiomyocytes and ATP level were decreased(P<0.05); LDH leakage and ROS production were increased(P<0.01); ΔΨm was decreased(P<0.01); LC3-Ⅱ/LC3-Ⅰ ratio, cleaved caspase-3 and NIX protein expression levels were increased(all P<0.05) in the model group. As compared with the model group, the activity of cells and ΔΨm were significantly increased(P<0.01); ATP level was increased(P<0.05); LDH leakage and ROS generation were decreased(P<0.01); LC3-Ⅱ/LC3-Ⅰ ratio was decreased(P<0.01); cleaved caspase-3 and NIX expression levels were decreased(P<0.05) in the salvianolic acid B group. The protective effect of salvianolic acid B on hypoxia/reoxygenation injury of H9 c2 cardiomyocytes may be associated with inhibiting mitochondrial auto-phagy. The specific mechanism may be related to inhibiting the activation of mitochondrial autophagy mediated by NIX, increasing ΔΨm, reducing ROS production, reducing the expression of cleaved caspase-3, LC3-Ⅱ, and increasing cell viability.
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Humanos , Apoptosis , Autofagia , Benzofuranos , Hipoxia de la Célula , Supervivencia Celular , Hipoxia , Miocitos CardíacosRESUMEN
Ischemic heart disease(IHD) is a common and frequently-occurring disease that causes serious harm to human health. Autophagy is a life process that maintains cell homeostasis by degrading macromolecules such as damaged organelles in cells. In the process of ischemic heart disease development, on the one hand, cardiomyocytes degrade macromolecules such as damaged organelles by autophagy to provide material basis for energy synthesis and maintain cell homeostasis; on the other hand, over-activated autophagy can also increase cardiomyocyte death. Ischemic heart disease has a complex pathological mechanism, and the occurrence of autophagy is closely related to the survival or death of myocardial cells, so the regulation of autophagy may be an important therapeutic target for ischemic heart disease. Traditional Chinese medicine(TCM) with obvious effects, unique advantages and great potential has been widely used in the treatment of ischemic heart disease. In recent years, more and more studies have found that TCM can protect myocardium by regulating autophagy of cardiomyocytes. In this review, we summarized recent studies on the regulation of autophagy in myocardial cells by traditional Chinese medicine in ischemic heart disease. The pharmacological mechanism of Chinese medicinein regulating autophagy to protect cardiomyocytes was reviewed through different ways(promoting or inhibiting autophagy) from three levels, i.e. active ingredient, as well as drug pair and compound. The specific mechanism of Chinese medicine in regulating autophagy to protect ischemic heart disease was explored to provide references or new ideas for clinical treatment and drug development of ischemic heart disease.
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Humanos , Autofagia , Medicina Tradicional China , Isquemia Miocárdica , Miocardio , Miocitos CardíacosRESUMEN
Ischemic heart disease is one of the most deadly diseases in the world, and new therapies and preventive measures are urgently needed. In general, cardiomyocytes rely on adenosine triphosphate(ATP) produced by mitochondrial oxidative phosphorylation to maintain their systolic and ion pump functions. Autophagy is a procedural degradation mechanism widely present in eukaryotic cells. It is a self-defense mechanism and self-repair process of the body tissues. It is also a way of apoptosis and a basic phenomenon to maintain the energy balance of human cells. Mitochondrial autophagy is a type of selective autophagy in cells. In fact, damaged mitochondria selectively remove damaged proteins and organelles with autophagy to maintain intracellular homeostasis. Mitochondrial autophagy is important for maintaining the homeostasis of cardiomyocytes. With the deepening of modern biological research, more and more traditional Chinese medicines(TCM) or their extracts have been proven to alleviate myocardial cell damage after ischemia/reperfusion through autophagy or regulation of mitochondrial function. This further inspires TCM workers to find effective treatment measures by targeting mitochondria. Under the above background, this paper reviews the effects of mitochondrial autophagy on ischemic heart disease and the intervention studies of TCM in this field.
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This study was to investigate the mechanism of safflower yellow injection for regulating inflammatory response against myocardial ischemia-reperfusion injury( MIRI) in rats. Male Wistar rats were randomly divided into sham operation group,model group,Hebeishuang group,safflower yellow injection high,medium and low dose groups. MIRI model was established by ligating left anterior descending coronary artery. Myocardial histopathological changes were observed by HE staining; myocardial infarct size was detected by TTC staining; content and changes of tumor necrosis factor-α( TNF-α) and interleukin-6( IL-6),serum creatine kinase( CK),aspartate aminotransferase( AST),and lactate dehydrogenase( LDH) were detected by biochemical method or enzyme-linked immunosorbent assay( ELISA). Western blot assay was used to detect the protein expression of Toll-like receptor 4( TLR4) and nuclear factor-κB( NF-κB p65) in myocardial tissues. The results showed that as compared with the sham operation group,the myocardial arrangement of the model group was disordered,with severe edemain the interstitial,significantly increased area of myocardial infarction,increased activities of AST,CK and LDH in serum,and significantly increased contents of TNF-α and IL-6; the expression levels of TLR4 and NF-κB( p65) protein in myocardial tissues were also increased. As compared with the model group,the myocardial tissues were arranged neatlyin the Hebeishuang group and safflower yellow injection high,medium and low dose groups; the edema was significantly reduced; the myocardial infarct size was significantly reduced; the serum AST,CK,LDH activity and TNF-α,IL-6 levels were significantly decreased,and the expression levels of TLR4 and NF-κB( p65) protein in myocardial tissues were decreased. As compared with the Hebeishuang group,the myocardial infarct size was larger in the safflower yellow injection high,medium and low dose groups; the activities of AST,CK and LDH in serum and the contents of TNF-α and IL-6 in serum were higher,but there was no statistically significant difference in the expression levels of TLR4 and NF-κB( p65) protein in tissues. It is suggested that safflower yellow injection has a significant anti-MIRI effect,and its mechanism may be related to the regulation of TLR-NF-κB pathway to inhibit inflammatory response.