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Ongoing inflammation in the heart is positively correlated with adverse remodeling, characterized by elevated levels of cytokines that stimulate activation of cardiac fibroblasts. It was found that CaMKIIδ response to Ang II or TAC triggers the accumulation of ROS in cardiomyocytes, which subsequently stimulates NF-κB/NLRP3 and leads to an increase in IL-6, IL-1ß, and IL-18. This is an important causative factor in the occurrence of adverse remodeling in heart failure. Sweroside is a biologically active natural iridoids extracted from Lonicerae Japonicae Flos. It shows potent anti-inflammatory and antioxidant activity in various cardiovascular diseases. In this study, we found that sweroside inhibited ROS-mediated NF-κB/NLRP3 in Ang II-treated cardiomyocytes by directly binding to CaMKIIδ. Knockdown of CaMK⠡δ abrogated the effect of sweroside regulation on NF-κB/NLRP3 in cardiomyocytes. AAV-CaMK⠡δ induced high expression of CaMK⠡δ in the myocardium of TAC/Ang II-mice, and the inhibitory effect of sweroside on TAC/Ang â ¡-induced elevation of NF-κB/NLRP3 was impeded. Sweroside showed significant inhibitory effects on CaMKIIδ/NF-κB/NLRP3 in cardiomyocytes from TAC/Ang â ¡-induced mice. This would be able to mitigate the adverse events of myocardial remodeling and contractile dysfunction at 8 weeks after the onset of the inflammatory response. Taken together, our findings have revealed the direct protein targets and molecular mechanisms by which sweroside improves heart failure, thereby supporting the further development of sweroside as a therapeutic agent for heart failure.
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Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina , Insuficiencia Cardíaca , Miocitos Cardíacos , FN-kappa B , Proteína con Dominio Pirina 3 de la Familia NLR , Especies Reactivas de Oxígeno , Animales , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/efectos de los fármacos , FN-kappa B/metabolismo , Insuficiencia Cardíaca/metabolismo , Insuficiencia Cardíaca/tratamiento farmacológico , Insuficiencia Cardíaca/etiología , Especies Reactivas de Oxígeno/metabolismo , Ratones , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Transducción de Señal/efectos de los fármacos , Masculino , Modelos Animales de EnfermedadRESUMEN
Heart failure is characterized by high incidence and mortality rates, and the search for effective treatment strategies for heart failure and the improvement of clinical outcomes have always been important research directions. Imbalanced inflammation has been proven to be one of the critical pathological factors in heart failure, positively correlated with adverse events such as impaired cardiac function and myocardial fibrosis. In recent years, studies have confirmed that the activation of the NOD-like receptor thermal protein domain-associated protein 3(NLRP3) inflammasome plays a common regulatory role in the inflammation imbalance induced by various factors in heart failure. Moreover, certain traditional Chinese medicine(TCM) and active components can significantly inhibit the activation of the NLRP3 inflammasome, thereby improving heart failure. This article first overviewed the basic information about the NLRP3 inflammasome, summarized the regulatory mechanisms of the NLRP3 inflammasome in heart failure induced by various factors, introduced recent research progress on TCM and active components that inhibited the NLRP3 inflammasome to improve heart failure, aiming to provide references for innovative drug research in the field of integrated Chinese and western medicine for the prevention and treatment of heart failure.
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Insuficiencia Cardíaca , Inflamasomas , Humanos , Inflamasomas/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Medicina Tradicional China , Insuficiencia Cardíaca/tratamiento farmacológico , InflamaciónRESUMEN
Jiming Powder is a traditional ancient prescription with good therapeutic effect in the treatment of heart failure, but its mechanism lacks further exploration. In this study, a mouse model of coronary artery ligation was used to evaluate the effect and mechanism of Jiming Powder on myocardial fibrosis in mice with myocardial infarction. The study constructed a mouse model of heart failure after myocardial infarction using the method of left anterior descending coronary artery ligation. The efficacy of Jiming Powder was evaluated from multiple angles, including ultrasound imaging, hematoxylin-eosin(HE) staining, Masson staining, Sirius Red staining, and serum myocardial enzyme spectrum detection. Western blot analysis was performed to detect key proteins involved in ventricular remodeling, including transforming growth factor-ß1(TGF-ß1), α-smooth muscle actin(α-SMA), wingless-type MMTV integration site family member 3a(Wnt3a), ß-catenin, matrix metallopeptidase 2(MMP2), matrix metallopeptidase 3(MMP3), TIMP metallopeptidase inhibitor 1(TIMP1), and TIMP metallopeptidase inhibitor 2(TIMP2). The results showed that compared with the model group, the high and low-dose Jiming Powder significantly reduced the left ventricular internal diameter in systole(LVID;s) and diastole(LVID;d), increased the left ventricular ejection fraction(LVEF) and left ventricular fractional shortening(LVFS), effectively improved cardiac function in mice after myocardial infarction, and effectively reduced the levels of myocardial injury markers such as creatine kinase(CK), creatine kinase isoenzyme(CK-MB), and lactic dehydrogenase(LDH), thus protecting ischemic myocardium. HE staining showed that Jiming Powder could attenuate myocardial inflammatory cell infiltration after myocardial infarction. Masson and Sirius Red staining demonstrated that Jiming Powder effectively inhibited myocardial fibrosis, reduced the collagen â /â ¢ ratio in myocardial tissues, and improved collagen remodeling after myocardial infarction. Western blot results showed that Jiming Powder reduced the expression of TGF-ß1, α-SMA, Wnt3a, and ß-catenin, decreased the levels of MMP2, MMP3, and TIMP2, and increased the level of TIMP1, suggesting its role in inhibiting cardiac fibroblast transformation, reducing extracellular matrix metabolism in myocardial cells, and lowering collagen â and α-SMA content, thus exerting an anti-myocardial fibrosis effect after myocardial infarction. This study revealed the role of Jiming Powder in improving ventricular remodeling and treating myocardial infarction, laying the foundation for further research on the pharmacological effect of Jiming Powder.
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Insuficiencia Cardíaca , Infarto del Miocardio , Ratones , Animales , Factor de Crecimiento Transformador beta1/metabolismo , Metaloproteinasa 2 de la Matriz/metabolismo , beta Catenina/metabolismo , Metaloproteinasa 3 de la Matriz/metabolismo , Metaloproteinasa 3 de la Matriz/uso terapéutico , Polvos , Remodelación Ventricular , Volumen Sistólico , Función Ventricular Izquierda , Infarto del Miocardio/tratamiento farmacológico , Miocardio/patología , Insuficiencia Cardíaca/metabolismo , Colágeno/metabolismo , Creatina Quinasa , FibrosisRESUMEN
In this study, untargeted metabolomics was conducted using the liquid chromatography-tandem mass spectrometry(LC-MS/MS) technique to analyze the potential biomarkers in the plasma of mice with heart failure with preserved ejection fraction(HFpEF) induced by a high-fat diet(HFD) and nitric oxide synthase inhibitor(Nω-nitro-L-arginine methyl ester hydrochloride, L-NAME) and explore the pharmacological effects and mechanism of Jiming Powder in improving HFpEF. Male C57BL/6N mice aged eight weeks were randomly assigned to a control group, a model group, an empagliflozin(10 mg·kg~(-1)·d~(-1)) group, and high-and low-dose Jiming Powder(14.3 and 7.15 g·kg~(-1)·d~(-1)) groups. Mice in the control group were fed on a low-fat diet, and mice in the model group and groups with drug intervention were fed on a high-fat diet. All mice had free access to water, with water in the model group and Jiming Powder groups being supplemented with L-NAME(0.5 g·L~(-1)). Drugs were administered on the first day of modeling, and 15 weeks later, blood pressure and cardiac function of the mice in each group were measured. Heart tissues were collected for hematoxylin-eosin(HE) staining to observe pathological changes and Masson's staining to observe myocardial collagen deposition. Untargeted metabolomics analysis was performed on the plasma collected from mice in each group, and metabolic pathway analysis was conducted using MetaboAnalyst 5.0. The results showed that the blood pressure was significantly lower and the myocardial concentric hypertrophy and left ventricular diastolic dysfunction were significantly improved in both the high-dose and low-dose Jiming Powder groups as compared with those in the model group. HE and Masson staining showed that both high-dose and low-dose Jiming Powder significantly alleviated myocardial fibrosis. In the metabolomics experiment, 23 potential biomarkers were identified and eight strongly correlated metabolic pathways were enriched, including linoleic acid metabolism, histidine metabolism, alpha-linolenic acid metabolism, glycerophospholipid metabolism, purine metabolism, porphyrin and chlorophyll metabolism, arachidonic acid metabolism, and pyrimidine metabolism. The study confirmed the pharmacological effects of Jiming Powder in lowering blood pressure and ameliorating HFpEF and revealed the mechanism of Jiming Powder using the metabolomics technique, providing experimental evidence for the clinical application of Jiming Powder in treating HFpEF and a new perspective for advancing and developing TCM therapy for HFpEF.
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Insuficiencia Cardíaca , Masculino , Ratones , Animales , Insuficiencia Cardíaca/tratamiento farmacológico , Insuficiencia Cardíaca/metabolismo , Polvos , Volumen Sistólico/fisiología , Cromatografía Liquida , NG-Nitroarginina Metil Éster/uso terapéutico , Ratones Endogámicos C57BL , Espectrometría de Masas en Tándem , Metabolómica , Biomarcadores , AguaRESUMEN
Introduction: Qishen Granule (QSG), a clinically approved traditional Chinese medicine, has been researched for treating heart failure (HF) for many years. However, the effect of QSG on intestinal microecology remains unconfirmed. Therefore, this study aimed to elucidate the possible mechanism of QSG regulating HF in rats based on intestinal microecological changes. Methods: A rat model with HF induced by myocardial infarction was prepared by left coronary artery ligation. Cardiac functions were assessed by echocardiography, pathological changes in the heart and ileum by hematoxylin-eosin (HE) and Masson staining, mitochondrial ultrastructure by transmission electron microscope, and gut microbiota by 16S rRNA sequencing. Results: QSG administration improved cardiac function, tightened cardiomyocytes alignment, decreased fibrous tissue and collagen deposition, and reduced inflammatory cell infiltration. Electron microscopic observation of mitochondria revealed that QSG could arrange mitochondria neatly, reduce swelling, and improve the structural integrity of the crest. Firmicutes were the dominant component in the model group, and QSG could significantly increase the abundance of Bacteroidetes and Prevotellaceae_NK3B31_group. Furthermore, QSG significantly reduced plasma lipopolysaccharide (LPS), improved intestinal structure, and recovered barrier protection function in rats with HF. Conclusion: These results demonstrated that QSG was able to improve cardiac function by regulating intestinal microecology in rats with HF, suggesting promising therapeutic targets for HF.
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Allosteric regulation by pathway products plays a vital role in amino acid metabolism. Homoserine dehydrogenase (HSD), the key enzyme for the biosynthesis of various aspartate family amino acids, is subject to feedback inhibition by l-threonine and l-isoleucine. The desensitized mutants with the potential for amino acid production remain limited. Herein, a semi-rational approach was proposed to relieve the feedback inhibition. HSD from Corynebacterium glutamicum (CgHSD) was first characterized as a homotetramer, and nine conservative sites at the tetramer interface were selected for saturation mutagenesis by structural simulations and sequence analysis. Then, we established a high-throughput screening (HTS) method based on resistance to l-threonine analog and successfully acquired two dominant mutants (I397V and A384D). Compared with the best-ever reported desensitized mutant G378E, both new mutants qualified the engineered strains with higher production of CgHSD-dependent amino acids. The mutant and wild-type enzymes were purified and assessed in the presence or absence of inhibitors. Both purified mutants maintained >90% activity with 10 mM l-threonine or 25 mM l-isoleucine. Moreover, they showed >50% higher specific activities than G378E without inhibitors. This work provides two competitive alternatives for constructing cell factories of CgHSD-related amino acids and derivatives. Moreover, the proposed approach can be applied to engineering other allosteric enzymes in the amino acid synthesis pathway.
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Recently, necroptosis has emerged as one of the important mechanisms of ischemia stroke. Necroptosis can be rapidly activated in endothelial cells to cause vascular damage and neuroinflammation. Panax notoginseng saponins (PNS), an ingredient extracted from the root of Panax notoginseng (Burk.) F.H. Chen, was commonly used for ischemic stroke, while its molecular mechanism and targets have not been fully clarified. Our study aimed to clarify the anti-necroptosis effect of PNS by regulating RIP1-RIP3-MLKL signaling pathway in brain microvascular endothelial cells (BMECs) subjected to transient oxygen-glucose deprivation (OGD/resupply [R]). In vitro, the necroptosis model of rat BMECs was established by testing the effect of OGD/R in the presence of the pan-caspase inhibitor z-VAD-FMK. After administration of PNS and Nec-1, cell viability, cell death modality, the expression of RIP1-RIP3-MLKL pathway and mitochondrial membrane potential (Δψm) level were investigated in BMECs upon OGD/R injury. The results showed that PNS significantly enhanced cell viability of BMECs determined by CCK-8 analysis, and protected BMECs from necroptosis by Flow cytometry and TEM. In addition, PNS inhibited the phosphorylation of RIP1, RIP3, MLKL and the downstream expression of PGAM5 and Drp1, while similar results were observed in Nec-1 intervention. We further investigated whether PNS prevented the Δψm depolarization. Our current findings showed that PNS effectively reduced the occurrence of necroptosis in BMECs exposed to OGD/R by inhibition of the RIP1-RIP3-MLK signaling pathway and mitigation of mitochondrial damage. This study provided a novel insight of PNS application in clinics.
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Panax notoginseng , Saponinas , Animales , Encéfalo/metabolismo , Caspasas/metabolismo , Caspasas/farmacología , Células Endoteliales/metabolismo , Glucosa/metabolismo , Necroptosis , Oxígeno/metabolismo , Panax notoginseng/química , Proteínas Quinasas/metabolismo , Ratas , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Saponinas/farmacología , Transducción de SeñalRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: "Qi deficiency and blood stasis" syndrome is one of the most common syndromes treated with Traditional Chinese Medicine among ischemic heart disease (IHD) patients in clinic. As a Chinese herbal formula with the function of tonifying Qi and activating blood, Yiqihuoxue Decoction (YQHX) has been frequently proven to be effective in the clinical treatment of IHD. AIM OF THE STUDY: The cardioprotective mechanisms of YQHX in treating ischemic heart disease were investigated, with emphasis on the key targets and pathways. MATERIALS AND METHODS: In the present study, the potential targets of compounds identified in YQHX were predicted using PharmMapper, Symmap, and STITCH databases, and a "herb-compound-target" network was constructed using Cytoscape. Subsequently, the GO and KEGG functional enrichment analyses were analyzed using the DAVID database. Furthermore, a protein-protein interaction network was constructed using STRING to obtain the key target information. Besides, we used a myocardial ischemia rat model to investigate the cardioprotective effects of YQHX. Transmission electron microscopy and Western blotting were used to observe apoptotic bodies and confirm protein expressions of key candidate targets, respectively. RESULTS: Network pharmacology showed that a total of 141 potential targets were obtained from these databases. The functional analysis results revealed that the targets of YQHX were largely associated with apoptosis, and the PI3K-AKT and MAPK pathways might represent key functional pathways. The hub genes of network include ALB, TP53, AKT1, TNF, VEGFA, EGFR, MAPK1, CASP3, JUN, FN1, MMP9, and MAPK8. In vivo, YQHX significantly improved cardiac function and suppressed apoptosis in ischemic rat myocardium. Furthermore, YQHX could significantly upregulate Nrf2 and HO-1 expression, and inhibit JNK phosphorylation. CONCLUSIONS: Based on network pharmacology and experimental evidence, this study proves that the cardioprotective effects and mechanisms of YQHX depend on multi-component, multi-target, and multi-pathway. In particular, YQHX exerts anti-apoptotic effects potentially by regulating the Nrf2/HO-1 and JNK-MAPK pathways.
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Cardiotónicos/farmacología , Medicamentos Herbarios Chinos/farmacología , Infarto del Miocardio/tratamiento farmacológico , Isquemia Miocárdica/tratamiento farmacológico , Animales , Apoptosis/efectos de los fármacos , Modelos Animales de Enfermedad , Hemo Oxigenasa (Desciclizante)/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Masculino , Infarto del Miocardio/genética , Infarto del Miocardio/fisiopatología , Isquemia Miocárdica/fisiopatología , Factor 2 Relacionado con NF-E2/metabolismo , Farmacología en Red , Mapas de Interacción de Proteínas , Ratas , Ratas Sprague-DawleyRESUMEN
Yi-Qi-Huo-Xue Decoction (YQHX) is the recombination of Dang-Gui-Bu-Xue Decoction (DBD), which is one of the well-known traditional Chinese Medicine (TCM) prescription, and has long been shown to have significant protective effects against myocardial ischemic injury. In previous studies, we found that YQHX could regulate lipid and glucose metabolism, promote angiogenesis, attenuate inflammatory response, and ameliorate left ventricular function in myocardial ischemia rat models. However, the underlying mechanism of how YQHX involves in lipid metabolism remains unclear so far. In this study, the underlying mechanism of YQHX in lipid metabolism disorders was elucidated in a myocardial ischemia rat model and a hypoxia-induced H9c2 cell injury model. YQHX (8.2 g·kg-1) and positive-control drug trimetazidine (10 mg·kg-1) were administered daily on the second day after left anterior descending (LAD) operation. At 7 days and 28 days after surgery, changes of cardiac morphology, structure, and function were evaluated by H&E staining and echocardiography, respectively. The plasma lipid levels and mitochondrial ATP content were also evaluated. Western blot and RT-PCR were used to determine the protein and mRNA expressions of AMPK, PGC-1α, CPT-1α, and PPARα. YQHX improved cardiac function and ameliorated lipid metabolism disorders. Furthermore, YQHX increased the expression of p-AMPK, PGC-1α, and CPT-1α without changing PPARα in ischemic rat myocardium. In vitro, YQHX activated the protein and mRNA expression of PGC-1α, CPT-1α, and PPARα in hypoxia-induced H9c2 cells injury, whereas AMPK inhibitor Compound c blocked the effects of YQHX. Taken together, the results suggest that YQHX reduces lipid metabolism disorders in myocardial ischemia via the AMPK-dependent signaling pathway.
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Medicamentos Herbarios Chinos/farmacología , Metabolismo de los Lípidos , Isquemia Miocárdica/tratamiento farmacológico , Transducción de Señal , Quinasas de la Proteína-Quinasa Activada por el AMP , Animales , Carnitina O-Palmitoiltransferasa , Línea Celular , Masculino , PPAR alfa , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma , Proteínas Quinasas , Ratas , Ratas Sprague-DawleyRESUMEN
Ischemic heart disease (IHD), a major global public health problem, is associated with high morbidity and mortality. Although the very best of modern approaches have proven effective in reducing morbidity and mortality, the poor prognosis of patients with IHD remains a major clinical concern. Cardiac energy metabolism is increasingly recognized as having a role in the pathogenesis of IHD, inducing metabolic substrate alterations, mitochondrial dysfunction, impaired function of the mitochondrial electron transport chain, and deprivation of cardiac energy. Factors involved in cardiac energy metabolism provide potential therapeutic targets for the treatment of IHD. Chinese herbal medicines (CHMs) have a long history of use in the prevention and treatment of cardiovascular diseases with multi-component, multi-target, and multi-signaling. Increasing evidence suggests that Chinese herbal medicines may improve myocardial ischemia through modulating cardiac energy metabolism. Here, we describe the possible targets and pathways of cardiac energy metabolism for CHMs, and appraise the modulatory effects of CHMs on energy metabolism in IHD. Especially, this review focuses on summarizing the metabolic effects and the underlying mechanisms of Chinese herbal medicines (including herbs, major bioactive components, and formulas) in IHD. In addition, we also discuss the current limitations and the major challenges for research investigating the use of CHMs in the treatment of cardiovascular diseases.
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BACKGROUND In traditional Chinese medicine, acupuncture has been used to treat angina due to coronary artery disease (CAD). The aim of this systematic review of the literature and meta-analysis was to identify published randomized controlled trials (RCTs) that quantified the effectiveness of adjunctive acupuncture treatment in patients with angina due to CAD who were also treated with Western or Chinese medicine. MATERIAL AND METHODS A systematic review of the literature included a search of the PubMed, Embase, Cochrane library, and China National Knowledge Infrastructure (CNKI) databases, from their inception to September 2018. Published findings from RCTs were included that investigated the effectiveness of acupuncture as an adjunctive treatment for angina due to CAD in combination with Western or traditional Chinese medicine. The odds ratio (OR) and 95% confidence interval (CI) were calculated using the random-effects model to determine the outcomes of markedly and moderately effective rates for the use of acupuncture. RESULTS Twenty-four published RCTs were identified that included 1,916 patients with CAD. Patients who received adjunctive acupuncture treatment had a significantly increased markedly effective rate. However, the moderately effective rate between adjunctive acupuncture combined with standard treatment for angina and standard treatment alone was not statistically significant. Sensitivity analysis showed that the pooled results for the markedly and moderately effective rates were robust. Subgroup analysis in most subsets supported the main findings. CONCLUSIONS Meta-analysis supported a positive treatment effect for the use of acupuncture when used as adjunctive therapy in patients with angina due to CAD.
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Terapia por Acupuntura/métodos , Angina de Pecho/terapia , Enfermedad de la Arteria Coronaria/terapia , China , Terapia Combinada , Femenino , Humanos , Masculino , Medicina Tradicional China/métodosRESUMEN
BACKGROUND: Mitochondrial dysfunction has been implicated in the pathogenesis of ischemic heart disease, exacerbating cardiomyocytes injury in myocardial infarction (MI). Peroxisome proliferator-activated receptor gamma co-activator (PGC-1α) has been recognized as the key regulator of mitochondrial biogenesis and energy metabolism. Yiqihuoxue decoction (YQHX), a Traditional Chinese Medicine (TCM) prescription, can prevent and treat ischemic heart disease. However, the mechanisms of YQHX on PGC-1α expression in the ischemic heart have remained unclear. METHODS: Myocardial ischemia rat model and ischemia/hypoxia injury model in the cardiomyocytes were used to minic human cardiovascular disease. Rats were randomly assigned into 4 groups: Sham, Model, YQHX (8.2 g/kg) and Trimetazidine (10 mg/kg) group. 28 days after MI, cardiac functions and morphology were detected by echocardiography and HE staining, respectively. In vitro, the effects of YQHX on H9c2 cell viability, LDH and ROS were detected, respectively. PGC-1α relevant proteins were evaluated by Western blotting. RESULTS: In vivo, echocardiography and HE staining results showed that YQHX improved cardiac functions and modified pathological changes. YQHX enhanced PGC-1α expression and improved the mitochondrial ultrastructure and functions in rats MI model for 4 weeks. Further, we explored its potential mechanisms in cardiomyocytes. In vitro, YQHX significantly enhanced cell viability and reduced LDH release and ROS production induced by hypoxia in cardiomyocytes. Interestingly, exposure of cardiomyocytes to hypoxic conditions for 12 h induced the downregulation of PGC-1α expression, but the expression levels nearly returned to the normal state after hypoxia for 24 h. YQHX significantly enhanced PGC-1α expression between 12 h and 24 h induced by hypoxia through a mechanism associated with the activation of AMPK phosphorylation in H9c2 cells. In addition, YQHX upregulated the expression of Tfam and NRF-1, while NRF-1 expression was completely blocked by an AMPK inhibitor. YQHX largely restored the mitochondrial morphology and increased mitochondrial membrane potential in hypoxia-induced injury. Furthermore, the UHPLC-LTQ-Orbitrap-MSn analysis found that there were 87 chemical constituents in YQHX. CONCLUSIONS: These results suggest that the protective effect of YQHX on cardiomyocytes against hypoxia-induced injury may be attributed to activation of PGC-1α and maintenance of mitochondrial functions through a mechanism involving the activation of AMPK phosphorylation.
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Cardiotónicos/farmacología , Medicamentos Herbarios Chinos/farmacología , Infarto del Miocardio/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma/metabolismo , Animales , Línea Celular , Corazón/efectos de los fármacos , Masculino , Infarto del Miocardio/patología , Miocardio/metabolismo , Miocardio/patología , Miocitos Cardíacos/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
OBJECTIVE: To investigate the effect of Yiqihuoxue prescription (YQHX) from Traditional Chinese Medicine (TCM) on myocardial glucose and lipid metabolism after myocardial infarction via the cross talk between the liver kinase B1 (LKB1)-dependent Notch1 and adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK). YQHX was prepared with substances with properties that benefit, to activate blood circulation based on the TCM theory. METHODS: Animal models of myocardial infarction were established by ligating Sprague Dawley rats' left anterior descending coronary arteries. The animals were randomly divided into a myocardial infarction (MI) group, a YQHX group, a perindopril group, a r-secretase inhibitor, Notch signal inhibitor (DAPT) group, a DAPT+YQHX group and a sham group. The related drugs were administered on the second day after operation, and changes in the relevant indexes were examined on weeks 1 and 4. Changes in cardiac structure and function were examined by echocardiography. The glucose and free fatty acids (FFA) were examined by ELISA. The expression of Notch, LKB1 and AMPK mRNA was examined by a real-time fluorescence quantitative method. The expression of glucose transporter 4 (GLUT4), and the expression of total acetyl-CoA carboxylase (ACC) and its phosphorylation were examined by western blotting. RESULTS: Compared with the sham group, the expression of Notch, LKB1 and AMPK mRNA in the MI group was lower. Compared with the MI group, the expression of these mRNAs in the YQHX and perindopril groups was higher, and their expression in the DAPT group was lower. At all time points, the protein expression of GLUT4 and pACC decreased in the MI group. On week 1, the expression of pACC protein was higher. In the DAPT group, the expression of pACC protein decreased. Compared with the YQHX group, the expression of pACC protein in the DAPT + YQHX group was lower. On week 4, compared with the MI group, the expression of GLUT4 protein in the YQHX group and the perindopril group was higher. The expression of GLUT4 protein in the DAPT group decreased. Compared with the YQHX group, the expression of GLUT4 protein in the DAPT+YQHX group was lower. There was no significant difference in the expression of ACC protein between the groups. CONCLUSION: YQHX promoted cross talk between the LKB1-dependent Notch1 and AMPK in myocardial tissue after myocardial infarction. Furthermore, it regulated the glucose and lipid metabolism of cardiomyocytes at different time points, thereby ameliorating the cardiac energy metabolism via different mechanisms and protecting the heart.
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ETHNOPHARMACOLOGICAL RELEVANCE: Fructus gardenia is widely used for treatment of stroke and infectious diseases in Chinese medicine. Geniposide is the key bioactive compound related to the pharmacodynamic actions of gardenia on ischemic stroke. The molecular mechanism by which geniposide improves the ischemic brain injury was observed in the study. AIM OF THE STUDY: Recent studies showed that geniposide had protective activities against the inflammatory response in ischemic stroke. However, the molecular mechanism of geniposide anti-inflammatory role has not yet been fully elucidated. In this study, we investigated the effect of geniposide on the expression of P2Y14 receptor and downstream signaling pathway in brain microvascular endothelial cells (BMECs). MATERIALS AND METHODS: An in vitro model of cerebral ischemia in BMECs was established by oxygen-glucose-deprivation (OGD). To further confirm the specific effect of geniposide on P2Y14 receptor and downstream signaling pathways, we set up a UDP-glucose (an agonist of the P2Y14 receptor) stimulated model. After administration of geniposide, the expression of P2Y14 receptor, phosphorylation of RAF-1, mitogen activated protein kinase kinase1/2 (MEK1/2), extracellular signal-regulated kinase 1/2 (ERK1/2), level of interleukin-8 (IL-8), interleukin-1ß (IL-1ß), monocyte chemotactic protein 1 (MCP-1) in BMECs were determined. RESULTS: The mRNA and protein expression of P2Y14 in the rat BMECs were up-regulated in OGD-induced injury. After administration of Geniposide, the expression of P2Y14 receptor was significantly down-regulated, the phosphorylation of RAF-1, MEK1/2, ERK1/2 were suppressed. Similar data were obtained in UDP-glc stimulated model. We also observed that geniposide markedly declined the production of IL-8, IL-1ß and MCP-1 in OGD-induced BMECs. CONCLUSION: Geniposide exerted anti-inflammatory effects by interfering with the expression of P2Y14 receptor, which subsequently inhibits the downstream ERK1/2 signaling pathways and the release of the pro-inflammatory cytokines IL-8, MCP-1, IL-1ß. Therefore, this study provides the evidence for gardenia's clinical application in cerebral ischemia.
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Células Endoteliales/efectos de los fármacos , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Inflamación/metabolismo , Iridoides/farmacología , Receptores Purinérgicos P2Y/metabolismo , Transducción de Señal/efectos de los fármacos , Animales , Células Cultivadas , Células Endoteliales/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/genética , Regulación de la Expresión Génica/efectos de los fármacos , Glucosa/metabolismo , Glucosa/farmacología , Oxígeno/metabolismo , Oxígeno/farmacología , Ratas , Ratas Sprague-Dawley , Receptores Purinérgicos P2Y/genéticaRESUMEN
OBJECTIVE: To explore the effect of Tongluojiunao injection (TLJN) prepared with Sanqi (Radix Notoginseng) and Zhizi (Fructus Gardeniae) on the interaction between brain microvascular endothelial cells (BMECs) and astrocytes in an in vitro ischemic model. METHODS: First, an in vitro model of cerebral ischemia in BMECs or astrocytes was established by oxygen-glucose deprivation (OGD). TLJN was used as a medicine of intervention. The OGD-injured BMECs were cultured in various astrocyte-conditioned media. Cell activity, alkaline phosphatase (AKP) and γ-glutamyl transpeptidase (γ-GT) activity, interleukin-1 beta (IL-1ß), and tumor necrosis factor alpha (TNF-α) content in BMECs were determined. Additionally, OGD-injured astrocytes were cultured in various BMEC-conditioned media. Cell activity, as well as expression of brain-derived neurotrophic factor (BDNF) and glial cell-derived neurotrophic factor (GDNF) in astrocytes, were detected. RESULTS: The results of paracrine signaling of normal BMECs or astrocytes showed a protective effect on each other: conditioned media from normal astrocytes improved cell viability, AKP, and γ-GT activity, and reduced IL-1ß and TNF-α content of injured BMECs; conditioned media from normal BMECs improved cell viability and expression of BDNF and GDNF in injured astrocytes. However, once the BMECs or astrocytes were injured by OGD, the protective effect decreased or disappeared. The above-mentioned protective induction was effectively recovered by TLJN intervention. CONCLUSION: The therapeutic benefit of TLJN was achieved by recovering two-way induction between BMECs and astrocytes, enhancing activity of injured BMECs and astrocytes, stabilizing enzymatic barriers, promoting expression of neurotrophic factors, and inhibiting inflammatory cytokines.
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Astrocitos/efectos de los fármacos , Isquemia Encefálica/tratamiento farmacológico , Medicamentos Herbarios Chinos/administración & dosificación , Células Endoteliales/efectos de los fármacos , Gardenia/química , Panax notoginseng/química , Animales , Astrocitos/metabolismo , Encéfalo/irrigación sanguínea , Encéfalo/efectos de los fármacos , Isquemia Encefálica/metabolismo , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Células Cultivadas , Células Endoteliales/metabolismo , Humanos , Interleucina-1beta/metabolismo , Masculino , Microvasos/citología , Microvasos/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Using a standard cellular fusion technique and indirect enzyme-linked immunosorbent assay (ELISA), a hybridoma cell line strain secreting anti-HBs monoclonal antibody (mAb) (defined G6 mAb) was obtained. The cells grew and secreted mAb stably. Antibody titers in the culture supernatant and ascites were 2.048×106 and 4.096×106, respectively. By applying the anti-HBs G6 mAb and horseradish peroxidase (HRP)-labeled goat anti-HBs antibody, we developed a sandwich ELISA (defined G6m ELISA) for detecting both wild-type and immune escape mutant HBsAgs (IEM HBsAg). The assay was performed to detect 17 species of genome recombinant expression HBsAg, including two wild-type species and 15 IEM HBsAg species, which varied in the "a" determinant, in a group of patients infected with hepatitis B virus (HBV). The patients previously had a lower ELISA detection signal [(absorbance of patients/absorbance of normal people (P/N): 1.0-4.5)]. The results demonstrated that the sensitivity of this assay to wild-type HBsAg was no less than 0.125 µg/L; 12 of 15 IEM HBsAg species (P/N⩾2.5) were positive for G6 mAb. Of the positive IEM HBsAg species, two had a low absorbance value at 450 nm (A 450), one had an intermediate A450 value and nine had a high A 450 value, which was 7.55%(mean), 59.4%and 92.1%-109.4% of the wild-type A 450 value, respectively. The two species with low A 450 value and the three negative species mutated at the bases 120-124 in the first loop of the HBV "a" determinant. Using the G6 ELISA and two commercial ELISA kits (A and B), 177 patients were tested. The G6 ELISA had a significantly higher detection rate than either commercial ELISAs (19.21% vs 14.89% and 6.21%, respectively; P<0.01, P<0.05, respectively).
RESUMEN
AIM: To prepare monoclonal antibodies against HBsAg and estimate its binding capacity to both wild-type and varies of immune escape mutant-type HBsAg. METHODS: Using self-made the anti-HBs G6 mAb and HRP-labeled goat anti-HBs antibody, A sandwich ELISA for detection both wild-type and varies of immune escape mutant-type HBsAg has been developed. Applying this assay, to detect 17 species of whole gene wild-type and recombination expressed HBsAg which varied in "a" determinant. to evaluate its clinical application characteristic of this assay, we used the other commercial reagents to Comparing with it. RESULTS: One strain hybridoma cell lin secreted anti-HBs mAb (defined G6), was obtained. It grew stably and the titers of the culture supernatant and hydroperitoneum were 2 048 and 4 096x10(3). The sensitivity to the wild-type HBsAg of this assay was less than 0.125 microg/L. This anti-HBs mAb can bind with 12 in 15 species mutant HBsAg (P/N> or =2.5), and 2 of them were low absorbent value at 450 nm(A(450)) group which was 7.55 percent of the wild-type, 1 of them was middle A(450) value group which was 59.40 percent of the wild-type, 9 of them were high A(450) value group which was 92.1-109.4 percent of the wild-type. The low A(450) value group and the negative group were diversified in 120-124 basyl in I loop of the HBV "a" determinant. We also used other commercial reagents, which was widely used in clinic, to detect partial species mutant HBsAg, the average A(450) value was less than foregoing assay. CONCLUSION: The G6 anti-HBs mAb was able to bind most of the immune escape mutant HBsAg in the test. And there would be some special regulations between the mutant site and the recombination expressed HBsAg binding capability.