OBJECTIVE To explore the effect of total flavonoids of Rhododendra simsii(TFR)on improving cerebral ischemia/reperfusion injury(CIRI)and its relationship with STIM/Orai-regulated operational Ca^(2+)influx(SOCE)pathway...OBJECTIVE To explore the effect of total flavonoids of Rhododendra simsii(TFR)on improving cerebral ischemia/reperfusion injury(CIRI)and its relationship with STIM/Orai-regulated operational Ca^(2+)influx(SOCE)pathway.METHODS Oxygen-glucose deprivation/reoxygenation(OGD/R)PC12 cells were used to simulate CIRI in vitro,and the intracellular Ca^(2+)concentration and apoptosis rate of PC12 cells were detected by laser confocal microscope and flow cytometry,respectively.The regulation of STIM/Orai on SOCE was analyzed by STIM/Orai gene silencing and STIM/O rai gene overexpression.The CIRI model was established by MCAO in SD rats.The activities of inflammatory cytokines IL^(-1),IL-6 and TNF-αin serum were detected by ELISA.The pathological changes of ischemic brain tissue and the infarction of rat brain tissue were detected by HE staining and TTC staining.The protein and mRNA expression levels of STIM1,STIM2,Orai1,caspase-3 and PKB in brain tissue were detected by Western blotting and RT-qPCR,respectively.RESULTS The results of in vitro experiment showed that the fluorescence intensity of Ca^(2+)and apoptosis rate in PC12 cells treated with TFR were significantly lower than those in OGD/R group,and this trend was enhanced by SOCE antagonist 2-APB.STIM1/STIM2/Orai1 gene silencing significantly reduced apoptosis and Ca^(2+)overload in OGD/R model,while TFR combined with overexpression of STIM1/STIM2/Orai1 aggravated apoptosis and Ca2+overload.In the in vivo experiment,TFR significantly reduced the brain histopathological damage,infarction of brain tissue,the contents of IL^(-1),IL-6 and TNF-αin the serum in MCAO rats and down-regulated the expression of STIM1,STIM2,Orai1 and caspase-3 protein and mRNA in the brain tissue,and up-regulated the expression of PKB.The above effects were enhanced by the addition of 2-APB.CONCLUSION The above results indicate that TFR may reduce the contents of inflammatory factors and apoptosis,decrease Ca2+overload and ameliorate brain injury by inhibiting SOCE pathway mediated by STIM and Orai,suggesting that it has a protective effect against subacute CIRI.展开更多
The morbidity and mortality of cardiovascular diseases are very high,which has attracted more and more attention all over the world.Common treatment methods for clinical treatment of acute myocardial infarction includ...The morbidity and mortality of cardiovascular diseases are very high,which has attracted more and more attention all over the world.Common treatment methods for clinical treatment of acute myocardial infarction include direct percutaneous coronary intervention and coronary artery bypass grafting,which can quickly restore blocked coronary blood flow and reduce the infarct size.However,the inevitable ischemia/reperfusion injury will occur during the recovery of coronary blood flow,its pathological mechanism is complicated,and the Western medicine countermeasures are very limited.Among the current drugs for the treatment of cardiovascular diseases,traditional Chinese medicine has become a research hotspot due to its multiple targets,safety,and low side effects.Ginger is the fresh rhizome of Zingiber officinale Rosc.,a perennial herbaceous plant in the ginger family.It is a dual-purpose resource of medicine and food.Ginger has the functions of relieving the appearance and dispelling cold,warming up and relieving vomiting,resolving phlegm and relieving cough,and relieving fish and crab poison.The chemical components of ginger mainly include volatile oil,gingerol,diphenylheptane,etc..Among them,6-gingerol,as the main active component of gingerols,has obvious pharmacological effects in myocardial protection,anti-oxidation,anti-inflammatory,etc..Studies have shown that 6-gingerol protects myocardium mainly through anti-oxidative stress,anti-inflammatory,inhibiting cell apoptosis,and preventing calcium influx.①Anti-oxidative stress:oxidative stress is a state where oxidation and anti-oxidation in the body are out of balance,and it is also an important factor leading to myocardial damage.Many studies have confirmed that 6-gingerol has an antioxidant effect,and it is considered a natural antioxidant.6-gingerol can significantly reduce the degree of oxidative stress and the level of reactive oxygen species caused by cardiomyocyte damage,and has a significant cardioprotective effect.②Anti-inflammatory:inflammation can cause substantial cell damage and organ dysfunction,which is another important cause of myocardial damage.6-gingerol can reduce the levels of inflammatory factors such as interleukin-6,interleukin-1β,and tumor necrosis factor-αin cardiomyocytes,and at the same time inhibit the TLR4/NF-κB signaling pathway,an important regulatory pathway of inflammation,showing that it may improve myocardial damage through anti-inflammatory effects.③Inhibition of apoptosis:apoptosis is a complex and orderly process in the autonomous biochemical process of cells,and one of the main mechanisms of myocardial injury.This process can be roughly divided into three pathways:mitochondria,endoplasmic reticulum,and death receptors.Among them,the mitochondrial pathway plays an important role,and Bcl-2 and Bax located upstream of this pathway can regulate the entire process of cell apoptosis by regulating the permeability of the mitochondrial membrane.Studies have found that the preventive application of 6-gingerol can reduce cell damage,reduce the number of apoptotic cells,reduce the activity of Bax and caspase-3,and increase the expression of Bcl-2.Therefore,6-gingerol pretreatment can reduce the damage of cardiomyocytes,and its mechanism may be related to the inhibition of apoptosis.④Prevent calcium influx:calcium overload is involved in the pathogenesis of myocardial ischemic injury,which may be related to excessive contracture,arrhythmia,and mitochondrial Ca2+accumulation that impairs myocardial function.6-gingerol inhibits the increase of intracellular Ca2+concentration by inhibiting L-type calcium current,thereby reducing extracellular Ca2+influx,thereby avoiding calcium overload and playing a cardioprotective effect.In summary,6-gingerol can effectively treat and improve myocardial ischemia/reperfusion injury,and it has great development potential in the fields of medicine and health products.展开更多
OBJECTIVE In this study we explored the role of Epac1-Rap1 pathway in the acute myocardial ischemia/reperfusion injury(MIRI) in vitro and in vivo.METHODS An acute myocardial ischemia/reperfusion injury model was estab...OBJECTIVE In this study we explored the role of Epac1-Rap1 pathway in the acute myocardial ischemia/reperfusion injury(MIRI) in vitro and in vivo.METHODS An acute myocardial ischemia/reperfusion injury model was established by the ligation of left anterior descending coronary.Myocardial architecture,fibers and apoptosis was evaluated by the Masson trichrome staining,Sirius red staining and TUNEL assay.H9c2 cells were subjected to hypoxia for 5 h followed by 1-h reoxygen.ation in vitro.Cell viability was measured by MTT assay and cellular injury was evaluated by measuring the release of lactate dehydrogenase(LDH).Western blot,real-time PCR and immunofluorescence were used to detect the expressions of Epac1 and relative downstream molecules.RESULTS Myocardial IR-induced cardiac apoptosis and accumulation of Epac1 and Rap1 in rat IR injury model.Direct Epac activation by 8-CPT(8-(4-chlorophenylthio)-2′-O-methyl-cAMP) exacerbated cardiomyocyte death and dysfunction following hypoxia-reoxygenation(H/R),selective activation of Epac in response to H/R was evident which enriched for cytosolic/membrane proteins and mRNA.Harmacological inhibitor of Epac(ESI-09) significantly ameliorated myocardial injury with the decline of Epac expression.Epac inhibitor and agonist studies also implicated the effect of Rap1,which is downstream of Epac in this pathway.The expression of Rap1 elevated when activated by Epac agonist and was blocked by Epac inhibitor.The same result was true for myocyte CaMK-II and intracellular calcium ions activation.Moreover,ESI-09 also increased ERK1/2 phosphorylation.CONCLUSION Our study reveal that Epac1/Rap1 signaling pathway is involved in the pathogenesis of myocardial I/R injury in rats,which provides evidence on the development of therapeutic strategies target this pathway for myocardial I/R injury.展开更多
OBJECTIVE To predict the potential targets of hyperoside(Hyp)on improving ischemia/reperfusion injury by network pharmacology,and explore its possible mechanism combined with related literature.METHODS The action targ...OBJECTIVE To predict the potential targets of hyperoside(Hyp)on improving ischemia/reperfusion injury by network pharmacology,and explore its possible mechanism combined with related literature.METHODS The action targets of Hyp and ischemia/reperfusion injury were obtained by TCMSP,Swiss Target Prediction,Pharm Mapper,Similarity ensemble approach,Online Mendelian Inheritance in Man,DisGENT and database.The common targets of drugs and diseases were screened by Omishare and STRING database respectively,and the protein-protein interaction(PPI)network map was constructed.Then the interaction network between Hyp and disease targets was constructed by Cytoscape software and topological cross-linking analysis was carried out.Then the interaction network between Hyp and disease targets was constructed and cross-linked analysis was carried out by using Cytoscape software.The gene ontology(GO)of the core target was analyzed by David database,and then the related pathways of the core target were enriched by KEGG database.RESULTS A total of 54 GO enrichment processes were obtained by GO enrichment analysis of 44 common genes,including 38 biological processes(BP),15 cell composition(CC)processes,and 1 molecular functional(MF)process.43 items were obtained by signal pathway enrichment analysis in KEGG database.CONCLUSION It is suggested that the mechanism of Hyp may be related to PI3K-Akt,RAP1,RAS,VEGF and other signal transduction pathways.The above results laid a theoretical foundation for the study of the mechanism and clinical application of the treatment of ischemia/reperfusion injury.展开更多
Aim Salvia miltiorrhiza Bunge （SM） and lignum dalbergiae odoriferae （DO） are both traditional Chi- nese medicine that have cardioprotective effects. Here, we further examined the combined effects of SM and DO on r...Aim Salvia miltiorrhiza Bunge （SM） and lignum dalbergiae odoriferae （DO） are both traditional Chi- nese medicine that have cardioprotective effects. Here, we further examined the combined effects of SM and DO on rat myocardial ischemia/reperfusion injury. The possible mechanism of SM and DO also were elucidated. Methods DO was divided into aqueous extract of lignum dalbergiae odoriferae （DOW） and lignum dalbergiae odoriferae oil （DOO）. Sprague-Dawley rats were randomized to seven groups： sham group, model group, treatment groups inclu- ding SM （10 g · kg^-1）, DOW （5 g · kg^-1）, DOO （0.5 ml · kg^-1）, SM ＋ DOW （10 g · kg^-1 ＋ 5 g · kg^-1）, SM ＋ DOO （ 10 g · kg^-1 ＋ 0. 5 ml · kg^-1）. Rats were pretreated with homologous drug for 7 days and then subjec- ted to 30 rain of ischemia followed by 180 rain of reperfusion. Electrocardiogram （ECG） and heart rate were moni- tored and recorded continuously. At the end of reperfusion, blood samples were collected to determine the serum levels of creatine kinase-MB （CK-MB） and lactate dehydrogenase （LDH）. Hearts were harvested to assess heart- body rate, infarct size and histopathological changes as well. Maximum and minimum effective points were deter- mined by measuring indicators associate with myocardial injury at different time-points of reperfusion （Smin, 15min, 30min, 45rain, 60min, 120min, 180min）. The potential therapeutic mechanism of SM and SM ＋ DOO were carried out by detecting superoxide dismutase （SOD）, malondialdehyde （MDA）, tumor necrosis factor-alpha （TNF-alpha） and interleukin 6 （IL-6）. Results The results showed SM and DO can ameliorate cardiac function respectively, and this cardioprotective effect was further strengthened by their combinations. Among all the combi- nations, SM ＋ DOO showed predominant potential to improve ECG and heart rate, reduce heart-body rate （28.5% ＋ 1.4% , P 〈 0.01 vs model） and myocardial infarct size （ 20.96% ＋ 1.61% , P 〈 0.01 vs model, P 〈 0.05 vs SM） , attenuate histopathological damage, decrease the levels of CK-MB and LDH （P 〈 0.01 vs model, P 〈 0.05 vs SM）. The maximum effective points of SM and SM ＋ DOO were 15min and 30rain respectively, and the minimum effective points of them were 180rain. In reducing serum level of MDA, TNF-alpha, IL-6 and increasing SOD activ- ity, SM ＋ DOO was similar to SM. Conclusion The results of this study indicated that SM ＋ DOO have combined effects that are highly effective than single pretreatment against myocardial ischemie reperfusion injury in rats. The possible mechanism of SM and DO were likely through its anti-oxidant and anti-inflammatory properties, and thus may be an effective and promising medicine for both prophylaxis and treatment of ischemic heart disease.展开更多
基金National Natural Science Foundation of China(81173596)and Major Project of Natural Science Foundation of the Department of Education of Anhui Province(KJ2019ZD32)。
文摘OBJECTIVE To explore the effect of total flavonoids of Rhododendra simsii(TFR)on improving cerebral ischemia/reperfusion injury(CIRI)and its relationship with STIM/Orai-regulated operational Ca^(2+)influx(SOCE)pathway.METHODS Oxygen-glucose deprivation/reoxygenation(OGD/R)PC12 cells were used to simulate CIRI in vitro,and the intracellular Ca^(2+)concentration and apoptosis rate of PC12 cells were detected by laser confocal microscope and flow cytometry,respectively.The regulation of STIM/Orai on SOCE was analyzed by STIM/Orai gene silencing and STIM/O rai gene overexpression.The CIRI model was established by MCAO in SD rats.The activities of inflammatory cytokines IL^(-1),IL-6 and TNF-αin serum were detected by ELISA.The pathological changes of ischemic brain tissue and the infarction of rat brain tissue were detected by HE staining and TTC staining.The protein and mRNA expression levels of STIM1,STIM2,Orai1,caspase-3 and PKB in brain tissue were detected by Western blotting and RT-qPCR,respectively.RESULTS The results of in vitro experiment showed that the fluorescence intensity of Ca^(2+)and apoptosis rate in PC12 cells treated with TFR were significantly lower than those in OGD/R group,and this trend was enhanced by SOCE antagonist 2-APB.STIM1/STIM2/Orai1 gene silencing significantly reduced apoptosis and Ca^(2+)overload in OGD/R model,while TFR combined with overexpression of STIM1/STIM2/Orai1 aggravated apoptosis and Ca2+overload.In the in vivo experiment,TFR significantly reduced the brain histopathological damage,infarction of brain tissue,the contents of IL^(-1),IL-6 and TNF-αin the serum in MCAO rats and down-regulated the expression of STIM1,STIM2,Orai1 and caspase-3 protein and mRNA in the brain tissue,and up-regulated the expression of PKB.The above effects were enhanced by the addition of 2-APB.CONCLUSION The above results indicate that TFR may reduce the contents of inflammatory factors and apoptosis,decrease Ca2+overload and ameliorate brain injury by inhibiting SOCE pathway mediated by STIM and Orai,suggesting that it has a protective effect against subacute CIRI.
基金Fund of Dean of Huachuang Institute of Areca Research-Hainan(HCBL2020YZ-012)。
文摘The morbidity and mortality of cardiovascular diseases are very high,which has attracted more and more attention all over the world.Common treatment methods for clinical treatment of acute myocardial infarction include direct percutaneous coronary intervention and coronary artery bypass grafting,which can quickly restore blocked coronary blood flow and reduce the infarct size.However,the inevitable ischemia/reperfusion injury will occur during the recovery of coronary blood flow,its pathological mechanism is complicated,and the Western medicine countermeasures are very limited.Among the current drugs for the treatment of cardiovascular diseases,traditional Chinese medicine has become a research hotspot due to its multiple targets,safety,and low side effects.Ginger is the fresh rhizome of Zingiber officinale Rosc.,a perennial herbaceous plant in the ginger family.It is a dual-purpose resource of medicine and food.Ginger has the functions of relieving the appearance and dispelling cold,warming up and relieving vomiting,resolving phlegm and relieving cough,and relieving fish and crab poison.The chemical components of ginger mainly include volatile oil,gingerol,diphenylheptane,etc..Among them,6-gingerol,as the main active component of gingerols,has obvious pharmacological effects in myocardial protection,anti-oxidation,anti-inflammatory,etc..Studies have shown that 6-gingerol protects myocardium mainly through anti-oxidative stress,anti-inflammatory,inhibiting cell apoptosis,and preventing calcium influx.①Anti-oxidative stress:oxidative stress is a state where oxidation and anti-oxidation in the body are out of balance,and it is also an important factor leading to myocardial damage.Many studies have confirmed that 6-gingerol has an antioxidant effect,and it is considered a natural antioxidant.6-gingerol can significantly reduce the degree of oxidative stress and the level of reactive oxygen species caused by cardiomyocyte damage,and has a significant cardioprotective effect.②Anti-inflammatory:inflammation can cause substantial cell damage and organ dysfunction,which is another important cause of myocardial damage.6-gingerol can reduce the levels of inflammatory factors such as interleukin-6,interleukin-1β,and tumor necrosis factor-αin cardiomyocytes,and at the same time inhibit the TLR4/NF-κB signaling pathway,an important regulatory pathway of inflammation,showing that it may improve myocardial damage through anti-inflammatory effects.③Inhibition of apoptosis:apoptosis is a complex and orderly process in the autonomous biochemical process of cells,and one of the main mechanisms of myocardial injury.This process can be roughly divided into three pathways:mitochondria,endoplasmic reticulum,and death receptors.Among them,the mitochondrial pathway plays an important role,and Bcl-2 and Bax located upstream of this pathway can regulate the entire process of cell apoptosis by regulating the permeability of the mitochondrial membrane.Studies have found that the preventive application of 6-gingerol can reduce cell damage,reduce the number of apoptotic cells,reduce the activity of Bax and caspase-3,and increase the expression of Bcl-2.Therefore,6-gingerol pretreatment can reduce the damage of cardiomyocytes,and its mechanism may be related to the inhibition of apoptosis.④Prevent calcium influx:calcium overload is involved in the pathogenesis of myocardial ischemic injury,which may be related to excessive contracture,arrhythmia,and mitochondrial Ca2+accumulation that impairs myocardial function.6-gingerol inhibits the increase of intracellular Ca2+concentration by inhibiting L-type calcium current,thereby reducing extracellular Ca2+influx,thereby avoiding calcium overload and playing a cardioprotective effect.In summary,6-gingerol can effectively treat and improve myocardial ischemia/reperfusion injury,and it has great development potential in the fields of medicine and health products.
基金This work supported by the National Natural Science Foundation of China (81470432) and Natural Science Foundation of Anhui Province Education Department (KJ2016A357).
文摘OBJECTIVE In this study we explored the role of Epac1-Rap1 pathway in the acute myocardial ischemia/reperfusion injury(MIRI) in vitro and in vivo.METHODS An acute myocardial ischemia/reperfusion injury model was established by the ligation of left anterior descending coronary.Myocardial architecture,fibers and apoptosis was evaluated by the Masson trichrome staining,Sirius red staining and TUNEL assay.H9c2 cells were subjected to hypoxia for 5 h followed by 1-h reoxygen.ation in vitro.Cell viability was measured by MTT assay and cellular injury was evaluated by measuring the release of lactate dehydrogenase(LDH).Western blot,real-time PCR and immunofluorescence were used to detect the expressions of Epac1 and relative downstream molecules.RESULTS Myocardial IR-induced cardiac apoptosis and accumulation of Epac1 and Rap1 in rat IR injury model.Direct Epac activation by 8-CPT(8-(4-chlorophenylthio)-2′-O-methyl-cAMP) exacerbated cardiomyocyte death and dysfunction following hypoxia-reoxygenation(H/R),selective activation of Epac in response to H/R was evident which enriched for cytosolic/membrane proteins and mRNA.Harmacological inhibitor of Epac(ESI-09) significantly ameliorated myocardial injury with the decline of Epac expression.Epac inhibitor and agonist studies also implicated the effect of Rap1,which is downstream of Epac in this pathway.The expression of Rap1 elevated when activated by Epac agonist and was blocked by Epac inhibitor.The same result was true for myocyte CaMK-II and intracellular calcium ions activation.Moreover,ESI-09 also increased ERK1/2 phosphorylation.CONCLUSION Our study reveal that Epac1/Rap1 signaling pathway is involved in the pathogenesis of myocardial I/R injury in rats,which provides evidence on the development of therapeutic strategies target this pathway for myocardial I/R injury.
基金National Natural Science Foundation of China(81170148)and Major Project of Natural Science Foundation of the Department of Education of Anhui Province(KJ2019ZD32)。
文摘OBJECTIVE To predict the potential targets of hyperoside(Hyp)on improving ischemia/reperfusion injury by network pharmacology,and explore its possible mechanism combined with related literature.METHODS The action targets of Hyp and ischemia/reperfusion injury were obtained by TCMSP,Swiss Target Prediction,Pharm Mapper,Similarity ensemble approach,Online Mendelian Inheritance in Man,DisGENT and database.The common targets of drugs and diseases were screened by Omishare and STRING database respectively,and the protein-protein interaction(PPI)network map was constructed.Then the interaction network between Hyp and disease targets was constructed by Cytoscape software and topological cross-linking analysis was carried out.Then the interaction network between Hyp and disease targets was constructed and cross-linked analysis was carried out by using Cytoscape software.The gene ontology(GO)of the core target was analyzed by David database,and then the related pathways of the core target were enriched by KEGG database.RESULTS A total of 54 GO enrichment processes were obtained by GO enrichment analysis of 44 common genes,including 38 biological processes(BP),15 cell composition(CC)processes,and 1 molecular functional(MF)process.43 items were obtained by signal pathway enrichment analysis in KEGG database.CONCLUSION It is suggested that the mechanism of Hyp may be related to PI3K-Akt,RAP1,RAS,VEGF and other signal transduction pathways.The above results laid a theoretical foundation for the study of the mechanism and clinical application of the treatment of ischemia/reperfusion injury.
文摘Aim Salvia miltiorrhiza Bunge （SM） and lignum dalbergiae odoriferae （DO） are both traditional Chi- nese medicine that have cardioprotective effects. Here, we further examined the combined effects of SM and DO on rat myocardial ischemia/reperfusion injury. The possible mechanism of SM and DO also were elucidated. Methods DO was divided into aqueous extract of lignum dalbergiae odoriferae （DOW） and lignum dalbergiae odoriferae oil （DOO）. Sprague-Dawley rats were randomized to seven groups： sham group, model group, treatment groups inclu- ding SM （10 g · kg^-1）, DOW （5 g · kg^-1）, DOO （0.5 ml · kg^-1）, SM ＋ DOW （10 g · kg^-1 ＋ 5 g · kg^-1）, SM ＋ DOO （ 10 g · kg^-1 ＋ 0. 5 ml · kg^-1）. Rats were pretreated with homologous drug for 7 days and then subjec- ted to 30 rain of ischemia followed by 180 rain of reperfusion. Electrocardiogram （ECG） and heart rate were moni- tored and recorded continuously. At the end of reperfusion, blood samples were collected to determine the serum levels of creatine kinase-MB （CK-MB） and lactate dehydrogenase （LDH）. Hearts were harvested to assess heart- body rate, infarct size and histopathological changes as well. Maximum and minimum effective points were deter- mined by measuring indicators associate with myocardial injury at different time-points of reperfusion （Smin, 15min, 30min, 45rain, 60min, 120min, 180min）. The potential therapeutic mechanism of SM and SM ＋ DOO were carried out by detecting superoxide dismutase （SOD）, malondialdehyde （MDA）, tumor necrosis factor-alpha （TNF-alpha） and interleukin 6 （IL-6）. Results The results showed SM and DO can ameliorate cardiac function respectively, and this cardioprotective effect was further strengthened by their combinations. Among all the combi- nations, SM ＋ DOO showed predominant potential to improve ECG and heart rate, reduce heart-body rate （28.5% ＋ 1.4% , P 〈 0.01 vs model） and myocardial infarct size （ 20.96% ＋ 1.61% , P 〈 0.01 vs model, P 〈 0.05 vs SM） , attenuate histopathological damage, decrease the levels of CK-MB and LDH （P 〈 0.01 vs model, P 〈 0.05 vs SM）. The maximum effective points of SM and SM ＋ DOO were 15min and 30rain respectively, and the minimum effective points of them were 180rain. In reducing serum level of MDA, TNF-alpha, IL-6 and increasing SOD activ- ity, SM ＋ DOO was similar to SM. Conclusion The results of this study indicated that SM ＋ DOO have combined effects that are highly effective than single pretreatment against myocardial ischemie reperfusion injury in rats. The possible mechanism of SM and DO were likely through its anti-oxidant and anti-inflammatory properties, and thus may be an effective and promising medicine for both prophylaxis and treatment of ischemic heart disease.
