Guizhi Fuling capsule(GFC), a traditional Chinese medicine(TCM) with effects of promoting blood circulation and dissipating blood stasis, has been widely used in the clinic. Because of the complex matrix and various c...Guizhi Fuling capsule(GFC), a traditional Chinese medicine(TCM) with effects of promoting blood circulation and dissipating blood stasis, has been widely used in the clinic. Because of the complex matrix and various chemical structure types, quality control of GFC remains great challenge. In the present study, an ultra performance liquid chromatography hybrid triple-quadrupole mass spectrometry(UPLC-QQQ MS) method with ultrafast positive/negative ionization switching was developed for simultaneous determination of 18 bioactive components in GFC, including methyl gallate, ethyl gallate, oxypaeoniflorin, benzoic acid, albiflorin, paeonolide, paeoniflorin, 1, 2, 3, 4, 6-pentagalloylglucose, mudanpioside C, benzoyloxypaeoniflorin, benzoylpaeoniflorin, pachymic acid, amygdalin, cinnamaldehyde, paeonol, cinnamic acid, 4-hydroxybenzoic acid, and gallic acid. Separation was performed on an Agilent Zorbax Extend–C18 column(2.1 mm × 50 mm, 1.8 μm), using a gradient elution with acetonitrile and water containing 0.1% formic acid. Cholic acid was selected as the internal standard. This newly developed method was fully validated for linearity, precision, accuracy, and stability, and then applied to quality assessment of GFC. Finally, the batch-to-batch reproducibility of GFC samples was evaluated by the cosine ration and Euclidean distance method, which showed high quality consistency. The results demonstrated that the developed method provided a reasonable and powerful manner for quality control of GFC.展开更多
Alzheimer’s disease(AD)is a neurodegenerative disorder disease mainly caused by extracellular senile plaques(SP)formed byβ-amyloid(Aβ1−42)protein deposits.Copper(Cu)is an essential metal involved in neural system,a...Alzheimer’s disease(AD)is a neurodegenerative disorder disease mainly caused by extracellular senile plaques(SP)formed byβ-amyloid(Aβ1−42)protein deposits.Copper(Cu)is an essential metal involved in neural system,and its homeostasis is the key to maintain its proper function.Herein,the subcellular locations of Cu(I)and Cu(II)in human neurodegenerative disease SH-SY5Y cells and AD mouse brains were imaged.We found that the content of Cu(II)decreased while that of Cu(I)increased under Aβexposure,which were further verified in the brain tissues of the AD mouse model,strongly suggesting the disruption of Cu homeostasis under Aβexposure or AD.Remarkably,the mitochondrial and lysosomal Cu(II)decreased significantly,whereas Cu(I)decreased in mitochondria but increased in lysosome.Lysosomes digested the damaged mitochondria via mitophagy to remove excess Cu(I)and maintain Cu homeostasis.The Aβinduced Cu(I)in mitochondria resulted in an overformation of reactive oxygen species and altered the morphology of this organelle.Due to the oxidative stress,glutathione(GSH)was converted into glutathione disulfide(GSSG),and Cu(I)bound with GSH was further released into the cytoplasm and absorbed by the lysosome.Transcriptomic analysis showed that genes(ATP7A/B)related to Cu transportation were upregulated,whereas genes related to mitochondrial complex were down-regulated,representing the damage of this organelle.This study demonstrated that Aβexposure caused the disruption of intracellular homeostasis by reducing Cu(II)to Cu(I)and damaging the mitochondria,which further triggered detoxification by the lysosome.Our finding provided new insights in Aβand AD induced Cu redox transformation and toxicity.展开更多
Acetylcholine(ACh)regulates inflammation viaα7 nicotinic acetylcholine receptor(α7 nAChR).Acetylcholinesterase(AChE),an enzyme hydrolyzing ACh,is expressed in immune cells suggesting non-classical function in inflam...Acetylcholine(ACh)regulates inflammation viaα7 nicotinic acetylcholine receptor(α7 nAChR).Acetylcholinesterase(AChE),an enzyme hydrolyzing ACh,is expressed in immune cells suggesting non-classical function in inflammatory responses.Here,the expression of PRiMA-linked G4 AChE was identified on the surface of macrophages.In lipopolysaccharide-induced inflammatory processes,AChE was upregulated by the binding of NF-κB onto the ACHE promotor.Conversely,the overexpression of G4 AChE inhibited ACh-suppressed cytokine release and cell migration,which was in contrast to that of applied AChE inhibitors.AChEmt,a DNA construct without enzymatic activity,was adopted to identify the protein role of AChE in immune system.Overexpression of G4 AChEmt induced cell migration and inhibited ACh-suppressed cell migration.The co-localization ofα7 nAChR and AChE was found in macrophases,suggesting the potential interaction ofα7 nAChR and AChE.Besides,immunoprecipitation showed a close association ofα7 nAChR and AChE protein in cell membrane.Hence,the novel function of AChE in macrophage by interacting withα7 nAChR was determined.Together with hydrolysis of ACh,AChE plays a direct role in the regulation of inflammatory response.As such,AChE could serve as a novel target to treat age-related diseases by antiinflammatory responses.展开更多
基金supported by the Specialized Research Fund for the Doctoral Program of Higher Education(No.20130096140001)the 111 Project(No.B16046)a project funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions(PAPD)
文摘Guizhi Fuling capsule(GFC), a traditional Chinese medicine(TCM) with effects of promoting blood circulation and dissipating blood stasis, has been widely used in the clinic. Because of the complex matrix and various chemical structure types, quality control of GFC remains great challenge. In the present study, an ultra performance liquid chromatography hybrid triple-quadrupole mass spectrometry(UPLC-QQQ MS) method with ultrafast positive/negative ionization switching was developed for simultaneous determination of 18 bioactive components in GFC, including methyl gallate, ethyl gallate, oxypaeoniflorin, benzoic acid, albiflorin, paeonolide, paeoniflorin, 1, 2, 3, 4, 6-pentagalloylglucose, mudanpioside C, benzoyloxypaeoniflorin, benzoylpaeoniflorin, pachymic acid, amygdalin, cinnamaldehyde, paeonol, cinnamic acid, 4-hydroxybenzoic acid, and gallic acid. Separation was performed on an Agilent Zorbax Extend–C18 column(2.1 mm × 50 mm, 1.8 μm), using a gradient elution with acetonitrile and water containing 0.1% formic acid. Cholic acid was selected as the internal standard. This newly developed method was fully validated for linearity, precision, accuracy, and stability, and then applied to quality assessment of GFC. Finally, the batch-to-batch reproducibility of GFC samples was evaluated by the cosine ration and Euclidean distance method, which showed high quality consistency. The results demonstrated that the developed method provided a reasonable and powerful manner for quality control of GFC.
文摘Alzheimer’s disease(AD)is a neurodegenerative disorder disease mainly caused by extracellular senile plaques(SP)formed byβ-amyloid(Aβ1−42)protein deposits.Copper(Cu)is an essential metal involved in neural system,and its homeostasis is the key to maintain its proper function.Herein,the subcellular locations of Cu(I)and Cu(II)in human neurodegenerative disease SH-SY5Y cells and AD mouse brains were imaged.We found that the content of Cu(II)decreased while that of Cu(I)increased under Aβexposure,which were further verified in the brain tissues of the AD mouse model,strongly suggesting the disruption of Cu homeostasis under Aβexposure or AD.Remarkably,the mitochondrial and lysosomal Cu(II)decreased significantly,whereas Cu(I)decreased in mitochondria but increased in lysosome.Lysosomes digested the damaged mitochondria via mitophagy to remove excess Cu(I)and maintain Cu homeostasis.The Aβinduced Cu(I)in mitochondria resulted in an overformation of reactive oxygen species and altered the morphology of this organelle.Due to the oxidative stress,glutathione(GSH)was converted into glutathione disulfide(GSSG),and Cu(I)bound with GSH was further released into the cytoplasm and absorbed by the lysosome.Transcriptomic analysis showed that genes(ATP7A/B)related to Cu transportation were upregulated,whereas genes related to mitochondrial complex were down-regulated,representing the damage of this organelle.This study demonstrated that Aβexposure caused the disruption of intracellular homeostasis by reducing Cu(II)to Cu(I)and damaging the mitochondria,which further triggered detoxification by the lysosome.Our finding provided new insights in Aβand AD induced Cu redox transformation and toxicity.
基金supported by Shenzhen Science and Technology Committee Research Grant(JCYJ20170413173747440,ZDSYS 201707281432317,JCYJ20180306174903174,CKFW2016082916015476,China)China Post-doctoral Science Foundation(2019M653087)+3 种基金Zhongshan Municipal Bureau of Science and Technology(ZSST20SC03,China)Guangzhou Science and Technology Committee Research Grant(GZSTI16SC02 and GZSTI17SC02,China)Hong Kong RGC Theme-based Research Scheme(T13-607/12R,China)Hong Kong Innovation Technology Fund(UIM/340,UIM/385,ITS/500/18FP,TCPD/17e9,PD18SC01 and HMRF18SC06,China)
文摘Acetylcholine(ACh)regulates inflammation viaα7 nicotinic acetylcholine receptor(α7 nAChR).Acetylcholinesterase(AChE),an enzyme hydrolyzing ACh,is expressed in immune cells suggesting non-classical function in inflammatory responses.Here,the expression of PRiMA-linked G4 AChE was identified on the surface of macrophages.In lipopolysaccharide-induced inflammatory processes,AChE was upregulated by the binding of NF-κB onto the ACHE promotor.Conversely,the overexpression of G4 AChE inhibited ACh-suppressed cytokine release and cell migration,which was in contrast to that of applied AChE inhibitors.AChEmt,a DNA construct without enzymatic activity,was adopted to identify the protein role of AChE in immune system.Overexpression of G4 AChEmt induced cell migration and inhibited ACh-suppressed cell migration.The co-localization ofα7 nAChR and AChE was found in macrophases,suggesting the potential interaction ofα7 nAChR and AChE.Besides,immunoprecipitation showed a close association ofα7 nAChR and AChE protein in cell membrane.Hence,the novel function of AChE in macrophage by interacting withα7 nAChR was determined.Together with hydrolysis of ACh,AChE plays a direct role in the regulation of inflammatory response.As such,AChE could serve as a novel target to treat age-related diseases by antiinflammatory responses.
