1.Calcium mobilizing effect of hawthorn leaf procyanidins in vascular endothelial cells.
Peng LI ; Jian-Nong WANG ; Jin-Cai HOU ; Jian-Hua FU ; Jian-Xun LIU
China Journal of Chinese Materia Medica 2018;43(12):2600-2606
The hawthorn leaves have the effect of activating blood, removing blood stasis, regulating qi through the veins, dissolving turbidity and lowering lipid. Procyanidinis is one of its main active components and plays an important role in regulating vasoactivity. Previous studies showed that the regulating effect of procyanidins was related to its regulation on nitric oxide secretion from vascular endothelial cells, and this effect was dependent on the extracellular calcium concentration, suggesting that the changes in intracellular calcium ion concentration in endothelial cells may play a key role in this process. However, the research on this issue is still insufficient so far. This study is aimed to observe the effect of hawthorn leaf oligomeric procyanidins (HLP) on calcium mobilization of vascular endothelial cells, and investigate the underlying mechanism. Human umbilical vein endothelial cells (HUVEC) were cultured and labeled with Fura-2. HUVEC were treated with HLP at concentrations of 6.25, 12.5, 25 and 50 mg·L⁻¹, and the intracellular calcium concentrations were measured with a living cell microscope for 30 min. HLP increased the intracellular calcium concentration of HUVEC in a concentration dependent manner; and the intracellular calcium concentrations in 25 and 50 mg·L⁻¹ HLP groups were significantly higher than that in the normal group. With the use of calcium-free incubation buffer, addition of calcium chelating agent EGTA in incubation buffer, or use of inhibitors for sodium calcium exchanger, the effect of HLP was significantly inhibited. On the other hand, the effect of HLP could also be weakened by inhibiting the calcium release from the intracellular storage. In conclusion, these results suggest that HLP can elicit calcium mobilization in vascular endothelial cells, which may be one of the mechanisms for its vascular modulatory activity; and this calcium mobilizing effect may be achieved through promoting both extracellular calcium influx and intracellular calcium release, additionally the former may be related to activating the reverse transport of Na⁺-Ca²⁺ exchangers on the cell membrane.
2.Protective effect of succinic acid on primary cardiomyocyte hypoxia/reoxygenation injury.
Xi-Lan TANG ; Jian-Xun LIU ; Peng LI ; Wei DONG ; Lei LI ; Yong-Qiu ZHENG ; Jin-Cai HOU
China Journal of Chinese Materia Medica 2013;38(21):3742-3746
To establish cardiomyocyte hypoxia/reoxygenation injury model by culturing primary cardiomyocytes from suckling SD rats, in order to study the effect of succinic acid on LDH leakage rate cardiomyocyte ischemia/reperfusion injury. Furthermore, flow cytometry and western blot were conducted to detect the effect of succinic acid on cardiomyocyte apoptosis, cleaved caspase-3 and p-Akt, and discuss the protective effect of succinic acid on primary cardiomyocyte hypoxia/reoxygenation injury of primary cardiomyocytes from neonatal SD rats. According to the findings of the study, succinic acid at the concentrations ranging between 31.25 mg x L(-1) and 500 mg x L(-1) had no significant effect on primary cardiomyocyte activity, and succinic acid at the concentrations of 400, 200, 100, 50 mg x L(-1) could notably reduce cardiomyocyte ischemia/reperfusion LDH leakage rate (P < 0.01 or P < 0.05, respectively). Succinic acid at the concentrations of 400 mg x L(-1) and 200 mg x L(-1) could significantly reduce the percentage of cardiomyocyte apoptosis (P < 0.05), and inhibit the protein expression of cleaved caspase-3 caused by cardiomyocyte ischemia/reperfusion (P < 0.05). Succinic acid at the concentration of 400 mg x L(-1) could remarkably increase the protein expression of cardiomyocyte Akt (P < 0.05), while succinic acid at the concentration of 200 mg x L(-1) had no obvious effect on the protein expression of cardiomyocyte Akt. Therefore, this study demonstrated that succinic acid could inhibit necrosis and apoptosis caused by cardiomyocyte hypoxia/reoxygenation by activating Akt phosphorylation.
Animals
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Apoptosis
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drug effects
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Caspase 3
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metabolism
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Cell Hypoxia
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drug effects
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Cell Survival
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drug effects
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Cells, Cultured
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Female
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Humans
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Hypoxia
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metabolism
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physiopathology
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prevention & control
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Male
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Myocardial Reperfusion Injury
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metabolism
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physiopathology
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prevention & control
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Myocytes, Cardiac
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cytology
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drug effects
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metabolism
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Oxygen
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metabolism
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Protective Agents
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pharmacology
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Proto-Oncogene Proteins c-akt
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metabolism
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Rats
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Rats, Sprague-Dawley
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Succinic Acid
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pharmacology
3.Network Pharmacology Approach to Explore Mechanisms of Qizhu Granules for Treatment of Diabetic Nephropathy
Ya-dong LIN ; Wen-jing GAO ; Min HOU ; Pan WANG ; Lei LEI ; Jun-guo REN
Chinese Journal of Experimental Traditional Medical Formulae 2020;26(24):161-168
Objective:To explore the mechanism of Qizhu granules in the treatment of diabetic nephropathy by using network pharmacology. Method:The Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform database （TCMSP） and The Encyclopedia of Traditional Chinese Medicine（ETCM） database were used to screen out the chemical constituents and protein targets of each drug in the Qizhu granules based on oral bioavailability and drug-like properties. The protein target was standardized into the corresponding gene name through the UniProt database. Online Mendelian Inheritance in Man（OMIM）， DisGeNET， Therapeutic Target Database （TTD）， ETCM database were used to search for related targets of diabetic nephropathy， after the intersection of the two， construct a protein interaction network through protein interaction database （STRING）， use Cytoscape to analyze the core target of the network， and the relevant targets were analyzed by KOBAS 3.0 database for Gene Ontology （GO） pathway enrichment analysis and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment analysis. Result:A total of 93 chemical components were obtained from Qizhu granules， involving 254 targets， and 607 targets related to diabetic nephropathy. After the intersection， 76 sputum granules were determined to treat diabetic nephropathy， including protein kinase B1 （Akt1）， vascular endothelial growth factor （VEGFA）， interleukin （IL）-6， tumor necrosis factor （TNF）， mitogen-activated protein kinase 1 （MAPK1）， matrix metalloproteinase （MMP）-9 and other core targets， after GO analysis and KEGG analysis， Qizhu granules can affect cellular response to nitrogen compound， regulation of reactive oxygen species metabolic process and other biological processes， regulate advanced glycation end product （AGE）/advanced glycation end product receptor （RAGE） signaling pathway in diabetic complications， fluid shear stress and atherosclerosis， IL-17 signaling pathways， HIF-1 signaling pathways TNF signaling pathways and other pathways. Conclusion:The therapeutic effect of Qizhu granules on diabetic nephropathy may affect Akt1，VEGFA， IL-6， TNF， MAPK1， MMP-9 and other targets， and regulate AGE/RAGE signaling pathway in diabetic complications， fluid shear stress and atherosclerosis， IL-17 signaling pathways， hypoxia-inducing factor-1（HIF-1）signaling pathways TNF signaling pathways and other pathways， which can provide a theoretical reference for further basic experimental research.
4.Mechanism of Astragali Radix Vesicle-like Nanoparticles for Reducing Blood Glucose in db/db Diabetic Mice by Regulating Gut Microbiota
Wen-jing GAO ; Min HOU ; Xiao-xiao CHEN ; Pan WANG ; Jun-guo REN ; Jian-xun LIU
Chinese Journal of Experimental Traditional Medical Formulae 2021;27(14):111-118
Objective:To study the extraction method and characteristics of vesicle-like nanoparticles （VLNs） in Astragali Radix decoction， and to explore the mechanism of the VLNs in reducing blood glucose by regulating the gut microbiota of db/db diabetic mice. Method:Ultracentrifugation and size exclusion chromatography were used to enrich VLNs from Astragali Radix decoction， and the morphology， particle size and concentration of the VLNs were analyzed by transmission electron microscope and nanoparticle tracking analyzer. The db/db diabetic mice were randomly divided into model group， Astragali Radix VLNs high-， medium- and low-dose （21.1， 10.6， 5.3 g·kg-1） groups and metformin group （0.25 g·kg-1） according to their blood glucose levels. There were 7 mice in each group， and another 7 C57BL/6 mice were set as the normal group. The mice were given intragastrically for 3 weeks （once a day）， and the changes of fasting blood glucose were observed every week. Hematoxylin-eosin （HE） staining was used to observe the pathological morphology of liver and pancreas of diabetic mice. The feces of mice were collected for 16S rRNA diversity detection of intestinal microbes. Result:The size of the nanoparticles obtained by the two methods was about 200 nm. Astragali Radix VLNs extracted by ultracentrifugation had a typical saucer-like shape with the concentration of 3.0×1011 particles·mL-1. The morphology of Astragali Radix VLNs obtained by size exclusion chromatography was relatively poor with the concentration of 2.2×1011 particles·mL-1. After 3 weeks of administration， compared with the model group， Astragali Radix VLNs high-， medium- and low-dose groups could significantly reduce the fasting blood glucose of diabetic mice （
5.Effect of Jiangtang Xiaozhi Tablet on Liver Circadian Clock-related Genes CLOCK， BMAL1， REV-ERBα， and REV-ERBβ in Mice with Metabolic Dysfunction-associated Fatty Liver Disease
Xiaoxiao CHEN ; Min HOU ; Pan WANG ; Junguo REN ; Jianxun LIU
Chinese Journal of Experimental Traditional Medical Formulae 2022;28(18):20-29
ObjectiveTo explore the effect of Jiangtang Xiaozhi tablet （JTXZT） on metabolic dysfunction-associated fatty liver disease and to study the mechanism from the perspective of circadian clock-related genes such as circadian locomotor output cycles kaput （CLOCK）， brain and muscle aryl hydrocarbon receptor nuclear translocator-like protein 1 （BMAL1）， reverse-eritroblastosis receptor （REV-ERB）α and β. MethodA total of 50 male SPF C57BL/6J mice were randomized into normal group （n=10） and modeling group （n=40）. The normal group was fed with normal diet， and the modeling group with high-fat diet for 4 weeks. Then the model mice were randomly classified into model group， high-dose （12.5 g·kg-1） and low-dose （6.25 g·kg-1） Jiangtang Xiaozhi tablet groups， and orlistat group （70 mg·kg-1）， with 10 mice in each group. The normal group and model group received equivalent volume of distilled water （8 weeks）. Then， the body weight of mice was measured， and the content of serum triglyceride （TG）， total cholesterol （TC）， low-density lipoprotein cholesterol （LDL-C）， high-density lipoprotein cholesterol （HDL-C）， aspartate aminotransferase （AST）， and alanine aminotransferase （ALT） was determined with biochemical method. Serum content of free fatty acid （FFA） and leptin was detected by enzyme-linked immunosorbent assay （ELISA）. Pathological changes of liver tissue and epididymal adipose tissue were observed based on hematoxylin-eosin （HE） staining. Liver fibrosis was examined based on Masson's trichrome staining， and changes of lipids based on oil red O staining. The expression of CLOCK， BMAL1， REV-ERBα， and REV-ERBβ was detected by Western blot and immunohistochemistry assay. ResultCompared with the normal group， the model group had high content of TG， TC， LDL-C， HDL-C， AST， ALT， FFA， and leptin （P<0.05， P<0.01）， showed ballooning degeneration and focal microvesicular steatosis of liver cells， enlarged adipocytes， and inflammatory cell clusters and fibrous tissue hyperplasia， and displayed increased protein expression of sterol regulatory element binding protein （SREBP） 1 and peroxisome proliferators-activated receptor （PPAR）γ （P<0.01） and decreased protein expression of PPARα （P<0.05）， CLOCK， BMAL1， REV-ERBα and β （P<0.05， P<0.01）. Compared with the model group， JTXZT-H group down-regulated the content of TG， TC， LDL-C， HDL-C， AST， ALT， FFA， and leptin in mice （P<0.05， P<0.01）， and the JTXZT groups demonstrated reduction in the degree and range of ballooning degeneration of liver tissue， alleviation of the compression of hepatic sinusoidal tissue， unobvious inflammatory cell infiltration and fibrous tissue proliferation， reduction in the expression of SREBP1 and PPARγ （P<0.05， P<0.01）， and rise of the protein expression of PPARα （P<0.01）， CLOCK， BMAL1， REV-ERBα， and REV-ERBβ （P<0.05， P<0.01）. ConclusionJTXZT can significantly alleviate the metabolic dysfunction-associated fatty liver disease in mice caused by high-fat diet. The mechanism is the likelihood that it regulates downstream related lipid metabolism proteins （such as SREBP1， PPARγ， and PPARα）.
6.Analysis on Medication Rules of Traditional Chinese Medicine for Treatment of Diabetic Peripheral Neuropathy Based on Clinical Literature
Fu-zhi ZHANG ; Ya-dong LIN ; Lei LEI ; Wen-jing GAO ; Min HOU ; Li KANG ; Jun-guo REN
Chinese Journal of Experimental Traditional Medical Formulae 2020;26(13):199-205
Objective:To investigate the medication rules of traditional Chinese medicine (TCM) for the treatment of diabetic peripheral neuropathy (DPN).Method:The literature published in China Knowledge Resource Integrated Database (CNKI), Wanfang Database, China Biomedical Literature Service System (SinoMed), VIP Database and PubMeb from 2008 to 2019 were retrieved by setting the topics of diabetic peripheral neuropathy and TCM. After screening, a database was established to analyze the medication rules (efficacy, frequency, flavor and meridian tropism, common couplet medicinals and core medicines) of TCM by frequency statistics, association rules and data statistical methods of constructing complex networks.Result:A total of 461 papers for treatment of DPN were included in this study, including 275 kinds of TCM and a total frequency of 6 361 times. Astragali Radix had the highest frequency. Among all kinds of medicinal materials, activating blood circulation and removing stasis was the most commonly used medicine, followed by Qi-invigorating medicine. Flavor of medicines was mainly sugariness and warm, and most of their meridian tropism was liver meridian. After the analysis by association rules, the couplet medicinals with the highest support was Astragali Radix-Angelicae Sinensis Radix. The core medicines obtained by complex network analysis were Astragali Radix, Angelicae Sinensis Radix, Chuanxiong Rhizoma, Spatholobi Caulis, Cinnamomi Ramulus, Carthami Flos, Pheretima, Paeoniae Radix Rubra, Salviae Miltiorrhizae Radix et Rhizoma and Persicae Semen.Conclusion:This study comprehensively analyzes the medication rules of TCM clinical treatment of DPN. The main treatment methods of TCM for DPN are invigorating Qi and blood, activating blood circulation and removing stasis, activating meridians to stop pain, which can provide guidance for the TCM clinical use and new Chinese medicines research and development of DPN.
7.Mdivi-1 protects oligodendrocytes through inhibiting apoptotic signaling pathway
Yanhua LI ; Xiaojuan ZHANG ; Siyu ZHANG ; Xiyuan HOU ; Ziyi LIU ; Xiao-Jing YU ; Nianping ZHANG
Chinese Journal of Pathophysiology 2024;40(3):527-534
AIM:To investigate the therapeutic effect of mitochondrial fission inhibitor-1(Mdivi-1)on experi-mental autoimmune encephalomyelitis(EAE)in mice,and to explore its mechanism.METHODS:The mice immunized with myelin oligodendrocyte glycoprotein peptide fragment 35-55(MOG35-55)were randomly divided into DMSO model group and Mdivi-1 intervention group.All mice were sacrificed on the 28th day after the first immunization.The demyelination was analyzed by Luxol fast blue staining.The protective mechanism of Mdivi-1 in the spinal cord tissue was investigated by immunofluorescence staining,TUNEL staining and the in vitro experiment with MO3.13 oligodendrocytes treated with staurosporine.The mitochondrial depolarization was detected by JC-1 staining,the cell injury was checked by LDH leakage,and the viability of MO3.13 oligodendrocytes was determined by MTT assay.RESULTS:Compared with DMSO model group,the demyelinating injury was alleviated and the proportion of apoptotic CC1+ oligodendrocytes in Mdivi-1 group was decreased.The cleaved caspase-3,caspase-9,cytochrome C and Bax protein expression levels in the spinal cord of Mdivi-1-treated mice was also attenuated.The in vitro MO3.13 cell experiments suggested that Mdivi-1 inhibited MO3.13 cell mitochondrial depolarization,attenuated the cell damage and increased the cell viability.CONCLUSION:Mdivi-1 pro-tects against the myelin injury in EAE mice,which may be related to the suppression of oligodendrocyte apoptosis.
8.Molecular Mechanism Analysis of Jiangtang Xiaozhi Tablets in Treatment of NAFLD
Min HOU ; Wen-jing GAO ; Yang DU ; Pan WANG ; Ju-qin PENG ; Ya-dong LIN ; Fu-zhi ZHANG ; Jun-guo REN ; Jian-xun LIU
Chinese Journal of Experimental Traditional Medical Formulae 2021;27(5):147-157
Objective:To explore the molecular mechanism of Jiangtang Xiaozhi tablets （JTXZT） in the treatment of non-alcoholic fatty liver disease （NAFLD） by means of network pharmacology and molecular docking. Method:With the help of traditional Chinese medicine （TCM） Systems Pharmacology Database and Analysis Platform （TCMSP）， TCMs Integrated Database （TCMID）， Encyclopedia of TCM （ETCM） and Bioinformatics Analysis Tool for Molecular Mechanism of TCM （BATMAN-TCM）， the chemical compositions of medicinal materials in JTXZT were obtained， the compound targets were predicted in SwissTargetPrediction database and STITCH database. The targets of NAFLD were searched by The Human Gene Database （GeneCards）， Online Mendelian Inheritance in Man （OMIM）， Therapeutic Target Database （TTD） and DisGeNET， and intersection analysis was performed with the targets of the active ingredients to obtain the targets of JTXZT for treatment of NAFLD. Based on STRING 11.0 database， the protein-protein interaction （PPI） network of therapeutic targets was constructed， and the enrichment analysis of therapeutic targets was carried out by DAVID 6.8. Finally， the interaction characteristics of key components and core therapeutic targets of JTXZT for treatment of NAFLD were verified based on molecular docking. Result:The key components of JTXZT for treatment of NAFLD were quercetin， luteolin， kaempferol， berberine， isorhamnetin， betulinic acid， oleanolic acid， ursolic acid. formononetin and hexitol， and the core targets of JTXZT for treatment of NAFLD were mitogen-activated protein kinase 1 （MAPK1）， Jun proto-oncogene， activator protein-1 （AP-1） transcription factor subunit （JUN）， MAPK3， protein kinase B1 （AKT1 or Akt1）， tumor protein p53 （TP53）， E1A binding protein p300 （EP300）， Fos proto-oncogene， AP-1 transcription factor subunit （FOS）， tumor necrosis factor （TNF），amyloid beta precursor protein （APP） and cytochrome P450 family 2 subfamily E member 1 （CYP2E1）. Biological function and pathway enrichment analysis showed that JTXZT mainly through xenobiotic metabolic process， oxidation-reduction process， cholesterol metabolic process and other biological processes， regulating phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt） signaling pathway， MAPK signaling pathway， NAFLD and insulin signaling pathway to play a role in the treatment of NAFLD. The results of molecular docking showed that the active components of JTXZT had a good affinity with the core targets of JTXZT for the treatment of NAFLD. Conclusion:JTXZT treats NAFLD through multiple active components， multiple key targets and multiple action pathways.
9.A network pharmacology approach to explore mechanisms of Buyang Huanwu decoction for treatment of cerebral infarction.
Nan LIU ; Yun-Yao JIANG ; Ting-Ting HUANG ; Jin-Cai HOU ; Jian-Xun LIU
China Journal of Chinese Materia Medica 2018;43(11):2190-2198
The point of this study is to explore and investigate mechanisms of Buyang Huanwu decoction for treatment of cerebral infarction (CI) using a network pharmacology approach. First, TCMSP database, DrugBank database and PharmMapper server were used and combined with oral bioavailability and drug analysis to screen the components of Buyang Hanwu decoction and predict the potential targets. Then, Cytoscape 3.5.1 software was used to construct compounds-targets network and the protein-protein interaction (PPI) networks for targets of compounds and CI-related targets and merge the two PPI networks to acquire active targets. Finally, gene ontology (GO) and KEGG pathway analysis of active targets were carried out by DAVID online analysis tool and KOBAS 3.0 software. In total of 150 screened compounds and 232 potential targets were obtained. And in total of 208 active targets were finally determined by merging networks. Results indicated that Buyang Huanwu decoction might have a role in treating CI by regulating some biological processes including response to drug, aging, response to hypoxia, and blood coagulation, and some molecular function, such as protein binding, enzyme binding and serine-type endopeptidase activity. The mechanisms might be concerned with PI3K-Akt signaling pathway, TNF signaling pathway, HIF-1 signaling pathway and cAMP signaling pathway. Among them, the regulation of PI3K-Akt signaling pathway might be one of the most crucial mechanisms.
10.Compatibility of geniposide and ginsenoside rgl: their regulating roles in secretion of anoxia induction injured microglia inflammatory cytokines.
Jun WANG ; Jin-Cai HOU ; Li-Hua XIANG ; Jie ZHANG ; Da-Hong JU
Chinese Journal of Integrated Traditional and Western Medicine 2014;34(1):91-95
OBJECTIVETo clarify the protective roles of compatibility of geniposide and ginsenoside (Rg1) in regulating ischemia injured microglia homeostasis by comparing the difference in regulatory roles of geniposide, Rg1, or ginsenoside + Rg1 in balancing secretion of oxygen glucose deprivation induced microglia inflammatory cytokines.
METHODSThe mimic ischemia injured microglia model was induced by oxygen-glucose deprivation (OGD). Then geniposide, Rg1, or ginsenoside + Rg1 (Tongluo Jiunao Injection, TJI) was respectively added. The NO content was determined by Griess Reagent. The cyto activity was detected using cell count kit. Contents of TNF-alpha and TGF-beta and their expression levels were detected by ELISA and Western blot.
RESULTSGeniposide + Rg1 could significantly inhibit the release of NO, elevate the TGF-beta level, and decrease the content of TNF-alpha without influencing the cell survival. The two active ingredients played different therapeutic roles. The compatible use was obviously superior to use any one of the two active ingredients alone.
CONCLUSIONSGeniposide, Rg1, or Ginsenoside + Rg1 had regulating roles in balancing ischemia injured microglia homeostasis. Its mechanisms might be related to up-regulating the TGF-beta expression and down-regulating TNF-alpha expression.
Animals ; Ginsenosides ; pharmacology ; Hypoxia ; metabolism ; Iridoids ; pharmacology ; Mice ; Microglia ; drug effects ; metabolism ; Nitric Oxide ; metabolism ; Transforming Growth Factor beta ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism
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