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ObjectiveTo study the possible molecular mechanism of baicalein （BAI）-mediated focal adhesion kinase （FAK） in the regulation of phosphatidylinositol 3-kinase/protein kinase B （PI3K/Akt） signaling pathway to inhibit the proliferation and migration of gastric cancer HGC-27 cells. MethodThe gastric epithelial GES-1 cells and gastric cancer HGC-27 cells were respectively treated with BAI （0， 5， 15， 25， and 50 μmol·L-1） for 48 h， and then methyl thiazolyl tetrazolium （MTT） assay was adopted to detect effect of BAI on cell proliferation. Western blot （WB） was employed to detect the expression of FAK and the proteins related to epithelial-mesenchymal transition （EMT） and PI3K signaling pathway after intervention with different concentrations of BAI. The HGC-27 cells stably overexpressing FAK were constructed with lentivirus-mediated transfection technique， and the transfection of FAK was detected through WB and green fluorescent protein （GFP）. The cells were divided into empty vector （NC） group， BAI group， FAK overexpression group， and BAI-treated FAK overexpression group， and cell proliferation activity was detected by MTT assay. The colony formation and cell migration were observed via colony formation assay and Transwell migration assay， respectively. The expression of proteins involved in EMT and PI3K signaling pathways were detected by Western blot. ResultCompared with the NC group， BAI （15， 25 and 50 μmol·L-1） inhibited the proliferation of HGC-27 cells in a dose-dependent manner （P<0.05， P<0.01） while did not affect that of GES-1 cells. BAI （5， 15 and 25 μmol·L-1） down-regulated the expression level of p-FAK （P<0.05， P<0.01）. Compared with NC group， FAK overexpression group showed up-regulated expression level of FAK in HGC-27 cells. The HGC-27 cells in both NC group and FAK overexpression group had green fluorescence. Compared with NC group， BAI inhibited the growth， colony formation， and migration， while FAK overexpression promoted those of HGC-27 cells. The treatment of FAK overexpression group with BAI inhibited the enhancement of cell proliferation and migration （P<0.05）. WB showed that compared with NC group， BAI （15， 25 μmol·L-1） significantly up-regulated the expression of E-cadherin protein and down-regulated that of Vimentin， Snail， p-PI3K， and p-Akt protein in HGC-27 cells （P<0.05， P<0.01）. Compared with NC group， FAK overexpression group showed down-regulated expression of E-cadherin， up-regulated expression of p-FAK， Vimentin， and Snail， and increased ratios of p-FAK/FAK， p-PI3K/PI3K and p-Akt/Akt （P<0.05）. This phenomenon would be reversed after BAI treatment. ConclusionBAI can affect the proliferation and migration of gastric cancer HGC-27 cells by mediating FAK to regulate PI3K/Akt signaling pathway.
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In this experiment, ultra high performance liquid chromatography-quadrupole time-of-flight mass spectrometry(UPLC-Q-TOF-MS) was used to analyze and identify chemical constituents of Ginseng-Douchi(GD) compound fermentation, and explore the conversion rules of ginsenosides and soybean isoflavones after compound fermentation. Waters Acquity UPLC BEH C_(18) column(2.1 mm×100 mm, 1.7 μm) was adopted, with 0.1% formic acid aqueous solution(A)-0.1% formic acid acetonitrile solution(B) as mobile phase for gradient elution; electrospray ion source(ESI) was used to collect data in positive and negative ion modes; according to the exact mass number, the secondary spectrum comparison of the database and the existing literature reports, Peakview 2.0/masterview 1.0 software was used to determine the common ion structure formula. Finally, a total of 133 chemical constituents were analyzed and identified from the GD. Ginseng saponins and isoflavone glycosides were significantly converted after fermentation. Among them, peak areas of prototype ginsenosides Rk_3, Rh_1, Rh_2, Rh_3, daidzin, glycitin and genistin decreased significantly; whereas peak areas of se-condary ginsenoside Rb_1, Rb_2, Rk_1, glycitein, genistein and daidzein increased significantly. In this experiment, liquid-mass spectrometry technique was used to investigate the conversion of active ingredients of GD compound fermented products after co-fermentation, so as to provide a scientific basis for elucidating pharmacodynamics material basis and quality control.
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Chromatography, High Pressure Liquid , Drugs, Chinese Herbal , Fermentation , Panax , Tandem Mass SpectrometryABSTRACT
Chronic unpredicted mild stress(CUMS) combined with isolated feeding was used to induce depressed rat model. The anti-depressant effects of Zhizichi Decoction(ZZCD) and its solid fermented product(ZZC) were analyzed by behavioral test and comparison of pathological tissues of hippocampus and liver, metabolic characteristics of intestinal flora, and relative abundance of species. The results showed that ZZC could increase sucrose preference, shorten the immobility time in the forced swim test and tail suspension test(P<0.05), and repair damaged hippocampus and liver tissues, and the effect was superior to that of ZZCD. The results of Biolog ECO plates showed that the average well color development(AWCD) of intestinal flora in the model group significantly decreased and the metabolic levels of sugar and amino acids were reduced, while the AWCD of the treatment groups increased. The metabolic levels of the two carbon sources were improved in the ZZC group, while only sugar metabolic level was elevated in the ZZCD group. Metagenomic analysis of intestinal flora showed that the ratio of Firmicutes/Bacteroidetes was 3.87 in the control group, 21.77 in the model group, 5.91 in the ZZC group, and 18.48 in the ZZCD group. Lactobacillus increased by 3.28 times, and Prevotella and Bacteroidetes decreased by 75.59% and 76.39%, respectively in the model group as compared with that in the control group. Lactobacillus decreased by 31.13%, and Prevotella and Bacteroidetes increased by more than three times in the ZZC group as compared with that in the model group, while the corresponding changes in the ZZCD group were not significant. ZZC could improve depression-like beha-viors by regulating the structure of intestinal flora and metabolic functions and repairing damaged hippocampus and liver tissues in depressed rats, showing an anti-depressant effect superior to that of ZZCD. This study is expected to provide a basis for the development of new anti-depressant food products.
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Animals , Rats , Depression/drug therapy , Disease Models, Animal , Fermentation , Gastrointestinal Microbiome , Hippocampus , Stress, PsychologicalABSTRACT
Objective: To investigate the effect of taraxerol on autophagy of breast cancer MCF-7 cells in vitro, and explore the related mechanisms. Method: The effect of various doses of taraxerol (12.5, 25, 50, 100, 200 μmol·L-1) on proliferation of MCF-7 cells was detected by methye thiazolye telrazlium (MTT) assay. The autophagy-inducing effect of taraxerol was observed by acridine orange staining, transmission electron microscope (TEM) and immunofluorescence. The expressions of autophagy-related proteins and the changes of mammalian target of rapamycin (mTOR) signaling pathway were determined by Western blot analysis. Result: The viability of MCF-7 cells was significantly inhibited by taraxerol. Acridine orange staining indicated that the acidic lysosomes increased significantly after treatment with taraxerol in MCF-7 cells. The autophagic structure in the treated group was observed by TEM. Immunofluorescence showed that the expression of microtubule-associated protein 1 light chain 3 (LC3) in the cells of the drug group was increased. Western blot demonstrated that the protein expressions of LC3-Ⅱ and Beclin-1 were increased in taraxerol-treated MCF-7 cells (PP-1 taraxerol group, combination group (taraxerol + 3-methyladenine, 3-MA) showed the down-regulation of LC3-Ⅱ in the MCF-7 cells (PPPConclusion: Taraxerol can induce autophagy in MCF-7 cells, which may be related to the inhibition of mTOR signaling pathway.
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AIM: To investigate the effects of baicalein(BAI)on the proliferation and migration of gastric cancer MGC-803 cells and the mechanisms.METHODS:After MGC-803 cells were treated with BAI at different concen-trations,the viability of the MGC-803 cells was tested by MTT assay.The cell colony formation ability were detected by plate colony formation assay.Wound-healing and Transwell cell migration assays were used to test the migration ability of the MGC-803 cells.The concentration of 12-hydroxyeicosatetraenoic acid(12-HETE)was measured by ELISA.The pro-tein levels of platelet type 12-lipoxygenase(p12-LOX),vascular endothelial growth factor(VEGF),p-ezrin and epithelial-mesenchymal transition(EMT)markers in MGC-803 cells were determined by Western blot.RESULTS:BAI significantly inhibited the proliferation,plate colony formation and migration abilities of the MGC-803 cells(P<0.05 or P<0.01), down-regulated the concentration of p12-LOX metabolite 12-HETE significantly(P<0.05 or P<0.01), decreased the protein levels of p12-LOX,VEGF,p-ezrin,vimentin and Snail(P<0.05 or P<0.01),and increased the protein expres-sion of E-cadherin(P<0.01).CONCLUSION:BAI suppresses the proliferation and migration abilities of gastric cancer MGC-803 cells effectively.These effects of BAI may be related to regulating the protein levels of p12-LOX,VEGF,p-ezrin and EMT-related proteins.
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In this study,the content of ethanol extraction of agarwood were performed following the method of Chinese Pharmacopoeia(ChP 2015 edition). The chromatographic fingerprints were established by GC-MS. Similarity Evaluation System for Chromatographic Fingerprint of traditional Chinese medicine（TCM）(version 2012) was employed to calculate the similarity of each chromatogram of agarwood. The ratios of sum peak area in the range of 170-270 min and 0-100 min of individual chromatogram were calculated using square peaks to normalization. AMDIS and RI were employed to identify the common and different peaks. Correlation coefficient P(corr) combined with Variable important in projection(VIP) value was employed to screen the different representative components based on OPLS-DA analysis. Grey related degree and TOPSIS were used to evaluate the quality of artificial agarwoods. The results showed that more than 10.0% of the ethanol extract content was found in 15 batches of artificial agarwoods among the total 18. The similarity of 18 batches artificial agarwoods was 0.439-0.779. The peak area ratios of two intervals were in the range of 0.307-13.254. The 9 common components and 8 different components were identified. Meanwhile, 2% salicylic acid is the best inducer based on grey related degree and TOPSIS. Grey related degree and TOPSIS can be used to evaluate the quality of artificial agarwoods rapidly. These results provide a reference data to evaluate the qualityof artificial agarwood.
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OBJECTIVE@#To investigate the effect of ischemic precondition to protect ischemia-reperfusion injury and reduce IL-6 expression in the rats liver transplantation.@*METHODS@#The rat portal vein infusion of autologous liver transplantation model were used. The rats were divided into ischemic preconditioning rats liver transplantation group (A group), the rats liver transplantation group (B group) and the normal rat control group (C group). Then we analyzed the changes of liver function, liver microstructure and the expression of IL-6, SOD and MDA within 48 h.@*RESULTS@#The pathology of liver in group A showed lobular architecture essentially normal, the liver cells was slightly swell and no significant changes in postoperative 12 h. In transmission electron microscope (46 000×), the mitochondria of liver cells in group A became swelling, elliptical can cristae partially broken. But there still has a small amount of arrangement. While that in group, the mitochondria were swollen, became round, serious visible crest reduce or ruptured. The result of over function test showed that the serum ALT and AST levels in group A and B were both higher than that in group C at each time period, but the serum ALT and AST levels in group A were lower than that in group B. The expression changes of IL-6 in group B were higher than that in group A and B (P<0.05). The expression of MDA in group A is more obvious than that in group B (P<0.05)@*CONCLUSIONS@#Ischemic precondition could alleviate part of ischemia-reperfusion injury in the rat liver transplantation, and also could reduce IL-6 expression to protect the liver cells against liver damage and inflammatory cytokine production.
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Animals , Female , Male , Rats , Disease Models, Animal , Hepatocytes , Pathology , Histocytochemistry , Interleukin-6 , Metabolism , Ischemic Preconditioning , Methods , Liver , Cell Biology , Metabolism , Pathology , Liver Transplantation , Methods , Malondialdehyde , Metabolism , Mitochondria, Liver , Metabolism , Pathology , Rats, Sprague-Dawley , Reperfusion Injury , Therapeutics , Superoxide Dismutase , MetabolismABSTRACT
Objective To study the effects of oxymatrine as inhibitor of hemorrhagic fever with renal syndrome virus (HFRSV) infection in vitro and in vivo.Methods In vitro studies,a dose of oxymatrine without cytotoxicity and 76-118 strain of HFRSV was taken to treat Vero cells in three ways:①After treated with oxymatrine for 48 h,Vero cells were attacked by HFRSV at dilution of 10-1 ～ 10-6,respectively for 24 h before changing to maintenance medium; ②Vero cells were first attacked by HFRSV of 10-1 ～ 10-6 dilution respectively,then oxymatrine was used for 48 h before changing to maintenance medium; ③Vero cells were attacked by HFRSV at dilution of 10-1 ～ 10-6 respectively,and meanwhile treated with oxymatrine for 48 h before changing to maintenance mcdium.Each dilution handled four porocytes,and four positive controls were set up at the same time.Indirect immunofluorescence assay (IFA) was performed to determine the inhibitory effect of oxymatrine in experimental group and positive control.In vivo studies,thirty 2-week-old hamsters,weighing about 30-40 g,were divided into experimental and control groups according to body weight,n =15.These aninals were inoculated intraperitoneally with HFRSV in 100TCID50(0.1 ml each); on days 4-13,0.1 ml of oxymatrine 1:100 were given to each hamster in experimental group daily by intraperitoneal injection,while the same amount of saline was given to the control ones.Lung tissue of hamsters was then dissected out to slice to be identified by immunofluorcscence stain.Results It was demonstrated that oxymatrine with the diluted fractions of 1:8 was safe in vitro.When the virus dilution of HFRSV was l0-4,compared with control groups,the differences were statistically significant in method 2 and 3 (z =-2.53,-2.53,all P ＜ 0.05),while no statistical significance in method 1 (z＝5.36,P＞ 0.05).When the virus dilution of HFRSV was 10-1 ～ 10-3,10-5,10-6,the differences were not statistically significant (z--0.00,-0.32,-0.19,4.21,4.21,all P ＞ 0.05).In vivo studies,compared with control group,the differences were statistically significant in experimental group (z =-3.85,P ＜ 0.05).Conclusion Oxymatrine significantly inhibites HFRSV.
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<p><b>OBJECTIVE</b>To investigate the minimally-invasive ablation of osteomas of the ethmoid sinuses endonasally.</p><p><b>METHODS</b>A retrospective analysis was done in 19 patients (15 male, 4 female, aged between 14 - 67, medium 37) diagnosed as osteomas of ethmoid sinuses hospitalized from April 2005 to October 2009. All patients underwent sixteen-detector row computed tomography scan and 3D reconstruction preoperatively. All underwent operation with the help of navigation system and nasal endoscope.</p><p><b>RESULTS</b>The ethmoid osteoma in all 19 patients was removed successfully with endoscope and navigation system. Two open procedures (1 through superciliary arch incision and 1 through labiogingival incision) were performed to assist the removal of the tumor, 17 tumors were removed under endoscopic and navigation guidance. In 5 patients whose osteoma was localized or with the diameter no more than 2 cm, these osteomas were removed endonasally with the help of navigation system. The osteomas in 2 patients was found to have narrow basilar part and relatively dissociative were removed from oral cavity after abscisin the basilar part. The osteomas in 12 patients were found to have basilar part connected with ante-meso skull base, lamina papyracea, orbital apex, cranialis opticus, fossa orbitalis bone, these osteomas were removed using electric drill with the guidance of navigation system. All patients were followed up from 8 to 64 months, and were asymptomatic (1 patient who suffered from amblyopia had the symptom disappeared, 2 patients suffered from prosopo-eminence, 1 patient was asymptomatic and 1 patient was feeling better). Two patients underwent removal of crista galli, 1 of them suffered from postoperative cerebro-spinal rhinorrhea, and recovered after endoscopic repairing procedure and iodoform gauze packing and recovered 15 days later. Two patients who underwent removal of crista galli suffered from anosmia and never recovered after 9 and 26 months follow-up. One patient with enormous osteoma suffered from repeated crusting and abnormal odor, and recovered after nasal flushing.</p><p><b>CONCLUSIONS</b>Endoscopic ablation of osteomas of the ethmoid sinuses with the guidance of navigation system is an accurate, secure, minimally-invasive procedure. Osteomas on median line and localized in ethmoid sinus is an indication of this operation. Preoperative CT scan is a safeguard for an accurate operation.</p>
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Adolescent , Adult , Aged , Female , Humans , Male , Middle Aged , Young Adult , Endoscopy , Ethmoid Sinus , General Surgery , Image Processing, Computer-Assisted , Osteoma , General Surgery , Paranasal Sinus Neoplasms , General Surgery , Retrospective Studies , Tomography, X-Ray ComputedABSTRACT
<p><b>OBJECTIVE</b>The objective of this research is to construct a clinic-usable genechip method for detection of hepatitis B virus lamivudine-resistant mutants and basal core promotor/Pre-C mutants, compare this method with DNA sequencing to investigate this genechip's character (sensity, specificity, stability and practicability in clinic) and apply it in clinic.</p><p><b>METHODS</b>This genechip detection method can detect the DNA and 8 mutative site of HBV, include 3 lamivudine-resistant mutation site(No. 180, 204, 207 site in DNA polymerase gene), 5 HBeAg escape-related mutation site (nt 1896, 1899, 1862, 1764,1762 site in BCP/Pre-C region).The results of genechip method was verified by DNA sequencing.</p><p><b>RESULTS</b>In detecting HBV DNA, the results of genechip were agree with 100% of the results of DNA sequencing. In detecting HBV mutants, 251 sites (in 32 samples, 256 sites) showed the same results using both methods, and only 5 sites were not completely match (P > 0.05). In these 5 sites, genechip methods got multi-infection results, but sequencing got single-infection results.</p><p><b>CONCLUSION</b>These results suggest that genechip method has the same positive rate and almost these same specificity with DNA sequencing method, and is better than DNA sequencing method in detecting multi-infected HBV strains. [Key words]</p>
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Humans , Antiviral Agents , Pharmacology , Base Sequence , Drug Resistance, Viral , Hepatitis B , Drug Therapy , Virology , Hepatitis B Core Antigens , Genetics , Hepatitis B virus , Genetics , Lamivudine , Pharmacology , Molecular Sequence Data , Mutation , Oligonucleotide Array Sequence Analysis , Methods , Promoter Regions, GeneticABSTRACT
Objective To evaluate the early outcomes of mini-open lumbar microdiscectomy. Methods There were 38 cases in each group of mini-open lumbar mierodiscectomy and conventional discecto- my.Operating time,blood loss,time of leaving the bed and length of hospital stay were compared in two groups.MacNab criteria were used to evaluate the outcomes.Results To compare the conventional discec- tomy group,microdiscectomy group spent similar operating time,but had less blood loss(P
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<p><b>OBJECTIVE</b>Apoptosis of the cells of liver cancer cell line HepG2 could be induced by TNF alpha and actinomycin D (Act D). In the current study, the molecular mechanism of the apoptosis protection of stronger neo-minophagen C (SNMC) to HepG2 cells was investigated.</p><p><b>METHODS</b>SNMC was added to the HepG2 cell culture medium when the cell concentration reached 0, 2, 20, 100, 200, 800 microg/ml 30 min before their apoptosis were inducted with TNF alpha and Act D. A flow cytometry assay was performed to detect the cell apoptosis rate; electromicroscopy was employed to visualize the subcellular structure after apoptosis. DNA ladder formation was checked with genomic DNA agarose electrophoresis. The expression pattern of apoptosis related protein Caspase-3, Bcl-2 and Bax was detected by Western blot.</p><p><b>RESULTS</b>After pretreatment with various concentrations of SNMC and 12 hours after treatment with TNF alpha and Act D, the HepG2 cell apoptosis rate and DNA ladder formation decreased dramatically when the SNMC concentration was higher in the media; the intracellular inactive form of Caspase-3 increased while the 17*10(3) active Caspase-3 decreased gradually. In addition, the expression of Bcl-2 increased and the expression of Bax decreased. Under the electromicroscope, the typical nucleolus condensation of HepG2 induced by TNF alpha and Act D was not seen among the 100 microg/ml SNMC treated cells.</p><p><b>CONCLUSION</b>SNMC inhibits TNF alpha and Act D induced HepG2 cell apoptosis. This protective action may be regulated by intracellular apoptosis related factors.</p>
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Humans , Apoptosis , Carcinoma, Hepatocellular , Pathology , Cell Line, Tumor , Cysteine , Pharmacology , Drug Combinations , Glycine , Pharmacology , Glycyrrhiza , Liver Neoplasms , Pathology , Oleanolic Acid , PharmacologyABSTRACT
<p><b>OBJECTIVE</b>To investigate the protective effect of stronger neo-minophagen C (SNMC) on fulminant liver failure (FLF).</p><p><b>METHODS</b>D-Gal N and LPS were injected once into the abdominal cavity of rats to establish an experimental model of FLF. The level of plasma ALT, Alb, TBil, TNFalpha, NO, ET-1, IL-6 and liver histopathology of the rats were examined.</p><p><b>RESULTS</b>In the D-Gal N and LPS model of FLF, there was an obvious decline of plasma TNFalpha (F = 52.84), NO (F = 15.81), ET-1 (F = 15.68), IL-6 (F = 15.32) and there was less hepatic tissue damage in SNMC-treated groups using different doses (high dose, medium dose, low dose) and at different times (pre-protection, simultaneous protection, post-protection) compared with those not treated with SNMC. These results indicated that SNMC could be used to treat FLF. It was better to use a low dose of SNMC and use it at the same time as inducing the FLF. There were no differences in the results of those treated with SNMC of different dosages and treated at different times.</p><p><b>CONCLUSION</b>SNMC can decrease the mortality of FLF by preventing hepatocyte apoptosis induced by D-Gal N and LPS and inhibit liver inflammation caused by all kinds of factors.</p>
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Animals , Female , Male , Mice , Anti-Inflammatory Agents, Non-Steroidal , Therapeutic Uses , Galactosamine , Glycyrrhizic Acid , Therapeutic Uses , Lipopolysaccharides , Liver Failure, Acute , Drug TherapyABSTRACT
<p><b>OBJECTIVE</b>Lamivudine resistant HBV strains in Shenzhen were detected at multiple sites and in large amounts to understand further the distribution of lamivudine resistant mutants.</p><p><b>METHODS</b>552 Hepatitis B patients's sera were examined using genechip method. Among them, 192 samples of lamivudine resistant mutant were further analyzed.</p><p><b>RESULTS</b>In those 192 lamivudine resistant samples, 191 were YMDD mutants, 124 mutants of codon 528 and 9 mutants of codon 555. 88% YMDD mutants were multi-mutants of YVDD and codon 528; single mutants of YIDD; multi-mutants of YIDD and codon 528. 91% codon of YMDD mutants were GTG, ATT; the other 9% were ATA, ATC.</p><p><b>CONCLUSIONS</b>These results suggest that mutants of codon 552 (YMDD) are core mutants. Mutants of codon 528 and 555 are incidental mutants, YVDD mutants always emerge with mutants of codon 528, but YIDD mutants appear differently. 9% YMDD mutants's codons are ATA or ATC. This may be the reason for the low positive rate shown by using the conventional PCR methods.</p>
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Humans , Amino Acid Motifs , Antiviral Agents , Pharmacology , Therapeutic Uses , Codon , Genetics , DNA-Directed DNA Polymerase , Genetics , Drug Resistance, Microbial , Genetics , Hepatitis B virus , Genetics , Hepatitis B, Chronic , Drug Therapy , Virology , Lamivudine , Pharmacology , Therapeutic Uses , Oligonucleotide Array Sequence Analysis , Point MutationABSTRACT
<p><b>BACKGROUND</b>To establish a genechip method for detection of hepatitis B virus (HBV) DNA, basal core promotor (BCP), and Pre-C mutants.</p><p><b>METHODS</b>This study used two kinds of technology (PCR, oligochip), which can detect five mutant hotspots including nt 1 896, nt 1 899, nt 1 862, nt 1 764 and nt 1 762. The results of genechip method was verified by DNA sequencing.</p><p><b>RESULTS</b>In detecting HBV DNA, the results of genechip were 100% consistent with those of DNA sequencing. In detecting HBV BCP and Pre-C mutants, 146 codons showed the same results using both methods, except for only 4 codons (P greater than 0.05).</p><p><b>CONCLUSION</b>This convenient high throughput genechip method could detect several BCP and Pre-C mutant codons at the same time. These results suggest that genechip method has the same positive rate and specificity with DNA sequencing method. It has more advantages than the latter in detecting mixed mutants and therefore may be used in clinical practice.</p>