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OBJECTIVE：To ex plore the effects of solamargine on the growth and apoptosis of human hepatocarcinoma cells HepG2 and its underlying mechanism. METHODS ：The effects of 0（blank group ）-12 μmol/L solamargine treatment of 24，48 h on survival rate of HepG 2 cells were investigated. The effects of 0（blank group ），6 μmol/L solamargine treatment of 10 days on cell clone formation were also investigated. The effects of 0（blank group ），4，6，8 μmol/L solamargine for 24 h on the apoptotic rate of cells，mRNA expression of Bcl- 2，Bax and caspase- 3， protein expression of Bcl- 2 and cleaved caspase- 3 as well as ratio of p-AMPKα to AMPKα were all tested. The effects of AMPK inhibitor as compound C on the protein expression of AMPKα and Bcl- 2 in cells were investigated after treated with 6 μmol/L solamargine for 24 h. RESULTS ：Compared with 020-39318678。E-mail：wujingjing6028@gzucm.edu.cn blank group ，1-12 μ mol/L solamargine for 24，48 h could significantly decrease the survival rates of cells （P＜0.05）in a concentration-dependent manner ；IC50 of them were 8.310 and 7.996 μmol/L，respectively；the rate of cell clone formation was decreased significantly after treated with 6 μmol/L solamargine for 10 days（P＜0.05）. The apoptotic rate of HepG 2 cells，mRNA expression of Bax and caspase- 3，protein expression of cleaved caspase-3（except for 8 μmol/L）as well as ratio of p-AMPKα to AMPKα（except for 8 μmol/L）were all increased significantly after treated with 6，8 μmol/L solamargine（P＜0.05）；mRNA and protein expression of Bcl- 2 were decreased significantly （P＜ 0.05）；the changes of some indexes were in a concentration-dependent manner. The compound C could inhibit protein expression of AMPKα，and reverse the inhibitory effect of solamargine on Bcl- 2 protein. CONCLUSIONS ：Solamargine can inhibit the proliferation of HepG 2 cells and induce apoptosis ，the mechanism of which may be associated with activating AMPK signaling pathway.
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OBJECTIVE:To study the effects of 3 extracts of Acanthopanax sessiliflorus fruits on the proliferation and apoptosis of human hepatocarcinoma cells SMMC-7721,and to provide reference for confirming the mechanism of anti-tumor effect. METHODS:MTT assay was adopted to investigate the effects of low-mass concentration,medium-mass concentration and high-mass concentration of ethanol extract(0.92,1.84,3.68 mg/mL),crude polysaccharide extract(0.06,0.12,0.24 mg/mL)and refined polysaccharide extract (0.04, 0.08, 0.16 mg/mL) from A. sessiliflorus fruits on the proliferation and apoptosis of SMMC-7721 cells after treated for 24,36,48 h,respectively. Flow cytometry was used to investigate the effects of 1.84 mg/mL ethanol extract,0.24 mg/mL crude polysaccharide extract and 0.16 mg/mL refined polysaccharide extract on cell cycle and cell apoptosis after treated for 24 h. The above tests were all negative control(only adding cells without drugs). RESULTS:Compared with negative control,3 extracts of A. sessiliflorus fruits could significantly inhibit the proliferation of SMMC-7721 cells (P<0.01),could significantly decrease the percentage of SMMC-7721 cells in G0/G1 and G2/M phase(P<0.01),could significantly increase the percentage of SMMC-7721 cells in S phase (P<0.01) and the apoptosis rate of SMMC-7721 cells (P<0.05);especially the effects of ethanol extract from A. sessiliflorus fruits were the most obvious. CONCLUSIONS:Three extracts of A.sessiliflorus fruits can inhibit the proliferation of human hepatocarcinoma SMMC-7721 cells,block SMMC-7721 cells in S phase and induce the apoptosis of SMMC-7721 cells.
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Objective:To analyze the effects of BANCR on proliferation,apoptosis,invasion and angiogenesis in human hepatocarcinoma cell line HepG2.Methods:The expression of BANCR was detected by quantitative real-time reverse transcription PCR (qRT-PCR).BANCR siRNA and Scramble was respectively transfected into human hepatocarcinoma cell line HepG2.Cell proliferation was detected by CCK-8.Flow cytometry was performed to analyze the apoptosis.Transwell assay was used to test the invasion.Angiogenesis was analyse by tube formation assay.Western blot was executed to check the expression of proliferating cell nuclear antigen (PCNA),caspase-3,matrix metalloprotein 9 (MMP-9),vascular endothelial growth factor(VEGF),acidic fibroblast growth factor (bFGF)and interferon-γ (IFN-γ).Results:The expression of BANCR in HepG2 was higher than L02 (P＜0.05).Compared with control group,the cell proliferation folds in BANCR siRNA was largely decreased.Besides,BANCR siRNA group had a higher apoptosis rate and less invasive cells (P＜0.05).Western blot showed that the expression level of caspase-3 and IFN-γwas obviously enhanced in BANCR siRNA group,and the expression of PCNA,MMP-9,Fn,Vimentin,VEGF and bFGF was distinctly surpressed in BANCR siRNA group compared to control group (P ＜ 0.05).Conclusion:siRNA interference of BANCR promotes apoptosis and represses proliferation,invasion and angiogenesis in human hepatocarcinoma cell line HepG2.
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Hepatitis C virus (HCV) is known to be a major cause of chronic hepatitis, liver cirrhosis and hepatocarcinoma. Therapeutic reagents are improving, but are still limited, and the protective vaccine against HCV is not available yet. However, the research of HCV life cycle and pathogenesis has been difficult due to obstacles, which are the lack of effective cell culture systems and small-animal models. Recently, breathtaking progress in terms of HCV replication system has been made using various forms of HCV clones and human hepatocarcinoma 7 cell lines (huh 7). The establishment of complete cell-culture system for HCV replication gave researchers opportunities to study the entire viral life cycle including entry, assembly, release of viral particles and the interaction with host cells. In fact, these efforts now appear to move into the identification and the development of innovative anti-HCV reagents. In this review, we go over the biological characters of HCV, a variety of in vitro cell culture, in vivo animal models of HCV infection, HCV immune-pathogenesis and the application of HCVcc system in terms of developing anti-HCV reagents.
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Humans , Cell Culture Techniques , Cell Line , Clone Cells , Hepacivirus , Hepatitis, Chronic , Indicators and Reagents , Life Cycle Stages , Liver Cirrhosis , Models, Animal , VirionABSTRACT
Objective To observe the role of human hepatocarcinoma HepG2 cell culture supernatant on T lymphocyte of the activation, proliferation, IL-2 secretion and inducing apoptosis in vitro. To explore the mechanism of immune escape in tumor.Methods Peripheral blood mononuclear cells (PBMC) were cultured with different concentration of tumor cell culture supernatant. Cell proliferation was examined with standard cytometry and MTT. The apoptotic rate and immune phenotype were analyzed by FACS. The level of IL-2 was determined with ELISA.Results Adding HepG2 supernatant, there was no significant influence in activation of lymphocyte, but the proliferation rat and the secretion of IL-2 were obviously decreased. The lymphocyte apoptosis rate was raised.Conclusion The tumor cell may produce a certain soluble protein, thus escape from the immune surveillance of tumor-bearing host.
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Objective To investigate the mechanism of Depression proliferation in human hepatocarcinoma cells by Abscisic acid.Methods To detect protein expression o of P53,Ki-67 and Cyclin D1 by immunocyte chemistry; detect mRNA expression of P53 and telomerase by RT-PCR. Results The protein expression level of mtP53, Cyclin D1, Ki-67 and the mRNA expression level of mtP53 and hTERT all decreased in cells treated by ABA,HMBA and ABA+ HMBA(P
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Objective To investingate the effect of 5-Aza2'-deoxycytidine(5-Aza-CdR)on cell growth and to explore the possibility of re-expression of the hypermethylated and silenced RUNX3 gene in hepatocarcinoma cell line HepG2.Methods The change in expression of the tumor suppressor gene RUNX3 mRNA in cultured HepG2 cells was observed by RT-PCR before and after 5-Aza-CdR treatment.Activity of cell growth was observed by MTT assay and colony-forming test.The cell cycle was analyzed by flow cytometry.Apoptotic morphology was observed by transmitting electron microscopy.Results The gene was reactivated by two different doses of 5-Aza-CdR treatment in HepG2 cell without expressing RUNX3.The hepatocarcinoma cell line treated with 5-Aza-CdR displayed a slowed growth rate in contrast to the control group.The colony formation rate of HepG2 cell treated with 5-Aza-CdR decreased dramatically(P
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Objective: To investigate the suppressive effects of nedaplatin on human liver cancer cell line SMMC7721 and the interactions between nedaplatin and adriamycin or mitomycin or fluorouracil. Methods: The cytotoxic index of nedaplatin alone or combined with other chemotherapy agents on SMMC cells were detected by MTT method. Results: SMMC was sensitive to nedaplatin with a positive correlation between cytotoxic index and nedaplatin concentration. There were significant synergism in the cytotoxic effects of nedaplatin combined with adriamycin or Mitomycin or Fluorouracil on SMMC cells. Conclusion: Nedapltin is a promising agent in the treatment of human liver cancer.
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Objective To study the mechanism of killing and apoptosis-inducing effects of Capparis spinosa alkaloid (CSA) on human hepatocarcinoma cell line HepG2. Methods The killing effect of CSA on human hepatocarcinoma cell line HepG2 was studied by MTT method. Morphological observation of HepG2 cells was carried out by fluorescence microscope. Results The CSA had obvious cytotoxicity on the HepG2 in a dose-dependent manner and its IC50 value was 142.82 ?g/mL. The HepG2 cells showed the characteristic morphologic changes of apoptosis by the function of CSA and the apoptosis percentage is higher than that of the natural one. The progress of cells cycle from S phase to G2 phase had been blocked by CSA. The intracellular Ca2+ level had been increased by the function of CSA, which was positively related with drug concentration. Conclusion CSA has obviously killing and apoptosis-inducing effects on human hepatocarcinoma cell line HepG2 and calcium overload might also be invovled in these events.
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Objective To investigate the method and optimum conditions of electric transfection,and the major influential factors of electransfection efficiency and the survival rate of dendritic cells. Methods RNA was extracted from human hepatocarcinoma cell line(Bel 7402).Purified monocytes as precursor DC-s(pDC-s) were separated from human peripheral blood cells(PBMCs) by density gradient centrifugalization with lymphocyte gradation fluid and adherence method,pDCs were incubated in RPMI-1640 medium containing rhGM-CSF(8?10~5IU/L) and rIL-4(5?10~5IU/L) for 7 days and made them fully differentiate into immature DCs(imDCs).The total RNA human hepatocarcinoma cell and green fluorescent protein(GFP) were electransfected respectively into imDCs by electroporation apparatus with different electric voltages,times of impulse,cell concentrations,temperatures and electroporation buffers.Numbers of green fluorescence positive cells and the total cell number were counted respectively under fluorescent microscope,and visible light microscope.One day after the electric transfection,the cells were stained with 0.4% trypan blue,and electransfection efficiency and the cell survival rate were counted. Results Electransfection efficiency was increased to the highest value,up to about 49.7% when imDCs with the concentration of 5?10~6 cells/ml were mixed with 40?g-total RNA of human hepatocarcinoma cell,the electric voltage of electroporation apparatus was set at 300V,and the time of impulse was 500Us.Conclusion Electric transfection provides a technical possibility to make human hepatocarcinoma RNA into imDCs.The major influential factors of the electransfection efficiency were electric voltage and impulsing time.As receptor cells,the imDCs growing condition was also an important influential factor.