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Objective: To investigate the activation of hepatic stellate cells (HSCs) cocultured with hepatoma carcinoma cells. Methods: MHCC97H cells and HSCs were cocultured by cell-cell contact method, fibrinogen-thrombin paste technique, conditioned media from MHCC97H(MHCC97H-CM) and Transwell coculture technique, respectively. MHCC97H cells and HSCs were inoculated s. c. into nude mice. The expression of α-SMA in HSCs was assessed by immunocytochemical staining and Western blotting. Proliferation and migration of HSCs was determined using Cell-Counting Kit-8 (CCK-8) and wound healing and Transwell technique, respectively. Results: The activation of HSCs was significantly increased in the all cocultured system. The expression of α-SMA was up-regulated by cell-cell contact method, MHCC97H-CM, Transwell coculture technique in vitro and in cancer-bearing mice in vivo. The increased chemotaxis of MHCC97H cells and HSCs was observed by cell-cell contact method, fibrinogen-thrombin paste technique and in cancer-bearing mice. The proliferation and migration abilities of HSCs were significantly enhanced. Conclusion: Hepatoma carcinoma cells can promote the activation, proliferation and migration of HSCs under the cocultured system in vitro and in vivo.
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Objective:To investigate the anticancer activity and mechanism of aconitine on cell proliferation,invasion and migration of hepatoma carcinoma cell(HCC).Methods:The effect of aconitine at different concentrations on proliferation was calculated by MTT assay.The effects of aconitine on invasion and migration of HCC were measured by Transwell and wound healing assay.Western blot was employed to detect the protein levels of P38MAPK signaling pathway-related proteins.Results:The concentrations of 5,10,20 μg/ml were selected according to the results of pre-experiment.Aconitine(10,20 μg/ml) inhibits proliferation and invasion of MHCC97 cells markedly after cells were treated with aconitine for 4 days.Treatment with aconitine down-regulated the ability of migration and decreased the ratio of p-P38/P38 and protein levels of p-MAPKAPK and p-HSP27.Conclusion:Aconitine inhibits prolif-eration,invasion and migration,and the mechanism may related with P38MAPK signaling pathway.
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Objective To investigate the effect of magnetic nanocomposites on proliferation ability of human hepatoma carcinoma (HCC)cells (HepG2 cell line).Methods Leucine-rich repeat-containing G protein-coupled receptor (LGR) 5-small interfering ribonucleic acid (siRNA ) was composited with polyethylenimine wrapped superparamagnetic iron oxide nanoparticle (PEI-SPIO)as the gene vector.PEI group was established by transfecting HepG2 cells when cell fusion reached 60% and SI group was established by transfecting HepG2 cells with equivalent simple LGR5-siRNA.Control (Ctrl)group was also established without transfecting.The efficiency of nanocomposites entering cells was scanned with MRI T2.The inhibition rate of cell proliferation was detected by (cell count kit,CCK)-8 assay.The expression level of messenger ribonucleic acid (mRNA)in LGR5 of cells was detected by reverse transcriptase polymerase chain reaction (RT-PCR)and the protein expressions of LGR5 and cyclin D1 were detected by western blotting.Results MRI T2 signal of HepG2 cells in PEI group decreased significantly.Compared with Ctrl group,the inhibition rate of cell proliferation of HepG2 cells in PEI group was significantly increased.The relative expression of LGR5 mRNA and the relative expression of LGR5 and cyclin D1 protein were both significantly decreased (all in P <0.05),while the corresponding indexes of the cells in SI group had no statistical significance (all in P>0.05 ).Conclusions Magnetic nanocomposites PEI-SPIO composited with LGR5-siRNA may effectively transfect HepG2 cells.Its mechanism may take effect through down-regulating the expression of cyclin D1 to inhibit the proliferation ability of hepatocellular carcinoma HepG2 cells.
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Objective: To investigate the antitumor effect of volatile oil from Sinapis Albae Semen (VOSAS) on H22-bearing mice and to determine the mechanism. Methods: To establish the H22 implanted hepatocellular carcinoma animal model which was used to analyze the effect of VOSAS on the growth of transplanted tumor. Mice were divided into five groups 24 h after modeling: model, cytoxan (CTX, 25 mg/kg) positive control, low-, mid-, and high-dose (20, 40, and 80 mg/kg) VOSAS groups. The mice were ip administered once daily for 10 d. Morphological changes in H22 solid tumor cells were observed by both Hematoxylin-eosin (HE) and acridine orange (AO) staining. The expression of Bax and Bcl-2 in the tumor tissue was determined using immunohistochemistry. Results: VOSAS could inhibit the tumor growth and extend the life span of H22-bearing mice (P < 0.01); and it could also raise the expression of Bax while suppress the expression of Bcl-2; the antitumor effect of VOSAS on H22-bearing mice demonstrated a good dose-effect relationship, but the high-dose group of the volatile oil has obvious toxicity and side effects on the mice. Conclusion: VOSAS could inhibit the growth of H22 tumor cells and the mechanism may be related to up-regulating the expression of Bax and down-regulating the expression of Bcl-2, and the induction of apoptosis.
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Objective To study the influences by a Cyelin-dependent kinase inhibitor Roscovitine on cell cycle in mitotic hepatoma carcinoma cell SMMC-7721. Methods Microscope,MTT, flow cytometry and R-T PCR were used to observe the effects of Roscovitine on morphology, proliferation, cell cycle, apoptosis and the mRNA expression of CDK2, Caspase-3, bcl-2 in SMMC-7721 cells. Results Roscovitine inhibited the proliferation of SMMC-7721 cells in dosage and time dependent manner and induced apoptosis. Flow cytometry showed the ratio of G0, G1 increased. R-T PCR showed that the expression of bcl-2 reduced, Caspase-3 increased. Conclusion Reseovitine can inhibit the growth, proliferation, block the cell cycle at G0/G1 and promotes apoptosis of mitotic SMMC-7721 cells, and the mechanism of apoptosis is dependent on the activity of bcl-2 and Caspase-3.
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Objective:To study the influence of GPC3 gene on the proliferation,adhesion and invasion of hepatoma cell line SK-Hep-1.Methods:SK-Hep-1 cells were transfected with pEGFP-N2-GPC3 using Lipofectamine2000.RT-PCR was used to examine the GPC3mRNA expression in GPC3-SK-Hep-1 cells.MTT assay was used to examine the proliferation and calculate the adhesion rate of SK-Hep-1 cells.Transwell system was used to assess the migration and invasion of the cells.Results:SK-Hep-1 cells were successfully transfected with pEGFP-N2-GPC palsmid.GPC3 mRNA was detected in SK-Hep-1 cells.Transfection with CPC3 significantly suppressed the growth of SK-Hep-1 cells(P
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Objective To construct a RhoC-siRNA expression vector, and study its role on the biological behaviors of hepatoma carcinoma cells. Methods RhoC-siRNA gene was synthesized and cloned into the expression vector pSilencer2.1. The constructed RhoC-siRNA expression vector was stably transfected into hepatoma carcinoma cell line SK-Hep1. The inhibitory effect of RhoC-siRNA on the expression of RhoC in transfected cells was detected by Western blotting. The morphous, growth velocity and the ability of cell adhesion, cell migration, cell invasion before and after transfection was observed. Results Enzyme digestion and DNA sequencing confirmed that the RhoC specific siRNA expression vector was constructed successfully. After transfection, RhoC expression was inhibited by 60%, and no marked difference was observed in cellular morphous and growth curve, while the ability of cell adhesion, cell migration, and cell invasion were markedly decreased. Conclusion The RhoC-siRNA expression vector can effectively suppress RhoC expression in transfected hepatoma carcinoma cells. Although having no effect on the morphous and growth velocity of hepatoma carcinoma cells, it decreases the potentiality of cell invasion and cell metastasis, which may provide a novel applicable strategy for gene therapy on hepatocellular carcinoma.
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Objective To observe the effect of rutin on growth and proliferation of human hepatic cancer line(HepG2).Methods HepG2 cells were cultured in vitro,then cocultured with 50 to 250 ?mol/L rutin for 24 h.The inhibition rate of rutin on growth and proliferation of HepG2 was determined by MTT,~(3)H-TdR,and apoptotic cells were observed in fluorescent staining by Olympus fluorescent microscopy,and cell cycle was analysed by flow cytometry.Results Rutin inhibited HepG2 cells from growth and proliferation,and evoked apoptosis.Flow cytometry showed that 50 to 250 ?mol/L rutin caused an increase at G_(0)/G_(1) phase and a decrease at G_(2)/M phase and arrest at G_(0)/G_(1) phase in the cell cycle.Conclusion Rutin markedly inhibits the proliferation of HepG2 cells and induces apoptosis in a concentration-dependent manner.
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OBJECTIVE:To investigate the anti-HBV activities of deoxynojirimycin(DNJ)derivatives N—benzyl—1—DNJ(P-DNJ)and N—nonyl—1—DNJ(NN-DNJ)in vitro.METHODS:Human hepatoma carcinoma cell lines HepG2 2,2,15,which were transfected from HBV DNA,were taken as target cells,cells were cultured with different concentrations of test drugs, with HBsAg,HBeAg and HBV DNA in the cultured supernatant determined by time-resolved immunofluorometric assay and fluorescent quantitation PCR assay on day 6th and 10th day,meanwhile,with the cytotoxicity of DNJ derivatives determined by MTT assay.RESULTS:No cytotoxic effects was noted when the test concentration of P-DNJ and NN-DNJ was within 5～125?g?mL-1,HBsAg and HBeAg levels and HBV DNA secretion decreased significantly at a concentration of 125?g?mL-1.CONCLUSION:P-DNJ and NN-DNJ showed anti-HBV activity in the in vitro cell culture experiment.