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Aim To screen the differential microRNA (miRNA) expression profiles of different metastatic po-tential liver cancer cell lines,and predict miRNAs-reg-ulated target genes and their functions. Methods To-tal RNA was extracted and the miRNA expression pro-files were obtained by miRNA microarray chip hybrid-ization. The miRNAs whose expression had significant difference were selected by analyzing the miRNA difference expression profiles of the two different meta-static potential liver cancer cell lines, namely MHCC-97H(high-metastasis) and Hep3B(non-metastasis), which were compared with normal hepatocytes L02 re-spectively. Moreover, we analyzed the miRNA differ-ential expression profile between liver cancer cell lines MHCC-97H and Hep3B. The miRNAs were verified by qPCR and target genes were predicted by four softwares (TargetScan, miRanda, miRWalk, miRDB). To un-derstand the biological functions of predicted target genes, bioinformatics analysis was performed. Results The miRNA microarray results showed that the ex-pression of miR-192-5p and miR-215-5p significantly increased in liver cancer cell lines (MHCC-97H, Hep3B) when compared with normal hepatocytes L02, while miR-130a-3p and miR-196a-5p were significantly reduced; compared with Hep3B, the expression of miR-224-5p markedly increased in liver cancer cell line MHCC-97H, while miR-146a-5p, miR-483-3p and miR-200b-3p were significantly reduced. The re-sults of qRT-PCR were consistent with chip results. Conclusion There are differences of miRNA expres-sion profiles in different metastatic potential liver canc-er cell lines MHCC-97H, Hep3B, and they may par-ticipate in regulating the development and invasion of hepatocellular carcinoma.
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Background and purpose: The previous work of this study has showed that the treatment of liver cancer cells with emodin could induce endoplasmic reticulum (ER) stress and apoptosis. Given the cross-talk between ER stress and autophagy, this study aimed to investigate whether blockage of autophagy, a defense mechanism against environmental stress, could improve the killing effect of emodin on liver cancer cells. Methods: The CYTO-ID auto-phagy detection kit and Western blot were used to determine autophagy in liver cancer cells. After combined treatment with chloroquine (CQ) and emodin, cancer cell survival was analyzed by ATPlite assay and clonogenic assay. Apoptosis was detected by both flow cytometry analysis and Western blot. Results: Autophagy could be induced in liver cancer cells after treatment with emodin. Inhibition of autophagy significantly increased growth-inhibitory effect of emodin on both HepG2 and Huh7 cancer cells. The combination treatment with CQ and emodin promoted remarkable apoptosis in liver cancer cells, evidenced by the increase in the percentage of cells in sub-G1 phase and the higher expression lever of cleaved caspase-3. Conclusion: Therapeutically targeting autophagy is capable of enhancing cytotoxicity of emodin in liver cancer cell lines.
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Objective To analyze inhibitory effect of taspine derivative TasD1-6 on human liver cancer cells of Hep3B, HepG2, and BEL7402, and to study their preliminary mechanism of anticancer action. Methods Effect of TasD1-6 on cell growth of Hep3B, HepG2, and BEL7402 were determined by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and real-time cellular analysis (RTCA) assay. Effect of TasD1-6 on Hep3B cell morphology was investigated with crystal violet staining. Next, Hep3B cell apoptosis was analyzed by Annexin V FITC/PI and Hoechst 33258 staining. Flow cytometry was used to determine Hep3B cell cycle. Effect of TasD1-6 on cell apoptosis protein expression in Hep3B cell was determined by Western blotting. Results MTT and RTCA assay demonstrated that TasD1-6 significantly inhibited the growth of Hep3B cell and the IC50 was (11.3 ± 1.6) μmol/L. Hep3B cell morphology indicated the shrinkage and obvious morphology changes. And TasD1-6 induced the Hep3B cell apoptosis in dose-dependent manner. Cell was stopped as G1 phase by TasD1-6. Western blotting analysis showed TasD1-6 could upregulate the protein expression of p53 and Bax and downregulate Mcl-1 protein expression (P < 0.05). Conclusion Taspine derivative TasD1-6 shows better inhibitory action on the growth of human liver cancer Hep3B cell than HepG2 and BEL7402 cells. At the same time, TasD1-6 can change cell cycle and induce cell apoptosis, which probably related to upregulation of p53 and Bax protein expression and downregulation of Mcl-1 protein expression.
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Objective:To study the expression levels of SAMHD1 in different liver cancer cells and the inhibition of HBV DNA production by SAMHD1. Methods:SAMHD1 mRNA and protein levels in Jurkat,THP-1,HepG2,HepG2. 2. 15 and Huh7 were detected by Real-time quantitative PCR and Western blot. SAMHD1 expression in HepG2. 2. 15 was increased by transfecting SAMHD1 over ex-pression vectors or was inhibited by transfecting SAMHD1 specific shRNA, and then HBV DNA levels were detected. The effect of different dNTP concentration on HBV DNA production was detected also. Results:SAMHD1 were highly expressed in liver cancer cells. Decreased SAMHD1 expression could enhance HBV DNA production by 2. 3-2. 8 times,and increased SAMHD1 expression could inhibit HBV DNA levels by 3 times(P<0. 01). Increased dNTP concentration could enhance HBV DNA production and counteract the inhibitory effect of SAMHD1. Conclusion:SAMHD1 can inhibit the production of HBV DNA in liver cancer cells by decreasing dNTP concentration.
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Objective:To study the expression levels of SAMHD1 in different liver cancer cells and the inhibition of HBV DNA production by SAMHD1. Methods:SAMHD1 mRNA and protein levels in Jurkat,THP-1,HepG2,HepG2. 2. 15 and Huh7 were detected by Real-time quantitative PCR and Western blot. SAMHD1 expression in HepG2. 2. 15 was increased by transfecting SAMHD1 over ex-pression vectors or was inhibited by transfecting SAMHD1 specific shRNA, and then HBV DNA levels were detected. The effect of different dNTP concentration on HBV DNA production was detected also. Results:SAMHD1 were highly expressed in liver cancer cells. Decreased SAMHD1 expression could enhance HBV DNA production by 2. 3-2. 8 times,and increased SAMHD1 expression could inhibit HBV DNA levels by 3 times(P<0. 01). Increased dNTP concentration could enhance HBV DNA production and counteract the inhibitory effect of SAMHD1. Conclusion:SAMHD1 can inhibit the production of HBV DNA in liver cancer cells by decreasing dNTP concentration.
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Objective:To investigate the silencing of P27RF-Rho gene with lenvirus targeting mediated technique,and to clarify its influence in the invasion of liver cancer cells.Methods:The P27RF-Rho RNAi lentivirus was constructed. The liver cancer BEL7402 cells were infected with lentivirus. The experiment was divided into P27RF Rho-siRNA group, scramble-siRNA group and BEL7402 group.The effect of silencing P27RF-Rho gene and the expression levels of hepatocellular carcinoma (HCC)associated proteins RhoA,RhoC, VEGF,P53 and PTEN were detected;the activities of matrix metalloproteinase (MMPs)associated with tumor invasion were analyzed by Gelatin zymography;the variation of transfer ability and invasion abilities were compared by Wound healing assay experiment and Transwell experiment.Results:The Western blotting results showed the expression levels of P27RF-Rho,RhoA,RhoC,and VEGF proteins in the BEL7402 cells in experiment group were significantly lower than those in two control groups (P<0.05),and the expression levels of P53 and PTEN were higher than those in two control groups (P<0.05).The results of Gelatin zymography showed the activities of MMPs in experiment group were significantly lower than those in two control groups (P<0.01 );Wound healing assay showed that the migration ability of the BEL7402 cells in experiment group was significantly inhibited (P<0.01);the number of cells passed through the Transwell Chambers in experiment group was significantly less than those in two control groups (P<0.01).Conclusion:Silenceing P27RF-Rho can weaken the invasion ability and migration ability of human HCC BEL7402 cells.
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OBJECTIVE: To design and synthesize two kinds of allyl-substituted quercetin and investigate their anti-oxidation and anti-tumor activities in vitro. METHODS: The target compounds 1 and 2 were first synthesized from quercetin by hydroxyl protection, allylation,Claisen rearrangement and deprotection. The anti-oxidation and anti-tumor activities of the target compounds and intermediate products were evaluated by DPPH and MTT assay. RESULTS: Eight compounds were synthesized,including six intermediates and two target compounds,in which four were new compounds. All of them were confirmed by 1H-NMR,13C-NMR and LC-MS spectra. The anti-oxidation and anti-tumor tests showed that compounds 1,2,5,6,8 and 9 had anti-oxidation activities and compound 5 inhibited A549 lung cancer cell proliferation. Compound 9 could inhibit the proliferation of lung cancer A549 cells and HepG2 cells. CONCLUSION: The compounds with electron-donating groups have significant anti-oxidant activity. When acetyl and methyl ether groups are used as the protecting groups,the position of introducing allyl group to quercetin has obvious impact on the anti-tumor activity.
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Two kinds of different functional groups modified RuBpy-doped silica fluorescent nanoprobes Probe A and B that conjugated with avidin were prepared for the recognition of liver cancer cells. Firstly RuBpy-doped silica nanoparticles were synthesized by reverse microemulsion and modified with different functional groups, then Probe A was prepared by the conjugation of avidin with carboxyl modified nanoparticles through covalent binding using 1-ethyl-3-( 3-dimethylamino propyl ) carbodiimide hydrochloride ( EDC )/sulfo-NHS, whereas Probe B was prepared by the conjugation of avidin with the polyethylene glycol ( PEG) linkers on the surface nanoparticles using cyanogen bromide method. Therefore, compared with Probe A, Probe B was obtained by coupling avidin to the nanoparticles through long-chain PEG molecules. The two probes were incubated with liver cancer cells respectively, and microscopic fluorescence imaging shows that Probe B which contained PEG molecules could be more effectively applied for the recognition of tumor marker carcinoembryonic antigen ( CEA) in liver cancer cells.
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Active fractions were obtained from the c(?)amworm homogenate by reverse-phase concentration, gel filtration and affinity column chromatography. The MTS/PMS assay was used in the screening of antimicrobial peptide in vitro. The MTS/PMS assay and ELISAAFP method were combined to evaluate the effects of antimicrobial peptide on the proliferation and ?-fetoprotein secretion of HepG2 cells as well as on the proliferation of normal mouse osteoblast MC3T3-E1. The results showed that the purified antimicrobial peptide from clamworm Murphysa sanguinea was a constitutive basic protein with MW 8.1 kDa and pI 8.6, which showed inhibition effects on the proliferation and ?-fetoprotein secretion of HepG2 cells at different levels. These effects have a positive correlation with the concentrations of antimicrobial peptide. No obvious inhibition and damage effects were observed on the normal osteoblast.
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Objective To observe the effects of rat tetramethylpyrazine (TMP)-containing serum on the proliferation of human liver cancer cells Hep G2. Methods Thirty SD male rats were divided into three groups randomly and the serum was collected after ip TMP with the dosage of 143.0, 71.5 mg/kg or NS 0.8 mL to prepare the TMP-containing serum in large- and small-dose groups and NS group as well. A reversed-phase high performance liquid chromatography (RP-HPLC) method was used to determine the TMP concentration in rat serum. Human liver cancer cells Hep G2 was treated with rat TMP-containing serum for 48 h. Inhibition of the TMP-containing serum on proliferation of Hep G2 was detected by MTT assay. Results The serum of large dose TMP (20%, 10%, and 5%) and small dose TMP (20% and 10%) could obviously inhibit Hep G2 multiplication (P
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In this article, the ultrastructure of the liver-original cancer cells and the cells adjacent to and far from the cancer was analysed quantitatively. We found that there were marked differences among some parameters of the nuclei and mitochondria of these three kinds of cells. The significance of the differences has been discussed.
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AIM To study the reversal effect of erythromycin on multidrug resistance of human liver cancer cells BEL 7402. METHODS Cultivated the parental cells BEL 7402 in the presence of doxorubicin to obtain a multidrug resistant subline designated BEL 7402/ADM. The expression of P glycopro tein in BEL 7402 and BEL 7402/ADM cells was determined by using cell ELASA method. The cytotoxicity of drugs was determined by using MTT assay, and the intracellular concentration of doxorubicin in BEL 7402/ADM was determined using a spectrofluorometer. RESULTS After growing BEL 7402 cells in the presence of ADM for some time, the resulting cells (BEL 7402/ADM) became not only resistant to ADM but cross resistant to VCR and MMC, the P gp on the subline cells' surface was highly expresssed. Erythromycin, which is a lipophilic antibiotic, enhanced the anti tumor functions of ADM, VCR and MMC to BEL 7402/ADM cells and increased the intracellular concentration of ADM with no effect on the expression of the P gp in BEL 7402/ADM cells. CONCLUSUON Erythromycin can significantly reverse the multidrug resistance in BEL 7402/ADM cells, it may do so by saturating the P gp pathway to reduce the efflux of intracellular concentration of the drugs.