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ObjectiveTo investigate the effect of Ganoderma lucidum polysaccharides （GLP） on the proliferation， migration， cycle， and apoptosis of hepatocellular carcinoma SKHEP1 and Huh7 cells and to explore the underlying mechanism. MethodSK-HEP-1 and Huh-7 cells were classified into the blank group and low-， medium-， and high-dose GLP groups （3.5， 7， 14 g·L-1）. The proliferation of the cells was examined by cell counting kit-8 （CCK8） assay， and the migration by scratch assay. Cell cycle was measured by flow cytometry and apoptosis was detected based on Hoechst33258 staining. In addition， the expression of phosphatidylinositol 3-kinase （PI3K）， protein kinase B （Akt）， phosphorylated PI3K （pPI3K）， and phosphorylated Akt （pAkt） in the cells was determined by Western blot. ResultCompared with the blank group， the three doses of GLP reduced the proliferation and migration of SKHEP1 and Huh7 cells （P<0.05）， increased the percentage of cells in G1 phase （P<0.05）， and decreased percentage of cells in S and G2 phase （P<0.05）. In addition， the three doses can induce apoptosis of both SK-HEP-1 and Huh-7 cells， particularly the high dose. Moreover， the three doses of GLP lowered the levels of pPI3K and pAkt （P<0.05）. ConclusionGLP significantly inhibited the malignant phenotype of SK-HEP-1 and Huh-7 cells through the PI3K/Akt signaling pathway.
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Objetive: To investigate the effects of silencing HOXA13 gene on the malignant phenotypes of hepatocellular carcinoma cell lines HepG2 and QGY-7703, and to provide a new molecular target for the diagnosis and treatment of hepatocellular carcinoma. Methods: The interference recombinant plasmid of plent-U6-GFP/si-HOXA13 was constructed, then it was stably transfected into the HepG2 and QGY-7703 cells. The interfering effects of HOXA13 gene were determined by RT-PCR and Western blotting methods. The HepG2 and QGY-7703 cells transfected with HOXA13 knockdown were used as experimental group, and the HepG2 and QGY-7703 cells transfected with empty plasmid were used as control group. Then the growth speed was examined by MTT assay, the cell doubling time was examined by double time assay, the colony formation ability was examined by colony formation assay, and the cell cycle was examined by flow cytometry. Results: The MTT assay results showed that compared with control group, the growth speeds of HepG2 and GY-7703 cells in experimental group were significantly decreased and the cell cycles were changed; the number of HepG2 and QGY-7703 cells in G1 phase was increased and the number of cells in S phase was decreased. The doubling time of HepG2 and QGY-7703 cells in control group and experimental group were (4.59 ± 0.27), (4.93 ± 0.17), (6.02 ± 0.86), and (6.43 ± 0.66) h, and the differences between control group and experimental group were significant (P<0.05). The colony number of HepG2 and QGY-7703 cells in control group and experimental group were 264.00 ± 12.62, 269.00 ± 4.55, 165.00±10.61, and 215.00 ± 4.43, and the differences between control group and experimental group were significant (P<0.01). Conclusion: HOXA13 can increase the proliferation, enhance the clone formation, decrease the number of cells at G1 phase and increase the number of cells at S phase in the hepatocellular carcinoma HepG2and QGY-7703 cells; and it may used as a new molecular target for diagnosis and treatment of hepatocellular carcinoma.
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Objective:To investigate the effects of silencing HOXA13 gene on the malignant phenotypes of hepatocellular carcinoma cell lines HepG2and QGY-7703,and to provide a new molecular target for the diagnosis and treatment of hepatocellular carcinoma.Methods:The interference recombinant plasmid of plent-U6-GFP/si-HOXA13 was constructed,then it was stably transfected into the HepG2and QGY-7703 cells.The interfering effects of HOXA13 gene were determined by RT-PCR and Western blotting methods.The HepG2and QGY-7703 cells transfected with HOXA13 knockdown were used as experimental group,and the HepG2and QGY-7703 cells transfected with empty plasmid were used as control group.Then the growth speed was examined by MTT assay, the cell doubling time was examined by double time assay,the colony formation ability was examined by colony formation assay,and the cell cycle was examined by flow cytometry.Results:The MTT assay results showed that compared with control group,the growth speeds of HepG2and GY-7703 cells in experimental group were significantly decreased and the cell cycles were changed;the number of HepG2and QGY-7703 cells in G1phase was increased and the number of cells in S phase was decreased.The doubling time of HepG2and QGY-7703 cells in control group and experimental group were(4.59±0.27),(4.93±0.17),(6.02±0.86),and(6.43±0.66)h, and the differences between control group and experimental group were significant(P<0.05).The colony number of HepG2and QGY-7703 cells in control group and experimental group were 264.00 ± 12.62,269.00 ± 4.55, 165.00± 10.61,and 215.00 ± 4.43,and the differences between control group and experimental group were significant(P<0.01).Conclusion:HOXA13 can increase the proliferation,enhance the clone formation,decrease the number of cells at G1phase and increase the number of cells at S phase in the hepatocellular carcinoma HepG2 and QGY-7703 cells;and it may used as a new molecular target for diagnosis and treatment of hepatocellular carcinoma.
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Objective The purpose of this study was to examine the inhibitory effect of ONO-AE3-208, an EP4 antagonist, on prostate cancer with bone metastasis in an animal model . Methods A PC3/LUC cell line was constructed by stably transfecting luciferin to prostate cancer PC 3 cells and inoculated into the left ventricle of nude mice to establish an animal model of prostate cancer with bone metastasis .After modeling , the animals in the experimental group and control groups were intraperitoneally given ONO -AE3-208 and double-distilled water, respectively, followed by examination of the metastasis loci and tumor burden by bioluminescence ima -ging and statistical analysis with survival curves . Results At 60 days after modeling , the animals in the control group exhibited sig-nificantly increased metastases and fluorescence burdens as compared with the experimental group (P<0.01), and the increase was in a time-dependent manner (P<0.01).At 60 days, the controls began to die while the experimental animals remained well alive , and at 180 days, the mice of the control group all died .The survival rate of the animals was significantly higher in the experimental group than in the control ( 13.3% vs 0%, P <0.01 ) and the median survival time remarkably longer in the former than in the latter group (162 d vs 116 d, P <0.01). Conclusion The EP4 antagonist ONO-AE3-208 inhibited the bone metastasis of prostate cancer and prolonged the survival time in the model mice .
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Objective To explore the effects and molecular mechanisms of suberoylanilide hydroxamic acid (SAHA) on ovarian carcinoma. Methods (1)Two groups of ovarian carcinoma cell lines (SKOV3 and SKOV3/DDP, HO8910 and HO8910-PM) were exposed to SAHA (1, 3, 5 and 7μmol/L SAHA,group 1-group 4). CCK-8 method was employed to eval-uate the inhibitory effects of SAHA.(2)Ovarian cancer cell lines treated with SAHA (2 or 5μmol/L SAHA) were used as 1 and 2 groups. Flow cytometry was performed following staining with Annexin V-FITC and PI for cell cycle and apoptosis.(3) Reverse transcription polymerase chain reaction (RT-PCR) and Western blot assay were used to assess the mRNA and pro-tein expression levels of phenotypic correlation factor. Results (1)After 48 h of SAHA treatment,the OD value of SKOV3, SKOV3/DDP,and HO8910 showed a trend of gradually reduce (P<0.05).(2)The apoptotic rates were significantly higher in SAHA 1 and SAHA 2 groups than those of control group (P<0.05). Compared with control group, after 48 h of SAHA treat-ment,S phase and G2/M phase of SKOV3 and SKOV3/DDP cells increased;G0/G1 phase of HO8910 and HO8910-PM cells increased in SAHA 1 and 2 groups (P<0.05).(3)The expression levels of CyclinB1 and Cdc2 (p34) mRNA were significant-ly lower in SAHA 1 and 2 groups than those of control group,while the expression levels of Caspase-3,p21 and p53 mRNA expression were significantly higher in SAHA 1 and 2 groups than those of control group. Furthermore,the expression of Ac-Histone H3,Ac-Histone H4,p53 protein were markedly improved,and CyclinB1,Cdc2(p34) protein decreased in SAHA 1-4 groups. Conclusion SAHA may suppress cell growth, induce apoptosis and cause cycle arrest in ovarian carcinoma cells by promoting histone acetylation or modulating their phenotype-related proteins of Caspase-3, p53, CyclinB1 and Cdc2(p34).
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Objective To explore the relationship between mitochondrial DNA deletion and malignant phenotypes of human lung cancer cells. Methods Two rho? derivatives of 95C and 95D were generated by treating the cultured cells with ethidium bromide. Agarose colony formation assays and Transwell invasion assays were carried out to detect the phenotypes of colony formation and invasiveness of the cultured cells, respectively. Cell growth was determined by MTT. Results The partially mtDNA-deleted cells exhibited stronger capacity of colony formation and invasiveness, and faster growth rates than their respective parental cell lines. Conclusion Mitochondrial DNA deletion might play a role in the formation of malignant phenotypes of human lung cancer.
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AIM:To study the effects of Se-methylselenocysteine(MSC) on the growth and apoptosis of human liver carcinoma cell line,HepG_2 cells and the molecular mechanism of effects.METHODS: After HepG_2 cells treated with various concentrations of MSC,survival and apoptosis were determined by MTT assay and light microscope,apoptosis and cell cycle were determined by flow cytometry.Malignant phenotype was determined by soft agarose growth assay.Cyclin D1 mRNA expression was determined by RT-PCR and Caspase-3 activity was measured by Caspase-3 activity assay kit.RESULTS: After being treated with 25(?mol?L~(-1)) MSC for 24 h,the survival of HepG_2 cells was decreased,a marked apoptosis and S phase arrest characteristic was observed in time-and dose-dependent manner.Soft agatose growth assay showed MSC inhibited HepG_2 cells growth in soft agarose.RT-PCR and Caspase-3 assay showed that HepG_2 cells treated with 25(?mol?L~(-1)) MSC for 24 and 48 hours,the expression of Cyclin D1 mRNA was down-regulated by 38% and 47%,while Caspase-3 activity was up-regulated by((39.61)?(6.65))%and((118.73)?(6.48))%.HepG_2 cells treated with 50(?mol?L~(-1)) MSC for 24 and 48 h,the expression of Cyclin D1 mRNA was down-regulated by 53% and 82%, whereas Caspase-3 activity was up-regulated by((80.66)?(9.31))% and((152.67)?(7.95))%.CONCLUSION: MSC can inhibit the growth of HepG_2 cells,and induce apoptosis and S phase arrest of HepG_2 cells.The phenotype alterations of HepG_2 cells might relate with inhibition of Cyclin D1 expression and Caspase-3 activity by MSC in the cells.
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The heterospecies hybrid cells(HL-N)from the fusion of human promyelocy-tic leukemia mutant cells(HL-60-AR)and mouse bone marrow nucleated red cellswere established in HAT selective medium.Malignant phenotype comparative analy-sis between parental tumor cells and hybrid cells showed that growth ability ofhybrid cells was decreased.The hybrid cells reduced their DNA synthesis rate andlost the ability of colony-forming in 0.3% soft agar medium.The cells lost tumor-producing ability when they were transplanted into nude mice also.Inhibition orreduction of c-myc oncogene expression was demonstrated by Northern molecularhybridization techniques.The ultrastructure of hybrid cells were also different fromtheir parental cells.These results mentioned above showed that the mouse bone mar-row nucleated red cells might provide some peculiar factors(both nuclear factorsand cytoplasmic factors)to inhibit the expression of HL-60-AR cell malignant phe-notypes.