Ginsenoside Rh2 On Human Hepatoma Cell Line Smmc-7721 Induced Differentiation Of Experimental Studies

Objective Study of ginsenoside Rh₂ on human hepatoma cell line SMMC-7721 induced differentiation, from cell inhibition, cell cycle and sub-cellular level (cell cytoskeleton, cell junction, nucleus matrix - lamin - intermediate filament system) point of view in inducing differentiation of the mechanisms.For further animal testing and research to explore the specific mechanism to provide a theoretical basis and experimental evidence.Methods In vitro human hepatoma cell line SMMC-7721 training, and using 10μg/ml, 20μg/ml, 40μg/ml its ginsenoside Rh₂ for 2days, 4days, 6days. Using tetrazolium blue (MTT) colorimetric assay with different concentrations of ginsenoside Rh₂ on SMMC-7721 tumor cell proliferation; by flow cytometry with different concentrations of ginsenoside Rh₂ on SMMC-7721 tumor cells role. According to MTT and flow cytometry on the role of ginsenoside Rh₂ inhibited SMMC-7721 rate and apoptosis rate of the measured results, it was found 20μg/ml and 40μg/ml Ginsenoside Rh₂ induced differentiation of a certain effect, it is divided into 20μg / ml, 40μg/ml, and the control group three groups. Using laser confocal microscopy observation of the control group, 20μg/ml, 40μg/ml Ginsenoside Rh₂ on the role of SMMC-7721 after the cytoskeleton; using laser confocal microscope in control group, 20μg/ml,40μg/mlgroups,SMMC-7721 Inter-communication of tumor cells; by whole mount electron microscope to observe the control group, 20μg/ml, 40μg/ml ginsenoside Rh₂ on SMMC-7721 tumor cell nuclear matrix - lamin - intermediate filament system changes.Conclusions 1. MTT assay showed that ginsenoside Rh₂ on SMMC-772 inhibit the growth of liver cells, and its role in drug concentration and time were dependent. One 10μg/ml, 20μg/ml, 40μg/ml group 4 days after the role of inhibition rate of 33.2%, 42.9%, 72.4%; 40ug/ml ginsenoside Rh₂ for 6 days, about 90% inhibition rate; while control cells no significant effect. Tips 10μg/ml, 20μg/ml, 40μg/ml for the effective dose, 2 days, 4 days, 6 days for the effective duration of action.2. Flow cytometry showed that ginsenoside Rh₂ on SMMC-7721 liver cancer cells induced apoptosis, apoptosis rate and the concentration of drug action time are dependent manner. The control group and 10μg/ml, 20μg/ml, 40μg/ml role of apoptosis 4 days were 4.99%, 6.35%, 10.75%, 22.42%. One group 20μg/ml, 40μg/ml group and the control group P <0.05, statistically significant, but 10μg/ml group and the control group P> 0.05 not statistically significant. Ginsenoside Rh₂ on cell cycle in a certain concentration of an impact, but after a certain concentration of tumor cells it can cause mortality, the result showed that 20μg/ml, 40μg/ml role of 4 days may hepatoma cells SMMC-7721 is ideal for induction of differentiation concentration and time.3. Were gathered microscope role of ginsenoside Rh₂ 40μg/ml group 4 days after SMMC-7721 cytoskeleton structure, visible actin polymerization, re-distribution, cell morphology extended to form stress fibers, along the longitudinal axis. Group in the role of 20μg/ml 4 days, showing slightly disordered arrangement of actin to form a little tension fiber distribution is not uniform; control group of cells are actin showed disorders, arranged irregularly, and no formation of stress fibers. The results showed that ginsenoside Rh₂ treatment group and the control group there were significant differences in changes. The role of ginsenoside Rh₂ in 20μg/ml may start to appear 4 days after the liver cancer cell SMMC-7721 cells to the normal signs; and 40μg/ml role may tend to 4 days after differentiation and maturation.4. Measured by fluorescence bleaching effect of different concentrations of ginsenoside Rh₂ in SMMC-7721 hepatoma cells 4 days after the clearance rate of fluorescence recovery. The results showed that ginsenoside Rh₂ on SMMC-7721 tumor cell communication to a certain extent. One control group, 20μg/ml, 40μg/ml each group, the role of fluorescence recovery after cells were 11.06±3.87,25.76±9.18,39.34±14.63, respectively compared with the control group after the P <0.05 and P <0.01 all have statistical significance, its 40μg/ml ginsenoside Rh₂ significant statistical significance. That ginsenoside Rh₂ on the hepatoma cell line SMMC-7721 cells with the control group after the role of gap fluorescence recovery at different speeds of change, this may be to increase the number of gap junctional, intercellular communication signal tends to normal cells.5. Helping You electron microscope effect of different concentrations of Ginsenoside Rh₂ SMMC-7721 tumor cell nuclear matrix - lamin - Intermediate Filaments. Can be found in the control group showed pleomorphic nucleoli, irregular, electron density. Nuclear matrix by the thickness of the uneven distribution of irregular fiber interwoven. Between the nuclear lamina between the cytoplasm and nuclear transfer, cytoplasmic intermediate filament from outside the nuclear lamina was irregular radial, but also focus on trends; and 40μg/ml group nucleolus rules, they showed mostly single, nuclear matrix quantity of fiber, rich layers, distributed, abundant cytoplasmic intermediate filament occurs, rules and uniform distribution; 20μg/ml group rules mostly nucleolar, nuclear matrix fibers mathematics is not obvious, clear cytoplasmic intermediate filament significantly unevenly distributed. The results showed that Ginsenoside Rh₂ on the hepatoma cell line SMMC-7721 after the role of nuclear matrix - lamin - there are differences among the system changes, 20μg/ml, 40μg/ml after the role of the trend to mature and differentiate.