| Objective: To study the effect of ginsenoside Rg3 on connexin26 gene (Cx26) expression and gap junctional intercellular communication (GJIC) in human breast cancer cell line MCF-7, cultured in vitro. It will be helpful to discover the mechanism of ginsenoside Rg3 in anti-tumor and lay a theory foundation for treating cancer.Methods: Human breast cancer cell line MCF-7 was exposed to ginsenoside Rg3 at differential concentrations respectively. The cell proliferation inhibition was measured by 3-〔4,5-dimethylthiazo-2-yl〕-2,5 diphenyl tetrazolium bromide(MTT) assay. The effect of ginsenoside Rg3 on Cx26 gene was measured by reverse transcriptase-polymerase chain reaction(RT-PCR) 24 hours after ginsenoside Rg3 exposure. The GJIC function of the cells was examined with scrape-loading dye transfer assay 24 hours after ginsenoside Rg3 treatment.Results:1. Inhibitory effect of ginsenoside Rg3 on proliferation of MCF-7 cells. Human breast cancer cell line MCF-7 was exposed to ginsenoside Rg3 at a concentration of 10μg/ml,20μg/ml,40μg/ml,80μg/ml,160μg/ml,320μg/ml respectively. The inhibition ratio was 3.1%,5.2%,16.0%,26.3%,29.1%,50.4% respectively after 24 hours, statistical significance can be found from 40μg/ml to 320μg/ml compare with the control group (p<0.05). The inhibition ratio was 6.0%,12.4%,30.3%,60.6%,69.1%,74.0% respectively after 48 hours, statistical significance can be found from 20μg/ml to 320μg/ml (p<0.05). Statistical significance can not be found between 24hours and 48 hours.. Under an inverted microscope , we can observed the cells adherent, fullness, polygon in control group, the cells amout sparse, suspended, quasi-round, and no refractional nature in experiment groups, the highter the concentration is, the greater themorphological change of cells will become. 2. The expression of Cx26 gene. Human breast cancer cell line MCF-7 cells were cultured between control group and experiment groups, extracted mRNA of all groups cells after 24 hours., then RT-PCR reaction. The genes Cx26 and internal reference GAPDH were expressioned in all groups cells. The expression of Cx26 gene was slight in control group compare with the experiment groups, but the highter the concentration is (40μg/ml,80μg/ml,160μg/ml), the greater the expression will become in experiment groups by Lab-work analyzed. There was no statistical significance between control group and 40μg/ml or 80μg/ml (p>0.05), but markedly Statistical significance can be found compare with 160μg/ml (p=0.01). There was markedly statistical significance between 160μg/ml and 40μg/ml or 80μg/ml (p<0.01), but There was no statistical significance between 40μg/ml and 80μg/ml (p>0.05). 3. Scrape-loading and dye transfer method. Under the fluorescent microscope, we can observed that Lucifer Yellow fluorescent staining localized the single cell in control group, Lucifer Yellow fluorescent transfered the adjacent cells by gap junction, then come into being flake fluorescent staining in experiment grops, but fluorescent staining was no transferred in the furthermore distant adjacent cells, the gap junctional intercellular communication was restored after cancer cells were exposed to ginsenoside Rg3.Conclusion: 1. Ginsenoside Rg3 has obvious anti-tumor effect on human breast cancer cell line MCF-7, the highter the concentration is, the greater the inhibitory effect on tumor cells proliferation will come. No statistical significance was found between 24 hours and 48 hours. 2. The expression of connexin 26 gene was lower in tumor compared with that of Rg3 groups, ginsenoside Rg3 can up regulate the expression of connexin26 gene. There was markedly statistical significance with the increased concentration of ginsenoside Rg3. 3. Gap junctional intercellular communication was restored gently after the tumor cells were exposed to ginsenoside Rg3. 4. Ginsenoside Rg3 can up regulate the expression of connexin 26 gene, and restore gap junctional intercellular communication , this may be one of important mechanisms related ginsenoside Rg3 in anti-tumor . |
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