| PurposeThis study aims to explore the inhibitory effect to colon cancer cells SW1116ofginsenoside Rh3in vitro. And briefly discusses the mechanism of growth of colon cancercells SW1116is inhibited by ginsenoside Rh3in vitro.Methods1. The effect of ginsenoside Rh3to colon cancer cells SW1116in vitro between theconcentration-response action research and cell morphology observation: take logarithm thegrowing period of SW1116colon cancer cells to4×103cells/well in96plate of vaccination.Will Rh3liquid storing diluted into480μ g/ml,240μg/ml,120μg/ml,60μg/ml,30μg/ml fivedifferent final concentrations to join has inoculated against cells in the well. Incubate in37℃,5%CO2for24h and take the morphological observation and MTT detection.2. The the limitation action research of ginsenoside Rh3to colon cancer cells SW1116in vitro: take logarithm the growing period of SW1116colon cancer cells of4×103cells/well in96plate of vaccination. Will Rh3liquid storing diluted into120μg/ml join cells inthe well,37℃and5%CO2respectively training0h,3h,6h,9h,12h,24h,48h, after that takeMTT detection.3. Use AO/EB detect the apoptosis of ginsenoside Rh3to colon cancer cells SW1116invitro: take logarithm the growing period of SW1116colon cancer cells to5×104cells/wellin cover glass of growth. Stick the wall, the experimental group to join120μg/mlginsenoside Rh3, the control to join DMEM medium,37℃,5%CO2training8h aftercollection of cells to climb. Join AO/EB mixture, in fluorescence microscope and takingpictures of the record.4. The detection of apoptosis ginsenoside Rh3to colon cancer cells SW1116in vitrowith TUNEL; take logarithm the growing period of SW1116colon cancer cells to5×104cells/well in cover glass of growth. Stick the wall, the experimental group to join120μg/ml ginsenoside Rh3, the control to join DMEM medium,37℃,5%CO2training8hafter collection of cells, fixed up. Use TUNEL kit for testing, then a microscope andtaking pictures of the record.5. To probe the mechanism of ginsenoside Rh3to colon cancer cells SW1116with RT-PCR in vitro: take logarithm the growing period of SW1116colon can-cer cells to1×106cells/well in6plate of vaccination. Stick the wall, to join the120μg/mlginsenoside Rh3, the control to join DMEM medium,37℃,5%CO2training8h afterextraction mRNA collects cells. Will the mRNA transcr-iptase are extracted and thenseparately with design good Caspase-3and GAPD-H primer for expansion. Take5μlelectrophoresis product amplified and taking pictures of the record.6. To probe the mechanism of ginsenoside Rh3to colon cancer cells SW1116in vitrowith immunohistochemical: take logarithm the growing period of SW1116colon cancercells to5×104cells/well in cover glass of growth. Stick the wall, the experimental group tojoin120μg/ml ginsenoside Rh3, the control to join DMEM medium,37℃,5%CO2training8h after collection of cells, fixed up. To Caspase-3antibodies as one resistant toimmunohistochemical study. After dyeing a microscope and taking pictures of the record.7. To probe the mechanism of ginsenoside Rh3to colon cancer cells SW1116with flowcytometric: take logarithm the growing period of SW1116colon cancer cells to1×106cells/well in6plate of vaccination. Stick the wall,join the120μg/ml ginsenoside Rh3,the control to join DMEM medium,37℃,5%CO2training8h after collecting cells, fixed.To Caspase-3antibodies as a resistance to flow cytometric detection.Result1. The inhibitory effect of ginsenoside Rh3ot colon cancer cells SW1116in vitro:ginsenoside Rh3colon cancer cells to the growth of SW1116inhibition.Concentratio-nresponse experimental results can be found Rh3ginseng saponins of coloncancer cells SW1116inhibitory effect with increased concentration strengthen, whenconcentration reach120μg/ml inhibition when to platform. Through to the colon cancercells SW1116morphological observation found that as ginseng saponins Rh3concentrationsincreasing cells form change significantly. Colon cancer cells gradually by the polygons goround, cell spacing is concentrated, nucleus, smaller. Aging test results showed that ginsengsaponins Rh3colon cancer cells to SW1116inhibition effect from Rh3after9h to work,after12h to platform period. Through the AO/EB and dyeing method of dual validationTUNEL, ginseng saponins Rh3can lead to colon cancer cells in vitro SW1116apoptosis.2. The mechanism ginsenoside Rh3inhibit the growth of colon cancer cells SW1116in vitro: ginseng saponins Rh3role after SW1116colon cancer cells can promoteCaspase-3expression. RT-PCR results show that NaCan gene expression in GAPDH issimilar, under the premise of the experimental group Caspase-3strip brightness was significantly higher than the control group. That group Caspase-3mRNA content of higherthan those in the control group. MRNA high content that can be indirectly Caspase-3increased protein expression. And further immunohistochemical and flow cytometryverified the results of more than that. From immunohistochemical can be seen in thepicture, and the control group was almost no cells were coloring, and the cells in the part ofthe visible light brown spot piece. Streaming results also showed a group Caspase-3increased protein expression.Conclusion1. Ginsenoside Rh3of colon cancer cells in vitro SW1116growth inhibition andinhibition of ginsenoside Rh3with increasing concentration and role increased with theextension of time. When the concentration reach120μg/ml, function of time for12h rolewhen to platform period.2. the ginsenoside Rh3can induce Caspase-3expression, start apoptosis way torestrain the growth of the SW1116colon cancer cells.The results suggest that ginsenoside Rh3can contribute to the SW1116colon cancercell apoptosis. Its potential to be developed as a treatment for colon cancer chemotherapydrugs, or as an adjunct therapy to reduce the toxic effects of chemotherapy drugs. |
