Effect Of Gynostemma Drug-containing Serum On Cell Proliferation,migration And Transforming Growth Factor ?₁?TGF-?₁?in Human Hepatocellular Carcinoma Cells

Objective: To study the effect of Gynostemma drug-containing serum on cell proliferation,migration and transforming growth factor beta 1(TGF-?1)in human hepatocellular carcinoma cell line huh 7.To discussion the possible mechanism of Gynostemma on proliferation and migration of human hepatocellular carcinoma cells.Methods:According to the principle of plasma pharmacology,healthy adult adult SD rats were treated with Gynostemma pentaphyllum decoction and normal white plasma control group.The plasma of Gynostemma pentaphyllum was collected by EDTA anticoagulant tube.Human hepatocellular carcinoma cell line Huh-7 was cultured in vitro and subjected to cell experiment.The control group was incubated with Huh-7 cells with 10% Gynostemma pentaphyllum plasma culture medium and Huh-7 cells in hepatocellular carcinoma.The control group was cultured with 10% concentration of blank plasma and Huh-7 cells of hepatocellular carcinoma,and the control group was established.The different concentrations of blank plasma were detected by CCK8 colorimetric assay,and the best experimental concentration was obtained.Then,the experiment was carried out with this concentration.The migration rate and migration inhibition rate of the control group and the experimental group were measured and calculated by cell scratches.The inhibitory effect of Gynostemma pentaphyllum on Huh-7 migration was observed.The concentration of OD and the inhibitory rate of cell proliferation were measured by CCK8 colorimetric assay.The effects of Gynostemma pentaphyllum on the expression of Huh-7 Cell proliferation inhibition.Effect of Gynostemma Pentaphyllum on the Expression of TGF-?1 in Hepatocellular Carcinoma Huh-7 Cells by Cell Immunofluorescence.The expression of TGF-?1 m RNA in hepatocellular carcinoma Huh-7 cells was detected by Western blotting.The expression of TGF-?1 protein in hepatocellular carcinoma Huh-7 cells was detected by Western blotting.The expression of TGF-?1 m RNA in hepatocellular carcinoma Huh-7 cells was detected by q RT-PCR.Results: 1.The results of CCK-8 shows that the inhibition rate of 1%,5%,10%,15% medicated blood plasma countering to the huh-7 cells of human' liver cancer are-14.13%?-32.08%?16.70% and 38.2% respectively.When the concentration is 1% and 5%,the cancer cells are proliferated;when the concentration is 10%,the blood plasma shows the weakest inhibitory effect on liver cancer cells,the concentration is moderate,and the drug concentration of medicated blood plasma is guaranteed.At experiment 3,gynostemma pentaphyllum blood plasma group shows obvious inhibition on the Huh-7 cells of liver cancer in vitro.The average inhibition rate at the 24 hours and 48 hours are respectively(P<0.05)?0.36(P<0.05).2.The results of scratch test show that the Huh-7 cells of liver cancer are cured in different methods,after being dosed in 0 hour,24 hours and 48 hours,the scratch difference of medicated blood plasma group increases when compared with blank serum group and blank control group,and blank serum group and blank control group show no difference(P>0.05),and the migration rate in 48 hous and that of in 24 hours decrease,which preliminary states that gynostemma pentaphyllum has inhibition and metastasis effects on Huh-7 cells.3.The results of fluorescence immunoassay show that the medicated blood plasma with gynostemma pentaphyllum makes the TGF-?1 at the Huh-7 cells of liver cancer present high expression level,while the TGF-?1 at blank serum control group and additive-free group present low expression level.4.The results of RT-PCR indicate that the medicated blood plasma with gynostemma pentaphyllum countering to the gene expression quantities of TGF-?1 at the cells of liver cancer is on downtrend,and it has signify-cant difference when contrasted to blank serum(P<0.05).5.The results of western blotting show that the medicated blood plasma with gynostemma pentaphyllum can reduce the protein level of TFG-?1,and it has significant difference when contrasted to blank serum(P<0.05).Conclusion: gynostemma pentaphyllum have obvious inhibitory effect on the activity,cell proliferation as well as invasion and migration of the Huh-7 cell of liver cancer.The best inhibitory concentration is 10%,and the inhibitory effect in 48 hours is better than that in 24 hours.Its mechanism may be related to the down-regulation expression level of TGF-beta 1.