Research The Effect Of Lupeol To Biological Behavior Of Liver Cancer Stem-like Cells In Human Hepatocarcinoma Cell Line SMMC-7721

Objective: Exploring the effect of lupeol to biological behavior of liver cancer stem-like cells in human hepatocarcinoma cell line SMMC-7721 in vitro.Method:1 To collect human liver cancer cells cultured in common culture media,then to obtain iver cancer stem-like cells by suspension culture with serum free medium.2 To use flow cytometry to detect different concentrations of lupeol(0,20,40,60?mol/L)of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells apoptosis rate and cell cycle distribution.3 To use of different concentrations of lupeol(0,20,40,60?mol/L)inhuman hepatocellular carcinoma cell line SMMC-7721 stem-like cells,respectively at 24 h,48h,72 h.The growth inhibition of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells were detected by MTS assy.4 To use of different concentrations of lupeol(0,20,40,60?mol/L)in human hepatocellular carcinoma cell line SMMC-7721 stem-like cells.To get observation of effects of lupeol on SMMC-7721 stem-like cell invasion and migration abilities by using Transwell inserts 24 hours later.5 To arrange human hepatocellular carcinoma cell line SMMC-7721 stem-like cells in the containing different concentrations of lupeol(0,20,40,60?mol/L)of suspension medium by 5-7 days,then to get observation of forming spheres inhibition of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells.6 To use of different concentrations of lupeol(0,20,40,60?mol/L)in human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,respectively at 20 min,40min,60 min,then to get observation of Cell matrix adhesion inhibition of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells.Results:1 The cell apoptosis and cell cycle of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells detected by flow cytometryFCM results showed that the different doses of lupeol(0,20,40,60?mol/L)impact on human hepatocellular carcinoma cell lineSMMC-7721stem-like cells,after 48 h apoptosis rates were 2.24±0.42%?16.27±2.4% ? 3.96±2.42% ? 27.37±1.93%,the difference was statistically significant(P<0.05).Cell cycle showed that with the increasing dose of lupeol,the number of cells which in the S phase had no significant(36.78±0.68?36.36±0.86?36.52±0.70?36.66±0.38,P>0.05),in the G2/M phase had no significant(27.58±1.81?27.82±1.33?27.50±0.90?28±0.32,P>0.05),in the G0/G1 phase had no significant(35.26±1.27 ? 35.42±0.93 ? 35.58±0.89 ?35.34±0.35,P>0.05).2 Comparison of proliferation ability of different concentrations of lupeol treatmentMTS assay showed that the different doses of lupeol(20,40,60?mol/L)impact on human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,after 24 h cell inhibition rates were 12.05%?23.37%?30.46%(P<0.05),after 48 h cell inhibition rates were 17.09%?26.74%?35.08%(P<0.05),after 72 h cell inhibition rates were 22.13%?33.23%?44.83%(P<0.05).Compared with the same concentration at different time,with time prolonging,the inhibition rate increased,there was significant difference between three groups.3 Comparison of invasion ability of different concentrations of lupeol treatmentMicroscope observation showed that the different doses of lupeol(20,40,60?mol/L)impact on human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,after 24 h cell invasion inhibition rates were 22.48%?27.54%?41.00%,there was a significant difference between the treatment groups and the control group(P<0.05).4 Comparison of migratory ability of different concentrations of lupeol treatmentMicroscope observation showed that the different doses of lupeol(20,40,60?mol/L)impact on human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,after 24 h cell migration inhibition rates were18.81%?31.93%?45.05%,there was a significant difference between the treatment groups and the control group(P<0.05).5 Comparison of forming spheres ability of different concentrations of lupeol treatmentMicroscope observation showed that the different doses of lupeol(20,40,60?mol/L)impact on human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,after 5-7 days cell forming sphere inhibition rates were 27.03%?36.49%?58.11%,there was a significant difference between the treatment groups and the control group(P<0.05).6 Comparison of Cell matrix adhesion ability of different concentrations of lupeol treatmentMicroscope observation showed the different doses of lupeol(20,40,60?mol/L)impact on human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,after 20 min cell adherent inhibition rates were 5.07% ? 20.96% ? 40.96%(P<0.05),after 40 min cell adherent inhibition rateswere 6.53% ? 27.50% ? 40.17%(P<0.05),after 60 min cell adherent inhibiti-on rates were 16.85%?29.15%?46.94%(P<0.05).Compared with the same concentration at different time,with time prolonging,the adherent inhibition rate increased,there was significant difference between three groups(P<0.05).Conclusion:1 Lupeol can be able to induce apoptosis of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,and the effect was associated with dose.But there was no significant effect on the cell cycle of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells.2 Lupeol can be able to inhibit the growth of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,and the effect was associated with dose and time.3 Lupeol can inhibit the invasion ability of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,and the effect was associated with dose.Lupeol can meanwhile inhibit the migratory ability of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,and the effect was associated with dose.4 Lupeol can inhibit the forming spheres ability of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,and the effect was associated with dose.5 Lupeol can inhibit the adhesive capacity of human hepatocellular carcinoma cell line SMMC-7721 stem-like cells,and the effect was associated with dose and time.