Identification And The Features Of Tumor Formation Of Colon Cancer Stem Cell And Effect Of Portulaca Oleracea’s Extraction On Colorectal Cancer Stem Cells

[Objectives] To study the features of tumor formation of colon cancer stem cell and to analysis it clinical significance; To study the inhibitory effect of portulaca oleracea extracts on the HT-29 cell and HT-29 stem cell and the mechanism of colorectal cancer in vitro.[Method] Portulaca oleracea was extracted by water and ethanol, the flavonoids content of the extractions were detected by UV.Colorectal cancer cells(Lovo cells、SW480 cells、HCT-116 cells、HCT-8 cells) were cultured in vitro, the proliferation ability of cultured cells in varying concentration of portulaca oleracea extracts were tested by CCK-8 analysis.HT-29 colon cancer cells were cultured without serum and detect the expression of CD133 and CD44 by flow cytometry; To detect the inhibition effect on HT-29 cancer cell and HT-29 cancer stem cell by 5-FU and to study the tumor formation features during different area in mice models; HT-29 cancer cell and HT-29 cancer stem cell were cultured and the proliferation ability of cultured cells in varying concentration of portulaca oleracea extracts were tested by CCK-8 analysis, and the cell apoptosis and cells cycle were detected with flow cytometry, and the gene expressing of B-catenin、Notch1、Notch2 were detect with Real-time PCR, the protein expressing of B-catenin、 Notch 1、Notch2 were detect with Western blot.[Results] The content of flavonoids extracted by water is no different with ethanol. Both of the water extraction and ethanol extraction (tested by CCK-8 analysis) inhibited the proliferation and showed a dose dependence and time dependence to colorectal cancer cells.The ethanol extraction inhibited colorectal cancer cells proliferation was more significant than the water extraction.HT-29 cancer stem cells were less stained in nuclear compared with HT-29 cancer cell; the positive expression of CD133 and CD44 in HT-29 was 44.6% and 0.6% respectively but in HT-29 cancer stem cells was 92.6% and 97.8% respectively. HT-29 cancer stem cell was drug resistance for 5-FU and its IC50 for 5-FU was 13.087ug/ml while the IC50 of HT-29 cancer cell was 1.394 ug/ml; The inhibition of Portulaca oleracea (tested by CCK-8 analysis) to HT-29 cell and HT-29 stem cell has dose dependence; Portulaca oleracea (detected with flow cytometry) can enhance the HT-29 cancer cell and HT-29 cancer stem cell cells’apoptosis rate, there is a Statistical significance compared the experimental group with the control group; Portulaca oleracea (detected by Real-time PCR) can down-regulate the gene expression of β-catenin、 Notchl of HT-29 cancer stem cell and up-regulate the gene expression of Notch2; the down-regulation of the genes expression of B-catenin、Notch1 were significant(p<0.05);Portulaca oleracea also can down-regulate the protein expression of β-catenin、 Notch1、Notch2 of HT-29 cancer stem cell.Compared with HT-29 cells, the down-regulation of the proteins expression of B-catenin、Notch1、Notch2 were significant.(p<0.05)[Conclusion] There were small proportion of cancer stem cell among HT-29 colon cancer cell line. Colon cancer stem cell can be isolated by culture without serum and the stem cell showed 5-FU drug resistance; Portulaca oleracea can significantly inhibit the proliferation of the HT-29 cells and HT-29 stem cells;The mechanism of inhibition is changing cell cycle and inducing cell apoptosis. And the mechanism is related to Portulaca oleracea can down-regulate the gene expression of β-catenin、Notch1、Notch2;And portulaca oleracea can down-regulate the protein expression of β-catenin、Notch1、Notch2.