AIM: To analyze the upregulated CD133 expression in tumorigenesis of primary colon cancer cells. METHODS: Upregulated CD133 expression in tumorigenesis of colorectal cancer cell lines (Lovo, Colo205, Caco-2, HCT116 an...AIM: To analyze the upregulated CD133 expression in tumorigenesis of primary colon cancer cells. METHODS: Upregulated CD133 expression in tumorigenesis of colorectal cancer cell lines (Lovo, Colo205, Caco-2, HCT116 and SW620) was analyzed by flow cytometry. Human colon cancer tissue samples were stained with anti-human CD133. SW620 cells were sorted according to the CD133 expression level measured by fluorescence-activated cell sorting. Spheroids of colorectal cancer cells were cultured with the hanging drop. Expression of CD133 and Lgr5 in spheroids of colorectal cancer cells and monolayer culture was detected by RT-qPCR. Spheroids of colorectal cancer cells were analyzed using anti-human CD133 with immunohistochemical staining. RESULTS: CD133 antigen was expressed in colorectal cancer cell lines (Lovo, Colo205, Caco-2, HCT116 and SW620) as well as in primary and metastatic human colon cancer tissues. However, the CD133 was differently expressed in these cell lines and tissues. The expression levels of CD133 and Lgr5 were significantly higher in spheroids of parental, CD133hi and CD133-cells than in their monolayer culture at the mRNA level (P < 0.05). Immunohistochemical staining of spheroids of CD133-cells showed that CD133 was highly expressed in colorectal cancer cell lines. CONCLUSION: Upregulated CD133 expression plays a role in tumorigenesis colorectal cancer cells, which may promote the expression of other critical genes that can drive tumorigenesis.展开更多
Objective:The aim of this study was to study the effect of Bcl-2 small hairpin RNA(shRNA) enhancing methotrexate(MTX)-induced apoptosis of Raji cells.Methods:Expression plasmid with Bcl-2 shRNA was transfected into Ra...Objective:The aim of this study was to study the effect of Bcl-2 small hairpin RNA(shRNA) enhancing methotrexate(MTX)-induced apoptosis of Raji cells.Methods:Expression plasmid with Bcl-2 shRNA was transfected into Raji cells by Lipofectmine 2000 and then treated with MTX.At 48 h of transfection,the expression level of Bcl-2 mRNA and protein was evaluated by RT-PCR and immunofluorescence.MTT assay was used to analyze cell proliferation at 24,48 and 72 h.Apoptosis was detected by Giemsa staining and flow cytometric cell cycle analysis.Results:After transfection with Bcl-2 shRNA,the expression levels of Bcl-2 mRNA and protein in Raji cells decreased(P < 0.05).Using Giemsa staining,cells transfected with Bcl-2 shRNA combined with MTX at 48 h displayed changes of apoptosis.MTX significantly inhibited the growth of cells after transfected with Bcl-2 shRNA(P < 0.05).Apoptotic rates of the Raji cells treated with Bcl-2 shRNA combined with MTX significantly increased(P < 0.05),compared with either control shRNA/MTX combination or MTX-treatment cells alone.Conclusion:Our results suggest the shRNA against Bcl-2 mRNA could increase MTX-induced apoptosis of Raji cells.展开更多
文摘AIM: To analyze the upregulated CD133 expression in tumorigenesis of primary colon cancer cells. METHODS: Upregulated CD133 expression in tumorigenesis of colorectal cancer cell lines (Lovo, Colo205, Caco-2, HCT116 and SW620) was analyzed by flow cytometry. Human colon cancer tissue samples were stained with anti-human CD133. SW620 cells were sorted according to the CD133 expression level measured by fluorescence-activated cell sorting. Spheroids of colorectal cancer cells were cultured with the hanging drop. Expression of CD133 and Lgr5 in spheroids of colorectal cancer cells and monolayer culture was detected by RT-qPCR. Spheroids of colorectal cancer cells were analyzed using anti-human CD133 with immunohistochemical staining. RESULTS: CD133 antigen was expressed in colorectal cancer cell lines (Lovo, Colo205, Caco-2, HCT116 and SW620) as well as in primary and metastatic human colon cancer tissues. However, the CD133 was differently expressed in these cell lines and tissues. The expression levels of CD133 and Lgr5 were significantly higher in spheroids of parental, CD133hi and CD133-cells than in their monolayer culture at the mRNA level (P < 0.05). Immunohistochemical staining of spheroids of CD133-cells showed that CD133 was highly expressed in colorectal cancer cell lines. CONCLUSION: Upregulated CD133 expression plays a role in tumorigenesis colorectal cancer cells, which may promote the expression of other critical genes that can drive tumorigenesis.
基金Supported by the grants from the Natural Science Program Foundation of Guangdong Province(No.04010446)the Overseas Chinese Affairs Office of the State Council Key Discipline Construction Fund (No.51205002)
文摘Objective:The aim of this study was to study the effect of Bcl-2 small hairpin RNA(shRNA) enhancing methotrexate(MTX)-induced apoptosis of Raji cells.Methods:Expression plasmid with Bcl-2 shRNA was transfected into Raji cells by Lipofectmine 2000 and then treated with MTX.At 48 h of transfection,the expression level of Bcl-2 mRNA and protein was evaluated by RT-PCR and immunofluorescence.MTT assay was used to analyze cell proliferation at 24,48 and 72 h.Apoptosis was detected by Giemsa staining and flow cytometric cell cycle analysis.Results:After transfection with Bcl-2 shRNA,the expression levels of Bcl-2 mRNA and protein in Raji cells decreased(P < 0.05).Using Giemsa staining,cells transfected with Bcl-2 shRNA combined with MTX at 48 h displayed changes of apoptosis.MTX significantly inhibited the growth of cells after transfected with Bcl-2 shRNA(P < 0.05).Apoptotic rates of the Raji cells treated with Bcl-2 shRNA combined with MTX significantly increased(P < 0.05),compared with either control shRNA/MTX combination or MTX-treatment cells alone.Conclusion:Our results suggest the shRNA against Bcl-2 mRNA could increase MTX-induced apoptosis of Raji cells.
