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目的从人结肠癌细胞系(HCT116)中分离鉴定结肠癌干细胞,并观察其体外生物学特性。方法利用RT-PCR、免疫荧光、流式细胞等技术检测HCT116细胞中CD133的表达情况;利用免疫磁珠分选技术分选CD133+细胞;采用无血清培养基培养分选后的CD133+细胞鉴定其体外增殖特性;采用含血清培养基诱导干细胞克隆球分化并采用免疫组化检测CK20表达情况鉴定其分化特性。结果HCT116细胞系中存在CD133+细胞,免疫磁珠能有效的分选CD133+细胞,分选后的阳性含量为91.92%,分选后的阳性细胞具有典型干细胞的体外增殖、分化特性。结论CD133+结肠癌干细胞在体外具有和普通干细胞类似的无限增殖、自我更新和分化能力,同时它们也表达ABCG2等耐药相关蛋白。
Objective To isolate and identify colon cancer stem cells from human colon cancer cell line (HCT116) and observe its biological characteristics in vitro. Methods The expression of CD133 in HCT116 cells was detected by RT-PCR, immunofluorescence and flow cytometry. The CD133 + cells were sorted by immunomagnetic bead sorting technique. The CD133 + cells were cultured in serum-free medium and identified in vitro Proliferation characteristics; Stem cell clone spheroid differentiation was induced by serum-containing medium and CK20 expression was detected by immunohistochemistry to identify its differentiation characteristics. Results There were CD133 + cells in HCT116 cell line. Immunomagnetic beads could effectively differentiate CD133 + cells. The positive percentage of sorted CD133 + cells was 91.92%. The sorted positive cells had typical proliferation and differentiation characteristics of stem cells. Conclusion CD133 + colon cancer stem cells have the similar ability of proliferation, self-renewal and differentiation as ordinary stem cells in vitro, meanwhile they also express ABCG2 and other drug-resistant proteins.