CD133阳性/阴性肺癌细胞的分选、鉴定及差异基因的筛选

BACKGROUND AND OBJECTIVE: It has been proven that cancer stem cell existed in variety of cancer, which an significant difference of biological characteristics was observed between the cancer stem cells and non-cancer stem cells. And CD133 is considered to be cancer stem cell marker. So there may be significant differences in CD133- positive cells and CD133-negative cells. The aim of this study is to isolate CD133+ cells and CD133- cells from lung cancer cell line A549, explore their biological characteristics and screen the metastasis-related genes. METHODS: MACS was applied to isolate CD133+ cells and CD133- cells from human lung cancer cell line A549. To observe the formation of sphere, CD133+ cells and CD133- cells were cultured in serum-free DMEM-F12 medium (containing EGF, bFGF) in vitro. The colony formatting efficiency of CD133+ cells, CD133- cells and cells without sorting was tested by colony-forming assay. The differentiation of sphere was induced by culturing in DMEM-F12 medium (containing serum). The metastasis-related genes (84 genes) of CD133+ cells and CD133- cells were detected by using DNA microarray. Immunohistochemistry was used to detect the expression of CD133 protein in Human lung cancer tissue. RESULTS: CD133+ cells formed sphere in serum-free DMEM-F12 medium, while the CD133- cells failed to form sphere. The rates of CD133+ cell colony formation (57.1%) was significantly higher than that of CD133- cells (3.3%). Sphere (CD133+/CK7-) was induced to differentiate, and CK7 expression was found in differentiated cells. The expression levels of 19 metastasis-related genes from CD133+ cells and CD133- cells were significant different. Little CD133 positive cells which distributing around the cancer nests were found in lung cancer tissue. The expression of CD133 was not related to tumor types, cell differentiation or TNM stage. CONCLUSION: CD133+ cells exhibit the characteristics of cancer stem cells. The difference of metastasis-related gene expression levels was discovered between CD133+ cells and CD133- cells. CD82 plays an important role in mechanism of tumor metastasis.