Establishment of Human Lung Adenocarcinoma Cell Line A549 with High Metastasis and Biological Characteristics of Its Cancer Stem Cells

Objective To investigate the biological characteristics of cancer stem cells derived from human lung adenocarcinoma cell line A549 with high metastasis.

Methods A549 cells were inoculated into subcutaneous tumor. The lung metastasis cells were obtained by mechanical separation method, and then inoculated into nude mice, repeated inoculation of nude mice to obtain a stable lung metastasis of the cell line A549-V13. Serum-free spheroid formation and PKH26 staining were used to determine whether there were cancer stem cells in A549-V13 cell line. Flow cytometry was performed to detect cancer stem cell marker CD133 expression in A549 and A549-V13 cells; CD133⁺ expressions were sorted to identify its biological characteristics in vivo and in vitro.

Results The high metastatic cell line A549-V13 are obtained. The single PKH26 positive cell was observed in spheroid cells after A549-V13 cells were cultured for 7 days. Compared with A549 cells, the proportion of cancer stem cell marker CD133 was significantly increased by 4.85 times in A549-V13 cells. The sorted CD133⁺ cells of A549-V13 showed higher self-renewal ability, resistance ability invasion ability was enhanced by 1.42 times and the IC₅₀ was increased by 1.26 times. Nude mice inoculated with 2×10² CD133⁺ cells of A549-V13 developed tumor after 3 months(4/6), whereas at least 2×10² CD133⁺ cells A549 formed tumor for 3 months(2/6).

Conclusion The high metastatic cell line A549-V13 are obtained. With the increase of the proportion of CD133⁺ cells, the biological function of CD133⁺ cells was significantly enhanced in vivo and in vitro.