Fig. 1: Establishment of brain metastasis NSCLC cell lines.

A Schematic representation of the route for the selected of brain metastatic derivatives in mice. PC9 and H1915 NSCLC cell lines (5 × 10⁵ cells) were inoculated intracardially into BALB/c nude mice. After 65-75 days, the brain metastasis of cancer cells were explored by in vivo bioluminescence imaging system (Bruker In Vivo). The brain-metastasized cancer cells were collected and cultured for another 30 days with Zeocin (50 mg/mL). The selection process was conducted twice, and we collected the cell lines as PC9-BrM and H1915-BrM. B The bioluminescence image of mice with the PC9-BrM and H1915-BrM cell brain metastasis. C Morphological difference between parental PC9 and H1915 cells and brain metastatic PC9-BrM and H1915-BrM cells was displayed a light microscope (Nikon Corporation) at ×100 or ×200 magnification. D To detect the metastatic capacity of cancer cell, EMT-related genes were measured between parental cell and brain metastasis cells. The protein level of EMT-related biomarkers (E-cadherin and Vimentin) was examined by the western blot assay. E The invasive ability of PC9/H1915 and PC9-BrM/H1915-BrM cells were validated by a Transwell assay. Data were presented as the means ± standard error of the mean of at least three independent experiments. ^**P < 0.01 and ^***P < 0.001.