Expressions And Significances Of S100B And EGFR In Highly Brain Metastatic Subclone Of A Human Lung Cancer Cell Line

BACKGROUND The brain metastasis is still the most common cause of lung cancer death at present. Routine brain scans of patients with newly diagnosed non-small cell lung cancer identify brain metastases in 3 to 10% of patients. In patient with small-cell lung cancer, the figure is about 15%. The median survival time of untreated patients is only about 3 months. Lung cancer is the most common primary tumor leading to brain metastases. About 39% brain metastases derived from lung cancer on diagnosis. Brain metastases are generally believed to result from hematogenous spreading of circulating tumor cells or frank tumor emboli. Unfortunately, the mechanisms of brain metastases are still unclear. The involvement of S100B in the brain metastasis rising from malignant melanoma is identified, in contrast, it is still perplexing for EGFR. Lack of fundamental research for molecular mechanism of brain metastases from lung cancer resulted in hindering treatment for lung cancer with brain metastasis. In our lab, a sub-clone, PC14/B, with increased potential of brain metastases was selected through repeated injection of human lung cancer cells into the tail vein of nude mice, which embodied biologic activities compared with PC14 which metastasize to brain occasionally and A549 which never metastasize to brain.OBJECTIVE Based on the cell lines: PC14/B, PC14, A549, we investigated the expression of two molecules (EGFR and S100B), trying to reveal the mechanism involved in the brain metastasis of lung adenocarcinoma.METHODS The cell lines A549, PC14, PC14/B were studied. For immunoblotting, the sample was prepared by cell lysis, and the concentration was determined by Bradford assay. The expression of EGFR and S100B was detected by western blotting and cell immunochemistry. Meanwhile, total RNA was isolated and cDNA was reversibly transcribed from the isolated mRNA. The expression of S100B was semi-quantified by RT-PCR (reverse transcription-polymerase chain reaction) additionally. The images were evaluated by image analysis software IPP (ImagePro Plus,version 6.0). Statistical analysis was performed using SPSS 12.0. LSD-t Test was employed for inter-group comparison. All data were expressed as mean±standard deviation (SD). Statistical significance was determined as P <0.05.RESULTS (1) the expressions of EGFR in the three cell lines In all the three cells, immunohistochemical signal of EGFR in the kytoplasm and the cell membrane were observed. The staining of A549 is stronger than that in PC14/B or PC14. In Western Blotting, we can see that the expression of EGFR in A549 is significantly higher than that in PC14/B and in PC14. The difference among the three cell lines have statistical significances( p ?0.05). (2)the expressions of S100B in the three cell lines The results of RT-PCR assay and cell immunochemistry staining indicate that the expression of S100B is significantly higher in PC14/B than that in PC14 and A549( p ?0.05). Additionally, in Western Blotting, we can observe that there are clear bands at 21KD in all the three cells and the intensity of them are similar. And bands of 32KD and 40KD were observed in PC14/B, in contrast, corresponding were observed in neither PC14 nor A549.CONCLUSIONS According to the expression of EGFR in these cells, it is concluded that wild type EGFR may not have significantly positive correlation with the process of brain metastases from lung adenocarcinoma. The two polymerides that contained S100B protein which cannot be detected in neither PC14(partial brain metastases) nor A549(no brain metastases)may correlate with brain metastases of lung adenocarcinoma cells. The immunocytochemical staining and mRNA expression of S100B are consistent with this concept. The S100B protein in serum or cerebrospinal fluie may derive partly from the tumor cells themselves.