DETECTION OF BRAIN-TUMOR INVASION AND MICROMETASTASIS IN-VIVO BY EXPRESSION OF ENHANCED GREEN FLUORESCENT PROTEIN

OBJECTIVE: To determine whether fluorescence from human brain tumor ce lls transfected with the enhanced green fluorescent protein (EGFP) gen e in vitro and xenotransplanted into the brain of nude mice would perm it the detection of brain tumor invasion and metastasis in vivo. METHO DS: Daoy medulloblastoma cells were transfected with a long terminal r epeat-based retroviral vector containing the EGFP gene. Stable EGFP-ex pressing clones were isolated and stereotactically injected into the f rontal cortex of nude mice. Four weeks later, whole brain sections wer e examined using fluorescence microscopy, immunohistochemistry, and ro utine hematoxylin and eosin staining for the visualization and detecti on of tumor cell invasion and metastasis. RESULTS: We demonstrate that EGFP-transduced Daoy cells maintain stable high-level EGFP expression in the central nervous system during their growth in vivo. EGFP fluor escence clearly demarcated the primary tumor margins and readily allow ed for the visualization of distant micrometastases and local invasion on the single-cell level. Small metastatic and locally invasive foci, including those immediately adjacent to the tumor's leading invasive edge, were virtually undetectable by routine hematoxylin and eosin sta ining and immunohistochemistry. EGFP expression also persisted in vitr o after cell reculture from brain tissue extracts. CONCLUSION: We show , for the first time, that EGFP-transduced human brain tumor cells can be visualized by fluorescence microscopy after intracerebral implanta tion. This method is superior to routine hematoxylin and eosin stainin g and immunohistochemistry for the detection and study of physiologica lly relevant patterns of brain tumor invasion and metastasis in vivo.