Biochemical and Biophysical Research Communications
Brain slice invasion model reveals genes differentially regulated in glioma invasion☆
Section snippets
Materials and methods
Generation of F98 cells expressing the enhanced green fluorescent protein (EGFP). The rat glioma cell line F98 was transfected with the pEGFP-N1-plasmid (Clontech, Palo Alto, USA) by using SuperFect Transfection Reagent (Qiagen, Hilden, Germany). Stably transfected cells were enriched by G418 selection (Invitrogen, Karlsruhe, Germany) and subsequent fluorescent activated cell sorting (FACS) analysis as reported previously [11].
Invasion-assay. Twenty-four hours prior to the preparation of the
The organotypic brain slice culture system as invasion model
We established an invasion model that enabled us to study gene expression differences in stationary and invasive glioma cells. Glioma cells attached to a porous membrane were confronted with a brain slice. We found that a portion of the cells had invaded the brain slice while a second population remained on the membrane. Since glioma cells were transfected with the fluorescent protein EGFP, we were able to separate the invading glioma cells from the brain cells using FACS. In addition, FACS
Discussion
We chose a combination of techniques for the identification of candidate genes involved in glioma invasion. Cultured mouse brain slices were used as matrix for tumor cell invasion. These slices contain extracellular matrix components known to be important in glioma cell invasion and thus provide a more natural environment than coated surfaces used in earlier studies. Organotypic brain cultures have been demonstrated to be viable for several weeks [9], [10]. Our invasion assay differs from most
Acknowledgments
We thank Kathrein Stichling and Dorothea Krupke for their technical assistance and Dr. Peter N. Robinson for critically reading the manuscript. We also thank Dr. Elena Elstner for kindly providing the human glioma cell lines. This work was supported by the Deutsche Forschungsgesellschaft.
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Supported by the Deutsche Forschungsgesellschaft SFB507.