Quantitative analysis of glioma cell invasion by confocal laser scanning microscopy in a novel brain slice model

To quantitatively analyze the spatial extent of glioma cell migration in an organotypic brain slice culture, we developed a new invasion model with th e aid of confocal laser scanning microscopy (CLSM). CLSM: allowed not only for three-dimensional visualization of the invasive pattern of human T98G g lioma cells in the living brain slice but also for serial analysis of the i nvasive process over several weeks. Twenty-four hours after the T98G: gliom a spheroid was initiated to coculture with a brain slice, the glioma cells detached themselves from the spheroid and spontaneously continued to migrat e on the surface of the brain slice, while they diffusely invaded into the slice by migrating to a deeper site. Immunohistochemical analysis revealed that these migrating glioma cells much more strongly immunostained for matr ix metalloproteinase (MMP)-2 and -9 than the tumor spheroid which remained at the implanted site. Treatment of the T98G glioma spheroid with 1,10-phen anthroline, a specific inhibitor of MMPs, significantly inhibited not only the cell migration on the surface of the brain slice but also the invasion of the glioma cells into the slice. The present version of the glioma invas ion model using CLSM makes it possible to spatially and serially analyze th e extent of glioma cell invasion in the living brain slice for several week s, making it a very useful tool for investigating the cellular and molecula r mechanisms of glioma invasion under conditions most analogous to those of normal brains in vivo. (C) 2000 Academic Press.