Abstract
Simple SummaryGlioblastomas are very malignant and essentially incurable brain tumors. One problem is the extensive penetration of tumor cells into the adjacent normal brain tissue. Thus, the testing of novel drugs requires appropriate tumor models, preferentially avoiding animal studies. This paper describes so-called brain tissue slice tandem-culture systems. They consist of a slice of normal brain tissue and a second layer of tumor tissue. The microscopic analysis of these slice tandem-cultures allows for the simultaneous assessment of single cells invading into the normal brain tissue and the space occupying growth of the total tumor mass. It is shown that the direct application of test drugs onto the slices exerts inhibitory effects on both mechanisms. We thus describe a system mimicking the situation in glioblastoma patients. It reduces animal studies, allows for the direct application of test drugs and the precise quantitation of their inhibitory effects on tumor growth and invasion.Glioblastomas (GBMs) are the most malignant brain tumors and are essentially incurable even after extensive surgery, radiotherapy, and chemotherapy, mainly because of extensive infiltration of tumor cells into the adjacent normal tissue. Thus, the evaluation of novel drugs in malignant glioma treatment requires sophisticated ex vivo models that approach the authentic interplay between tumor and host environment while avoiding extensive in vivo studies in animals. This paper describes the standardized setup of an organotypic brain tissue slice tandem-culture system, comprising of normal brain tissue from adult mice and tumor tissue from human glioblastoma xenografts, and explore its utility for assessing inhibitory effects of test drugs. The microscopic analysis of vertical sections of the slice tandem-cultures allows for the simultaneous assessment of (i) the invasive potential of single cells or cell aggregates and (ii) the space occupying growth of the bulk tumor mass, both contributing to malignant tumor progression. The comparison of tissue slice co-cultures with spheroids vs. tissue slice tandem-cultures using tumor xenograft slices demonstrates advantages of the xenograft tandem approach. The direct and facile application of test drugs is shown to exert inhibitory effects on bulk tumor growth and/or tumor cell invasion, and allows their precise quantitation. In conclusion, we describe a straightforward ex vivo system mimicking the in vivo situation of the tumor mass and the normal brain in GBM patients. It reduces animal studies and allows for the direct and reproducible application of test drugs and the precise quantitation of their effects on the bulk tumor mass and on the tumor’s invasive properties.
Highlights
Gliomas are the most common primary brain tumors, with glioblastoma being most detrimental and associated with a median survival of only 12–15 months after diagnosis [1,2].Its poor prognosis that leaves it essentially incurable even after extensive surgery, radioand chemotherapy is mainly due to the extensive infiltration of tumor cells into the adjacent normal tissue [3]
Cortical brain segments were selected as the basal layer underneath glioblastoma xenograft tissue slices or spheroids
While cultured spheroids were already comparable in size, glioblastoma xenograft tissue slices were processed into equal sizes by excising circular segments of 2 mm in diameter from peripheral, non-necrotic tumor areas using a biopsy punch
Summary
Gliomas are the most common primary brain tumors, with glioblastoma (grade IV glioma) being most detrimental and associated with a median survival of only 12–15 months after diagnosis [1,2].Its poor prognosis that leaves it essentially incurable even after extensive surgery, radioand chemotherapy is mainly due to the extensive infiltration of tumor cells into the adjacent normal tissue [3]. Rather than single tumor cells in suspension, which would preferentially grow on the tissue surface, spheroids were implanted onto the brain slice surface or into the brain slice tissue [15,16] While this approach readily provides some essential features of the host tissue like an authentic ECM, glial-neuronal contact and interaction as well as neuronal connectivity, the spheroids represent only partially the in vivo tumor situation due to the lack of intact tumor tissue structures, which may well affect tumor cell mobility, migration, and invasive potential. Combining the advantages of the intact host microenvironment of organotypic brain tissue slices with an intact tumor tissue, we have developed a glioblastoma tissue slice tandem co-culture setting based on intact tumor tissue derived from mouse xenografts This allows for ex vivo studies on the effects of drugs, not requiring animals with the exception of the tissue donors and representing an alternative to the use of animals for scientific purposes. Tumor xenograft material was used (with a very small number of animals required since the tumor tissues are propagated into many ex vivo samples), this may be even extended toward primary tumors from patients
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