Abstract
Primary brain tumors are hallmarked for their destructive activity on the microenvironment and vasculature. However, solely few experimental techniques exist to access the tumor microenvironment under anatomical intact conditions with remaining cellular and extracellular composition. Here, we detail an ex vivo vascular glioma impact method (VOGIM) to investigate the influence of gliomas and chemotherapeutics on the tumor microenvironment and angiogenesis under conditions that closely resemble the in vivo situation. We generated organotypic brain slice cultures from rats and transgenic mice and implanted glioma cells expressing fluorescent reporter proteins. In the VOGIM, tumor-induced vessels presented the whole range of vascular pathologies and tumor zones as found in human primary brain tumor specimens. In contrast, non-transformed cells such as primary astrocytes do not alter the vessel architecture. Vascular characteristics with vessel branching, junctions and vessel meshes are quantitatively assessable as well as the peritumoral zone. In particular, the VOGIM resembles the brain tumor microenvironment with alterations of neurons, microglia and cell survival. Hence, this method allows live cell monitoring of virtually any fluorescence-reporter expressing cell. We further analyzed the vasculature and microglia under the influence of tumor cells and chemotherapeutics such as Temozolamide (Temodal/Temcad®). Noteworthy, temozolomide normalized vasculare junctions and branches as well as microglial distribution in tumor-implanted brains. Moreover, VOGIM can be facilitated for implementing the 3Rs in experimentations. In summary, the VOGIM represents a versatile and robust technique which allows the assessment of the brain tumor microenvironment with parameters such as angiogenesis, neuronal cell death and microglial activity at the morphological and quantitative level.
Highlights
Brain vasculature and the angiogenic process play both key roles during development and function of the brain [1] [2]
We describe the establishment of the Vascular Organotypic Glioma Impact Model (VOGIM) presenting a robust and reliable tool to investigate physiological and pathological angiogenesis
Any brain tissue from wild type or transgenic mice or rodents can be facilitated for the organotypic brain slice assay
Summary
Brain vasculature and the angiogenic process play both key roles during development and function of the brain [1] [2]. There are intrinsic advantages in evaluating the angiogenic process and the vascular morphology in organotypic 3D culture compared to 2D cell cultures Cell culture assays such as the tube formation, retina, aortic ring and endothelial spheroid assays which use dissociated endothelial cells (mainly human umbilical vein endothelial cells [HUVEC], brain endothelial cells or aortic endothelial cells) or tissue pieces (retina or arterial tissues) are to a certain extent redundant or reflect ectopic www.impactjournals.com/oncotarget angiogenesis independent of the particular conditions present in the brain microenvironment [20, 21]. Such cell culture-based models lost their tissue integrity and are deprived from their physiologic micromilieu, connectivitydependent signals and their organotypic environment which are important determinants in in vivo processes
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