AbstractsBiology & Animal Science

Investigating the properties of glioma-associated microglia/macrophages

by Frank Szulzewsky

Institution: Freie Universität Berlin
Department: FB Biologie, Chemie, Pharmazie
Degree: PhD
Year: 2015
Record ID: 1113995
Full text PDF: http://edocs.fu-berlin.de/diss/receive/FUDISS_thesis_000000099023


Malignant glioma belong to the most aggressive neoplasms in humans with no successful treatment available. Patients suffering from glioblastoma multiforme (GBM), the highest-grade glioma, have an average survival time of only 12-15 months after diagnosis. Both microglia and peripheral macrophages/monocytes accumulate within and around glioma, but fail to exert effective anti-tumor activity and even support tumor growth. Here I investigated the properties and functions of glioma-associated microglia/macrophages (GAMs) using different molecular and experimental approaches. In the first project I used microarray analysis to compare the expression profiles of GAMs and naive control cells. Samples were generated from CD11b+ MACS-isolated cells from naïve and GL261-implanted C57BL/6 mouse brains. Around 1000 genes were more than 2-fold up- or downregulated in GAMs when compared to control cells. Comparison to published data sets of M1/M2a,b,c-polarized macrophages revealed a gene expression pattern that has only partial overlap with any of the M1 or M2 gene expression patterns. Samples for the qRT-PCR validation of selected M1 and M2a,b,c-specific genes were generated from two different glioma mouse models and isolated by flow cytometry to distinguish between resident microglia and invading macrophages/monocytes. In both models the unique GAMs phenotype including a mixture of M1 and M2a,b,c-specific genes could be confirmed. To validate the expression of these genes in human samples, I MACS-isolated CD11b+ microglia/macrophages from glioblastoma multiforme (GBM), lower grade brain tumors, and control specimens. Apart from the M1/M2 gene analysis, I demonstrate that Gpnmb and Spp1 are highly expressed in both murine and human GAMs. High expression of these genes has been associated with worsened prognosis in human GBM, as indicated by patient outcome linked to gene expression data. I also show that GAMs are the predominant source of these transcripts in murine and human GBM. In the second project, I investigated the role of Cx3cr1 in the microglia-dependent support of glioma growth by employing a monocyte-free organotypic brain slice model. In Cx3cr1GFP/GFP slices, lacking Cx3cr1 expression, the growth of GL261 tumors was significantly enhanced, when compared to Cx3cr1GFP/wt or Cx3cr1wt/wt slices. In another glioma model the loss of Cx3cr1 did not significantly impact tumor growth when compared to Cx3cr1GFP/wt or Cx3cr1wt/wt slices. In both models a trend toward a higher microglia migration toward tumors could be observed in Cx3cr1GFP/GFP slices when compared to Cx3cr1GFP/wt. This indicates that CX3CR1 signaling attenuates a glioma-directed migration of microglia and subsequent glioma-supporting functions of these cells. The findings of my thesis provide new insights in the phenotype and function of GAMs and will help to better understand their role for glioma development and progression and to define new potential targets for future anti-glioma therapy. Maligne Gliome gehören zu den aggressivsten Neoplasmen im…