MOESM5 of In situ characterization of stem cells-like biomarkers in meningiomas

Additional file 5: Figure S3. A. Representative immunofluorescence images for consecutive sections for the grade I Jed64_MN meningioma. Sections were double stained for Ki67 (red) with Nestin (green), SOX2 (red) with CD133 (green), Vimentin (green) with FZD9 (red), SSEA4 (green) with SOX2 (red), and SSEA4 (green) with Olig2 (red), and each section was stained with DAPI (blue). Single staining of GFAP (red) or BIIITubulin (red) is also shown. All images were taken at 20x. B. A grid used as a repository of information for categorical staining is shown with a color-coded legend and size dimensions for sub-areas

MOESM3 of In situ characterization of stem cells-like biomarkers in meningiomas

Additional file 3: Table S2. Differentially expressed cancer driver genes [66] in individual tumors compared with three normal brain sample data sets, referenced in Gene Expression Omnibus (GEO) submission GSE77259. Values were generated from previously published data sets [64, 65] using Transcriptome Analysis Console v. 4.0

MOESM1 of In situ characterization of stem cells-like biomarkers in meningiomas

Additional file 1: Table S1. The clinical profiles for the included patients and their tumors’ histopathological features

MOESM4 of In situ characterization of stem cells-like biomarkers in meningiomas

Additional file 4: Figure S2. A bar graph showing averages of counts for Ki67 stained sections collected using manual counting or automated counting in Image J software

MOESM2 of In situ characterization of stem cells-like biomarkers in meningiomas

Additional file 2: Figure S1. H&E images for different morphological variants and atypical features for meningiomas used in this study. A. Images showing WHO classified morphological features for different meningioma variants. Meningothelial (Jed39_MN) with neoplastic growth of syncytial epithelial cells with indistinct cell borders arranged in whorls; fibroblastic (Jed40_MN) showing spindle cells with indistinct cell boundaries running in fascicle; transitional (Jed38_MN) with ratios of meningothelial to fibroblastic patterns 40:60; psammomatous (Jed43_MN) composed of whorled clusters of spindle cells with numerous psammoma bodies; chordoid (Jed79_MN), Cords of epithelioid cells with focal clear to foamy cytoplasm on myxoid stroma.; rhabdoid (Jed29_MN) showing hypercellular sheets with rhabdoid morphology. B. Tumors with atypical features. Images show patternless growth (sheeting) in Jed72_MN, necrosis and small cells with high nuclear to cytoplasm ratio in Jed58_MN, and brain invasion in Jed13_MN. Magnifications are indicated above images

MOESM6 of In situ characterization of stem cells-like biomarkers in meningiomas

Additional file 6: Figure S4. A. Representative immunofluorescence images for consecutive sections for the grade III Jed29_MN meningioma. Sections were double stained for Ki67 (red) with Nestin (green), SOX2 (red) with CD133 (green), Vimentin (green) with FZD9 (red), SSEA4 (green) with SOX2 (red), and SSEA4 (green) with Olig2 (red), and each section was stained with DAPI (blue). Single staining of GFAP (red) or BIIITubulin (red) is also shown. All images were taken at 20x. B. A grid used as a repository of information for categorical staining is shown with a color-coded legend and size dimensions for sub-areas

MOESM7 of In situ characterization of stem cells-like biomarkers in meningiomas

Additional file 7: Table S3. All combinations of markers observed in consecutive sections and their frequencies in all 15 meningioma samples

Microarray Expression Data Identify <i>DCC</i> as a Candidate Gene for Early Meningioma Progression

<div><p>Meningiomas are the most common primary brain tumors bearing in a minority of cases an aggressive phenotype. Although meningiomas are stratified according to their histology and clinical behavior, the underlying molecular genetics predicting aggressiveness are not thoroughly understood. We performed whole transcript expression profiling in 10 grade I and four grade II meningiomas, three of which invaded the brain. Microarray expression analysis identified deleted in colorectal cancer (<i>DCC</i>) as a differentially expressed gene (DEG) enabling us to cluster meningiomas into <i>DCC</i> low expression (3 grade I and 3 grade II tumors), <i>DCC</i> medium expression (2 grade I and 1 grade II tumors), and <i>DCC</i> high expression (5 grade I tumors) groups. Comparison between the <i>DCC</i> low expression and <i>DCC</i> high expression groups resulted in 416 DEGs (<i>p</i>-value < 0.05; fold change > 2). The most significantly downregulated genes in the <i>DCC</i> low expression group comprised <i>DCC</i>, phosphodiesterase 1C (<i>PDE1C</i>), calmodulin-dependent 70kDa olfactomedin 2 (<i>OLFM2</i>), glutathione S-transferase mu 5 (<i>GSTM5</i>), phosphotyrosine interaction domain containing 1 (<i>PID1</i>), sema domain, transmembrane domain (TM) and cytoplasmic domain, (semaphorin) 6D (<i>SEMA6D</i>), and indolethylamine N-methyltransferase (<i>INMT</i>). The most significantly upregulated genes comprised chromosome 5 open reading frame 63 (<i>C5orf63</i>), homeodomain interacting protein kinase 2 (<i>HIPK2</i>), and basic helix-loop-helix family, member e40 (<i>BHLHE40</i>). Biofunctional analysis identified as predicted top upstream regulators beta-estradiol, TGFB1, Tgf beta complex, LY294002, and dexamethasone and as predicted top regulator effectors NFkB, PIK3R1, and CREBBP. The microarray expression data served also for a comparison between meningiomas from female and male patients and for a comparison between brain invasive and non-invasive meningiomas resulting in a number of significant DEGs and related biofunctions. In conclusion, based on its expression levels, <i>DCC</i> may constitute a valid biomarker to identify those benign meningiomas at risk for progression.</p></div

Unsupervised hierarchical cluster analysis of 416 genes that were differentially expressed (<i>p</i>-value < 0.05; fold change > 2.0) between the three <i>DCC</i> expression groups.

<p>BN samples are included in cluster analysis. A number of genes is represented by more than one transcript. Meningiomas are clustering into two main branches, one of which contains the <i>DCC</i> low expression samples and a <i>DCC</i> medium expression sample that was a brain invasive case. Color scheme bar indicates comparably higher and lower expression values in red and blue color, respectively. Color scheme for samples: yellow, <i>DCC</i> low expression; green, <i>DCC</i> medium expression; orange, <i>DCC</i> high expression; BN samples, red.</p

The predicted top upstream regulators in the comparison group female <i>vs</i>. male meningioma patients are tacrolimus, glutathione, ITPR, (E)-2,3',4,5'-tetramethoxystilbene, and SLC39A4 with a <i>p</i>-value of overlap of 1.16E-03, 1.55E-03, 2.02E-03, 2.02E-03, and 2.02E-03, respectively.

<p>Target genes are <i>APOD</i>, <i>KLRC4-KLRK1</i>/<i>KLRK1</i>, <i>MYH10</i>, <i>TNC</i>, <i>SLC7A11</i>, <i>CYP1B1</i>, and <i>NELL1</i>. Upregulated and downregulated genes in red and blue color, respectively. Asterisk indicates a gene that is represented in the dataset by more than one transcript.</p