?(Fig

?(Fig.1F).1F). subpopulation earnings considerably from this treatment. Methods The aim of our study was to determine the expression of VEGF-A and its (co-) receptors by immunohistochemistry and to test the association with patient survival in 350 glioma patients. Additionally, VEGF-A expression was analyzed by in-situ hybridization. In 18 patients, the protein expression was compared with the bevacizumab response according to extended and altered RANO criteria. Results We found a heterogeneous expression pattern of VEGF and its receptors in glioblastoma patients with significantly lower levels in WHO grade II and III tumors and normal-appearing brain Lappaconite HBr tissue ( .001). Pilocytic astrocytomas (WHO grade I) showed significantly higher VEGFR-1, -2 and neuropilin-1 levels as compared to WHO grade II and III astrocytomas ( .01) but at lower levels than glioblastomas. The expression of neuropilin-2 was low in all tumors. There was neither a significant correlation between protein expression and patient survival nor between protein levels and bevacizumab response after altered RANO criteria. Conclusion Since our data show that beneficial response to bevacizumab treatment is usually independent of the expression of VEGF-A and its (co-) receptors, further investigation is needed to decipher the underlying mechanisms of antiangiogenic treatment response. = 39) and normal-appearing gray matter (= 62) or white matter (= 19) were also included. Stereotactic biopsies were excluded from the study due to small sample sizes. Individual samples with mainly necrotic tissue or samples with predominantly reactive changes were also excluded. The statistical analysis was based on tissue microarrays (TMAs). Representative whole mount sections of randomly selected patients (at least 5 cases of each entity; data not shown) were Lappaconite HBr investigated to validate the TMA data. To rule out intraindividual differences, repeated cores of the same patients were included in the TMAs. Correlation analyses were performed for staining scores of repetitive cores. Identical expression scores for the assessed factors were obtained in 60% of all repetitive cores. Only 7% of all repetitive cores displayed a score difference 3. The first core of each patient was utilized for statistical analyses to avoid subjective bias. All samples were examined neuropathologically according to WHO criteria by 2 board-certified neuropathologists (P.N.H., M.Mi.)1 All samples were assessed for IDH-1_R132H-, p53-, Ki67-, and pHH3-expression. The study was approved by the ethics Lappaconite HBr committee of the University or college Hospital of Frankfurt and the University or college Cancer Center (UCT) Frankfurt/Main (EC number 4/09, project SNO_SNO_01C08). Table 1. Summary of tissue specimens and individual data = .05C.01 *; .01C.001 **; .001 ***). Statistical analysis was performed using JMP 8.0 and JMP 11.0 software (SAS) and GraphPad Prism 5 (GraphPad Software Inc.). Photographic paperwork was performed using an Olympus BX50 light microscope. Results VEGF-A Is usually Upregulated in Glioblastomas as Compared With Lower-grade Gliomas at Protein Level and Correlates With mRNA Expression In GBM, VEGF-A protein was observed on tumor cells around hypoxic or necrotic foci and also on tumor vessels in the same areas (Fig. ?(Fig.1A1A and C). VEGF-A mRNA and protein expression overlapped to a high extent and showed tumor cells as the main source and blood vessels as a minor source for VEGF-A expression (Fig. ?(Fig.1B1B and D). Although VEGF-A protein expression on tumor cells and vessels in GBM was still low, reaching a median of 1 1 (range: 0C9), it was significantly higher as compared with lower grade astrocytomas ( .001) (Fig. ?(Fig.2A).2A). Furthermore, VEGF-A levels were significantly higher in the tumor center than in corresponding infiltration zones or surrounding normal-appearing gray and white matter of the same patients ( .001) (Supplementary material, Fig. S1A). Open in a separate windows Fig. 1. VEGF-A, VEGFR-1, -2, -3 and NRP-1 and -2 expression in human astrocytomas by in-situ hybridization ISH) and immunohistochemistry (IHC). VEGF-A: (A) IHC-staining of a glioblastoma with high VEGF-A levels on tumor cells and tumor vessels. (B) Lappaconite HBr Corresponding ISH of the same area on a serial section showing similar mRNA signals. (C and D) Higher magnification of corresponding areas of the same tumor. Level bars (A) and (B) = 1000 m, (C) and (D) = 200 m. VEGFR-1: (E) Pilocytic WHO grade I astrocytoma showing moderate-to-strong expression of VEGFR-1 on endothelial cells (arrowheads). (F) WHO grade II astrocytoma exhibiting VEGFR-1 unfavorable vessels (arrows). (G) WHO grade III Mouse monoclonal to CD14.4AW4 reacts with CD14, a 53-55 kDa molecule. CD14 is a human high affinity cell-surface receptor for complexes of lipopolysaccharide (LPS-endotoxin) and serum LPS-binding protein (LPB). CD14 antigen has a strong presence on the surface of monocytes/macrophages, is weakly expressed on granulocytes, but not expressed by myeloid progenitor cells. CD14 functions as a receptor for endotoxin; when the monocytes become activated they release cytokines such as TNF, and up-regulate cell surface molecules including adhesion molecules.This clone is cross reactive with non-human primate astrocytoma with poor endothelial staining for VEGFR-1 (arrowheads). (H) Representative vital tumor center of a glioblastoma with VEGFR-1 Lappaconite HBr positive endothelial cells (arrow-heads). VEGFR-2: (J) Pilocytic astrocytoma WHO grade I showing.