Extracellular matrix differences in glioblastoma patients with different prognoses

Abstract

Glioblastoma is the most common malignant central nervous system tumor. Patient outcome remains poor despite the development of therapy and increased understanding of the disease in the past decades. Glioma cells invade the peritumoral brain, which results in inevitable tumor recurrence. Previous studies have demonstrated that the extracellular matrix (ECM) is altered in gliomas and serves a major role in glioma invasion. The present study focuses on differences in the ECM composition of tumors in patients with poor and improved prognosis. The mRNA and protein expression of 16 invasion-associated ECM molecules was determined using reverse trascription-quantitiative polymerase chain reaction and immunohistochemistry, respectively. Clinical factors of patients with different prognoses was also analyzed. It was determined that age and postoperative Karnofsky performance score were associated with patient survival. Furthermore, Fms-related tyrosine kinase 4/vascular endothelial growth factor receptor 3 (FLT4/VEGFR3), murine double minute 2 (MDM2) and matrix metallopeptidase 2 (MMP2) mRNA levels were significantly different between the two prognostic groups. Additionally, brevican, cluster of differentiation 44, hyaluronan mediated motility receptor, integrin-αV and -β1, and MDM2 protein expression were indicated to be significantly different in immunohistochemistry slides. Using the expression profile, including the invasion spectrum of the samples, it was possible to identify the prognostic group of the sample with high efficacy, particularly in cases with poor prognosis. In conclusion, it was determined that ECM components exhibit different expression levels in tumors with different prognoses and thus the invasion spectrum can be used as a prognostic factor in glioblastoma.

Keywords: expression; extracellular matrix; glioblastoma; prognosis; survival.

Figure 1.

Kaplan-Meier survival analysis of patients with glioblastoma with different disease prognoses. Patients in group A had significantly reduced OS time compared with those in group B. Group A, OS <24 months; group B, OS >24 months; OS, overall survival.

Figure 2.

Invasion spectrum (the mRNA expression pattern of invasion-associated extracellular matrix components) differs in patients with ‘worse’ and ‘better’ prognoses. mRNA expression measurements were performed twice for each gene to confirm the data. A longer bar on the logarithmic scale indicates reduced expression. *P<0.05 vs. group A (Mann-Whitney U test). Group A, OS <24 months; group B, OS >24 months; BCAN, brevican; CD44, cluster of differentiation 44; CSPG5, chondroitin sulfate proteoglycan 5; EGFR, epidermal growth factor receptor; FLT4, Fms-related tyrosine kinase 4; HMMR, hyaluronan-mediated motility receptor; IDH1, isocitrate dehydrogenase 1; ITGAV, integrin-αV; MDM2, murine double minute 2; MMP-2, matrix metallopeptidase 2; NCAN, neurocan; PDGFA, platelet-derived growth factor α; TNC, tenascin C; VCAN, versican; OS, overall survival.

Figure 3.

A total of 3 invasion-associated extracellular matrix components demonstrated significantly different expression levels in the samples from different prognostic groups. All 3 molecules had increased expression in patients in group A, indicating that the level of these molecules was associated with tumor invasiveness and patient survival. Group A, OS <24 months; group B, OS >24 month.; FLT4, Fms-related tyrosine kinase 4; MDM2, murine double minute 2; MMP-2, matrix metallopeptidase 2; rel., relative; OS, overall survival.

Figure 4.

Immunohistochemical images of glioblastoma stained for FLT4/VEGF-3. (A) Overexpression of FLT4/VEGF-3 in patients with glioblastoma with a worse prognosis. (B) Moderate positivity in patients with a better prognosis (staining is notably reduced compared with that in patients with a worse prognosis). (C) Negative control. All images depicted are at ×20 magnification. FLT4, Fms-related tyrosine kinase 4; VEGF, vascular endothelial growth factor.

Figure 5.

Immunohistochemical images of glioblastoma stained for MDM2. (A) Immunhistochemical images from patients with a worse prognosis demonstrated significantly increased positivity for MDM2, compared with (B) those with a better prognosis. (C) Negative control. All images are depicted at ×20 magnification. MDM2, murine double minute 2.

Figure 6.

Immunohistochemical images of glioblastoma stained for MMP-2. (A) MMP-2 positivity was notably strong in immunohistochemical images from patients with a worse prognosis, confirming the mRNA expression results. (B) Patients with a better prognosis expressed an increased amount of MMP-2, as the strong positivity indicates, but the immunohistochemical scores are significantly reduced. (C) Negative control slide. All images are depicted at ×20 magnification. MMP-2, matrix metallopeptidase 2.