Diffuse Low-Grade Astrocytomas: P53-Mediated Inhibition of Angiogenesis

Abstract

Angiogenesis is playing a crucial role in the growth and progression of astrocytomas. P53 a tumor suppressor gene located on chromosome 17p has been implicated to the regulation of cell death, particularly apoptosis, proliferation and also angiogenesis. In this in vitro study we evaluated the association of p53 gene status (wild-type or mutated) with micro vascular density in a set of astrocytomas. Immunohistochemistry for CD31, a surface marker expressed on endothelial cells, was performed on 23 diffuse astrocytomas (WHO Grade II). Mutation status of the p53 gene was identified by PCR amplification with consequent sequencing of genomic DNA extracted from each tumor tissue. Intratumoural or peritumoural microvascular hot spots were assessed and images taken at a 200× fold magnification. Microvessel count was performed with a modern automatic image analyses algorithm by using these images. P53 mutation occurred in 11 out of 23 (47%) astrocytomas. In p53 mutated gliomas the micro-vascular density and the absolute vessel number was significantly higher compared to p53 wild-type gliomas, thereby supporting the hypothesis of a p53-mediated regulation on angiogenesis in diffuse low-grade astrocytomas. To analyze a possible molecular mechanisms between these two factors, LN229, a glioma cell line, harbouring a p53 mutation, was transfected with p53 wild-type and empty vector, as a negative control. A protein array analysis provided evidence that Thrombospondin-1, Coagulation factor III, Serpin E1 and MMP-9 are potential p53 targets and important key players in regulating angiogenesis in gliomas. Our results support the hypotheses that p53 regulates angiogenesis in low grade astrocytomas.