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J Biomol Tech. 2007 February; 18(1): 11–12.
PMCID: PMC2291864

P31-S Identification of Protein Biomarkers Associated with Hypoxia in Human Malignant Glioma Cell Lines Using Proteomics Technologies

Abstract

The most common type of primary malignant brain tumors are glioblastomas. These highly aggressive, rapidly growing tumors are exposed to hypoxia, which occurs as a consequence of inadequate blood supply. Hypoxia exerts a variety of influences on tumor cell biology. Among these are activation of signal transduction pathways, adapting hypoxic tumor cells to an anaerobic environment.

As a complementary approach to gene expression profiling, we aimed to investigate changes in the overall protein pattern of malignant glioma cell lines after hypoxia treatment compared to normoxic controls utilizing various proteomics techniques.

The human malignant glioma cell line LNT-229 was initially used in our studies. Three individual samples representing three cell cultures grown under identical conditions were taken at four different time-points of hypoxia treatment (control, 8, 24 and 72 h). Proteins from total cell lysates were separated by high-resolution 2D gel electrophoresis and visualized by silver staining. Computer-assisted image analysis of the gels allowed the detection of differentially expressed proteins.

Proteins spots that were recognized to be up- or down-regulated were identified by mass spectrometry in preparative gels. We have found 14 proteins up-regulated and 6 proteins down-regulated in hypoxia compared to normoxia based on the image analysis of the corresponding 2D gels. These results are currently being verified by Western-blot analysis of samples from three different glioma cell lines—LN-18, U87, and LNT229. Furthermore, all candidate proteins will be evaluated on primary cell cultures from human gliomas, and immunohistochemically on glioma tissue microarrays.


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