Differential patterns of Notch1-4 receptor expression are markers of glioma cell differentiation.

Background. Notch signaling is deregulated in human gliomas and may play a role in their malignancy. However, the role of each Notch receptor in glioma cell differentiation and progression is not clear. We examined the expression pattern of Notch receptors and compared it with differentiation markers in gliomas' i) cell lines, ii) primary human cultures, and iii) biopsies of different grade. Furthermore, the effects of a ?-secretase-inhibitor (GSI) on cell survival were assessed. Methods. Notch receptors and markers of cellular differentiation were analyzed by Reverse-Transcriptase Polymerase Chain Reaction (RT-PCR) , Western blotting, immunohistochemistry and immunocytochemistry. GSI sensitivity was assessed in both cell lines and primary cultures grown as monolayer or tumorspheres, by MTT assay. Results. In cell lines Notch1 and Notch2/4 levels paralleled those of GFAP and vimentin, respectively. In human gliomas and in primary cultures Notch1 was moderate/strong in low grade, but weak in GBMs. Conversely, Notch4 increased from AstrII to GBMs. Primary GBM cultures grown in serum (monolayer) showed moderate/high levels of CD133, nestin, vimentin and Notch4 and very low levels of GFAP and Notch1, which were reduced in tumorspheres. This effect was drastic for Notch4. GSI reduced cell survival with stronger effect in serum, whilst human primary cultures showed different sensitivity. Conclusion. Data from cell lines and human gliomas suggest a correlation between expression of Notch receptors and cell differentiation. Namely, Notch1 and Notch4 are markers of differentiated and less differentiated glioma cells, respectively. We propose Notch receptors as markers of glioma grading and possible prognostic factors.