Gliomas are the most common neoplasms of the Central Nervous System (CNS) and a frequent cause of mental impairment and death. Gliomas are grouped into four clinical World Health Organization (WHO) grades according to their degree of malignancy. Grade I tumors are biologically "benign" and can be surgically cured, if resectable, at the time of diagnosis; grade II and III are more malignant and progressively transforming into grade IV glioblastoma multiforme (GBM) the most malignant form of tumor lethal within 9-12 months. Despite the improved technology for glioma molecular characterization, diagnosis and grading is still based on histological criteria. In the past few years, it has become apparent that Notch signalling, an important player in cell fate decision, proliferation, differentiation and normal development of CNS is often misregulated in GBM (1). The Notch family of transmembrane receptors comprises Notch-1, -2, -3 and -4. Notch signalling, a ligand-receptor initiated pathway, activates a diverse repertoire of genes regulating different cellular functions and for this reason has been called "gatekeeper against differentiation". In addition, recently it has been observed that Notch signalling is essential for the maintenance of neural stem cells (NSC) (2) and that promotes the formation of cancer stem cells (CSCs) in gliomas (3). Starting from these evidences we decided to examine the expression Notch1-4 receptors in 1) glioma cell lines and human primary glioma cell cultures grown in the presence of serum or in neurosphere-forming medium and in biopsies of different grade human gliomas. The expression of Notch was compared to the expression of proteins such as nestin, CD133, vimentin, and GFAP both in cultures and in human specimens. Results obtained show a differential expression of Notch receptors among the diverse glioma cell lines. In fact, Notch1 was more expressed in cell lines classified as astrocytoma that express moderate to high GFAP level and low vimentin level. Notch 4 expression was high in glioma cell lines, which derive from highly invasive Grade III and IV human tumors, that express very low levels of GFAP and moderate to high vimentin levels. Similar results were obtained in human glioma biopsies analyzed by Western blot: in glioma tissue of grade II Notch1 expression was moderate, while was low in grade IV samples. Notch4 expression was low in grade II sample and moderate to very high in high grade ones. These results were corroborated by scoring Noch1-4 immunohistochemistry in human glioma specimens. Notch1 immunoreactivity (IR) was strong in low grade gliomas but low in high grade ones. In contrast, Notch4 IR was increasing from AstrII to GBM. In primary cultures, obtained from human freshly resected gliomas, grown in the presence of serum, the expression of Notch1 and Notch4 was similar to that observed in the biopsies. In contrast, when cultures were maintained in neurosphere forming medium Notch4 expression was drastically reduced. Interestingly, levels of CD133 and nestin were moderate to high in serum and low in neurospheres by both Western blot and immunocytochemistry analysis. Our data suggest that Notch1 is a marker of glioma cell differentiation whereas Notch4 is a marker of less differentiated gliomas. In addition, glioma cell cultures grown in the presence of serum exhibit molecular signature of less differentiated cells compared to neurosphere. 1)Dell'Albani P (2008) Neurochem Res 33: 2407-2415. 2)Hitoshi S. et al. (2002) Genes & Dev., 16: 846-858. 3)Shih AH, Holland EC (2006) Neoplasia 8(12): 1072-82.
Notch receptors as marker of glioma grading
Dell'Albani P;Pellitteri R;Rodolico M;Albanese V;Catania MV
2011
Abstract
Gliomas are the most common neoplasms of the Central Nervous System (CNS) and a frequent cause of mental impairment and death. Gliomas are grouped into four clinical World Health Organization (WHO) grades according to their degree of malignancy. Grade I tumors are biologically "benign" and can be surgically cured, if resectable, at the time of diagnosis; grade II and III are more malignant and progressively transforming into grade IV glioblastoma multiforme (GBM) the most malignant form of tumor lethal within 9-12 months. Despite the improved technology for glioma molecular characterization, diagnosis and grading is still based on histological criteria. In the past few years, it has become apparent that Notch signalling, an important player in cell fate decision, proliferation, differentiation and normal development of CNS is often misregulated in GBM (1). The Notch family of transmembrane receptors comprises Notch-1, -2, -3 and -4. Notch signalling, a ligand-receptor initiated pathway, activates a diverse repertoire of genes regulating different cellular functions and for this reason has been called "gatekeeper against differentiation". In addition, recently it has been observed that Notch signalling is essential for the maintenance of neural stem cells (NSC) (2) and that promotes the formation of cancer stem cells (CSCs) in gliomas (3). Starting from these evidences we decided to examine the expression Notch1-4 receptors in 1) glioma cell lines and human primary glioma cell cultures grown in the presence of serum or in neurosphere-forming medium and in biopsies of different grade human gliomas. The expression of Notch was compared to the expression of proteins such as nestin, CD133, vimentin, and GFAP both in cultures and in human specimens. Results obtained show a differential expression of Notch receptors among the diverse glioma cell lines. In fact, Notch1 was more expressed in cell lines classified as astrocytoma that express moderate to high GFAP level and low vimentin level. Notch 4 expression was high in glioma cell lines, which derive from highly invasive Grade III and IV human tumors, that express very low levels of GFAP and moderate to high vimentin levels. Similar results were obtained in human glioma biopsies analyzed by Western blot: in glioma tissue of grade II Notch1 expression was moderate, while was low in grade IV samples. Notch4 expression was low in grade II sample and moderate to very high in high grade ones. These results were corroborated by scoring Noch1-4 immunohistochemistry in human glioma specimens. Notch1 immunoreactivity (IR) was strong in low grade gliomas but low in high grade ones. In contrast, Notch4 IR was increasing from AstrII to GBM. In primary cultures, obtained from human freshly resected gliomas, grown in the presence of serum, the expression of Notch1 and Notch4 was similar to that observed in the biopsies. In contrast, when cultures were maintained in neurosphere forming medium Notch4 expression was drastically reduced. Interestingly, levels of CD133 and nestin were moderate to high in serum and low in neurospheres by both Western blot and immunocytochemistry analysis. Our data suggest that Notch1 is a marker of glioma cell differentiation whereas Notch4 is a marker of less differentiated gliomas. In addition, glioma cell cultures grown in the presence of serum exhibit molecular signature of less differentiated cells compared to neurosphere. 1)Dell'Albani P (2008) Neurochem Res 33: 2407-2415. 2)Hitoshi S. et al. (2002) Genes & Dev., 16: 846-858. 3)Shih AH, Holland EC (2006) Neoplasia 8(12): 1072-82.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
