Neuroepithelial stem cell marker nestin regulates the migration, invasion and growth of human gliomas

Abstract

Abstract. Nestin, a class VI intermediate filament protein, was originally described as a neuronal stem cell marker during central nervous system development. Nestin is expressed in gliomas, and its expression levels are higher in gliomas with high WHO histopathological classification grades than in those with low grades. In the present study, we examined whether nestin regulates the biological activities of human glioma cells. Immunohistochemically, the nestin expression patterns in 10 human glioblastoma patients were examined. The expression levels of nestin in A172, a human high-grade glioma cell line, and KG-1-C, a human low-grade glioma cell line, were examined using real-time PCR, Western blot and immunofluorescence analyses. An expression vector carrying a short hairpin RNA targeting nestin was stably transfected into A172 (Sh) cells. The effects of decreased expression levels of nestin in Sh cells on cell growth, migration, invasion, adhesion to extracellular matrices and fibrillar actin expression on three-dimensional culture plates were examined. The nestin expression vector was transiently transfected into KG-1-C (Nes) cells, and the effects of the nestin overexpression on cell growth and migration were examined. Nestin was expressed in the cytoplasm of the glioblastoma cells in all cases examined. Sh cells showed marked decreases in the expression levels of nestin mRNA and protein, and the growth rate of Sh cells was lower than that of sham (Sc) cells. In contrast, the adhesion activity of Sh cells to types I and IV collagens, fibronectin and laminin was higher than that of Sc cells. Fibrillar actin was clearly detected at the periphery of colonies of Sh cells at the attachment sites on three-dimensional culture plates. The migration and invasion of Sh cells were markedly inhibited compared with those of Sc cells. In contrast, the levels of nestin expression markedly increased in the Nes cells, which were transiently transfected with the nestin expression vector. The growth rate and motility of Nes cells were higher than those of the mock cells. In conclusion, nestin plays important roles in cell growth, migration, invasion and adhesion to extra-cellular matrices in glioma cells. Nestin may serve as a novel candidate for molecular-targeted therapy for gliomas, including glioblastomas.

Selective CREB-dependent cyclin expression mediated by the PI3K and MAPK pathways supports glioma cell proliferation

“…In one GBM tumour examined, strong pCREB-positive cells were clustered in foci exhibiting strong nestin co-labelling (Figure 1b and vi). Nestin, an intermediate filament protein, is highly expressed in immature NSPCs and, similar to pCREB, is associated with brain tumour grade 43, 44, 45 This suggests that CREB may contribute to immature tumour cell biology by regulating CREB target genes involved in tumour cell growth and suppression of differentiation/promotion of immaturity; this is supported by studies showing that PI3K regulation of CREB is crucial for NSPC function. 46 Our data also shows that less aggressive GBM tumour subtypes, known to be associated with better survival, express lower levels of activated CREB.…”

Expression and role of nestin in human cervical intraepithelial neoplasia and cervical cancer

“…Then, cDNA synthesis was performed using High Capacity cDNA Reverse Transcription kit following the manufacturer’s protocol (Applied Biosystems, Carlsbad, CA). Real-time PCR for nestin and 18S rRNA was performed using the StepOnePlus real-time PCR system (Applied Biosystems) with specific primers for nestin (Hs00707120_s1) and 18S rRNA (Hs03928990_g1) and a TaqMan probe (7). …”

“…Nestin contains a short N-terminus and an unusually long C-terminus. It is assembled into IF by forming heterodimers with vimentin and desmin, but nestin cannot form homopolymers (7–10). …”

Cancer stem cell contribution to glioblastoma invasiveness

“…Cells at the leading edge of the tumor have been found to be positive for putative stem cell markers such as L1CAM [12], nucleostemin [13], and nestin [14-16], supporting the notion that CSCs are indeed responsible for GBM invasion. A recent paper provided new insights into the role of SOX2 as a novel determinant of the invasive and migration properties of GBM CSCs and glioma cell lines [17].…”

“…Several molecules have been used to identify invading GBM cells (for example, nestin [15,16] and vimentin [64]), but their expression could be altered by the grade of differentiation of the tumors. NSC discovery in the adult brain and the strong relationship between NSCs and CSCs have led to the consideration that these two types of cells may share common markers as well.…”