Neural Stem Cells Adopt Tumorigenic Properties by Constitutively Activated NF-κB and Subsequent VEGF Up-Regulation

Abstract

One of the challenges in stem cell research is to avoid transformation during cultivation. We studied high passage subventricular zone derived neural stem cells (NSCs) cultures of adult rats in the absence of growth factors epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF). We termed this culture exogenous growth factor independent neural stem cells (GiNSCs). GiNSCs expressed stemness markers, displayed a high constitutive NF-kappaB activity and an increased, aberrant, polyploid DNA content. GiNSCs showed a tumorigenic phenotype and formed colonies in a soft agar assay. Microarray analysis showed the up-regulation of the NF-kappaB target gene vascular endothelial growth factor (VEGF). In contrast, proneuronal genes were down-regulated. Under neuronal differentiation conditions GiNSCs adopted a glioma-like phenotype, with nuclear p53, preserving high amounts of Nestin positive cells and prolonged proliferation. Neutralization of VEGF strongly inhibited proliferation and induced differentiation. In a gain of function approach, the transfection of NSCs with constitutively active upstream kinase IKK-2 led to constitutively activated NF-kappaB, proliferation in absence of growth factors and augmented VEGF secretion. In a rescue experiment a reduction of NF-kappaB activity by overexpression of IkappaB-AA1 was able to shift the morphology toward an elongated cell form, increased cell death, and decreased proliferation. Thus GiNSCs may provide a potent tool in cancer research, as their exogenous cytokine independent proliferation and their constitutively high NF-kappaB expression presumes cancerous properties observed in gliomas. In addition, this study might add a novel mechanism for detecting oncogenic transformation in therapeutic stem cell cultures.

Characterization and comparison of telomere length, telomerase and reverse transcriptase activity and gene expression in human mesenchymal stem cells and cancer cells of various origins

“…Since NF-κB regulates the expression of numerous genes that are involved in carcinogenesis, the expression of these genes might be suppressed through the inhibition of NF-κB activation (Prasad et al 2010). Most importantly, NF-κB regulates the expression of VEGF, a growth factor necessary for angiogenesis and an abundant cytokine in the tumor microenvironment (Kaus et al 2010). In addition, various activation pathways of NF-κB might cause the expression of proteins that promote apoptosis (e.g., p53) or inhibit apoptosis (e.g., BCL-2-like proteins; Prasad et al 2010).…”

IKK/NF-κB signaling contributes to glioblastoma stem cell maintenance

“…In normal neural stem cells (NSCs), TNFα has been shown to induce proliferation through NF-κB [22]. Kaus et al [23] have shown that as NSCs acquire the ability to proliferate independent of exogenous growth factors, these cells demonstrate increased NF-κB activity. In GBM, NF-κB has been reported to regulate survival, invasion, and resistance to both radiation and chemotherapy [24–28].…”

Prolonged cultivation of hippocampal neural precursor cells shifts their differentiation potential and selects for aneuploid cells

“…Stem cells from the adult rat SVZ were likewise shown to transform into tumorigenic cell lines after their expansion in vitro, as demonstrated by multiple acquired chromosomal aberrations and malignant tumor formation after syngeneic transplantation into the brain of adult rats (Siebzehnrubl et al, 2009). As reported by Kaus et al (2010), long-term culture of adult rat NSCs not only results in aberrant, aneuploid DNA contents, and malignant tumor formation after in vivo transplantation but also in cell growth independent to exogenous growth factors. Substantiating these findings, we observed abnormal chromosome numbers as well as growth factor-independent proliferation in NPCs from murine adult hippocampus after long-and intermediate-term culture.…”

“…Metaphase chromosomes of NPCs were prepared according as described by Kaus et al (2010). In particular, 1 × 10 7 NPCs of respective passages (p6, p12, p19, and p27) as well as MEFs were metaphase-arrested, treated with pre-warmed hypotonic solution, and progressively fixed in methanol/acetate.…”