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Microglia are increasingly implicated as a source of non-neural regulation of postnatal neurogenesis and neuronal development. To evaluate better the contributions of microglia to neural stem cells (NSCs) of the subventricular neuraxis, we employed an adherent culture system that models the continuing proliferation and differentiation of the dissociated neuropoietic subventricular tissues. In this model, neuropoietic cells retain the ability to self-renew and form multipotent neurospheres, but progressively lose the ability to generate committed neuroblasts with continued culture. Neurogenesis in highly expanded NSCs can be rescued by coculture with microglial cells or microglia-conditioned medium, indicating that microglia provide secreted factor(s) essential for neurogenesis, but not NSC maintenance, self-renewal, or propagation. Our findings suggest an instructive role for microglial cells in contributing to postnatal neurogenesis in the largest neurogenic niche of the mammalian brain.

The tyrosine kinase c-Met promotes the formation and malignant progression of multiple cancers. It is well known that c-Met hyperactivation increases tumorigenicity and tumor cell resistance to DNA damaging agents, properties associated with tumor-initiating stem cells. However, a link between c-Met signaling and the formation and/or maintenance of neoplastic stem cells has not been previously identified. Here, we show that c-Met is activated and functional in glioblastoma (GBM) neurospheres enriched for glioblastoma tumorinitiating stem cells and that c-Met expression/function correlates with stem cell marker expression and the neoplastic stem cell phenotype in glioblastoma neurospheres and clinical glioblastoma specimens. c-Met activation was found to induce the expression of reprogramming transcription factors (RFs) known to support embryonic stem cells and induce differentiated cells to form pluripotent stem (iPS) cells, and c-Met activation counteracted the effects of forced differentiation in glioblastoma neurospheres. Expression of the reprogramming transcription factor Nanog by glioblastoma cells is shown to mediate the ability of c-Met to induce the stem cell characteristics of neurosphere formation and neurosphere cell self-renewal. These findings show that c-Met enhances the population of glioblastoma stem cells (GBM SCs) via a mechanism requiring Nanog and potentially other c-Met-responsive reprogramming transcription factors.cancer stem cell | hepatocyte growth factor | Sox2 | Oct4 | Klf4 G lioblastomas (GBMs) are heterogeneous aggressive neoplasms containing neoplastic stem-like cells (1). These cells commonly referred to as glioblastoma stem cells (GBM SCs), exhibit the capacity for unlimited growth as multicellular spheres in defined medium, multilineage differentiation, and efficient tumor initiation in immune-deficient animals. GBM SCs are currently believed to play a leading role in therapeutic resistance and tumor recurrence (2). Defining the origin(s) of GBM SCs and the biochemical/molecular pathways that support the stem-like tumor-initiating phenotype is of major importance.Transcription factors such as Sox2, c-Myc, Klf4, Oct4, and Nanog have an essential role in sustaining the growth and selfrenewal of embryonic stem (ES) cells. Introducing these transcription factors into mouse and human differentiated somatic cells results in their reprogramming into pluripotent ES-like cells called induced pluripotent stem (iPS) cells (3). Remarkable similarities exist between stem cell reprogramming and oncogenesis. Both processes are supported by alterations in the expression/function of similar collaborating genes perpetuating subpopulations of cells capable of indefinite self-renewal (4). Reprogramming transcription factors (RFs) display varying degrees of oncogenic potential, are overexpressed in human cancers, and their expression levels have been correlated with malignant progression and poor prognosis (5, 6). Loss of tumor suppressors such as p53 enhances the efficiency of iPS cell generation b...

The modern concept of neurogenesis in the adult brain is predicated on the premise that multipotent glial cells give rise to new neurons throughout life. Although extensive evidence exists indicating that this is the case, the transition from glial to neuronal phenotype remains poorly understood. A unique monolayer cellculture system was developed to induce, expose, and recapitulate the entire developmental series of events of subventricular zone (SVZ) neurogenesis. We show here, using immunophentoypic, ultrastructural, electrophysiological, and time-lapse analyses, that SVZ-derived glial fibrillary acidic protein low ͞A2B5 ؉ ͞nestin ؉ candidate founder cells undergo metamorphosis to eventually generate large numbers of fully differentiated interneuron phenotypes. A model of postnatal neurogenesis is considered in light of known embryonic events and reveals a limited developmental potential of SVZ stem͞progenitor cells, whereby ancestral cells in both embryonic and postnatal͞adult settings give rise to glia and GABAergic interneurons.adult stem cells ͉ electrophysiology ͉ in vitro ͉ neurogenesis ͉ subventricular zone A ttempts to trace the cellular source of neurogenesis in the adult CNS have recently led to the surprising conclusion that dedicated glial cells give rise to new neurons throughout life (1-5). Even though neurons and glia are both derived from the embryonic neuroepithelium, sharing common signaling pathways and downstream transcription factors during development (6), it is difficult to imagine how one major cell class in the adult brain can transpose into the other. Postnatal neurogenesis in the subventricular zone (SVZ) of rodents proceeds as a characteristic series of events, where multipotent glial cells (referred to as type-B cells) can divide to form colonies of neuroblasts (type-A cells) through a transit-amplifying cell population (type-C cells) (7). Newborn neuroblasts migrate from the SVZ through the rostral migratory stream and mature to GABAergic granule cells and periglomerular cells, which, 3-4 weeks after generation, integrate as inhibitory interneurons into the olfactory bulb of rodents (8-10). Certain features, such as nestinand glial-fibrillary acidic protein (GFAP) expression, are ascribed to the founder cells of postnatal neurogenesis (3,(11)(12)(13)(14), but their distinctive antigenic and functional profiles remain elusive. Traditional approaches for the isolation and characterization of persistent neurogenesis have relied on the in vitro neurosphere (NS) assay (15, 16) or on post hoc identification, depending on the incorporation of BrdUrd and͞or retroviral constructs to label precursors during cell division. However, neither of these methods affords the recognition of the dynamic processes involved in the maturation of individual cells en route from stem cells to fully differentiated neural phenotypes.Here, we present an alternative culture model that closely recapitulates in vivo postnatal͞adult SVZ neurogenesis, allowing us to monitor the entire sequence of hierarchical eve...

Summary: Background and Purpose: Focal cortical dysplasia of Taylor's balloon-cell type (FCD-BC) are a frequent cause of pharmacoresistant epilepsy in young patients. In order to characterize FCD-BC, we coupled MRI and histopathology, and analyzed the clinical outcome following epilepsy surgery.Methods: From an epilepsy data bank with 547 histological specimens, 17 FCD-BC were re-evaluated of which high resolution MRI was available. Five additional FCD-BC were prospectively identified by MRI. Histopathological and immunohistochemical features were related to MRI. Outcome following lesionectomy was analyzed as determined on routine examinations 3, 6 and 12 months following surgery.Results: All but one lesion were located outside the temporal lobe. A markedly hyperintense funnel-shaped subcortical zone tapering towards the lateral ventricle was the characteristic finding on FLAIR MRI. Histopathologically, the subcortical zone of the FCD-BC displayed hypomyelinated white matter with radially oriented balloon cells and gliosis. Dysplastic neurons were found in the adjacent, disorganized cortex. All patients with complete lesionectomy were seizure free one year following surgery. Conclusion: Focal cortical dysplasias of Taylor's ballooncell type (FCD-BC) have characteristic MRI and histopathological findings. MRI recognition is important, since outcome following resective surgery is favorable. Key words: focal cortical dysplasia-balloon cells-epilepsy surgery-MRI Malformations of the cerebral cortex are a frequent cause of pharmacoresistant epilepsies and developmental disorders (1). Various terms have been introduced to classify the underlying pathology such as cortical dysgenesis (2,3), focal cortical dysplasias (4) or glioneuronal hamartias/hamartomas (5,6). Unfortunately, the nomenclature is not uniform, and different diagnostic terms are even used for malformative lesions that appear identical on histological specimens. One of these peculiar lesions is characterized by focal cortical disorganization with enlarged, dysplastic neurons and the presence of bizarre balloon cells within the cortex and subcortical white matter. Taylor et al. first described it in 1971 (7) detailing the history of ten patients suffering from longstanding epilepsy: Six of ten lesions contained "grotesque cells, probably of glial origin," within the cortex and subcortical white matter, and four lesions did not.Both lesion types were initially merged and denominated as focal cortical dysplasia of Taylor. Other neuropathological terms for the lesions containing balloon cells include focal cortical dysplasia (2), focal transmantle cortical dysplasia (8), focal cortical dysplasia of Taylor, balloon cell subtype (9), severe cortical dysplasia, type II (10), type II focal cortical dysplasia (11), forme fruste of tuberous sclerosis or type III focal cortical dysplasia (11), balloon cell changes (4) and glioneuronal hamartoma with TS cells (5).With the recent progress in magnetic resonance imaging (MRI), malformations of the cerebral cortex are i...

Purpose: Glioblastoma is a highly malignant, invariably fatal brain tumor for which effective pharmacotherapy remains an unmet medical need.Experimental Design: Screening of a compound library of 160 synthetic and natural toxic substances identified the antihelmintic niclosamide as a previously unrecognized candidate for clinical development. Considering the cellular and interindividual heterogeneity of glioblastoma, a portfolio of short-term expanded primary human glioblastoma cells (pGBM; n ¼ 21), common glioma lines (n ¼ 5), and noncancer human control cells (n ¼ 3) was applied as a discovery platform and for preclinical validation. Pharmacodynamic analysis, study of cell-cycle progression, apoptosis, cell migration, proliferation, and on the frequency of multipotent/self-renewing pGBM cells were conducted in vitro, and orthotopic xenotransplantation was used to confirm anticancer effects in vivo.Results: Niclosamide led to cytostatic, cytotoxic, and antimigratory effects, strongly reduced the frequencies of multipotent/self-renewing cells in vitro, and after exposure significantly diminished the pGBMs' malignant potential in vivo. Mechanism of action analysis revealed that niclosamide simultaneously inhibited intracellular WNT/CTNNB1-, NOTCH-, mTOR-, and NF-kB signaling cascades. Furthermore, combinatorial drug testing established that a heterozygous deletion of the NFKBIA locus in glioblastoma samples could serve as a genomic biomarker for predicting a synergistic activity of niclosamide with temozolomide, the current standard in glioblastoma therapy.Conclusions: Together, our data advocate the use of pGBMs for exploration of compound libraries to reveal unexpected leads, for example, niclosamide that might be suited for further development toward personalized clinical application. Clin Cancer Res; 19(15); 4124-36. Ó2013 AACR.

Glioblastoma (GBM) remains the most pervasive and lethal of all brain malignancies. One factor that contributes to this poor prognosis is the highly invasive character of the tumor. GBM is characterized by microscopic infiltration of tumor cells throughout the brain, whereas non-neural metastases, as well as select lower grade gliomas, develop as self-contained and clearly delineated lesions. Illustrated by rodent xenograft tumor models as well as pathological human patient specimens, we present evidence that one fundamental switch between these two distinct pathologies-invasion and noninvasion-is mediated through the tumor extracellular matrix. Specifically, noninvasive lesions are associated with a rich matrix containing substantial amounts of glycosylated chondroitin sulfate proteoglycans (CSPGs), whereas glycosylated CSPGs are essentially absent from diffusely infiltrating tumors. CSPGs, acting as central organizers of the tumor microenvironment, dramatically influence resident reactive astrocytes, inducing their exodus from the tumor mass and the resultant encapsulation of noninvasive lesions. Additionally, CSPGs induce activation of tumor-associated microglia. We demonstrate that the astrogliotic capsule can directly inhibit tumor invasion, and its absence from GBM presents an environment favorable to diffuse infiltration. We also identify the leukocyte common antigen-related phosphatase receptor (PTPRF) as a putative intermediary between extracellular glycosylated CSPGs and noninvasive tumor cells. In all, we present CSPGs as critical regulators of brain tumor histopathology and help to clarify the role of the tumor microenvironment in brain tumor invasion.

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