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Identifying cellular programs that drive cancers to be stem-like and treatment resistant is critical to improving outcomes in patients. Here, we demonstrate that constitutive extracellular signal-regulated kinase 1/2 (ERK1/2) activation sustains a stem-like state in glioblastoma (GBM), the most common primary malignant brain tumor. Pharmacological inhibition of ERK1/2 activation restores neurogenesis during murine astrocytoma formation, inducing neuronal differentiation in tumorspheres. Constitutive ERK1/2 activation globally regulates miRNA expression in murine and human GBMs, while neuronal differentiation of GBM tumorspheres following the inhibition of ERK1/2 activation requires the functional expression of miR-124 and the depletion of its target gene SOX9. Overexpression of miR124 depletes SOX9 in vivo and promotes a stem-like-to-neuronal transition, with reduced tumorigenicity and increased radiation sensitivity. Providing a rationale for reports demonstrating miR-124-induced abrogation of GBM aggressiveness, we conclude that reversal of an ERK1/2-miR-124-SOX9 axis induces a neuronal phenotype and that enforcing neuronal differentiation represents a therapeutic strategy to improve outcomes in GBM., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)

Slow-cycling and treatment-resistant cancer cells escape therapy, providing a rationale for regrowth and recurrence in patients. Much interest has focused on identifying the properties of slow-cycling tumor cells in glioblastoma (GBM), the most common and lethal primary brain tumor. Despite aggressive ionizing radiation (IR) and treatment with the alkylating agent temozolomide (TMZ), GBM patients invariably relapse and ultimately succumb to the disease. In patient biopsies, we demonstrated that GBM cells expressing the proliferation markers Ki67 and MCM2 displayed a larger cell volume compared to rare slow-cycling tumor cells. In optimized density gradients, we isolated a minor fraction of slow-cycling GBM cells in patient biopsies and tumorsphere cultures. Transcriptional profiling, self-renewal, and tumorigenicity assays reflected the slow-cycling state of high-density GBM cells (HDGCs) compared to the tumor bulk of low-density GBM cells (LDGCs). Slow-cycling HDGCs enriched for stem cell antigens proliferated a few days after isolation to generate LDGCs. Both in vitro and in vivo, we demonstrated that HDGCs show increased treatment-resistance to IR and TMZ treatment compared to LDGCs. In conclusion, density gradients represent a non-marker based approach to isolate slow-cycling and treatment-resistant GBM cells across GBM subgroups., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Mechanical cues regulate neuronal function and reactivity of glial cells, the origin of gliomas. In this issue of Neuron, Chen et al. (2018) uncover a feedforward loop mediated by the mechanosensitive ion channel Piezo1 and tissue stiffness that drives glioma aggression., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The weak immunogenicity of gliomas presents a barrier for effective immunotherapy. Na/H exchanger isoform 1 (NHE1) maintains alkaline intracellular pH (pH i ) of glioma cells and acidic microenvironment. In addition, NHE1 is expressed in tumor-associated microglia and tumor-associated macrophages (TAMs) and involved in protumoral communications between glioma and TAMs. Therefore, we hypothesize that NHE1 plays a role in developing tumor resistance and immunosuppressive tumor microenvironment. In this study, we investigated the efficacy of pharmacological inhibition of NHE1 on combinatorial therapies. Here we show that temozolomide (TMZ) treatment stimulates NHE1 protein expression in two intracranial syngeneic mouse glioma models (SB28, GL26). Pharmacological inhibition of NHE1 potentiated the cytotoxic effects of TMZ, leading to reduced tumor growth and increased median survival of mice. Blockade of NHE1 stimulated proinflammatory activation of TAM and increased cytotoxic T cell infiltration into tumors. Combining TMZ, anti-PD-1 antibody treatment with NHE1 blockade significantly prolonged the median survival in the mouse glioma model. These results demonstrate that pharmacological inhibition of NHE1 protein presents a new strategy for potentiating TMZ-induced cytotoxicity and increasing tumor immunogenicity for immunotherapy to improve glioma therapy.

The goal of this research was to elucidate the relationship between WHO 2016 molecular classifications of newly diagnosed, nonenhancing lower grade gliomas (LrGG), tissue sample histopathology, and magnetic resonance (MR) parameters derived from diffusion, perfusion, and 1 H spectroscopic imaging from the tissue sample locations and the entire tumor. A total of 135 patients were scanned prior to initial surgery, with tumor cellularity scores obtained from 88 image-guided tissue samples. MR parameters were obtained from corresponding sample locations, and histograms of normalized MR parameters within the T2 fluid-attenuated inversion recovery lesion were analyzed in order to evaluate differences between subgroups. For tissue samples, higher tumor scores were related to increased normalized apparent diffusion coefficient (nADC), lower fractional anisotropy (nFA), lower cerebral blood volume (nCBV), higher choline (nCho), and lower N-acetylaspartate (nNAA). Within the T2 lesion, higher tumor grade was associated with higher nADC, lower nFA, and higher Cho to NAA index. Pathological analysis confirmed that diffusion and metabolic parameters increased and perfusion decreased with tumor cellularity. This information can be used to select targets for tissue sampling and to aid in making decisions about treating residual disease., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

Gliomas comprise heterogeneous malignant glial and stromal cells. While blood vessel co-option is a potential mechanism to escape anti-angiogenic therapy, the relevance of glial phenotype in this process is unclear. We show that Olig2 + oligodendrocyte precursor-like glioma cells invade by single-cell vessel co-option and preserve the blood-brain barrier (BBB). Conversely, Olig2-negative glioma cells form dense perivascular collections and promote angiogenesis and BBB breakdown, leading to innate immune cell activation. Experimentally, Olig2 promotes Wnt7b expression, a finding that correlates in human glioma profiling. Targeted Wnt7a/7b deletion or pharmacologic Wnt inhibition blocks Olig2 + glioma single-cell vessel co-option and enhances responses to temozolomide. Finally, Olig2 and Wnt7 become upregulated after anti-VEGF treatment in preclinical models and patients. Thus, glial-encoded pathways regulate distinct glioma-vascular microenvironmental interactions., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Interstitial fluid pressure (IFP) presents a barrier to drug uptake in solid tumors, including the aggressive primary brain tumor glioblastoma (GBM). It remains unclear how fluid dynamics impacts tumor progression and can be targeted therapeutically. To address this issue, a novel telemetry-based approach was developed to measure changes in IFP during progression of GBM xenografts. Antisecretory factor (AF) is an endogenous protein that displays antisecretory effects in animals and patients. Here, endogenous induction of AF protein or exogenous administration of AF peptide reduced IFP and increased drug uptake in GBM xenografts. AF inhibited cell volume regulation of GBM cells, an effect that was phenocopied in vitro by the sodium-potassium-chloride cotransporter 1 (SLC12A2/NKCC1) inhibitor bumetanide. As a result, AF induced apoptosis and increased survival in GBM models. In vitro , the ability of AF to reduce GBM cell proliferation was phenocopied by bumetanide and NKCC1 knockdown. Next, AF's ability to sensitize GBM cells to the alkylating agent temozolomide, standard of care in GBM patients, was evaluated. Importantly, combination of AF induction and temozolomide treatment blocked regrowth in GBM xenografts. Thus, AF-mediated inhibition of cell volume regulation represents a novel strategy to increase drug uptake and improve outcome in GBM. Mol Cancer Res; 16(5); 777-90. ©2018 AACR ., (©2018 American Association for Cancer Research.)

Glioblastoma (GBM) is the most common primary malignant brain tumor of adults and confers a poor prognosis due, in part, to diffuse invasion of tumor cells. Heparan sulfate (HS) glycosaminoglycans, present on the cell surface and in the extracellular matrix, regulate cell signaling pathways and cell-microenvironment interactions. In GBM, the expression of HS glycosaminoglycans and the enzymes that regulate their function are altered, but the actual HS content and structure are unknown. However, inhibition of HS glycosaminoglycan function is emerging as a promising therapeutic strategy for some cancers. In this study, we use liquid chromatography-mass spectrometry analysis to demonstrate differences in HS disaccharide content and structure across four patient-derived tumorsphere lines (GBM1, 5, 6, 43) and between two murine tumorsphere lines derived from murine GBM with enrichment of mesenchymal and proneural gene expression (mMES and mPN, respectively) markers. In GBM, the heterogeneous HS content and structure across patient-derived tumorsphere lines suggested diverse functions in the GBM tumor microenvironment. In GBM5 and mPN, elevated expression of sulfatase 2 (SULF2), an extracellular enzyme that alters ligand binding to HS, was associated with low trisulfated HS disaccharides, a substrate of SULF2. In contrast, other primary tumorsphere lines had elevated expression of the HS-modifying enzyme heparanase (HPSE). Using gene editing strategies to inhibit HPSE, a role for HPSE in promoting tumor cell adhesion and invasion was identified. These studies characterize the heterogeneity in HS glycosaminoglycan content and structure across GBM and reveal their role in tumor cell invasion. Implications: HS-interacting factors promote GBM invasion and are potential therapeutic targets. Mol Cancer Res; 15(11); 1623-33. ©2017 AACR ., (©2017 American Association for Cancer Research.)

Cranial radiation severely affects brain health and function, including glial cell production and myelination. Recent studies indicate that voluntary exercise has beneficial effects on oligodendrogenesis and myelination. Here, we hypothesized that voluntary running would increase oligodendrocyte numbers in the corpus callosum after irradiation of the juvenile mouse brain. The brains of C57Bl/6J male mice were 6 Gy irradiated on postnatal day 9 during the main gliogenic developmental phase, resulting in a loss of oligodendrocyte precursor cells. Upon adulthood, the mice were injected with bromodeoxyuridine and allowed to exercise on a running wheel for four weeks. Cell proliferation and survival, Ascl1 + oligodendrocyte precursor and Olig2 + oligodendrocyte cell numbers as well as CC1 + mature oligodendrocytes were quantified using immunohistology. Radiation induced a reduction in the number of Olig2 + oligodendrocytes by nearly 50% without affecting production or survival of new Olig2 + cells. Ascl1 + cells earlier in the oligodendroglial cell lineage were also profoundly affected, with numbers reduced by half. By three weeks of age, Olig2 + cell numbers had not recovered, and this was paralleled by a volumetric loss in the corpus callosum. The deficiency of Olig2 + oligodendrocytes persisted into adulthood. Additionally, the depletion of Ascl1 + progenitor cells was irreversible, and was even more pronounced at 12 weeks postirradiation compared to day 2 postirradiation. Furthermore, the overall number of CC1 + mature oligodendrocytes decreased by 28%. The depletion of Olig2 + cells in irradiated animals was reversed by 4 weeks of voluntary exercise. Moreover, voluntary exercise also increased the number of Ascl1 + progenitor cells in irradiated animals. Taken together, these results demonstrate that exercise in adulthood significantly ameliorates the profound and long-lasting effects of moderate exposure to immature oligodendrocytes during postnatal development.

High-grade gliomas in children are different from those that arise in adults. Recent collaborative molecular analyses of these rare cancers have revealed previously unappreciated connections among chromatin regulation, developmental signaling, and tumorigenesis. As we begin to unravel the unique developmental origins and distinct biological drivers of this heterogeneous group of tumors, clinical trials need to keep pace. It is important to avoid therapeutic strategies developed purely using data obtained from studies on adult glioblastoma. This approach has resulted in repetitive trials and ineffective treatments being applied to these children, with limited improvement in clinical outcome. The authors of this perspective, comprising biology and clinical expertise in the disease, recently convened to discuss the most effective ways to translate the emerging molecular insights into patient benefit. This article reviews our current understanding of pediatric high-grade glioma and suggests approaches for innovative clinical management., (© The Author(s) 2016. Published by Oxford University Press on behalf of the Society for Neuro-Oncology.)

Increased overall survival for patients with glioma brain tumours is associated with mutations in the metabolic regulator isocitrate dehydrogenase 1 (IDH1). Gliomas develop within a mechanically challenged microenvironment that is characterized by a dense extracellular matrix (ECM) that compromises vascular integrity to induce hypoxia and activate HIF1α. We found that glioma aggression and patient prognosis correlate with HIF1α levels and the stiffness of a tenascin C (TNC)-enriched ECM. Gain- and loss-of-function xenograft manipulations demonstrated that a mutant IDH1 restricts glioma aggression by reducing HIF1α-dependent TNC expression to decrease ECM stiffness and mechanosignalling. Recurrent IDH1-mutant patient gliomas had a stiffer TNC-enriched ECM that our studies attributed to reduced miR-203 suppression of HIF1α and TNC mediated via a tension-dependent positive feedback loop. Thus, our work suggests that elevated ECM stiffness can independently foster glioblastoma aggression and contribute to glioblastoma recurrence via bypassing the protective activity of IDH1 mutational status.

Glioblastoma multiforme (GBM) carries a poor prognosis and continues to lack effective treatments. Glioblastoma stem cells (GSCs) drive tumor formation, invasion, and drug resistance and, as such, are the focus of studies to identify new therapies for disease control. Here, we identify the involvement of IKK and NF-κB signaling in the maintenance of GSCs. Inhibition of this pathway impairs self-renewal as analyzed in tumorsphere formation and GBM expansion as analyzed in brain slice culture. Interestingly, both the canonical and non-canonical branches of the NF-κB pathway are shown to contribute to this phenotype. One source of NF-κB activation in GBM involves the TGF-β/TAK1 signaling axis. Together, our results demonstrate a role for the NF-κB pathway in GSCs and provide a mechanistic basis for its potential as a therapeutic target in glioblastoma.

High grade gliomas (HGG) are classified into four subgroups based on transcriptional signatures and phenotypic characteristics. In particular, the proneural-to-mesenchymal transition (PMT) is associated with increased malignancy, poor prognosis, and disease recurrence, but the underlying causes of PMT are still unclear. In this study, we investigated whether radiotherapy promotes PMT using a genetically engineered mouse model of proneural HGG. We found that cranial ionizing radiation induced robust and durable PMT in tumors. Additionally, we isolated primary proneural HGG cells from mouse and human tumors and demonstrate that radiation induced a sustained cell-intrinsic mesenchymal transition associated with increased invasiveness and resistance to the alkylating agent temozolomide. Expectedly, irradiation-induced PMT was also associated with activation of the STAT3 transcription factor, and the combination of STAT3 blockade using JAK2 inhibitors with radiation abrogated the mesenchymal transition and extended survival of mice. Taken together, our data suggest that clinical JAK2 inhibitors should be tested in conjunction with radiation in patients with proneural HGG as a new strategy for blocking the emergence of therapy-resistant mesenchymal tumors at relapse., (©2015 American Association for Cancer Research.)

Glioblastoma (GBM) is the most common malignant primary brain tumor. Radiotherapy fails to eliminate subpopulations of stem-like tumor-propagating cells (TPC), resulting in tumor regrowth. To identify kinases that promote TPC self-renewal rather than increasing proliferation in human GBM cultures, we screened a library of 54 nonselective tool compounds and determined their kinase inhibitor profiles in vitro. Most compounds inhibited aurora kinase (AURK) activity and blocked TPC self-renewal, while inducing GBM cell polynucleation and apoptosis. To prevent regrowth by TPCs, we used a priming dose of radiation followed by incubation with the pan-AURK inhibitor VX680 to block self-renewal and induce apoptosis in GBM cultures. In mice xenografted with human GBM cells, radiotherapy followed by VX680 treatment resulted in reduced tumor growth and increased survival relative to either monotherapy alone or VX680 treatment before radiation. Our results indicate that AURK inhibition, subsequent to radiation, may enhance the efficacy of radiotherapy by targeting radioresistant TPCs in human GBMs., (©2014 American Association for Cancer Research.)

Neurogenesis, the generation of new neurons, is deregulated in neural stem cell (NSC)- and progenitor-derived murine models of malignant medulloblastoma and glioma, the most common brain tumors of children and adults, respectively. Molecular characterization of human malignant brain tumors, and in particular brain tumor stem cells (BTSCs), has identified neurodevelopmental transcription factors, microRNAs, and epigenetic factors known to inhibit neuronal and glial differentiation. We are starting to understand how these factors are regulated by the major oncogenic drivers in malignant brain tumors. In this review, we will focus on the molecular switches that block normal neuronal differentiation and induce brain tumor formation. Genetic or pharmacological manipulation of these switches in BTSCs has been shown to restore the ability of tumor cells to differentiate. We will discuss potential brain tumor therapies that will promote differentiation in order to reduce treatment resistance, suppress tumor growth, and prevent recurrence in patients.

Brain tumors have frequently been associated with a neural stem cell (NSC) origin and contain stem-like tumor cells, so-called brain tumor stem cells (BTSCs) that share many features with normal NSCs. A stem cell state of BTSCs confers resistance to radiotherapy and treatment with alkylating agents. It is also a hallmark of aggressive brain tumors and is maintained by transcriptional networks that are also active in embryonic stem cells. Advances in reprogramming of somatic cells into induced pluripotent stem (iPS) cells have further identified genes that drive stemness. In this review, we will highlight the possible drivers of stemness in medulloblastoma and glioma, the most frequent types of primary malignant brain cancer in children and adults, respectively. Signals that drive expansion of developmentally defined neural precursor cells are also active in corresponding brain tumors. Transcriptomal subgroups of human medulloblastoma and glioma match features of NSCs but also more restricted progenitors. Lessons from genetically-engineered mouse (GEM) models show that temporally and regionally defined NSCs can give rise to distinct subgroups of medulloblastoma and glioma. We will further discuss how acquisition of stem cell features may drive brain tumorigenesis from a non-NSC origin. Genetic alterations, signaling pathways, and therapy-induced changes in the tumor microenvironment can drive reprogramming networks and induce stemness in brain tumors. Finally, we propose a model where dysregulation of microRNAs (miRNAs) that normally provide barriers against reprogramming plays an integral role in promoting stemness in brain tumors., (Copyright © 2013 Elsevier Inc. All rights reserved.)

The modified phonon dispersion is of importance for understanding the origin of the reduced heat conductivity in nanowires. We have measured the phonon dispersion for 50 nm diameter InSb (111) nanowires using time-resolved X-ray diffraction. By comparing the sound speed of the bulk (3880 m/s) and that of a classical thin rod (3600 m/s) to our measurement (2880 m/s), we conclude that the origin of the reduced sound speed and thereby to the reduced heat conductivity is that the C44 elastic constant is reduced by 35% compared to the bulk material.

Glioma is the most common primary malignant brain tumor and arises throughout the central nervous system. Recent focus on stem-like glioma cells has implicated neural stem cells (NSCs), a minor precursor population restricted to germinal zones, as a potential source of gliomas. In this review, we focus on the relationship between oligodendrocyte progenitor cells (OPCs), the largest population of cycling glial progenitors in the postnatal brain, and gliomagenesis. OPCs can give rise to gliomas, with signaling pathways associated with NSCs also playing key roles during OPC lineage development. Gliomas can also undergo a switch from progenitor- to stem-like phenotype after therapy, consistent with an OPC-origin even for stem-like gliomas. Future in-depth studies of OPC biology may shed light on the etiology of OPC-derived gliomas and reveal new therapeutic avenues., (© 2014 Elsevier Inc. All rights reserved.)

Based on clinical presentation, glioblastoma (GBM) is stratified into primary and secondary types. The protein 53 (p53) pathway is functionally incapacitated in most GBMs by distinctive type-specific mechanisms. To model human gliomagenesis, we used a GFAP-HRas(V12) mouse model crossed into the p53ER(TAM) background, such that either one or both copies of endogenous p53 is replaced by a conditional p53ER(TAM) allele. The p53ER(TAM) protein can be toggled reversibly in vivo between wild-type and inactive conformations by administration or withdrawal of 4-hydroxytamoxifen (4-OHT), respectively. Surprisingly, gliomas that develop in GFAP-HRas(V12);p53(+/KI) mice abrogate the p53 pathway by mutating p19(ARF)/MDM2 while retaining wild-type p53 allele. Consequently, such tumors are unaffected by restoration of their p53ER(TAM) allele. By contrast, gliomas arising in GFAP-HRas(V12);p53(KI/KI) mice develop in the absence of functional p53. Such tumors retain a functional p19(ARF)/MDM2-signaling pathway, and restoration of p53ER(TAM) allele triggers p53-tumor-suppressor activity. Congruently, growth inhibition upon normalization of mutant p53 by a small molecule, Prima-1, in human GBM cultures also requires p14(ARF)/MDM2 functionality. Notably, the antitumoral efficacy of p53 restoration in tumor-bearing GFAP-HRas(V12);p53(KI/KI) animals depends on the duration and frequency of p53 restoration. Thus, intermittent exposure to p53ER(TAM) activity mitigated the selective pressure to inactivate the p19(ARF)/MDM2/p53 pathway as a means of resistance, extending progression-free survival. Our results suggest that intermittent dosing regimes of drugs that restore wild-type tumor-suppressor function onto mutant, inactive p53 proteins will prove to be more efficacious than traditional chronic dosing by similarly reducing adaptive resistance.

The proto-oncogene MYCN is mis-expressed in various types of human brain tumors. To clarify how developmental and regional differences influence transformation, we transduced wild-type or mutationally stabilized murine N-myc(T58A) into neural stem cells (NSCs) from perinatal murine cerebellum, brain stem, and forebrain. Transplantation of N-myc(WT) NSCs was insufficient for tumor formation. N-myc(T58A) cerebellar and brain stem NSCs generated medulloblastoma/primitive neuroectodermal tumors, whereas forebrain NSCs developed diffuse glioma. Expression analyses distinguished tumors generated from these different regions, with tumors from embryonic versus postnatal cerebellar NSCs demonstrating Sonic Hedgehog (SHH) dependence and SHH independence, respectively. These differences were regulated in part by the transcription factor SOX9, activated in the SHH subclass of human medulloblastoma. Our results demonstrate context-dependent transformation of NSCs in response to a common oncogenic signal., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Postnatal oligodendrocyte progenitor cells (OPC) self-renew, generate mature oligodendrocytes, and are a cellular origin of oligodendrogliomas. We show that the proteoglycan NG2 segregates asymmetrically during mitosis to generate OPC cells of distinct fate. NG2 is required for asymmetric segregation of EGFR to the NG2(+) progeny, which consequently activates EGFR and undergoes EGF-dependent proliferation and self-renewal. In contrast, the NG2(-) progeny differentiates. In a mouse model, decreased NG2 asymmetry coincides with premalignant, abnormal self-renewal rather than differentiation and with tumor-initiating potential. Asymmetric division of human NG2(+) cells is prevalent in non-neoplastic tissue but is decreased in oligodendrogliomas. Regulators of asymmetric cell division are misexpressed in low-grade oligodendrogliomas. Our results identify loss of asymmetric division associated with the neoplastic transformation of OPC., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Malignant astrocytic brain tumors are among the most lethal cancers. Quiescent and therapy-resistant neural stem cell (NSC)-like cells in astrocytomas are likely to contribute to poor outcome. Malignant oligodendroglial brain tumors, in contrast, are therapy sensitive. Using magnetic resonance imaging (MRI) and detailed developmental analyses, we demonstrated that murine oligodendroglioma cells show characteristics of oligodendrocyte progenitor cells (OPCs) and are therapy sensitive, and that OPC rather than NSC markers enriched for tumor formation. MRI of human oligodendroglioma also suggested a white matter (WM) origin, with markers for OPCs rather than NSCs similarly enriching for tumor formation. Our results suggest that oligodendroglioma cells show hallmarks of OPCs, and that a progenitor rather than a NSC origin underlies improved prognosis in patients with this tumor., (Copyright © 2010 Elsevier Inc. All rights reserved.)

The ability to measure and understand heat flow in nanowire composites is crucial for applications ranging from high-speed electronics to thermoelectrics. Here we demonstrate the measurement of the thermal conductance of nanowire composites consisting of regular arrays of InAs nanowires embedded in PMMA using time-domain thermoreflectance (TDTR). On the basis of a proposed model for heat flow in the composite, we can, as a consistency check, extract the thermal conductivity Lambda of the InAs nanowires and find Lambda(NW) = 5.3 +/- 1.5 W m(-1) K(-1), in good agreement with theory and previous measurements of individual nanowires.

Nanowires are important candidates for use in future electronics, photonics and thermoelectrics applications. We focus here in particular on nanowires for use in thermoelectric power generation and present a method of fabricating dense uniform InAs nanowire arrays amenable to future incorporation of advanced heterostructures that could further increase the thermoelectric performance of these nanowires. In these applications it will be important to have the nanowires densely packed in order to give an appreciable amount of power output. Here we present the fabrication of such dense arrays, using metal-particle seeded growth and chemical beam epitaxy, where the metal particles are defined by electron beam lithography, metal evaporation and lift-off. We evaluate the potential of chemical beam epitaxy for the growth of dense, freestanding InAs nanowire arrays and describe the process that enabled us to achieve areal packing densities of up to 19% with a variation of only a few per cent in nanowire diameter and height. We close by discussing how even higher areal packing densities can be achieved.

In this issue of Cell Stem Stell, Bleau et al. report that activity of the drug transporter ABCG2 defines a tumorigenic stem-like side population (SP) isolated from glioma. Surprisingly, temozolomide, the first-line chemotherapeutic used for treatment of glioma, increased this side population, and even more so when PTEN was deleted.

When a quantum dot is subjected to a thermal gradient, the temperature of electrons entering the dot can be determined from the dot's thermocurrent if the conductance spectrum and background temperature are known. We demonstrate this technique by measuring the temperature difference across a 15 nm quantum dot embedded in a nanowire. This technique can be used when the dot's energy states are separated by many kT and will enable future quantitative investigations of electron-phonon interaction, nonlinear thermoelectric effects, and the efficiency of thermoelectric energy conversion in quantum dots.

Background: Glioblastoma multiforme (GBM) is an invariably fatal central nervous system tumor despite treatment with surgery, radiation, and chemotherapy. Further insights into the molecular and cellular mechanisms that drive GBM formation are required to improve patient outcome. MicroRNAs are emerging as important regulators of cellular differentiation and proliferation, and have been implicated in the etiology of a variety of cancers, yet the role of microRNAs in GBM remains poorly understood. In this study, we investigated the role of microRNAs in regulating the differentiation and proliferation of neural stem cells and glioblastoma-multiforme tumor cells., Methods: We used quantitative RT-PCR to assess microRNA expression in high-grade astrocytomas and adult mouse neural stem cells. To assess the function of candidate microRNAs in high-grade astrocytomas, we transfected miR mimics to cultured-mouse neural stem cells, -mouse oligodendroglioma-derived stem cells, -human glioblastoma multiforme-derived stem cells and -glioblastoma multiforme cell lines. Cellular differentiation was assessed by immunostaining, and cellular proliferation was determined using fluorescence-activated cell sorting., Results: Our studies revealed that expression levels of microRNA-124 and microRNA-137 were significantly decreased in anaplastic astrocytomas (World Health Organization grade III) and glioblastoma multiforme (World Health Organization grade IV) relative to non-neoplastic brain tissue (P < 0.01), and were increased 8- to 20-fold during differentiation of cultured mouse neural stem cells following growth factor withdrawal. Expression of microRNA-137 was increased 3- to 12-fold in glioblastoma multiforme cell lines U87 and U251 following inhibition of DNA methylation with 5-aza-2'-deoxycytidine (5-aza-dC). Transfection of microRNA-124 or microRNA-137 induced morphological changes and marker expressions consistent with neuronal differentiation in mouse neural stem cells, mouse oligodendroglioma-derived stem cells derived from S100 beta-v-erbB tumors and cluster of differentiation 133+ human glioblastoma multiforme-derived stem cells (SF6969). Transfection of microRNA-124 or microRNA-137 also induced G1 cell cycle arrest in U251 and SF6969 glioblastoma multiforme cells, which was associated with decreased expression of cyclin-dependent kinase 6 and phosphorylated retinoblastoma (pSer 807/811) proteins., Conclusion: microRNA-124 and microRNA-137 induce differentiation of adult mouse neural stem cells, mouse oligodendroglioma-derived stem cells and human glioblastoma multiforme-derived stem cells and induce glioblastoma multiforme cell cycle arrest. These results suggest that targeted delivery of microRNA-124 and/or microRNA-137 to glioblastoma multiforme tumor cells may be therapeutically efficacious for the treatment of this disease.

An n-type InAs/InAsP heterostructure nanowire field-effect transistor has been fabricated and compared with a homogeneous InAs field-effect transistor. For the same device geometry, by introduction of the heterostructure, the threshold voltage is shifted 4 V, the maximum current on-off ratio is enhanced by a factor of 10,000, and the subthreshold swing is lowered by a factor 4 compared to the homogeneous transistor. At the same time, the drive current remains constant for a fixed gate overdrive. A single nanowire heterostructure transistor has a transconductance of 5 muA/V at a low source-drain voltage of 0.3 V. For the homogeneous InAs transistor, we deduced a high electron mobility of 1500 cm2/Vs.

Growth factors and peptides playing important roles during early development of the central nervous system have also been shown to maintain their regulation of cell genesis in the adult brain. We have previously described that endogenous opioids, expressed in the developing hippocampus, regulate proliferation and differentiation in the adult rat hippocampus. The aim of this study was to investigate the effects of the opioid beta-endorphin on gene expression and glial differentiation in cultures of adult rat hippocampal progenitors (AHPs). Changes in gene expression after stimulation of AHPs with beta-endorphin for 48 h were investigated using cDNA arrays. Confirmation experiments verified that stimulation with beta-endorphin increased the mRNA levels of myelin basic protein, glutathione S-transferase pi, c-junD and rab16 (P < 0.05), genes that are associated with oligodendrogenesis. Furthermore, beta-endorphin increased the levels of Id1, but not Id3, mRNA on the arrays. Incubation of AHPs with beta-endorphin resulted in a threefold increase in oligodendrogenesis (P < 0.01) but no significant change in astrogliogenesis. No effect on oligodendrogenesis was observed in the presence of the opioid antagonist naloxone. Coincubation of beta-endorphin with Id1 antisense oligonucleotides for 10 days also entirely blocked the induced oligodendrogenesis in our AHP cultures. Moreover, a subpopulation of AHPs (25%) showed nuclear expression of the proneural transcriptional activator Mash1 that was reduced to approximately 5% of the cells when exposed to beta-endorphin. We suggest a requirement for Id1 in opioid-induced oligodendrogenesis in cultured AHPs possibly acting on opioid-responsive AHPs expressing the proneural transcriptional activator Mash1.

It has been postulated that opiates induce addictive behaviour via changes in gene expression. PC12 cells were stably transfected with the recombinant human mu-opioid receptor (MOR) to study opioid-induced gene expression. Expression was verified by binding assay, immunocytochemistry, and immunblotting experiments. Forskolin-induced cAMP formation was inhibited by [D-Ala(2), N-Me-Phe(4), Gly(5)-ol]-enkephalin (DAMGO 1 microM), a specific MOR agonist. This effect was completely antagonized by naloxone. By using cDNA arrays, including approximately 1,200 well-defined genes normally expressed in neural tissue, we monitored semi-quantitative changes in gene expression after 3 h short-term exposure to DAMGO. Incubation with DAMGO increased mRNA levels for 13 genes and down-regulated 13 other genes. Annexin V, RGS4 and CREB genes showed pronounced increase in expression after stimulation with DAMGO. Quantitative RT-PCR confirmed that DAMGO increased mRNA levels of Annexin V, an apoptosis-induced gene. We suggest that the PC12 cell transfected with the recombinant human MOR is a useful tool for identification of opioid-induced genes that may provide information on opiate effects of relevance for dependence.

We present the growth of homogeneous InAs(1-x)P(x) nanowires as well as InAs(1-x)P(x) heterostructure segments in InAs nanowires with P concentrations varying from 22% to 100%. The incorporation of P has been studied as a function of TBP/TBAs ratio, temperature, and diameter of the wires. The crystal structure of the InAs as well as the InAs(1-x)P(x) segments were found to be wurtzite as determined from high-resolution transmission electron microscopy. Furthermore, temperature-dependent electrical transport measurements were performed on individual heterostructured wires to extract the conduction band offset of InAs(1-x)P(x) relative to InAs as a function of composition. From these measurements we extract a value of the linear coefficient of the conduction band versus x of 0.6 eV and a nonlinear coefficient, or bowing parameter, of 0.2 eV. Finally, homogeneous InAs(0.8)P(0.2) nanowires were shown to have a nondegenerate n-type doping and function as field-effect transistors at room temperature.

We report on spectrally resolved photocurrent measurements on single self-assembled nanowire heterostructures. The wires, typically 3 microm long with an average diameter of 85 nm, consist of InAs with a 1 microm central part of InAsP. Two different sets of wires were prepared with phosphorus contents of 15+/-3% and 35+/-3%, respectively, as determined by energy-dispersive spectroscopy measurements made in transmission electron microscopy. Ohmic contacts are fabricated to the InAs ends of the wire using e-beam lithography. The conduction band offset between the InAs and InAsP regions virtually removes the dark current through the wires at low temperature. In the optical experiments, interband excitation in the phosphorus-rich part of the wires results in a photocurrent with threshold energies of about 0.65 and 0.82 eV, respectively, in qualitative agreement with the expected band gap of the two compositions. Furthermore, a strong polarization dependence is observed with an order of magnitude larger photocurrent for light polarized parallel to the wire than for light polarized perpendicular to the wire. We believe that these wires form promising candidates as nanoscale infrared polarization-sensitive photodetectors.

It has previously been suggested that exogenous growth hormone (GH) affect quality of life and higher brain functions through the endogenous opioid system. Recently, we showed that GH down-regulate 72 and 48 kDa delta opioid receptor (DOR) proteins in the adult rat cerebral cortex and cerebellum. In the present study, we found that an antiserum raised against the N-terminus of the DOR also recognizes a 36 kDa protein, not recognized by a C-terminus-directed antiserum. We aimed to investigate the identity of the 72, 48 and 36 kDa proteins and to further study the effects of GH on their expression in different brain regions. The expression was studied in hypophysectomized (Hx) and untreated normal female rats. One subgroup of Hx rats received GH as a daily subcutaneous injection for 19 days. Our data show that treatment with GH in Hx rats normalized the expression of the 72 kDa protein in the cerebral cortex, whereas no significant effect were observed for the 48 or 36 kDa proteins. However, GH significantly reduced the ratio between the 72 and 36 kDa proteins in different brain regions of Hx rats. Our data suggest that GH reduces the levels of a 72 kDa DOR that likely represents a dimeric form of a 36 kDa DOR post-translationally truncated at the C-terminus, and that altered receptor dimerization may be involved in GH induced effects in the central nervous system.

Previous work has shown that voluntary running increases cell proliferation and neurogenesis in the dentate gyrus of the adult hippocampus. Here we report that long-term running for 24 days results in a down-regulation of hippocampal progenitor proliferation to one-half the level of nonrunning controls compared with a fivefold increase in progenitor proliferation seen after 9 days of voluntary running (short-term running). The negative effects seen on proliferation after 24 days of running were prevented by restricting daily running distances (by 30-50%) during 24 days. Long-term running for 24 days increases the response of the hypothalamic-pituitary-adrenal axis, with an increase in adrenal gland weight and increased plasma corticosterone levels, as well as decreased thymus weight, indicating a stress response as a possible mediator of decreased progenitor proliferation. Furthermore, the negative effects seen on the observed stress response after 24 days of running were prevented by restricting daily running distance. Short-term running did not alter these stress parameters compared with nonrunning controls. However, it increased phosphorylated cyclic AMP response element binding protein (pCREB) in the dentate gyrus, an increase that was not seen in nonrunning controls or after 24 days of running. Taken together, these data suggest that voluntary running does not always enhance proliferation and that the decrease in progenitor proliferation seen in long-term running is possibly mediated by mechanisms involving a stress response in the animal. However, a moderate level of long-term running was able to prevent the negative stress-related changes seen in unrestricted long-term running.

Controllable production of nanometre-sized structures is an important field of research, and synthesis of one-dimensional objects, such as nanowires, is a rapidly expanding area with numerous applications, for example, in electronics, photonics, biology and medicine. Nanoscale electronic devices created inside nanowires, such as p-n junctions, were reported ten years ago. More recently, hetero-structure devices with clear quantum-mechanical behaviour have been reported, for example the double-barrier resonant tunnelling diode and the single-electron transistor. The generally accepted theory of semiconductor nanowire growth is the vapour-liquid-solid (VLS) growth mechanism, based on growth from a liquid metal seed particle. In this letter we suggest the existence of a growth regime quite different from VLS. We show that this new growth regime is based on a solid-phase diffusion mechanism of a single component through a gold seed particle, as shown by in situ heating experiments of GaAs nanowires in a transmission electron microscope, and supported by highly resolved chemical analysis and finite element calculations of the mass transport and composition profiles.

We have investigated the presence of different glycoforms of cystatin C secreted by adult hippocampal rat-derived stem/progenitor cells (AHPs) into conditioned medium. A glycosylated form of cystatin C (CCg) has been identified previously in conditioned medium from AHPs as an autocrine/paracrine cofactor. Fibroblast growth factor-2 (FGF2) requires cooperation with CCg to support AHP survival at low density in vitro. The purpose of the present study was to investigate further if cystatin C consists of one glycoform or if several different glycoforms are secreted by AHPs in vitro. The presence of the glycoforms was studied using enzymatic deglycosylation in conjunction with gel electrophoresis and Western blotting. The glycoforms of cystatin C were isolated with a combination of gel electrophoresis and electroelution, yielding the intact glycoforms in liquid phase before enzymatic deglycosylation. Our results revealed several novel glycoforms, in contrast to previous publication. The results suggest that N- and O-linked glycans with sialic acid are attached to cystatin C. Furthermore, we have demonstrated that all glycoforms are present in conditioned medium after only 48 hr of culturing and that all nestin-positive AHPs are immunopositive against cystatin C. These findings suggest secretion of the glycoforms by cultured AHPs., (Copyright 2004 Wiley-Liss, Inc.)

Voluntary running in mice and forced treadmill running in rats have been shown to increase the amount of proliferating cells in the hippocampus. Little is known as yet about the mechanisms involved in these processes. It is well known that the endogenous opioid system is affected during running and other forms of physical exercise. In this study, we evaluated the involvement of the endogenous opioids in the regulation of hippocampal proliferation in non-running and voluntary running rats. Nine days of wheel running was compared with non-running in spontaneously hypertensive rats (SHR), a rat strain known to run voluntarily. On the last 2 days of the experimental period all rats received two daily injections of the opioid receptor antagonists naltrexone or naltrindole together with injections of bromodeoxyuridine to label dividing cells. Brain sections from the running rats showed approximately a five-fold increase in newly generated cells in the hippocampus, and this increase was partly reduced by naltrexone but not by naltrindole. By contrast, both naltrexone and naltrindole increased hippocampal proliferation in non-running rats. In non-running rats the administration of naltrexone decreased corticosterone levels and adrenal gland weights, whereas no significant effects on these parameters could be detected for naltrindole. However, adrenal gland weights were increased in naltrexone- but not in naltrindole-administered running rats. In addition, in voluntary running rats there was a three-fold increase in the hippocampal levels of Met-enkephalin-Arg-Phe compared with non-runners, indicating an increase in opioid activity in the hippocampus during running. These data suggest an involvement of endogenous opioids in the regulation of hippocampal proliferation in non-running rats, probably through hypothalamic-pituitary-adrenal axis modulation. During voluntary running in SHR naltrexone altered hippocampal proliferation via as yet unknown mechanisms.

Along with rapidly developing nanotechnology, new types of analytical instruments and techniques are needed. Here we report an alternative procedure for electrical measurements on semiconductor nanowhiskers, allowing precise selection and visual control at close to atomic resolution. We use a combination of two powerful microscope techniques, scanning tunneling microscopy (STM) and simultaneous viewing in a transmission electron microscope (TEM). The STM is mounted in the sample holder for the TEM. We describe here a method for creating an ohmic contact between the STM tip and the nanowhisker. We examine three different types of STM tips and present a technique for cleaning the STM tip in situ. Measurements on 1-microm-tall and 40-nm-thick epitaxially grown InAs nanowhiskers show an ohmic contact and a resistance of down to 7 kOmega.

Administration of opioid agonists or antagonists has been reported to regulate proliferation or survival of neural progenitors in vivo. Here we report that beta-endorphin and selective mu-opioid receptor (MOR) and delta-opioid receptor (DOR) agonists stimulate proliferation of isolated rat adult hippocampal progenitors (AHPs). The AHPs were found to express DORs and MORs, but not kappa-opioid receptors. Incubation with beta-endorphin for 48 h increased the number of AHPs found in mitosis, the total DNA content, and the expression of proliferating cell nuclear antigen. This proliferative effect from beta-endorphin on AHPs was antagonized by naloxone. The beta-endorphin-induced proliferation was mediated through phosphorylation of extracellular signal-regulated kinases 1 and 2 and dependent on phosphatidylinositol 3-kinase and both intra- and extracellular calcium. These data suggest a role for the opioid system in the regulation of proliferation in progenitors from the adult hippocampus.

Opioids have previously been shown to affect proliferation and differentiation in various neural cell types. In the present study, cultured rat adult hippocampal progenitors (AHPs) were shown to release beta-endorphin. Membrane preparations of AHPs were found to bind [125I]beta-endorphin, and immunoreactivity for mu- and delta-opioid receptors (MORs and DORs), but not for kappa-opioid receptors (KORs), was found on cells in culture. Both DNA content and [3H]thymidine incorporation were reduced after a 48-h incubation with 100 microM naloxone, 10 micro m naltrindole or 10 microM beta-funaltrexamine, but not nor-binaltorphimine, suggesting proliferative actions of endogenous opioids against MORs and DORs on AHPs. Furthermore, analysis of gene and protein expression after incubation with MOR and DOR antagonists for 48 h using RT-PCR and Western blotting suggested decreased signalling through the mitogen-activated protein kinase (MAPK) pathway and lowered levels of genes and proteins that are important in cell cycling. Cultures were incubated with naloxone (10 or 100 microM) for 10 days to study the effects on differentiation. This resulted in an approximately threefold increase in neurogenesis, a threefold decrease in astrogliogenesis and a 50% decrease in oligodendrogenesis. In conclusion, this study suggests that reduced signalling through MORs and DORs decreases proliferation in rat AHPs, increases the number of in vitro-generated neurons and reduces the number of astrocytes and oligodendrocytes in culture.

Hormones released from the pituitary have been shown to regulate the expression of different proteins in the central nervous system. We wanted to examine whether peripheral administration of bovine growth hormone (bGH) regulates the expression of delta-opioid receptor (DOR) in the cerebral cortex and cerebellum. Expression of the DOR protein was quantified using Western blot densitometry. DOR mRNA was quantified with a solution hybridization RNase protection assay. Hypophysectomized (Hx) and untreated normal female rats were included in the study. All Hx rats were hormonally treated with cortisol (400 microg/kg/day) and L-thyroxine (10 microg/kg/day) for 19 days. Hypophysectomy resulted in a threefold increase in cerebral cortex and a twofold increase in cerebellum of the DOR protein compared with normal rats. One subgroup of Hx rats received bGH (1 mg/kg body weight) as a daily subcutaneous injection for 19 days. This treatment normalized the levels of DOR protein in the cerebral cortex and cerebellum. Immunohistochemical experiments showed that GH decreased DOR expression especially in layers II-VI in cerebral cortex and in stratum moleculare in cerebellum. Quantification of DOR mRNA by solution hybridization RNase protection assay corresponded to the DOR protein measurements. We conclude that the expression of DORs in cerebral cortex and cerebellum is regulated by GH., (Copyright 2002 Wiley-Liss, Inc.)

It is known that proliferation and survival of neural stem/progenitor cells in vitro not only depend on exogenous factors, but also on autocrine factors secreted into the conditioned medium. It is also well known that the identification of bioactive proteins secreted into the conditioned medium poses a substantial challenge. Recently, neural stem/progenitor cells were shown to secrete a survival factor, cystatin C, into the conditioned medium. Here, we demonstrate an approach to identify other low molecular weight proteins in conditioned medium from cultured adult rat hippocampal progenitor cells. A combination of preparative two-dimensional gel electrophoresis (2-DE) and mass spectrometry was utilized in the analysis. We were able to identify a number of proteins, which include Rho-guanine nucleotide dissociation inhibitor 1, phosphatidylethanolamine binding protein (PEBP), also termed Raf-1 kinase interacting protein, polyubiquitin, immunophilin FK506 binding protein 12 (FKBP12) and cystatin C. The presence of PEBP and FKBP12 in conditioned medium was confirmed immunologically. All nestin-positive progenitor cells showed immunoreactivity for antibodies against PEBP and FKBP12. To our knowledge we are the first to use this preparative proteomic approach to search for stem cell factors in conditioned medium. The method could be used to identify novel bioactive proteins secreted by stem/progenitor cells in vitro. Identification of bioactive proteins in vitro is of potential importance for the understanding of the regulatory mechanisms of the cells in vivo., (Copyright 2003 John Wiley & Sons, Ltd.)