Search

Within the glioblastoma cellular niche, glioma stem cells (GSCs) can give rise to differentiated glioma cells (DGCs) and, when necessary, DGCs can reciprocally give rise to GSCs to maintain the cellular equilibrium necessary for optimal tumor growth. Here, using ribosome profiling, transcriptome and m6A RNA sequencing, we show that GSCs from patients with different subtypes of glioblastoma share a set of transcripts, which exhibit a pattern of m6A loss and increased protein translation during differentiation. The target sequences of a group of miRNAs overlap the canonical RRACH m6A motifs of these transcripts, many of which confer a survival advantage in glioblastoma. Ectopic expression of the RRACH-binding miR-145 induces loss of m6A, formation of FTO/AGO1/ILF3/miR-145 complexes on a clinically relevant tumor suppressor gene (CLIP3) and significant increase in its nascent translation. Inhibition of miR-145 maintains RRACH m6A levels of CLIP3 and inhibits its nascent translation. This study highlights a critical role of miRNAs in assembling complexes for m6A demethylation and induction of protein translation during GSC state transition. Author summary: Cellular plasticity and epigenetic adaptation of human glioblastoma stem cells to the tumor microenvironment is a hallmark of this devastating disease. With our present work, we discover the relationship between miRNAs and the RNA methylation machinery in human glioblastoma and show how miRNA-induced loss of m6A results in increase in protein translation of clinically important transcripts during glioblastoma stem cell differentiation. Leveraging the dynamic functions of these miRNAs can be important in the design of optimal therapeutics targeted at cancer cell plasticity. [ABSTRACT FROM AUTHOR]

Gliomas are highly invasive forms of brain cancer comprising more than 50% of brain tumor cases in adults, and astrocytomas account for ∼60-70% of all gliomas. As a result of multiple factors, including enhanced migratory properties and extracellular matrix remodeling, even with current standards of care, mean survival time for patients is only ∼12 months. Because glioblastoma multiforme (GBM) cells arise from astrocytes, there is great interest in elucidating the interactions of these two cell types in vivo. Previous work performed on two-dimensional assays (i.e., tissue culture plastic and Boyden chamber assays) utilizes substrates that lack the complexities of the natural microenvironment. Here, we employed a three-dimensional, electrospun poly-(caprolactone) (PCL) nanofiber system (NFS) to mimic some features of topographical properties evidenced in vivo. Co-cultures of human GBM cells and rat astrocytes, as performed on the NFS, showed a significant increase in astrocyte GFAP expression, particularly in the presence of extracellular matrix (ECM) deposited by GBM cells. In addition, GBM migration increased in the presence of astrocytes or soluble factors (i.e., conditioned media). However, the presence of fixed astrocytes acted as an antagonist, lowering GBM migration rates. This data suggests that astrocytes and GBM cells interact through a multitude of pathways, including soluble factors and direct contact. This work demonstrates the potential of the NFS to duplicate some topographical features of the GBM tumor microenvironment, permitting analysis of topographical effects in GBM migration. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:1406-1415, 2015 [ABSTRACT FROM AUTHOR]

Cells sense and respond to the rigidity of their microenvironment by altering their morphology and migration behavior. To examine this response, hydrogels with a range of moduli or mechanical gradients have been developed. Here, we show that edge effects inherent in hydrogels supported on rigid substrates also influence cell behavior. A Matrigel hydrogel was supported on a rigid glass substrate, an interface which computational techniques revealed to yield relative stiffening close to the rigid substrate support. To explore the influence of these gradients in 3D, hydrogels of varying Matrigel content were synthesized and the morphology, spreading, actin organization, and migration of glioblastoma multiforme (GBM) tumor cells were examined at the lowest (<50 mm) and highest (>500 mm) gel positions. GBMs adopted bipolar morphologies, displayed actin stress fiber formation, and evidenced fast, mesenchymal migration close to the substrate, whereas away from the interface, they adopted more rounded or ellipsoid morphologies, displayed poor actin architecture, and evidenced slow migration with some amoeboid characteristics. Mechanical gradients produced via edge effects could be observed with other hydrogels and substrates and permit observation of responses to multiple mechanical environments in a single hydrogel. Thus, hydrogel-support edge effects could be used to explore mechanosensitivity in a single 3D hydrogel system and should be considered in 3D hydrogel cell culture systems. [ABSTRACT FROM AUTHOR]

Abstract: Glioblastoma multiforme (GBM), one of the deadliest forms of human cancer, is characterized by its high infiltration capacity, partially regulated by the neural extracellular matrix (ECM). A major limitation in developing effective treatments is the lack of in vitro models that mimic features of GBM migration highways. Ideally, these models would permit tunable control of mechanics and chemistry to allow the unique role of each of these components to be examined. To address this need, we developed aligned nanofiber biomaterials via core–shell electrospinning that permit systematic study of mechanical and chemical influences on cell adhesion and migration. These models mimic the topography of white matter tracts, a major GBM migration ‘highway’. To independently investigate the influence of chemistry and mechanics on GBM behaviors, nanofiber mechanics were modulated by using different polymers (i.e., gelatin, poly(ethersulfone), poly(dimethylsiloxane)) in the ‘core’ while employing a common poly(ε-caprolactone) (PCL) ‘shell’ to conserve surface chemistry. These materials revealed GBM sensitivity to nanofiber mechanics, with single cell morphology (Feret diameter), migration speed, focal adhesion kinase (FAK) and myosin light chain 2 (MLC2) expression all showing a strong dependence on nanofiber modulus. Similarly, modulating nanofiber chemistry using extracellular matrix molecules (i.e., hyaluronic acid (HA), collagen, and Matrigel) in the ‘shell’ material with a common PCL ‘core’ to conserve mechanical properties revealed GBM sensitivity to HA; specifically, a negative effect on migration. This system, which mimics the topographical features of white matter tracts, should allow further examination of the complex interplay of mechanics, chemistry, and topography in regulating brain tumor behaviors. [Copyright &y& Elsevier]