Modulation in the expression levels and protein localization of cell volume and actin cytoskeleton associated proteins by the Slit/ Robo signaling pathway in Glioblastoma multiforme.

Honorable Mention Winner Glioblastoma multiforme (GBM) is the most aggressive and malignant primary brain tumor in adults1. Even after surgical resection, chemotherapy, and radiation, the average patient survival rate is 14 months2. Recent studies indicate that GBM tumors that contact the lateral ventricles within the subventricular zone (SVZ) tend to migrate and recur at distal locations after surgical resection3. One important signaling pathway in Neural Progenitor Cells (NPCs) migration from the SVZ to the olfactory bulbs is the Slit/Robo signaling pathway4. Slit2 proteins are produced by the choroid plexus and create a gradient across ependymal cells. Slit2 proteins then interact with the Roundabout (Robo) receptor on NPCs and acts as a chemo-repelling agent5. This Slit2/ Robo1 signaling pathway is a possible mechanism that contributes to GBM migration from the SVZ to distal locations. The intracellular changes when the Slit/Robo pathway is activated includes changes in cell volume regulation and actin cytoskeleton dynamics6. We then examined how the Slit2/ Robo1 signaling pathway causes changes in the expression of cell volume regulatory genes (AQP4, KCC1, NKCC1, NCC), actin cytoskeleton genes regulatory genes (p-ERM, p-Cofilin), and the localization of actin-associated proteins (FAK, Paxillin, N-WASP, Vinculin). It was found that the Slit/Robo signaling pathway increases the expression levels of genes that contribute to cell volume regulation in GBM cells, and the Slit/Robo signaling pathway increases the phosphorylation of p-Cofilin and p-ERM. This indicates that the Slit2/ Robo1 pathway participates in the regulation of cell volume and actin cytoskeleton dynamics in GBM cells

Determining glioblastoma proteome changes in response to lateral ventricle neural stem cells

Glioblastoma (GBM) is the most common and malignant primary tumor in adults. When GBM tumors are located close to the lateral ventricle they display a more aggressive recurrence pattern and negatively impact patient survival. These findings suggest the involvement of the subventricular zone neurogenic niche in GBM malignancy. To define the inter-cellular communication between neural stem cells and GBM cells, we optimized a tool to determine cell-specific proteomic changes of GBM cells in response to neural stem cell proximity. We cloned the mutated methionyl-tRNA synthetase (MetRS) gene into the lentiviral plasmid MetRS puro. MetRS allows for incorporation of azide-tagged methionine analog azidonorleucine (ANL) into newly formed proteins, effectively labeling proteins synthesized by expressing cells. We utilized the pLKO.1 vector backbone allowing puromycin resistance as a selection method. The MetRS L274 modification was confirmed, as only MetRS-transduced cells of both commercial HEK and primary GBM1A cell lines selectively incorporated ANL. Following verification, we successfully packaged the plasmid into a lentivirus. We transduced primary human fetal neural stem cell (hfNSC) and GBM lines and selected the MetRS-expressing cells by puromycin exposure. After 96 hours, wild type (WT) cells died while successfully transduced cells exhibited resistance and the ANL-compatible MetRS enzyme. Co-cultures consisting of MetRS-transduced GBM and WT hfNSCs were used to simulate a similar environment of glioblastoma neighboring lateral ventricles. Proteome Profiler results showed a significant downregulation of an angiogenesis inhibitor and upregulation of malignancy promoting proteins in GBM1A. Going forward, this analysis method will be used for cell-specific proteomics in vivo

The generation of oligodendroglial cells is preserved in the rostral migratory stream during aging.

The subventricular zone (SVZ) is the largest source of newly generated cells in the adult mammalian brain. SVZ-derived neuroblasts migrate via the rostral migratory stream (RMS) to the olfactory bulb (OB), where they differentiate into mature neurons. Additionally, a small proportion of SVZ-derived cells contribute to the generation of myelinating oligodendrocytes. The production of new cells in the SVZ decreases during aging, affecting the incorporation of new neurons into the OB. However, the age-related changes that occur across the RMS are not fully understood. In this study we evaluate how aging affects the cellular organization of migrating neuroblast chains, the proliferation, and the fate of the newly generated cells in the SVZ-OB system. By using electron microscopy and immunostaining, we found that the RMS path becomes discontinuous and its cytoarchitecture is disorganized in aged mice (24-month-old mice). Subsequently, OB neurogenesis was impaired in the aged brain while the production of oligodendrocytes was not compromised. These findings provide new insight into oligodendrocyte preservation throughout life. Further exploration of this matter could help the development of new strategies to prevent neurological disorders associated with senescence

Use of Mesenchymal Stem Cells in Pre-Clinical Models of Spinal Cord Injury

Spinal Cord Injury (SCI) is a devastating disease that causes disruption of sensorimotor function below the site of injury. Current management is based on surgical decompression of the neural tissue and pharmacotherapy; however, there is no gold standard treatment readily available for patients in the clinic. This indicates that novel therapeutic strategies for the treatment are still needed in the clinical setting. There are several alternatives that are currently under investigation for the treatment of this disease, with increasing focus in regenerative medicine treatments. Mesenchymal stem cells (MSCs) are one of the most promising candidates for stem cell therapy in SCI, as they are easily obtained, have high safety profiles, and help with neural regeneration in SCI mainly via release of trophic factors, neovascularization, and immunomodulation. In this work, authors provide an insight of the available MSC for neural regeneration, their therapeutic role, and the potential MSC-based therapies for SCI

Shear Forces during Blast, Not Abrupt Changes in Pressure Alone, Generate Calcium Activity in Human Brain Cells

Blast-Induced Traumatic Brain Injury (bTBI) describes a spectrum of injuries caused by an explosive force that results in changes in brain function. The mechanism responsible for primary bTBI following a blast shockwave remains unknown. We have developed a pneumatic device that delivers shockwaves, similar to those known to induce bTBI, within a chamber optimal for fluorescence microscopy. Abrupt changes in pressure can be created with and without the presence of shear forces at the surface of cells. In primary cultures of human central nervous system cells, the cellular calcium response to shockwaves alone was negligible. Even when the applied pressure reached 15 atm, there was no damage or excitation, unless concomitant shear forces, peaking between 0.3 to 0.7 Pa, were present at the cell surface. The probability of cellular injury in response to a shockwave was low and cell survival was unaffected 20 hours after shockwave exposure

Brief report: Robo1 regulates the migration of human subventricular zone neural progenitor cells during development

Guerrero-Cazares, Hugo et al.Human neural progenitor cell (NPC) migration within the subventricular zone (SVZ) of the lateral ganglionic eminence is an active process throughout early brain development. The migration of human NPCs from the SVZ to the olfactory bulb during fetal stages resembles what occurs in adult rodents. As the human brain develops during infancy, this migratory stream is drastically reduced in cell number and becomes barely evident in adults. The mechanisms regulating human NPC migration are unknown. The Slit–Robo signaling pathway has been defined as a chemorepulsive cue involved in axon guidance and neuroblast migration in rodents. Slit and Robo proteins expressed in the rodent brain help guide neuroblast migration from the SVZ through the rostral migratory stream to the olfactory bulb. Here, we present the first study on the role that Slit and Robo proteins play in human-derived fetal neural progenitor cell migration (hfNPC). We describe that Robo1 and Robo2 isoforms are expressed in the human fetal SVZ. Furthermore, we demonstrate that Slit2 is able to induce a chemorepellent effect on the migration of hfNPCs derived from the human fetal SVZ. In addition, when Robo1 expression is inhibited, hfNPCs are unable to migrate to the olfactory bulb of mice when injected in the anterior SVZ. Our findings indicate that the migration of human NPCs from the SVZ is partially regulated by the Slit–Robo axis. This pathway could be regulated to direct the migration of NPCs in human endogenous neural cell therapy.H.G.C., A.Q.H., V.C.G., and P.S. were supported by the National Institutes of Health (R01NS070024); H.G.C. and E.L. were supported by the National Cancer Institute (R21CA199295) and the Maryland Stem Cells Research Fund; L.C. was supported by the Howard Hughes Medical Institute; V.C.G. was supported by the Fundacion Progreso y Salud of the Andalusian Regional Ministry of Health (PI01092014).Peer Reviewe

Editorial: Neural Stem Cells of the Subventricular Zone: From Neurogenesis to Glioblastoma Origin

EM is funded by VI Plan Propio de Investigación (Universidad de Sevilla) Grant number 2020/0000081. NZ and HG-C are funded by the NIH-NINDS K01NS110930-03 and the Neuro oncology convergence. CC is funded by the Integrated Territorial Investment Operational Programme of the European Commission and by the Department of Department of Health and Families (Consejerıa de Salud y Familias) of the Regional Government of Andalusia. Project reference: ITI-0042-2019: ITI Cadiz 2019Ye

Sex-Specific Differences in Glioblastoma

Sex differences have been well identified in many brain tumors. Even though glioblastoma (GBM) is the most common primary malignant brain tumor in adults and has the worst outcome, well-established differences between men and women are limited to incidence and outcome. Little is known about sex differences in GBM at the disease phenotype and genetical/molecular level. This review focuses on a deep understanding of the pathophysiology of GBM, including hormones, metabolic pathways, the immune system, and molecular changes, along with differences between men and women and how these dimorphisms affect disease outcome. The information analyzed in this review shows a greater incidence and worse outcome in male patients with GBM compared with female patients. We highlight the protective role of estrogen and the upregulation of androgen receptors and testosterone having detrimental effects on GBM. Moreover, hormones and the immune system work in synergy to directly affect the GBM microenvironment. Genetic and molecular differences have also recently been identified. Specific genes and molecular pathways, either upregulated or downregulated depending on sex, could potentially directly dictate GBM outcome differences. It appears that sexual dimorphism in GBM affects patient outcome and requires an individualized approach to management considering the sex of the patient, especially in relation to differences at the molecular level

Influence of basement membrane proteins and endothelial cell-derived factors on the morphology of human fetal-derived astrocytes in 2D.

Astrocytes are the most prevalent type of glial cell in the brain, participating in a variety of diverse functions from regulating cerebral blood flow to controlling synapse formation. Astrocytes and astrocyte-conditioned media are widely used in models of the blood-brain barrier (BBB), however, very little is known about astrocyte culture in 2D. To test the hypothesis that surface coating and soluble factors influence astrocyte morphology in 2D, we quantitatively analyzed the morphology of human fetal derived astrocytes on glass, matrigel, fibronectin, collagen IV, and collagen I, and after the addition soluble factors including platelet-derived growth factor (PDGF), laminin, basic fibroblast growth factor (bFGF), and leukemia inhibitory factor (LIF). Matrigel surface coatings, as well as addition of leukemia inhibitory factor (LIF) to the media, were found to have the strongest effects on 2D astrocyte morphology, and may be important in improving existing BBB models. In addition, the novel set of quantitative parameters proposed in this paper provide a test for determining the influence of compounds on astrocyte morphology, both to screen for new endothelial cell-secreted factors that influence astrocytes, and to determine in a high-throughput way which factors are important for translation to more complex, 3D BBB models