Neural stem cells traffic functional mitochondria via extracellular vesicles.

Neural stem cell (NSC) transplantation induces recovery in animal models of central nervous system (CNS) diseases. Although the replacement of lost endogenous cells was originally proposed as the primary healing mechanism of NSC grafts, it is now clear that transplanted NSCs operate via multiple mechanisms, including the horizontal exchange of therapeutic cargoes to host cells via extracellular vesicles (EVs). EVs are membrane particles trafficking nucleic acids, proteins, metabolites and metabolic enzymes, lipids, and entire organelles. However, the function and the contribution of these cargoes to the broad therapeutic effects of NSCs are yet to be fully understood. Mitochondrial dysfunction is an established feature of several inflammatory and degenerative CNS disorders, most of which are potentially treatable with exogenous stem cell therapeutics. Herein, we investigated the hypothesis that NSCs release and traffic functional mitochondria via EVs to restore mitochondrial function in target cells. Untargeted proteomics revealed a significant enrichment of mitochondrial proteins spontaneously released by NSCs in EVs. Morphological and functional analyses confirmed the presence of ultrastructurally intact mitochondria within EVs with conserved membrane potential and respiration. We found that the transfer of these mitochondria from EVs to mtDNA-deficient L929 Rho0 cells rescued mitochondrial function and increased Rho0 cell survival. Furthermore, the incorporation of mitochondria from EVs into inflammatory mononuclear phagocytes restored normal mitochondrial dynamics and cellular metabolism and reduced the expression of pro-inflammatory markers in target cells. When transplanted in an animal model of multiple sclerosis, exogenous NSCs actively transferred mitochondria to mononuclear phagocytes and induced a significant amelioration of clinical deficits. Our data provide the first evidence that NSCs deliver functional mitochondria to target cells via EVs, paving the way for the development of novel (a)cellular approaches aimed at restoring mitochondrial dysfunction not only in multiple sclerosis, but also in degenerative neurological diseases.

Selective Ablation of BDNF from Microglia Reveals Novel Roles in Self-Renewal and Hippocampal Neurogenesis.

Microglia, the resident immune cells of the CNS, have emerged as key regulators of neural precursor cell activity in the adult brain. However, the microglia-derived factors that mediate these effects remain largely unknown. In the present study, we investigated a role for microglial brain-derived neurotrophic factor (BDNF), a neurotrophic factor with well known effects on neuronal survival and plasticity. Surprisingly, we found that selective genetic ablation of BDNF from microglia increased the production of newborn neurons under both physiological and inflammatory conditions (e.g., LPS-induced infection and traumatic brain injury). Genetic ablation of BDNF from microglia otherwise also interfered with self-renewal/proliferation, reducing their overall density. In conclusion, we identify microglial BDNF as an important factor regulating microglia population dynamics and states, which in turn influences neurogenesis under both homeostatic and pathologic conditions.SIGNIFICANCE STATEMENT (1) Microglial BDNF contributes to self-renewal and density of microglia in the brain. (2) Selective ablation of BDNF in microglia stimulates neural precursor proliferation. (3) Loss of microglial BDNF augments working memory following traumatic brain injury. (4) Benefits of repopulating microglia on brain injury are not mediated via microglial BDNF.

Nanotechnology Facilitated Cultured Neuronal Network and Its Applications.

The development of a biomimetic neuronal network from neural cells is a big challenge for researchers. Recent advances in nanotechnology, on the other hand, have enabled unprecedented tools and techniques for guiding and directing neural stem cell proliferation and differentiation in vitro to construct an in vivo-like neuronal network. Nanotechnology allows control over neural stem cells by means of scaffolds that guide neurons to reform synaptic networks in suitable directions in 3D architecture, surface modification/nanopatterning to decide cell fate and stimulate/record signals from neurons to find out the relationships between neuronal circuit connectivity and their pathophysiological functions. Overall, nanotechnology-mediated methods facilitate precise physiochemical controls essential to develop tools appropriate for applications in neuroscience. This review emphasizes the newest applications of nanotechnology for examining central nervous system (CNS) roles and, therefore, provides an insight into how these technologies can be tested in vitro before being used in preclinical and clinical research and their potential role in regenerative medicine and tissue engineering.

Dynamic Changes in Ultrastructure of the Primary Cilium in Migrating Neuroblasts in the Postnatal Brain.

New neurons, referred to as neuroblasts, are continuously generated in the ventricular-subventricular zone of the brain throughout an animal's life. These neuroblasts are characterized by their unique potential for proliferation, formation of chain-like cell aggregates, and long-distance and high-speed migration through the rostral migratory stream (RMS) toward the olfactory bulb (OB), where they decelerate and differentiate into mature interneurons. The dynamic changes of ultrastructural features in postnatal-born neuroblasts during migration are not yet fully understood. Here we report the presence of a primary cilium, and its ultrastructural morphology and spatiotemporal dynamics, in migrating neuroblasts in the postnatal RMS and OB. The primary cilium was observed in migrating neuroblasts in the postnatal RMS and OB in male and female mice and zebrafish, and a male rhesus monkey. Inhibition of intraflagellar transport molecules in migrating neuroblasts impaired their ciliogenesis and rostral migration toward the OB. Serial section transmission electron microscopy revealed that each migrating neuroblast possesses either a pair of centrioles or a basal body with an immature or mature primary cilium. Using immunohistochemistry, live imaging, and serial block-face scanning electron microscopy, we demonstrate that the localization and orientation of the primary cilium are altered depending on the mitotic state, saltatory migration, and deceleration of neuroblasts. Together, our results highlight a close mutual relationship between spatiotemporal regulation of the primary cilium and efficient chain migration of neuroblasts in the postnatal brain.SIGNIFICANCE STATEMENT Immature neurons (neuroblasts) generated in the postnatal brain have a mitotic potential and migrate in chain-like cell aggregates toward the olfactory bulb. Here we report that migrating neuroblasts possess a tiny cellular protrusion called a primary cilium. Immunohistochemical studies with zebrafish, mouse, and monkey brains suggest that the presence of the primary cilium in migrating neuroblasts is evolutionarily conserved. Ciliogenesis in migrating neuroblasts in the rostral migratory stream is suppressed during mitosis and promoted after cell cycle exit. Moreover, live imaging and 3D electron microscopy revealed that ciliary localization and orientation change during saltatory movement of neuroblasts. Our results reveal highly organized dynamics in maturation and positioning of the primary cilium during neuroblast migration that underlie saltatory movement of postnatal-born neuroblasts.

Cadherin-11 promotes neural crest cell spreading by reducing intracellular tension-Mapping adhesion and mechanics in neural crest explants by atomic force microscopy.

During development cranial neural crest cells (NCCs) display a striking transition from collective to single-cell migration, but the mechanisms enabling individual NCCs to separate from the neural crest tissue are still incompletely understood. In this study we have employed atomic force microscopy (AFM) to investigate potential adhesive and mechanical changes associated with the dissociation of individual cells from cohesive Xenopus NCC explants at early stages of migration. AFM-based single-cell force spectroscopy (SCFS) revealed a uniform distribution of cell-cell adhesion forces within NCC explants, including semi-detached leader cells in the process of delaminating from the explant edge. This suggested that dissociation from the cell sheet may not require prior weakening of cell-cell contacts. However, mapping NCC sheet elasticity by AFM microbead indentation demonstrated strongly reduced cell stiffness in semi-detached leader cells compared to neighbouring cells in the NCC sheet periphery. Reduced leader cell stiffness coincided with enhanced cell spreading and high substrate traction, indicating a possible mechano-regulation of leader cell delamination. In support, AFM elasticity measurements of individual NCCs in optical side view mode demonstrated that reducing cell tension by inhibiting actomyosin contractility induces rapid spreading, possibly maximizing cell-substrate interactions as a result. Depletion of cadherin-11, a classical cadherin with an essential role in NCC migration and substrate adhesion, prevented the tension reduction necessary for NCC spreading, both in individual cells and at the edge of explanted sheets. In contrast, overexpression of cadherin-11 accelerated spreading of both individual cells and delaminating leader cells. As cadherin-11 expression increases strongly during NCC migration, this suggests an important role of cadherin-11 in regulating NCC elasticity and spreading at later stages of NCC migration. We therefore propose a model in which high tension at the NCC sheet periphery prevents premature NCC spreading and delamination during early stages of migration, while a cadherin-11-dependent local decrease in cell tension promotes leader cell spreading and delamination at later stages of migration.

Potential of ovine Wharton jelly derived mesenchymal stem cells to transdifferentiate into neuronal phenotype for application in neuroregenerative therapy.

Introduction: The transdifferentiation potential of mesenchymal stem cells (MSCs) is not limited to mesodermal derivatives but also to other cell types such as neuronal cells under appropriate cell culture conditions.Materials and methods: The present study characterizes the differentiation of Wharton's jelly (WJ) derived MSCs using neuronal conditioned medium (NCM) collected from cultured foetal brain cells.Results: After induction with NCM to neuronal stem cells (NSC), the WJ MSCs showed profound morphological changes showing multiple neurites extending from the cell body containing reminiscent of Nissl substance and single long axon-like processes. In RT PCR and immunocytochemistry, the induced neuronal cells showed a strong positive expression of neuronal markers Nestin, ß III tubulin and GFAP indicated that, the cells were reactive to NCM for differentiation. A significant (p < 0.01) increase in the level of secretome BDNF was observed in NCM suggests that the BDNF could play a key role in the transdifferentiation of WJMSCs to NSCs.Conclusion: These results support the potential of ovine MSCs isolated from umbilical cord WJ of abattoir derived foetuses to differentiate into neuronal stem cells and also provide a valuable experimental data for NSC transplant research in veterinary medicine.

Visualizing the Synaptic and Cellular Ultrastructure in Neurons Differentiated from Human Induced Neural Stem Cells-An Optimized Protocol.

The size of the synaptic subcomponents falls below the limits of visible light microscopy. Despite new developments in advanced microscopy techniques, the resolution of transmission electron microscopy (TEM) remains unsurpassed. The requirements of tissue preservation are very high, and human post mortem material often does not offer adequate quality. However, new reprogramming techniques that generate human neurons in vitro provide samples that can easily fulfill these requirements. The objective of this study was to identify the culture technique with the best ultrastructural preservation in combination with the best embedding and contrasting technique for visualizing neuronal elements. Two induced neural stem cell lines derived from healthy control subjects underwent differentiation either adherent on glass coverslips, embedded in a droplet of highly concentrated Matrigel, or as a compact neurosphere. Afterward, they were fixed using a combination of glutaraldehyde (GA) and paraformaldehyde (PFA) followed by three approaches (standard stain, Ruthenium red stain, high contrast en-bloc stain) using different combinations of membrane enhancing and contrasting steps before ultrathin sectioning and imaging by TEM. The compact free-floating neurospheres exhibited the best ultrastructural preservation. High-contrast en-bloc stain offered particularly sharp staining of membrane structures and the highest quality visualization of neuronal structures. In conclusion, compact neurospheres growing under free-floating conditions in combination with a high contrast en-bloc staining protocol, offer the optimal preservation and contrast with a particular focus on visualizing membrane structures as required for analyzing synaptic structures.

High glucose concentrations mask cellular phenotypes in a stem cell model of tuberous sclerosis complex.

Tuberous sclerosis complex (TSC) is a neurodevelopmental disorder caused by deletions in the TSC1 or TSC2 genes that is associated with epilepsy in up to 90% of patients. Seizures are suggested to start in benign brain tumors, cortical tubers, or in the perituberal tissue making these tubers an interesting target for further research into mechanisms underlying epileptogenesis in TSC. Animal models of TSC insufficiently capture the neurodevelopmental biology of cortical tubers, and hence, human stem cell-based in vitro models of TSC are being increasingly explored in attempts to recapitulate tuber development and epileptogenesis in TSC. However, in vitro culture conditions for stem cell-derived neurons do not necessarily mimic physiological conditions. For example, very high glucose concentrations of up to 25 mM are common in culture media formulations. As TSC is potentially caused by a disruption of the mechanistic target of rapamycin (mTOR) pathway, a main integrator of metabolic information and intracellular signaling, we aimed to examine the impact of different glucose concentrations in the culture media on cellular phenotypes implicated in tuber characteristics. Here, we present preliminary data from a pilot study exploring cortical neuronal differentiation on human embryonic stem cells (hES) harboring a TSC2 knockout mutation (TSC2-/-) and an isogenic control line (TSC2+/+). We show that the commonly used high glucose media profoundly mask cellular phenotypes in TSC2-/- cultures during neuronal differentiation. These phenotypes only become apparent when differentiating TSC2+/+ and TSC2-/- cultures in more physiologically relevant conditions of 5 mM glucose suggesting that the careful consideration of culture conditions is vital to ensuring biological relevance and translatability of stem cell models for neurological disorders such as TSC. This article is part of the Special Issue "Proceedings of the 7th London-Innsbruck Colloquium on Status Epilepticus and Acute Seizures".

Development of a neurotoxicity assay that is tuned to detect mitochondrial toxicants.

Many neurotoxicants affect energy metabolism in man, but currently available test methods may still fail to predict mito- and neurotoxicity. We addressed this issue using LUHMES cells, i.e., human neuronal precursors that easily differentiate into mature neurons. Within the NeuriTox assay, they have been used to screen for neurotoxicants. Our new approach is based on culturing the cells in either glucose or galactose (Glc-Gal-NeuriTox) as the main carbohydrate source during toxicity testing. Using this Glc-Gal-NeuriTox assay, 52 mitochondrial and non-mitochondrial toxicants were tested. The panel of chemicals comprised 11 inhibitors of mitochondrial respiratory chain complex I (cI), 4 inhibitors of cII, 8 of cIII, and 2 of cIV; 8 toxicants were included as they are assumed to be mitochondrial uncouplers. In galactose, cells became more dependent on mitochondrial function, which made them 2-3 orders of magnitude more sensitive to various mitotoxicants. Moreover, galactose enhanced the specific neurotoxicity (destruction of neurites) compared to a general cytotoxicity (plasma membrane lysis) of the toxicants. The Glc-Gal-NeuriTox assay worked particularly well for inhibitors of cI and cIII, while the toxicity of uncouplers and non-mitochondrial toxicants did not differ significantly upon glucose ↔ galactose exchange. As a secondary assay, we developed a method to quantify the inhibition of all mitochondrial respiratory chain functions/complexes in LUHMES cells. The combination of the Glc-Gal-NeuriTox neurotoxicity screening assay with the mechanistic follow up of target site identification allowed both, a more sensitive detection of neurotoxicants and a sharper definition of the mode of action of mitochondrial toxicants.

Cranial grafting of stem cell-derived microvesicles improves cognition and reduces neuropathology in the irradiated brain.

Cancer survivors face a variety of challenges as they cope with disease recurrence and a myriad of normal tissue complications brought on by radio- and chemotherapeutic treatment regimens. For patients subjected to cranial irradiation for the control of CNS malignancy, progressive and debilitating cognitive dysfunction remains a pressing unmet medical need. Although this problem has been recognized for decades, few if any satisfactory long-term solutions exist to resolve this serious unintended side effect of radiotherapy. Past work from our laboratory has demonstrated the neurocognitive benefits of human neural stem cell (hNSC) grafting in the irradiated brain, where intrahippocampal transplantation of hNSC ameliorated radiation-induced cognitive deficits. Using a similar strategy, we now provide, to our knowledge, the first evidence that cranial grafting of microvesicles secreted from hNSC affords similar neuroprotective phenotypes after head-only irradiation. Cortical- and hippocampal-based deficits found 1 mo after irradiation were completely resolved in animals cranially grafted with microvesicles. Microvesicle treatment was found to attenuate neuroinflammation and preserve host neuronal morphology in distinct regions of the brain. These data suggest that the neuroprotective properties of microvesicles act through a trophic support mechanism that reduces inflammation and preserves the structural integrity of the irradiated microenvironment.

Valproic acid promotes the neuronal differentiation of spiral ganglion neural stem cells with robust axonal growth.

Hearing loss occurs with the loss of hair cells of the cochlea and subsequent degeneration of spiral ganglion neurons (SGNs). Regeneration of SGNs is a potentially promising therapeutic approach to hearing loss in addition to the use of a cochlear implant (CI), because this device stimulates SGNs directly to restore hearing bypassing the missing hair cells. The presence of SGN-neural stem cells (NSCs) has been reported in adult human and mice. These cells have the potential to become SGNs and thus represent a cellular foundation for regeneration therapies for hearing loss. Valproic acid (VPA) has been shown to influence the neural differentiation of NSCs through multiple signaling pathways involving glycogen synthase kinase3ß (GSK3ß). Our present study therefore aimed to modulate the neural differentiation potential of SGN-NSCs by treatment with VPA. We here report that a clinically relevant concentration of 1 mM VPA induced the differentiation of basic fibroblast growth factor (bFGF)-treated P1- and P14-SGN-NSCs into neuronal and glial cells, confirmed by neuronal marker (Tuj1 and MAP2) and glial cell marker (GFAP and S100ß) detection. VPA-treated cells also promoted much longer neurite outgrowth compared to differentiated cells cultured without bFGF. The effects of VPA on the regulation of differentiation may be related to the activation of the Wnt/ß-catenin signaling pathway, but not the inhibition of histone deacetylases (HDACs). We propose that VPA has the potential to convert SGN-NSCs into SGNs and thereby restore hearing when combined with a CI.

Ghrelin protects adult rat hippocampal neural stem cells from excessive autophagy during oxygen-glucose deprivation.

Ghrelin functions as a neuroprotective agent and saves neurons from various insults include ischemic injury. However, it remains to be elucidated whether ghrelin protects neuronal cells against ischemic injury-induced excessive autophagy. Autophagy is required for the maintenance of neural stem cell homeostasis. However, regarding autophagic cell death, it is commonly assumed that excessive autophagy leads to self-elimination of mammalian cells. The purpose of this study was to investigate the potential neuroprotection effects of ghrelin from excessive autophagy in adult rat hippocampal neural stem cells (NSCs). Oxygen-Glucose Deprivation (OGD) strongly induces autophagy in adult rat hippocampal NSCs. Ghrelin treatment inhibited OGD-induced cell death of adult rat hippocampal NSCs assessed by cell-counting-kit-8 assay. Ghrelin also suppressed OGD-induced excessive autophagy activity. The protective effect of ghrelin was accompanied by an increased expression levels of Bcl-2, p-62 and decreased expression level of LC3-II, Beclin-1 by Western blot. Furthermore, ghrelin reduced autophagosome formation and number of GFP-LC3 transfected puncta. In conclusion, our data suggest that ghrelin protects adult rat hippocampal NSCs from excessive autophagy in experimental stroke (oxygen-glucose deprivation) model. Regulating autophagic activity may be a potential optimizing target for promoting adult rat hippocampal NSCs based therapy for stroke.

Deformation of Mitochondrial Cristae in Human Neural Progenitor Cells Exposed to Valproic Acid.

Neural development represents a dynamic process where mitochondrial integrity is decisive for neuronal activity. Structural changes in these organelles may be related to neurological disorders. Valproic acid (VPA) is an anticonvulsive drug commonly used for epilepsy treatment and its use is associated to increased risk of neuropsychiatric disorders. Recently we showed changes in shape and membrane potential in mitochondria from human neural progenitor cells (NPCs) exposed to VPA (da Costa et al. 2015). Here, we applied transmission electron microscopy and electron tomography to evaluate mitochondrial damage caused by VPA in NPCs. Results showed mitochondrial cristae disorganization in a dose dependent manner. Disturbance in mitochondrial ultrastructure may influence metabolism, leading to synaptic plasticity and neurogenesis impairment. These data contribute to understanding VPA exposure potential effects on brain development.

Resistance of glia-like central and peripheral neural stem cells to genetically induced mitochondrial dysfunction--differential effects on neurogenesis.

Mitochondria play a central role in stem cell homeostasis. Reversible switching between aerobic and anaerobic metabolism is critical for stem cell quiescence, multipotency, and differentiation, as well as for cell reprogramming. However, the effect of mitochondrial dysfunction on neural stem cell (NSC) function is unstudied. We have generated an animal model with homozygous deletion of the succinate dehydrogenase subunit D gene restricted to cells of glial fibrillary acidic protein lineage (hGFAP-SDHD mouse). Genetic mitochondrial damage did not alter the generation, maintenance, or multipotency of glia-like central NSCs. However, differentiation to neurons and oligodendrocytes (but not to astrocytes) was impaired and, hence, hGFAP-SDHD mice showed extensive brain atrophy. Peripheral neuronal populations were normal in hGFAP-SDHD mice, thus highlighting their non-glial (non hGFAP(+)) lineage. An exception to this was the carotid body, an arterial chemoreceptor organ atrophied in hGFAP-SDHD mice. The carotid body contains glia-like adult stem cells, which, as for brain NSCs, are resistant to genetic mitochondrial damage.

Isolation and characterization of neural stem cells from dystrophic mdx mouse.

The blood-brain barrier (BBB) is altered in mdx mouse, an animal model to study Duchenne muscular dystrophy (DMD). Our previous work demonstrated that perivascular glial endfeet control the selective exchanges between blood and neuropil as well as the BBB development and integrity; the alterations of dystrophin and dystrophin-associated protein complex (DAPs) in the glial cells of mdx mouse, parallel damages of the BBB and increase in vascular permeability. The aim of this study was to improve our knowledge about brain cellular components in the mdx mouse through the isolation, for the first time, of the adult neural stem cells (ANSCs). We characterized them by FACS, electron microscopy, confocal immunofluorescence microscopy, Real Time-PCR and western blotting, and we studied the expression of the DAPs aquaporin-4 (AQP4), potassium channel Kir4.1, α- and ß-dystroglycan (αDG, ßDG), α-syntrophin (αSyn), and short dystrophin isoform Dp71 proteins. The results showed that the mdx ANSCs expressed CD133 and Nestin receptor as the control ones, but showed a reduction in Notch receptor and altered cell proliferation with an increment in the apoptotic nuclei. Ultrastructurally, they appeared 50% size reduced compared to control ones, with a few cytoplasmic organelles. Moreover, the mdx ANSCs are devoid in full length dystrophin 427, and they expressed post-transcriptional reduction in the Dp71 in parallel with the ubiquitin proteasome activation, and decrement of DAPs proteins which appeared diffused in the cytoplasm and not polarized on the stem cells plasmamembrane, as prevalently observed in the controls. Overall, these results indicate that structural and molecular alterations affect the neural stem cells in the dystrophic brain, whose increased apoptosis and reduced Dp71 and DAPs proteins expression, together with loss in Dp427 dystrophin, could be responsible of the altered mdx glial maintenance and differentiation and consequent failure in the vessels barrier control occurring in the adult dystrophic brain.

Metformin activation of AMPK-dependent pathways is neuroprotective in human neural stem cells against Amyloid-beta-induced mitochondrial dysfunction.

Alzheimer's disease (AD) is the general consequence of dementia and is diagnostic neuropathology by the cumulation of amyloid-beta (Aß) protein aggregates, which are thought to promote mitochondrial dysfunction processes leading to neurodegeneration. AMP-activated protein kinase (AMPK), a critical regulator of energy homeostasis and a major player in lipid and glucose metabolism, is potentially implied in the mitochondrial deficiency of AD. Metformin, one of the widespread used anti- metabolic disease drugs, use its actions in part by stimulation of AMPK. While the mechanisms of AD are well established, the neuronal roles for AMPK in AD are still not well understood. In the present study, human neural stem cells (hNSCs) exposed to Aß had significantly reduced cell viability, which correlated with decreased AMPK, neuroprotective genes (Bcl-2 and CREB) and mitochondria associated genes (PGC1α, NRF-1 and Tfam) expressions, as well as increased activation of caspase 3/9 activity and cytosolic cytochrome c. Co-treatment with metformin distinct abolished the Aß-caused actions in hNSCs. Metformin also significantly rescued hNSCs from Aß-mediated mitochondrial deficiency (lower D-loop level, mitochondrial mass, maximal respiratory function, COX activity, and mitochondrial membrane potential). Importantly, co-treatment with metformin significantly restored fragmented mitochondria to almost normal morphology in the hNSCs with Aß. These findings extend our understanding of the central role of AMPK in Aß-related neuronal impairment. Thus, a better understanding of AMPK might assist in both the recognition of its critical effects and the implementation of new therapeutic strategies in the treatment of AD.

Primary cilia are required in a unique subpopulation of neural progenitors.

The apical domain of embryonic (radial glia) and adult (B1 cells) neural stem cells (NSCs) contains a primary cilium. This organelle has been suggested to function as an antenna for the detection of morphogens or growth factors. In particular, primary cilia are essential for Hedgehog (Hh) signaling, which plays key roles in brain development. Their unique location facing the ventricular lumen suggests that primary cilia in NSCs could play an important role in reception of signals within the cerebrospinal fluid. Surprisingly, ablation of primary cilia using conditional alleles for genes essential for intraflagellar transport [kinesin family member 3A (Kif3a) and intraflagellar transport 88 (Ift88)] and Cre drivers that are activated at early [Nestin; embryonic day 10.5 (E10.5)] and late [human glial fibrillary acidic protein (hGFAP); E13.5] stages of mouse neural development resulted in no apparent developmental defects. Neurogenesis in the ventricular-subventricular zone (V-SVZ) shortly after birth was also largely unaffected, except for a restricted ventral domain previously known to be regulated by Hh signaling. However, Kif3a and Ift88 genetic ablation also disrupts ependymal cilia, resulting in hydrocephalus by postnatal day 4. To directly study the role of B1 cells' primary cilia without the confounding effects of hydrocephalus, we stereotaxically targeted elimination of Kif3a from a subpopulation of radial glia, which resulted in ablation of primary cilia in a subset of B1 cells. Again, this experiment resulted in decreased neurogenesis only in the ventral V-SVZ. Primary cilia ablation led to disruption of Hh signaling in this subdomain. We conclude that primary cilia are required in a specific Hh-regulated subregion of the postnatal V-SVZ.

IL-1ß induces hypomyelination in the periventricular white matter through inhibition of oligodendrocyte progenitor cell maturation via FYN/MEK/ERK signaling pathway in septic neonatal rats.

Neuroinflammation elicited by microglia plays a key role in periventricular white matter (PWM) damage (PWMD) induced by infectious exposure. This study aimed to determine if microglia-derived interleukin-1ß (IL-1ß) would induce hypomyelination through suppression of maturation of oligodendrocyte progenitor cells (OPCs) in the developing PWM. Sprague-Dawley rats (1-day old) were injected with lipopolysaccharide (LPS) (1 mg/kg) intraperitoneally, following which upregulated expression of IL-1ß and IL-1 receptor 1 (IL-1R1 ) was observed. This was coupled with enhanced apoptosis and suppressed proliferation of OPCs in the PWM. The number of PDGFR-α and NG2-positive OPCs was significantly decreased in the PWM at 24 h and 3 days after injection of LPS, whereas it was increased at 14 days and 28 days. The protein expression of Olig1, Olig2, and Nkx2.2 was significantly reduced, and mRNA expression of Tcf4 and Axin2 was upregulated in the developing PWM after LPS injection. The expression of myelin basic protein (MBP) and 2',3'-cyclic-nucleotide 3"-phosphodiesterase (CNPase) was downregulated in the PWM at 14 days and 28 days after LPS injection; this was linked to reduction of the proportion of myelinated axons and thinner myelin sheath as revealed by electron microscopy. Primary cultured OPCs treated with IL-1ß showed the failure of maturation and proliferation. Furthermore, FYN/MEK/ERK signaling pathway was involved in suppression of maturation of primary OPCs induced by IL-1ß administration. Our results suggest that following LPS injection, microglia are activated and produce IL-1ß in the PWM in the neonatal rats. Excess IL-1ß inhibits the maturation of OPCs via suppression of FYN/MEK/ERK phosphorylation thereby leading to axonal hypomyelination.

Differential vascular permeability along the forebrain ventricular neurogenic niche in the adult murine brain.

Adult neurogenesis is influenced by blood-borne factors. In this context, greater or lesser vascular permeability along neurogenic niches would expose differentially neural stem cells (NSCs), transit amplifying cells (TACs), and neuroblasts to such factors. Here we evaluate endothelial cell morphology and vascular permeability along the forebrain neurogenic niche in the adult brain. Our results confirm that the subventricular zone (SVZ) contains highly permeable, discontinuous blood vessels, some of which allow the extravasation of molecules larger than those previously reported. In contrast, the rostral migratory stream (RMS) and the olfactory bulb core (OBc) display mostly impermeable, continuous blood vessels. These results imply that NSCs, TACs, and neuroblasts located within the SVZ are exposed more readily to blood-borne molecules, including those with very high molecular weights, than those positioned along the RMS and the OBc, subregions in which every stage of neurogenesis also takes place. These observations suggest that the existence of specialized vascular niches is not a precondition for neurogenesis to occur; specialized vascular beds might be essential for keeping high rates of proliferation and/or differential differentiation of neural precursors located at distinct domains.

Juxtacerebral Tissue Regeneration Potential: Telocytes Contribution.

It is well proved already that neurogenesis does take place in mammals' brain, including human brain. However, neurogenesis by itself is not able to compensate for brain tissue loss in serious neurological diseases, such as stroke, brain trauma or neurodegenerative disorders. Recent evidences show that neural stem cell niches are present not only in classical locations, such as subventricularor subgranular zones, but in other areas as well, including tissues contiguous to the brain (meninges and choroid plexus).In this chapter we revise the relationship of neural stem cells with interstitial cells (mainly telocytes), which we think is significant, and we describe what is known about the juxtacerebral tissue neurogenesis potential.