Survival Of Neural Stem Cells Research Articles

Involvement of Wnt7a in the role of M2c microglia in neural stem cell oligodendrogenesis

BackgroundThe participation of microglia in CNS development and homeostasis indicate that these cells are pivotal for the regeneration that occurs after demyelination. The clearance of myelin debris and the inflammatory-dependent activation of local oligodendrocyte progenitor cells in a demyelinated lesion is dependent on the activation of M2c microglia, which display both phagocytic and healing functions. Emerging interest has been raised about the role of Wnt/β-catenin signaling in oligodendrogenesis and myelination. Besides, cytokines and growth factors released by microglia can control the survival, proliferation, migration, and differentiation of neural stem cells (NSCs), contributing to remyelination through the oligodendrocyte specification of this adult neurogenic niche.MethodsTMEV-IDD model was used to study the contribution of dorsal SVZ stem cells to newly born oligodendrocytes in the corpus callosum following demyelination by (i) en-face dorsal SVZ preparations; (ii) immunohistochemistry; and (iii) cellular tracking. By RT-PCR, we analyzed the expression of Wnt proteins in demyelinated and remyelinating corpus callosum. Using in vitro approaches with microglia cultures and embryonic NSCs, we studied the role of purified myelin, Wnt proteins, and polarized microglia-conditioned medium to NSC proliferation and differentiation. One-way ANOVA followed by Bonferroni’s post-hoc test, or a Student’s t test were used to establish statistical significance.ResultsThe demyelination caused by TMEV infection is paralleled by an increase in B1 cells and pinwheels in the dorsal SVZ, resulting in the mobilization of SVZ proliferative progenitors and their differentiation into mature oligodendrocytes. Demyelination decreased the gene expression of Wnt5a and Wnt7a, which was restored during remyelination. In vitro approaches show that Wnt3a enhances NSC proliferation, while Wnt7a and myelin debris promotes oligodendrogenesis from NSCs. As phagocytic M2c microglia secrete Wnt 7a, their conditioned media was found to induce Wnt/β-Catenin signaling in NSCs promoting an oligodendroglial fate.ConclusionsWe define here the contribution of microglia to Wnt production depending on their activation state, with M1 microglia secreting the Wnt5a protein and M2c microglia secreting Wnt7a. Collectively, our data reveal the role of reparative microglia in NSC oligodendrogenesis with the involvement of Wnt7a.

Impact of composite scaffold degradation rate on neural stem cell persistence in the glioblastoma surgical resection cavity.

Tumoricidal neural stem cells (NSCs) are an emerging therapy to combat glioblastoma (GBM). This therapy employs genetically engineered NSCs that secrete tumoricidal agents to seek out and kill tumor foci remaining after GBM surgical resection. Biomaterial scaffolds have previously been utilized to deliver NSCs to the resection cavity. Here, we investigated the impact of scaffold degradation rate on NSC persistence in the brain resection cavity. Composite acetalated dextran (Ace-DEX) gelatin electrospun scaffolds were fabricated with two distinct degradation profiles created by changing the ratio of cyclic to acyclic acetal coverage of Ace-DEX. In vitro, fast degrading scaffolds were fully degraded by one week, whereas slow degrading scaffolds had a half-life of >56days. The scaffolds also retained distinct degradation profiles in vivo. Two different NSC lines readily adhered to and remained viable on Ace-DEX gelatin scaffolds, in vitro. Therapeutic NSCs secreting tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) had the same TRAIL output as tissue culture treated polystyrene (TCPS) when seeded on both scaffolds. Furthermore, secreted TRAIL was found to be highly potent against the human derived GBM cell line, GBM8, in vitro. Firefly luciferase expressing NSCs were seeded on scaffolds, implanted in a surgical resection cavity and their persistence in the brain was monitored by bioluminescent imaging (BLI). NSC loaded scaffolds were compared to a direct injection (DI) of NSCs in suspension, which is the current clinical approach to NSC therapy for GBM. Fast and slow degrading scaffolds enhanced NSC implantation efficiency 2.87 and 3.08-fold over DI, respectively. Interestingly, scaffold degradation profile did not significantly impact NSC persistence. However, persistence and long-term survival of NSCs was significantly greater for both scaffolds compared to DI, with scaffold implanted NSCs still detected by BLI at day 120 in most mice. Overall, these results highlight the benefit of utilizing a scaffold for application of tumoricidal NSC therapy for GBM.

The effect of Matrigel as scaffold material for neural stem cell transplantation for treating spinal cord injury

Traumatic injury to the spinal cord causes permanent loss of function and major personal, social, and economic problems. Cell-based delivery strategies is a promising approach for treating spinal cord injury (SCI). However, the inhospitable microenvironment in the injured spinal cord results in poor cell survival and uncontrolled differentiation of the transplanted stem cells. The combination of a scaffold with cells has been developed with a tendency for achieving greater survival and integration with the host tissue. We investigated the effect of Matrigel combined with neural stem cells (NSCs) in vitro and in vivo. We compared the effect of different types of scaffold on the survival and differentiation of brain-derived NSCs in an in vitro culture. Subsequently, NSCs were transplanted subcutaneously into nude mice to detect graft survival and differentiation in vivo. Finally, phosphate-buffered saline (PBS), Matrigel alone, or Matrigel seeded with NSCs was injected into 48 subacute, clinically relevant rat models of SCI (16 rats per group). Matrigel supported cell survival and differentiation efficiently in vitro and in vivo. SCI rats transplanted with NSCs in Matrigel showed improved behavioral recovery and neuronal and reactive astrocyte marker expression levels compared to PBS- or Matrigel-transplanted rats. Functional repair and neuronal and reactive astrocyte marker expression was slightly improved in the Matrigel-alone group relative to the PBS group, but not statistically significantly. These data suggest that Matrigel is a promising scaffold material for cell transplantation to the injured spinal cord.

Neuroinflammation and Neurogenesis in Alzheimer's Disease and Potential Therapeutic Approaches.

In adult brain, new neurons are generated throughout adulthood in the subventricular zone and the dentate gyrus; this process is commonly known as adult neurogenesis. The regulation or modulation of adult neurogenesis includes various intrinsic pathways (signal transduction pathway and epigenetic or genetic modulation pathways) or extrinsic pathways (metabolic growth factor modulation, vascular, and immune system pathways). Altered neurogenesis has been identified in Alzheimer’s disease (AD), in both human AD brains and AD rodent models. The exact mechanism of the dysregulation of adult neurogenesis in AD has not been completely elucidated. However, neuroinflammation has been demonstrated to alter adult neurogenesis. The presence of various inflammatory components, such as immune cells, cytokines, or chemokines, plays a role in regulating the survival, proliferation, and maturation of neural stem cells. Neuroinflammation has also been considered as a hallmark neuropathological feature of AD. In this review, we summarize current, state-of-the art perspectives on adult neurogenesis, neuroinflammation, and the relationship between these two phenomena in AD. Furthermore, we discuss the potential therapeutic approaches, focusing on the anti-inflammatory and proneurogenic interventions that have been reported in this field.

Can Mesenchymal Stem Cells Act Multipotential in Traumatic Brain Injury?

Traumatic brain injury (TBI), a leading cause of morbidity and mortality throughout the world, will probably become the third cause of death in the world by the year 2020. Lack of effective treatments approved for TBI is a major health problem. TBI is a heterogeneous disease due to the different mechanisms of injury. Therefore, it requires combination therapies or multipotential therapy that can affect multiple targets. In recent years, mesenchymal stem cells (MSCs) transplantation has considered one of the most promising therapeutic strategies to repair of brain injuries including TBI. In these studies, it has been shown that MSCs can migrate to the site of injury and differentiate into the cells secreting growth factors and anti-inflammatory cytokines. The reduction in brain edema, neuroinflammation, microglia accumulation, apoptosis, ischemia, the improvement of motor and cognitive function, and the enhancement in neurogenesis, angiogenesis, and neural stem cells survival, proliferation, and differentiation have been indicated in these studies. However, translation of MSCs research in TBI into a clinical setting will require additional preclinical trials.

Canine mesenchymal stromal cell-conditioned medium promotes survival and neurite outgrowth of neural stem cells.

We examined the paracrine action of canine mesenchymal stromal cells (MSCs) derived from bone marrow on the survival and differentiation of neural stem cells (NSCs) in vitro. MSCs were collected from the proximal end of the diaphysis of femur of healthy beagle dogs. The 70–80% confluent MSCs were re-fed with serum-free DMEM. The MSCs were incubated for 48 hr and the supernatant was collected as the conditioned medium (MSC-CM). The survival rate of NSCs in MSC-CM was significantly greater than in the medium without MSC-CM. The percentage of differentiated neurons and neurite length in MSC-CM was also significantly higher than in the medium without MSC-CM. These results suggested that canine MSC-CM promotes stem cell survival and neural differentiation of NSCs.

Retinoic Acid Is Required for Oligodendrocyte Precursor Cell Production and Differentiation in the Postnatal Mouse Corpus Callosum.

Myelination of the CNS relies on the production and differentiation of oligodendrocyte (OL) precursor cells (OPCs) into mature OLs. During the first month of postnatal life, OPCs that populate the corpus callosum (CC) arise from neural stem cells (NSCs) in the subcallosal subventricular zone (SVZ), and then differentiate to generate myelinating OLs. However, the signals that regulate these processes are not fully understood. In this study, we show that endogenous expression of the retinoic acid (RA)-synthesizing enzyme retinaldehyde dehydrogenase 2 (RALDH2) is required for OPC generation and differentiation in the postnatal subcortical white matter. In male and female pups, conditional deletion of Raldh2 reduced OPC numbers and differentiation. Moreover, decreased OPC numbers coincided with reductions in NSC survival and expression of the sonic hedgehog (SHH) signaling effector protein Gli1 in the SVZ. Additionally, GFAP expression in the CC was decreased, and cortical neuron numbers were altered. Our work suggests a role for endogenous RALDH2-dependent RA synthesis in OPC production and differentiation in the CC, as well as in the development of other cell types derived from NSCs in the embryonic ventricular zone (VZ) and SVZ, as well as the postnatal subcallosal SVZ.

PDE9 inhibition promotes proliferation of neural stem cells via cGMP-PKG pathway following oxygen-glucose deprivation/reoxygenation injury in vitro

Cerebral ischemia is one of leading causes of death and long-term disability worldwide. Stem cell-based therapy is promising some valuable strategies for the structural and functional recovery after ischemic insult. The inhibition of phosphodiesterases (PDEs) has wide spectrum neuroprotective properties by stimulating proliferation of neural stem cells (NSCs). However, the potential role of PDE9 on NSCs proliferation after cerebral ischemia is not well investigated. The present study aimed to assess the contribution of PDE9 inhibition on the proliferation of NSCs and to determine the details of its underlying mechanisms against cerebral ischemia. The survival and proliferation of NSCs were assessed by CCK-8 assay and BrdU immunofluorescence staining, respectively. PDE9 activity and cGMP level were measured by ELISA kits. The protein expression of PKG and BDNF was detected by Western blot. Exposing NSCs of cultured primary hippocampus to oxygen-glucose deprivation/reoxygenation (OGD/R) significantly decreased the survival rate, but increased the proliferation of NSCs. Meanwhile, PDE9 activity was decreased, cGMP level was increased, PKG and BDNF protein expression was increased. PF-04447953, a PDE9 inhibitor, increased the survival rate of NSCs, moreover, PDE9 activity reduced more, and NSCs proliferation, cGMP level, PKG and BDNF protein expression were increased further, compared with OGD/R model group. These effects of PF-04447953, except for PDE9 activity and cGMP level, were reversed by treatment with KT5823, a PKG inhibitor. Taken together, the inhibition of PDE9 can promote the proliferation of NSCs following OGD/R injury, which may be, at least partly, mediated by cGMP-PKG pathway.

Minimally Invasive Delivery of Microbeads with Encapsulated, Viable and Quiescent Neural Stem Cells to the Adult Subventricular Zone

Stem cell therapies demonstrate promising results as treatment for neurological disease and injury, owing to their innate ability to enhance endogenous neural tissue repair and promote functional recovery. However, delivery of undifferentiated and viable neuronal stem cells requires an engineered delivery system that promotes integration of transplanted cells into the inflamed and cytotoxic region of damaged tissue. Within the brain, endothelial cells (EC) of the subventricular zone play a critical role in neural stem cell (NSC) maintenance, quiescence and survival. Therefore, here, we describe the use of polyethylene glycol microbeads for the coincident delivery of EC and NSC as a means of enhancing appropriate NSC quiescence and survival during transplantation into the mouse brain. We demonstrate that EC and NSC co-encapsulation maintained NSC quiescence, enhanced NSC viability, and facilitated NSC extravasation in vitro, as compared to NSC encapsulated alone. In addition, co-encapsulated cells delivered to an in vivo non-injury model reduced inflammatory response compared to freely injected NSC. These results suggest the strong potential of a biomimetic engineered niche for NSC delivery into the brain following neurological injury.

Identifying the Zika Virus Target Cell in Malignant Glioma

Abstract The Zika Virus epidemic of 2015–2016 was associated with striking failure of forebrain development in infants born to infected mothers, resulting in microcephaly. Studies from numerous labs subsequently confirmed a selective effect of the virus on neural stem cell survival, self-renewal and differentiation. The glioma stem cells which drive glioblastoma and other malignant gliomas depend on neural stem cell transcription programs, so that understanding Zika infection in these cells promises valuable insights into future therapy. We describe here: 1) Study of low passage patient-derived glioblastoma cell lines and normal neural stem cells (CRUK Glioma Cellular Genetics Resource) in adherent culture to address the influence of glioma subtype on infectability 2) Application of genetically modified mCherry reporter Zika Virus strains to address the Zika target cell in glioblastoma. 3) Development of a cerebral organoid model to interrogate the differential effects of the virus on tumour cells and surrounding normal brain. 4) Demonstration of Zika infection on primary patient derived tissue in slice culture format Using these tools we find that Zika targets a common stem cell population across tumour subtypes and neural stem cell controls, and in embryonic and primary tumour explants, and that infection is influenced by activity of cholesterol biosynthesis pathways.

Dysfunctions of mitochondrial fatty acid β-oxidation in rare and common diseases

Dysfunctions of mitochondrial fatty acid ß-oxidation (ß-FAO) in various tissues represent a hallmark of many common disorders, and are acknowledged to play an essential role in the pathogenesis of diabetes, obesity, and cardiac diseases. Moreover, inborn defects in ß-FAO form a large family of rare diseases with variable phenotypes, ranging from fatal multi-organ failure in the newborn to isolated adult onset myopathy. These pathologies highlight the critical role of ß-FAO in many tissues with high-energy demand (heart, muscle, liver, kidney). Furthermore, and unexpectedly, very recent data unveiled the possible involvement of ß-FAO in instructing complex non energy-related functions, such as chromatin modification, control of neural stem cell activity, or survival and fate of cancer cells. Pharmacological targeting of ß-FAO by small molecules might therefore open new avenues for the treatment of various rare or common diseases.

Tissue-specific roles of p73 in development and homeostasis.

p73 (TP73) belongs to the p53 family of transcription factors. Its gene locus encodes two opposing types of isoforms, the transcriptionally active TAp73 class and the dominant-negative DNp73 class, which both play critical roles in development and homeostasis in an astonishingly diverse array of biological systems within specific tissues. While p73 has functions in cancer, this Review focuses on the non-oncogenic activities of p73. In the central and peripheral nervous system, both isoforms cooperate in complex ways to regulate neural stem cell survival, self-renewal and terminal differentiation. In airways, oviduct and to a lesser extent in brain ependyma, TAp73 is the master transcriptional regulator of multiciliogenesis, enabling fluid and germ cell transport across tissue surfaces. In male and female reproduction, TAp73 regulates gene networks that control cell-cell adhesion programs within germinal epithelium to enable germ cell maturation. Finally, p73 participates in the control of angiogenesis in development and cancer. While many open questions remain, we discuss here key findings that provide insight into the complex functions of this gene at the organismal, cellular and molecular level.

Chronic poly-drug administration damages adult mouse brain neural stem cells

Cocaine and ethanol are two commonly co-abused substances; however, the neuropathology following chronic dual consumption is poorly understood. Neural stem cells (NSCs) are a subpopulation of cells within the adult brain that are integral to brain maintenance and repair making them an appealing target to reverse neurodegeneration associated with abused substances. Yet, knowledge about NSC response to chronic poly-drug administration of ethanol and cocaine is minimal. Here, we developed a novel chronic poly-drug administration paradigm of ethanol and cocaine using a transgenic mouse model to trace endogenous NSC survival and differentiation in three brain regions from both male and female mice. We report significant and distinct patterns of NSC survival and differentiation among brain regions, as well as between sexes. Additionally, poly-drug administration had synergistic effects on NSC survival. Altered cognitive and hedonic behaviors were also observed, however the extent of these behavioral changes was not proportional to the NSC changes. With this mouse model we can effectively examine cognitive and behavioral changes and correlate them with pathological changes in the brain in response to chronic poly-drug administration, which is of great value in understanding the progression of neurodegeneration in polysubstance use disorders and evaluation potential therapeutics on neuroregeneration.

Knockdown of long noncoding RNA WNT5A-AS restores the fate of neural stem cells exposed to sevoflurane via inhibiting WNT5A/Ryk-ROS signaling

Long noncoding RNAs (lncRNAs) have been implicated in neurogenesis. LncRNA WNT5A-AS is upregulated in neural stem cells (NSCs), the proliferation of which is inhibited by sevoflurane. Thus, we hypothesized that knocking down of lncRNA WNT5A-AS may restore the fate of NSCs exposed to sevoflurane. To test this hypothesis, NSCs obtained from postnatal Sprague-Dawley rats were exposed to 2.4% sevoflurane or control gas for 6 h. Bioinformatics analysis, quantitative PCR and RNA interference technology were used to identify the properties of lncRNA WNT5A-AS. Cell proliferation was assessed using counting a Cell Counting Kit-cell 8 assay, a 5-ethynyl-2′-deoxyuridine incorporation assay, and a plate cloning assay. Cell survival was detected by flow cytometry, which was also used to examine the levels of reactive oxygen species (ROS) and the cell cycle. The levels of WNT5A and receptor tyrosine kinase (Ryk) were measured via Western blotting. LncRNA WNT5A-AS was identified to have low coding potency and to be located on the antisense strand of WNT5A. The level of upregulated lncRNA WNT5A-AS was positively correlated with that of WNT5A in response to sevoflurane exposure. The knockdown of lncRNA WNT5A-AS promoted the proliferation and survival of NSCs, whereas it suppressed the WNT5A/Ryk-ROS signaling and drove cell cycle processes. Taken together, findings strongly suggest that the inhibition of lncRNA WNT5A-AS can rescue the fate of NSCs. In addition, WNT5A/Ryk-ROS signaling might be a downstream target of lncRNA WNT5A-AS.

Lentivirus-mediated microRNA-26a-modified neural stem cells improve brain injury in rats with cerebral palsy.

This study is launched to investigate the effect of lentivirus-mediated microRNA-26a (miR-26a)-modified neural stem cells (NSCs) in brain injury in rats with cerebral palsy (CP). The successfully constructed miR-26a lentivirus expression vector and empty vector virus were used to modify NSCs. The model of CP with ischemia and anoxia was established in rats. NSCs and miR-26a-NSCs were stereoscopically injected into the cerebral cortex of the modeled rats, respectively. The survival and migration of NSCs infected with recombinant lentivirus expressing green fluorescence in vivo was observed under a light microscope. The neurobehavioral functions, morphology, and ultrastructure of cerebral cortex and hippocampus, apoptosis of brain cells, expression of apoptosis-related protein caspase-3 and Bax, together with the expression of the glial fibrillary acidic protein (GFAP) in cerebral cortex and hippocampus were determined. Expression of miR-26a in NSCs infected with plVTHM-miR-26a increased significantly. After NSCs transplantation, the neurobehavioral status of CP rats was improved, the degree of brain pathological injury was alleviated, the apoptotic index of cells in cerebral cortex and hippocampus and the expression of the apoptotic protein (caspase-3 and Bax) were decreased, the expression of GFAP were significantly decreased. After miR-26a-NSCs transplantation, these aforementioned results further improved or decreased. Our study suggests that miR-26a-modified NSCs mediated by lentivirus can improve brain injury, inhibit apoptosis of brain cells and activation of astrocytes in CP rats.

Substrate elasticity induces quiescence and promotes neurogenesis of primary neural stem cells-A biophysical in vitro model of the physiological cerebral milieu.

In the brain, neural stem cells (NSC) are tightly regulated by external signals and biophysical cues mediated by the local microenvironment or "niche." In particular, the influence of tissue elasticity, known to fundamentally affect the function of various cell types in the body, on NSC remains poorly understood. We, accordingly, aimed to characterize the effects of elastic substrates on critical NSC functions. Primary rat NSC were grown as monolayers on polydimethylsiloxane- (PDMS-) based gels. PDMS-coated cell culture plates, simulating the physiological microenvironment of the living brain, were generated in various degrees of elasticity, ranging from 1 to 50kPa; additionally, results were compared with regular glass plates as usually used in cell culture work. Survival of NSC on the PDMS-based substrates was unimpaired. The proliferation rate on 1kPa PDMS decreased by 45% compared with stiffer PMDS substrates of 50kPa (p<0.05) whereas expression of cyclin-dependent kinase inhibitor 1B/p27Kip1 increased more than two fold (p<0.01), suggesting NSC quiescence. NSC differentiation was accelerated on softer substrates and favored the generation of neurons (42% neurons on 1kPa PDMS vs. 25% on 50kPa PDMS; p<0.05). Neurons generated on 1kPa PDMS showed 29% longer neurites compared with those on stiffer PDMS substrates (p<0.05), suggesting optimized neuronal maturation and an accelerated generation of neuronal networks. Data show that primary NSC are significantly affected by the mechanical properties of their microenvironment. Culturing NSC on a substrate of brain-like elasticity keeps them in their physiological, quiescent state and increases their neurogenic potential.

Photosensitive Hydrogel Creates Favorable Biologic Niches to Promote Spinal Cord Injury Repair.

Photochemistry is considered to be a promising strategy for hydrogels to mimic the complex and dynamic properties of natural extracellular matrix. However, it is seldom applied in 3D tissue engineering and regenerative medicine due to the attenuation of light. In this study, phenyl azide photchemistry and optical fiber technology are first used to localize adhesive protein on the inner surface of the nerve guidance conduit in a 3D hydrogel scaffold. In vitro coculture assay of neural stem cells (NSCs) shows that photoimmobilization of collagen significantly improves the adhesion and survival of NSCs in the conduit, and exhibits synergistic effect with the sustainable release of growth factor. After implantation in transected spinal cord, the optimized hydrogel scaffold is found to improve the locomotion recovery of rats 12 weeks after spinal cord injury (SCI). Histological analysis suggests that the designed hydrogel scaffold provides a favorable biological niche for neuronal regeneration, thus producing directional neuron tissue and promoting the repair of SCI. This study demonstrates a promising hydrogel scaffold for SCI repair and provides the first understanding of the photoimmobilization of adhesive protein in a 3D hydrogel conduit concerning its functions on spinal cord tissue restoration.

Distinct gene-selective roles for a network of core promoter factors in Drosophila neural stem cell identity.

ABSTRACTThe transcriptional mechanisms that allow neural stem cells (NSC) to balance self-renewal with differentiation are not well understood. Employing an in vivo RNAi screen we identify here NSC-TAFs, a subset of nine TATA-binding protein associated factors (TAFs), as NSC identity genes in Drosophila. We found that depletion of NSC-TAFs results in decreased NSC clone size, reduced proliferation, defective cell polarity and increased hypersensitivity to cell cycle perturbation, without affecting NSC survival. Integrated gene expression and genomic binding analyses revealed that NSC-TAFs function with both TBP and TRF2, and that NSC-TAF-TBP and NSC-TAF-TRF2 shared target genes encode different subsets of transcription factors and RNA-binding proteins with established or emerging roles in NSC identity and brain development. Taken together, our results demonstrate that core promoter factors are selectively required for NSC identity in vivo by promoting cell cycle progression and NSC cell polarity. Because pathogenic variants in a subset of TAFs have all been linked to human neurological disorders, this work may stimulate and inform future animal models of TAF-linked neurological disorders.

Methionine-Choline Deprivation Impairs Adult Hippocampal Neurogenesis in C57BL/6 Mice.

Methionine and choline, which are essential nutrients for mammalian animals, are important for cell composition, as metabolic factors, and for the synthesis of other biochemical compounds for cell metabolism. Methionine and choline, which are methyl group donors, play key roles in the homocysteine cycle and neuronal development and maintenance. In this study, we investigated the effects of methionine and choline deficiency on adult hippocampal neurogenesis and neural stem cell (NSC) lineage in the adult stage. For this study, we divided C57BL/6 mice into three groups as follows: normal chow (NC)-fed, methionine choline sufficient (MCS) diet-fed, and methionine choline deficient (MCD) diet-fed mice. The mice were fed the NC, MCS, and MCD diets for 4 weeks from the age of 8 weeks. MCD diet-fed mice showed significantly decreased proliferation and differentiation of NSCs when compared with the NC diet-fed or MCS diet-fed mice. In addition, the survival of newly generated neurons was critically impaired in the MCD diet-fed mice. We confirmed a decrease in the proliferation and differentiation of NSCs after 4 weeks of MCD diet administration, compared with that in NC- and MCS diet-fed mice. MCD diet critically impaired NSCs survival and survival of neurons during the 4 weeks. The number of phosphorylated cyclic AMP response element binding (pCREB) protein immunoreactive nuclei was decreased in the MCD diet-fed mice compared with that in the NC- or MCS diet-fed group. These results suggest that suitable levels of methionine and choline are essential for the maintenance of hippocampal neurogenesis in mice and affect NSC proliferation and differentiation through phosphorylation of CREB.

Chronic Intrahippocampal Infusion of HIV-1 Neurotoxic Proteins: A Novel Mouse Model of HIV-1 Associated Inflammation and Neural Stem Cell Dysfunction.

HIV-1 infection causes chronic neuroinflammation resulting in cognitive decline associated with diminution of survival of neural stem cells (NSC). In part, this is attributable to production of toxic viral proteins (gp120 and tat) by infected cells in the brain that can activate microglia. Here, we evaluated a novel model for HIV-1 neuropathogenesis by direct administration of viral proteins into the hippocampus. Chronic administration of either HIV-1 gp120 or tat over 14days significantly decreased NSC proliferation, survival and neuroblast formation (by 32-37%) within the hippocampal subgranular zone as detected by doublecortin/BrdU or Ki67-positive cells. Intrahippocampal administration of gp120 or tat induced microglial activation within the hippocampus as determined by increases in microglial number and increases in the volume of the microglia (2.5-3-fold, evaluated by double IBA-1/CD68 staining). We further assessed inflammatory responses within the hippocampus by RNAseq and Ingenuity Pathway Analysis. There was a significant mRNA upregulation of numerous inflammatory mediators including Il1b, Icam1, Il12a, Ccl2, and Ccl4. These data suggest that chronic administration induces a prolonged inflammatory state within the hippocampus that negatively affects NSC survival potentially leading to cognitive dysfunction. Graphical Abstract.