Hippocampal Precursor Cells Research Articles

Melanocortin-receptor 4 activation modulates proliferation and differentiation of rat postnatal hippocampal neural precursor cells

Postnatal hippocampal neurogenesis is essential for learning and memory. Hippocampal neural precursor cells (NPCs) can be induced to proliferate and differentiate into either glial cells or dentate granule cells. Notably, hippocampal neurogenesis decreases dramatically with age, partly due to a reduction in the NPC pool and a decrease in their proliferative activity. Alpha-melanocyte-stimulating hormone (α-MSH) improves learning, memory, neuronal survival and plasticity. Here, we used postnatally-isolated hippocampal NPCs from Wistar rat pups (male and female combined) to determine the role of the melanocortin analog [Nle4, D-Phe7]-α-MSH (NDP-MSH) in proliferation and fate acquisition of NPCs. Incubation of growth-factor deprived NPCs with 10 nM NDP-MSH for 6 days increased the proportion of Ki-67- and 5-bromo-2′-deoxyuridine (BrdU)-positive cells, compared to the control group, and these effects were blocked by the MC4R antagonist JKC-363. NDP-MSH also increased the proportion of glial fibrillar acidic protein (GFAP)/Ki-67, GFAP/sex-determining region Y-box2 (SOX2) and neuroepithelial stem cell protein (NESTIN)/Ki-67-double positive cells (type-1 and type-2 precursors). Finally, NDP-MSH induced peroxisome proliferator-activated receptor (PPAR)-γ protein expression, and co-incubation with the PPAR-γ inhibitor GW9662 prevented the effect of NDP-MSH on NPC proliferation and differentiation. Our results indicate that in vitro activation of MC4R in growth-factor-deprived postnatal hippocampal NPCs induces proliferation and promotes the relative expansion of the type-1 and type-2 NPC pool through a PPAR-γ-dependent mechanism. These results shed new light on the mechanisms underlying the beneficial effects of melanocortins in hippocampal plasticity and provide evidence linking the MC4R and PPAR-γ pathways in modulation of hippocampal NPC proliferation and differentiation.

A Poly-D-lysine-Coated Coralline Matrix Promotes Hippocampal Neural Precursor Cells' Differentiation into GFAP-Positive Astrocytes.

A major goal of regenerative medicine of the central nervous system is to accelerate the regeneration of nerve tissue, where astrocytes, despite their positive and negative roles, play a critical role. Thus, scaffolds capable of producing astrocytes from neural precursor cells (NPCs) are most desirable. Our study shows that NPCs are converted into reactive astrocytes upon cultivation on coralline-derived calcium carbonate coated with poly-D-lysine (PDL-CS). As shown via nuclei staining, the adhesion of neurospheres containing hundreds of hippocampal neural cells to PDL-CS resulted in disaggregation of the cell cluster as well as the radial migration of dozens of cells away from the neurosphere core. Migrating cells per neurosphere averaged 100 on PDL-CS, significantly higher than on uncoated CS (28), PDL-coated glass (65), or uncoated glass (20). After 3 days of culture on PDL-CS, cell migration plateaued and remained stable for four more days. In addition, NPCs expressing nestin underwent continuous morphological changes from round to spiky, extending and elongating their processes, resembling activated astrocytes. The extension of the process increased continuously during the maturation of the culture and doubled after 7 days compared to day 1, whereas bifurcation increased by twofold during the first 3 days before plateauing. In addition, nestin positive cells' shape, measured through the opposite circularity level correlation, decreased approximately twofold after three days, indicating spiky transformation. Moreover, nestin-positive cells co-expressing GFAP increased by 2.2 from day 1 to 7, reaching 40% of the NPC population on day 7. In this way, PDL-CS promotes NPC differentiation into reactive astrocytes, which could accelerate the repair of neural tissue.

Platelet-derived exerkine CXCL4/platelet factor 4 rejuvenates hippocampal neurogenesis and restores cognitive function in aged mice

The beneficial effects of physical activity on brain ageing are well recognised, with exerkines, factors that are secreted into the circulation in response to exercise, emerging as likely mediators of this response. However, the source and identity of these exerkines remain unclear. Here we provide evidence that an anti-geronic exerkine is secreted by platelets. We show that platelets are activated by exercise and are required for the exercise-induced increase in hippocampal precursor cell proliferation in aged mice. We also demonstrate that increasing the systemic levels of the platelet-derived exerkine CXCL4/platelet factor 4 (PF4) ameliorates age-related regenerative and cognitive impairments in a hippocampal neurogenesis-dependent manner. Together these findings highlight the role of platelets in mediating the rejuvenating effects of exercise during physiological brain ageing.

Mechanism of Molecular Activity of Yolkin—a Polypeptide Complex Derived from Hen Egg Yolk—in PC12 Cells and Immortalized Hippocampal Precursor Cells H19-7

Food-derived bioactive peptides able to regulate neuronal function have been intensively searched and studied for their potential therapeutic application. Our previous study showed that a polypeptide complex yolkin, isolated from hen egg yolk as a fraction accompanying immunoglobulin Y (IgY), improved memory and cognitive functions in rats. However, the mechanism activated by the yolkin is not explained. The goal of the present study was to examine what molecular mechanism regulating brain-derived neurotrophic factor (BDNF) expression is activated by the yolkin complex, using in vitro models of PC12 cell line and fetal rat hippocampal cell line H19-7. It was shown that yolkin increased the proliferative activity of rat hippocampal precursor cells H19-7 cells and upregulated the expression/production of BDNF in a cyclic adenosine monophosphate (cAMP)-response element-binding protein (CREB)-dependent manner. Additionally the upregulation of carboxypeptidase E/neurotrophic factor–α1 (CPE/(NF-α1) expression was shown. It was also determined that upregulation of CREB phosphorylation by yolkin is dependent on cyclic adenosine monophosphate/protein kinase A (cAMP/PKA) and phosphoinositide 3-kinases/protein kinase B (PI3K/Akt) signaling pathway activation. Moreover, the impact of yolkin on the level of intracellular Ca2+, nitric oxide, and activation of extracellular signal-regulated kinases 1/2 (ERK 1/2 kinase) was excluded. These results emphasize that yolkin can act comprehensively and in many directions and may participate in the regulation of neurons’ survival and activity. Therefore, it seems that the yolkin specimen can be used in the future as a safe, bioavailable, natural nutraceutical helping to improve the cognition of older people.

Zinc Water Prevents Autism-Like Behaviors in the BTBR Mice

This study aims to explore the effects of zinc water on autism-like behavior, convulsion threshold, and neurogenesis in ASD model animals. This study used the young BTBR ASD mouse model to explore the effect of a 6-week zinc water supplementation on ASD-like behaviors such as repetitive behavior and social communication disorder, seizure threshold, and the correlation with excitability regulation. The mice were divided into four groups of normal controls (B6) and models (BTBR) who did and did not receive zinc supplementation in water (B6, B6 + zinc, BTBR, and BTBR + zinc). For morphological changes in the hippocampus, we selected two indicators: hippocampal mossy fiber sprouting and neurogenesis. ASD-like behavior testing, seizure threshold determination, Timm staining, and neurogenesis-related assays—represented by Ki67 and DCX—were performed after 6 weeks of zinc supplementation. Our results show that zinc water can prevent autism-like behavior, reduce susceptibility to convulsions, and increase the proliferation of hippocampal progenitor cells in BTBR mice but has less effect on mossy fiber sprouting and neural progenitor cell differentiation. Zinc water reduces autism-like behavior in a partially inherited autism model mice—BTBR—which may be associated with hippocampal neural precursor cell proliferation and reversed hyperexcitability.

FOXG1 Contributes Adult Hippocampal Neurogenesis in Mice

Strategies to enhance hippocampal precursor cells efficiently differentiate into neurons could be crucial for structural repair after neurodegenerative damage. FOXG1 has been shown to play an important role in pattern formation, cell proliferation, and cell specification during embryonic and early postnatal neurogenesis. Thus far, the role of FOXG1 in adult hippocampal neurogenesis is largely unknown. Utilizing CAG-loxp-stop-loxp-Foxg1-IRES-EGFP (Foxg1fl/fl), a specific mouse line combined with CreAAV infusion, we successfully forced FOXG1 overexpressed in the hippocampal dentate gyrus (DG) of the genotype mice. Thereafter, we explored the function of FOXG1 on neuronal lineage progression and hippocampal neurogenesis in adult mice. By inhibiting p21cip1 expression, FOXG1-regulated activities enable the expansion of the precursor cell population. Besides, FOXG1 induced quiescent radial-glia like type I neural progenitor, giving rise to intermediate progenitor cells, neuroblasts in the hippocampal DG. Through increasing the length of G1 phase, FOXG1 promoted lineage-committed cells to exit the cell cycle and differentiate into mature neurons. The present results suggest that FOXG1 likely promotes neuronal lineage progression and thereby contributes to adult hippocampal neurogenesis. Elevating FOXG1 levels either pharmacologically or through other means could present a therapeutic strategy for disease related with neuronal loss.

CircRNA Acbd6 promotes neural stem cell differentiation into cholinergic neurons via the miR-320-5p-Osbpl2 axis

Neural stem cells (NSCs) persist in the dentate gyrus of the hippocampus into adulthood and are essential for both neurogenesis and neural circuit integration. Exosomes have also been shown to play vital roles in regulating biological processes of receptor cells as a medium for cell-to-cell communication signaling molecules. The precise molecular mechanisms of exosome-mediated signaling, however, remain largely unknown. Here, we found that exosomes produced by denervated hippocampi following fimbria–fornix transection could promote the differentiation of hippocampal neural precursor cells into cholinergic neurons in coculture with NSCs. Furthermore, we found that 14 circular RNAs (circRNAs) were upregulated in hippocampal exosomes after fimbria–fornix transection using high-throughput RNA-Seq technology. We further characterized the function and mechanism by which the upregulated circRNA Acbd6 (acyl-CoA-binding domain–containing 6) promoted the differentiation of NSCs into cholinergic neurons using RT–quantitative PCR, Western blot, ELISA, flow cytometry, immunohistochemistry, and immunofluorescence assay. By luciferase reporter assay, we demonstrated that circAcbd6 functioned as an endogenous miR-320-5p sponge to inhibit miR-320-5p activity, resulting in increased oxysterol-binding protein–related protein 2 expression with subsequent facilitation of NSC differentiation. Taken together, our results suggest that circAcbd6 promotes differentiation of NSCs into cholinergic neurons via miR-320-5p/oxysterol-binding protein–related protein 2 axis, which contribute important insights to our understanding of how circRNAs regulate neurogenesis.

Selenium mediates exercise-induced adult neurogenesis and reverses learning deficits induced by hippocampal injury and aging

Although the neurogenesis-enhancing effects of exercise have been extensively studied, the molecular mechanisms underlying this response remain unclear. Here, we propose that this is mediated by the exercise-induced systemic release of the antioxidant selenium transport protein, selenoprotein P (SEPP1). Using knockout mouse models, we confirmed that SEPP1 and its receptor low-density lipoprotein receptor-related protein 8 (LRP8) are required for the exercise-induced increase in adult hippocampal neurogenesis. Invivo selenium infusion increased hippocampal neural precursor cell (NPC) proliferation and adult neurogenesis. Mimicking the effect of exercise through dietary selenium supplementation restored neurogenesis and reversed the cognitive decline associated with aging and hippocampal injury, suggesting potential therapeutic relevance. These results provide a molecular mechanism linking exercise-induced changes in the systemic environment to the activation of quiescent hippocampal NPCs and their subsequent recruitment into the neurogenic trajectory.

Isolation and Culture of Adult Hippocampal Precursor Cells as Free-Floating Neurospheres.

The neurosphere assay is the most widely used in vitro tool to determine the proliferative and differentiation potential of adult neural precursor cells in rodents. Although originally developed for, and predominantly applied to, the growth of embryonic and adult subventricular zone-derived stem cells, hippocampal neurospheres are now routinely cultured by many laboratories. As hippocampal neurospheres are fewer in number, on average smaller in size, and more slowly growing than their ventricular counterparts, the methodology traditionally used to isolate and culture neurospheres from the subventricular zone is not optimal for hippocampal neurosphere growth. Here, we provide a detailed description of an optimized protocol for the microdissection, dissociation, and neurosphere generation from adult hippocampal dentate gyrus tissue. We also outline the protocols required to perform downstream passaging, differentiation, and immunohistological determination of the multipotentiality of hippocampal neurospheres.

Phenolic constituents isolated from Senna tora sprouts and their neuroprotective effects against glutamate-induced oxidative stress in HT22 and R28 cells

The consumption of sprouts has been steadily increasing due to their being an excellent source of nutrition. It is known that the bioactive constituents of legumes can be increased after germination. In this study, the extract from Senna tora sprouts is shown to exhibit improved radical scavenging activities and better neuroprotective effects in HT22 hippocampal neuronal (HT22) and R28 retina precursor (R28) cells than those from seeds due to an increased content of phenolic constituents, especially compounds 1 and 3–6. A phytochemical investigation of S. tora sprouts resulted in the isolation of two new naphthopyrone glycosides (1–2) with 27 previously reported compounds. Their structures were determined via interpreting spectroscopic data. Compounds 1 and 3–6 were found to possess radical scavenging activities and neuroprotective effects against oxidative stress in both neuronal cells. Hence, Senna tora sprouts and their constituents may be developed as natural neuroprotective agents via antioxidative effects.

Increased hippocampal TrkA expression ameliorates cranial radiation‑induced neurogenesis impairment and cognitive deficit via PI3K/AKT signaling.

Cognitive deficit is one of the most serious complications of cranial radiotherapy of head and neck cancers. However, the underlying mechanism of this cognitive impairment remains unclear. In the present study, the role of tropomyosin receptor kinase A (TrkA) and its ligand neurotrophin nerve growth factor (NGF) were investigated following whole-brain irradiation (WBI). Young male Sprague-Dawley rats underwent WBI at a single dose of 10 Gy. WBI was determined to result in notable memory decline and substantial neurogenesis impairment in the hippocampus 3 months post-irradiation. Compared with the control group, TrkA protein expression was greater in irradiated rats 1 week after WBI, which then decreased significantly by the 3-month time-point. However, no difference in NGF expression was observed from 1 day to 3 months post-WBI. Overexpression of hippocampal TrkA in rats using adeno-associated virus ameliorated memory decline induced by irradiation. Additionally, upregulating TrkA expression rescued irradiation-induced hippocampal precursor cell proliferation and promoted neurogenesis. PI3K, Akt and ERK1/2 phosphorylation were also revealed to be significantly inhibited by WBI, which was ameliorated by TrkA overexpression. Findings of the present study indicated that the TrkA-dependent signaling pathway may serve a critical role in radiotherapy-induced cognitive deficit and impairments in neurogenesis.

Antisense Oligonucleotide Therapeutic Approach for Suppression of Ataxin-1 Expression: A Safety Assessment

Spinocerebellar ataxia type 1 (SCA1) is a lethal, autosomal dominant neurodegenerative disease caused by a polyglutamine expansion in the ATAXIN-1 (ATXN1) protein. Preclinical studies demonstrate the therapeutic efficacy of approaches that target and reduce Atxn1 expression in a non-allele-specific manner. However, studies using Atxn1−/− mice raise cautionary notes that therapeutic reductions of ATXN1 might lead to undesirable effects such as reduction in the activity of the tumor suppressor Capicua (CIC), activation of the protease β-secretase 1 (BACE1) and subsequent increased amyloidogenic cleavage of the amyloid precursor protein (APP), or a reduction in hippocampal neuronal precursor cells that would impact hippocampal function. Here, we tested whether an antisense oligonucleotide (ASO)-mediated reduction of Atxn1 produced unwanted effects involving BACE1, CIC activity, or reduction in hippocampal neuronal precursor cells. Notably, no effects on BACE1, CIC tumor suppressor function, or number of hippocampal neuronal precursor cells were found in mice subjected to a chronic in vivo ASO-mediated reduction of Atxn1. These data provide further support for targeted reductions of ATXN1 as a therapeutic approach for SCA1.

Gintonin influences the morphology and motility of adult brain neurons via LPA receptors

BackgroundGintonin is an exogenous ginseng-derived G-protein-coupled lysophosphatidic acid (LPA) receptor ligand. LPA induces in vitro morphological changes and migration through neuronal LPA1 receptor. Recently, we reported that systemic administration of gintonin increases blood-brain barrier (BBB) permeability via the paracellular pathway and its binding to brain neurons. However, little is known about the influences of gintonin on in vivo neuron morphology and migration in the brain. Materials and methodsWe examined the effects of gintonin on in vitro migration and morphology using primary hippocampal neural precursor cells (hNPC) and in vivo effects of gintonin on adult brain neurons using real time microscopic analysis and immunohistochemical analysis to observe the morphological and locational changes induced by gintonin treatment. ResultsWe found that treating hNPCs with gintonin induced morphological changes with a cell rounding following cell aggregation and return to individual neurons with time relapses. However, the in vitro effects of gintonin on hNPCs were blocked by the LPA1/3 receptor antagonist, Ki16425, and Rho kinase inhibitor, Y27632. We also examined the in vivo effects of gintonin on the morphological changes and migration of neurons in adult mouse brains using anti-NeuN and -neurofilament H antibodies. We found that acute intravenous administration of gintonin induced morphological and migrational changes in brain neurons. Gintonin induced some migrations of neurons with shortened neurofilament H in the cortex. The in vivo effects of gintonin were also blocked by Ki16425. ConclusionThe present report raises the possibility that gintonin could enter the brain and exert its influences on the migration and morphology of adult mouse brain neurons and possibly explains the therapeutic effects of neurological diseases behind the gintonin administration.

Aβ40 Oligomers Promote Survival and Early Neuronal Differentiation of Dentate Gyrus-Isolated Precursor Cells Through Activation of the Akt Signaling Pathway.

The amyloid beta-peptide (Aβ) is the low-abundance product of amyloid precursor protein (APP), which is produced lifelong in the healthy brain. The functional properties of Aβ40 and Aβ42 peptides have not been completely elucidated to date. Although, several studies suggest that these peptides have a number of neurotrophic and neurotoxic properties, respectively. Interestingly, low concentrations of Aβ40 and Aβ42 regulate synaptic plasticity and improve cognitive functions, whereas the accumulation of Aβ42, coupled with the effects of age, can cause dysregulation of synaptic function, as is shown in Alzheimer's disease. Additionally, several studies suggest that both peptides, Aβ40 and Aβ42, are involved in neurogenic processes; however, these results are still controversial. Moreover, existing data indicate a direct relationship between the physicochemical characteristics of the peptides and their effects. Herein, we evaluated the effect of Aβ40 oligomers on hippocampal precursor cells isolated from the dentate gyrus of adult female C57Bl6 mice (mADGPCs). To this end, mADGPCs were treated with nanomolar and micromolar range concentrations of oligomeric forms of Aβ40 for 24, 48, and 72h to evaluate their effects on several events in the neurogenic process in vitro, including viability, proliferation, and early differentiation. The results indicate that Aβ40 favors mADGPC proliferation, survival, and neuronal differentiation following a mechanism that involves activation of the Akt signaling pathway. Thus, this study provides evidence about the positive effects of Aβ40 oligomers on the neurogenic process in adult mouse hippocampal precursor cells in vitro.

Dexmedetomidine attenuates ethanol-induced inhibition of hippocampal neurogenesis in neonatal mice

Background/AimsEthanol (EtOH) exposure during a period comparable to the third trimester in human results in obvious neurotoxicity in the developing hippocampus and persistent deficits in hippocampal neurogenesis. Dexmedetomidine (DEX), a highly selective α-2-adrenergic agonist has been demonstrated to restore the impaired neurogenesis and neuronal plasticity in the dentate gyrus (DG) that follows neurological insult. However, the protective roles of DEX in the EtOH-induced deficits of postnatal neurogenesis in the hippocampus are still unknown. MethodsMice were pretreated with DEX prior to EtOH exposure to determine its protective effects on impaired postnatal hippocampal neurogenesis. Six-day-old neonatal mice were treated with DEX (125 μg/kg) or saline, followed by EtOH at a total of 5 g/kg or an equivalent volume of saline on P7. Immunohistochemistry and immunofluorescence were used to evaluate the neurogenesis and activated microglia in the DG. Quantitative real time PCR (qRT-PCR) was utilized to assess the expression of inflammatory factors in the hippocampus. ResultsDEX pretreatment attenuated the inhibition of EtOH-mediated hippocampal neurogenesis and the reduction of hippocampal neural precursor cells (NPCs). We further confirmed that DEX pretreatment reversed the EtOH-induced microglia activation in the DG as well as the upregulation of the hippocampal TNFα, MCP-1, IL-6, and IL-1β mRNA levels. ConclusionOur findings indicate that DEX pretreatment protects against EtOH-mediated inhibition of hippocampal neurogenesis in postnatal mice and reverses EtOH-induced neuroinflammation via repressing microglia activation and the expression of inflammatory cytokines.

Bcl-2 Overexpression Induces Neurite Outgrowth via the Bmp4/Tbx3/NeuroD1 Cascade in H19-7 Cells.

Bcl-2 is overexpressed in the nervous system during neural development and plays an important role in modulating cell survival. In addition to its anti-apoptotic function, it has been suggested previously that Bcl-2 might act as a mediator of neuronal differentiation. However, the mechanism by which Bcl-2 might influence neurogenesis is not sufficiently understood. In this study, we aimed to determine the non-apoptotic functions of Bcl-2 during neuronal differentiation. First, we used microarrays to analyze the whole-genome expression patterns of rat neural stem cells overexpressing Bcl-2 and found that Bcl-2 overexpression induced the expression of various neurogenic genes. Moreover, Bcl-2 overexpression increased the neurite length as well as expression of Bmp4, Tbx3, and proneural basic helix-loop-helix genes, such as NeuroD1, NeuroD2, and Mash1, in H19-7 rat hippocampal precursor cells. To determine the hierarchy of these molecules, we selectively depleted Bmp4, Tbx3, and NeuroD1 in Bcl-2-overexpressing cells. Bmp4 depletion suppressed the upregulation of Tbx3 and NeuroD1 as well as neurite outgrowth, which was induced by Bcl-2 overexpression. Although Tbx3 knockdown repressed Bcl-2-mediated neurite elaboration and downregulated NeuroD1 expression, it did not affect Bcl-2-induced Bmp4 expression. While the depletion of NeuroD1 had no effect on the expression of Bcl-2, Bmp4, or Tbx3, Bcl-2-mediated neurite outgrowth was suppressed. Taken together, these results demonstrate that Bcl-2 regulates neurite outgrowth through the Bmp4/Tbx3/NeuroD1 cascade in H19-7 cells, indicating that Bcl-2 may have a direct role in neuronal development in addition to its well-known anti-apoptotic function in response to environmental insults.

The systemic exercise-released chemokine lymphotactin/XCL1 modulates in vitro adult hippocampal precursor cell proliferation and neuronal differentiation

Physical exercise has well-established anti-inflammatory effects, with neuro-immunological crosstalk being proposed as a mechanism underlying the beneficial effects of exercise on brain health. Here, we used physical exercise, a strong positive modulator of adult hippocampal neurogenesis, as a model to identify immune molecules that are secreted into the blood stream, which could potentially mediate this process. Proteomic profiling of mouse plasma showed that levels of the chemokine lymphotactin (XCL1) were elevated after four days of running. We found that XCL1 treatment of primary cells isolated from both the dentate gyrus and the subventricular zone of the adult mice led to an increase in the number of neurospheres and neuronal differentiation in neurospheres derived from the dentate gyrus. In contrast, primary dentate gyrus cells isolated from XCL1 knockout mice formed fewer neurospheres and exhibited a reduced neuronal differentiation potential. XCL1 supplementation in a dentate gyrus-derived neural precursor cell line promoted neuronal differentiation and resulted in lower cell motility and a reduced number of cells in the S phase of the cell cycle. This work suggests an additional function of the chemokine XCL1 in the brain and underpins the complexity of neuro-immune interactions that contribute to the regulation of adult hippocampal neurogenesis.

Anti-Myelin Proteolipid Protein Peptide Monoclonal Antibodies Recognize Cell Surface Proteins on Developing Neurons and Inhibit Their Differentiation

Using a panel of monoclonal antibodies (mAbs) to myelin proteolipid protein (PLP) peptides, we found that in addition to CNS myelin, mAbs to external face but not cytoplasmic face epitopes immunostained neurons in immature human CNS tissues and in adult hippocampal dentate gyrus and olfactory bulbs, that is neural stem cell niches (NSCN). To explore the pathobiological significance of these observations, we assessed the mAb effects on neurodifferentiation in vitro. The mAbs to PLP 50–69 (IgG1κ and IgG2aκ), and 178–191 and 200–219 (both IgG1κ) immunostained live cell surfaces and inhibited neurite outgrowth of E18 rat hippocampal precursor cells and of PC12 cells, which do not express PLP. Proteins immunoprecipitated from PC12 cell extracts and captured by mAb-coated magnetic beads were identified by GeLC-MS/MS. Each neurite outgrowth-inhibiting mAb captured a distinct set of neurodifferentiation molecules including sequence-similar M6 proteins and other unrelated membrane and extracellular matrix proteins, for example integrins, Eph receptors, NCAM-1, and protocadherins. These molecules are expressed in adult human NSCN and are implicated in the pathogenesis of many chronic CNS disease processes. Thus, diverse anti-PLP epitope autoantibodies may inhibit neuronal precursor cell differentiation via multispecific recognition of cell surface molecules thereby potentially impeding endogenous neuroregeneration in NSCN and in vivo differentiation of exogenous neural stem cells.

Potassium bisperoxo (1,10-phenanthroline) oxovanadate suppresses proliferation of hippocampal neuronal cell lines by increasing DNA methyltransferases.

Bisperoxo (1,10-phenanthroline) oxovanadate (BpV) can reportedly block the cell cycle. The present study examined whether BpV alters gene expression by affecting DNA methyltransferases (DNMTs), which would impact the cell cycle. Immortalized mouse hippocampal neuronal precursor cells (HT22) were treated with 0.3 or 3 μM BpV. Proliferation, morphology, and viability of HT22 cells were detected with an IncuCyte real-time video imaging system or inverted microscope and 3-(4,5-dimethylthiazol-2-yl)-5(3-carboxymethonyphenol)-2-(4-sulfophenyl)-2H-tetrazolium, respectively. mRNA and protein expression of DNMTs and p21 in HT22 cells was detected by real-time polymerase chain reaction and immunoblotting, respectively. In addition, DNMT activity was measured with an enzyme-linked immunosorbent assay. Effects of BpV on the cell cycle were analyzed using flow cytometry. Results demonstrated that treatment with 0.3 μM BpV did not affect cell proliferation, morphology, or viability; however, treatment with 3 μM BpV decreased cell viability, increased expression of both DNMT3B mRNA and protein, and inhibited the proliferation of HT22 cells; and 3 μM BpV also blocked the cell cycle and increased expression of the regulatory factor p21 by increasing DNMT expression in mouse hippocampal neurons.

EphA4 Regulates Hippocampal Neural Precursor Proliferation in the Adult Mouse Brain by d-Serine Modulation of N-Methyl-d-Aspartate Receptor Signaling.

The hippocampal dentate gyrus (DG) is a major region of the adult rodent brain in which neurogenesis occurs throughout life. The EphA4 receptor, which regulates neurogenesis and boundary formation in the developing brain, is also expressed in the adult DG, but whether it regulates adult hippocampal neurogenesis is not known. Here, we show that, in the adult mouse brain, EphA4 inhibits hippocampal precursor cell proliferation but does not affect precursor differentiation or survival. Genetic deletion or pharmacological inhibition of EphA4 significantly increased hippocampal precursor proliferation in vivo and in vitro, by blocking EphA4 forward signaling. EphA4 was expressed by mature hippocampal DG neurons but not neural precursor cells, and an EphA4 antagonist, EphA4-Fc, did not activate clonal cultures of precursors until they were co-cultured with non-precursor cells, indicating an indirect effect of EphA4 on the regulation of precursor activity. Supplementation with d-serine blocked the increased precursor proliferation induced by EphA4 inhibition, whereas blocking the interaction between d-serine and N-methyl-d-aspartate receptors (NMDARs) promoted precursor activity, even at the clonal level. Collectively, these findings demonstrate that EphA4 indirectly regulates adult hippocampal precursor proliferation and thus plays a role in neurogenesis via d-serine-regulated NMDAR signaling.