Promotion of neurite outgrowth by 3,5,7,3',4'-pentamethoxyflavone is mediated through ERK signaling pathway in Neuro2a cells.

In this study, the effects of 3,5,7,3',4'-pentamethoxyflavone (KP1), a major bioactive ingredient isolated from the Kaempferia parviflora rhizomes, on a neurite outgrowth in Neuro2a cells and its mechanism have been investigated. KP1 increased concentration-dependently the percentage of neurite-bearing cells. KP1 showed a remarkable capability to elicit neurite outgrowth in Neuro2a cells, as evidenced by morphological alterations and immunostaining using anti-class III ß-tubulin and anti-NeuN antibodies. KP1 also displayed a higher neurogenic activity than retinoic acid (RA), a promoter of neurite outgrowth in Neuro2a cells. KP1 treatment caused significant elevation in phosphorylation of extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase (p38 MAPK) and glycogen synthase kinase-3ß (GSK-3ß). However, KP1-triggered neurite outgrowth was markedly inhibited by treatment with the ERK inhibitor U0126, whereas p38 MAPK inhibitor SB203580 and GSK-3ß inhibitor SB216763 did not influence KP1-induced neurite outgrowth. These results demonstrate that KP1 elicits neurite outgrowth and triggers cell differentiation of Neuro2a cells through ERK signal pathway.

Poria cocos (Schw.) Wolf, a Traditional Chinese Edible Medicinal Herb, Promotes Neuronal Differentiation, and the Morphological Maturation of Newborn Neurons in Neural Stem/Progenitor Cells.

Neurogenesis in the adult brain comprises the entire set of events of neuronal development. It begins with the division of precursor cells to form a mature, integrated, and functioning neuronal network. Adult neurogenesis is believed to play an important role in animals' cognitive abilities, including learning and memory. In the present study, significant neuronal differentiation-promoting activity of 80% (v/v) ethanol extract of P. cocos (EEPC) was found in Neuro-2a cells and mouse cortical neural stem/progenitor cells (NSPCs). Subsequently, a total of 97 compounds in EEPC were identified by UHPLC-Q-Exactive-MS/MS. Among them, four major compounds-Adenosine; Choline; Ethyl palmitoleate; and L-(-)-arabinitol-were further studied for their neuronal differentiation-promoting activity. Of which, choline has the most significant neuronal differentiation-promoting activity, indicating that choline, as the main bioactive compound in P. cocos, may have a positive effect on learning and memory functions. Compared with similar research literature, this is the first time that the neuronal differentiation-promoting effects of P. cocos extract have been studied.

Kaempferia parviflora Extracts Protect Neural Stem Cells from Amyloid Peptide-Mediated Inflammation in Co-Culture Model with Microglia.

The existence of neuroinflammation and oxidative stress surrounding amyloid beta (Aß) plaques, a hallmark of Alzheimer's disease (AD), has been demonstrated and may result in the activation of neuronal death and inhibition of neurogenesis. Therefore, dysregulation of neuroinflammation and oxidative stress is one possible therapeutic target for AD. Kaempferia parviflora Wall. ex Baker (KP), a member of the Zingiberaceae family, possesses health-promoting benefits including anti-oxidative stress and anti-inflammation in vitro and in vivo with a high level of safety; however, the role of KP in suppressing Aß-mediated neuroinflammation and neuronal differentiation has not yet been investigated. The neuroprotective effects of KP extract against Aß42 have been examined in both monoculture and co-culture systems of mouse neuroectodermal (NE-4C) stem cells and BV-2 microglia cells. Our results showed that fractions of KP extract containing 5,7-dimethoxyflavone, 5,7,4'-trimethoxyflavone, and 3,5,7,3',4'-pentamethoxyflavone protected neural stem cells (both undifferentiated and differentiated) and microglia activation from Aß42-induced neuroinflammation and oxidative stress in both monoculture and co-culture system of microglia and neuronal stem cells. Interestingly, KP extracts also prevented Aß42-suppressed neurogenesis, possibly due to the contained methoxyflavone derivatives. Our data indicated the promising role of KP in treating AD through the suppression of neuroinflammation and oxidative stress induced by Aß peptides.

The role of NAD metabolism in neuronal differentiation.

BACKGROUND: Nicotinamide adenine dinucleotide (NAD) metabolism is involved in redox and non-redox reactions that regulate several processes including differentiation of cells of different origins. Here, the role of NAD metabolism in neuronal differentiation, which remains elusive so far, was investigated. MATERIAL AND METHODS: A protein-protein interaction network between neurotrophin signaling and NAD metabolic pathways was built. Expression of NAD biosynthetic enzymes in SH-SY5Y cells during retinoic acid (RA)/brain derived neurotrophic factor (BDNF) differentiation, was evaluated. The effects of NAD biosynthetic enzymes QPRT and NAPRT inhibition in neurite outgrowth, cell viability, NAD availability and histone deacetylase (HDAC) activity, were analyzed in RA- and BDNF-differentiated cells. RESULTS: Bioinformatics analysis revealed the interaction between NAD biosynthetic enzyme NMNAT1 and NTRK2, a receptor activated by RA/BDNF sequential treatment. Differences were found in the expression of NAD biosynthetic enzymes during neuronal differentiation, namely, increased QPRT gene expression along the course of RA/BDNF treatment and NAPRT protein expression after a 5-day treatment with RA. QPRT inhibition in BDNF-differentiated SH-SY5Y cells resulted in less neuritic length per cell, decreased expression of the neuronal marker ß-III Tubulin and also decreased NAD+ levels and HDAC activity. NAPRT inhibition had no effect in neuritic length per cell, NAD+ levels and HDAC activity. Of note, NAD supplementation along with RA, but not with BDNF, resulted in considerable cell death. CONCLUSIONS: Taken together, our results show the involvement of NAD metabolism in neuronal differentiation, specifically, the importance of QPRT-mediated NAD biosynthesis in BDNF-associated SH-SY5Y differentiation and suggest additional roles for NAPRT beyond NAD production in RA-differentiated cells.

Basic Fibroblast Growth Factor Induces Cholinergic Differentiation of Tonsil-Derived Mesenchymal Stem Cells.

BACKGROUND: Mesenchymal stem cells (MSCs) are considered a potential tool for regenerating damaged tissues due to their great multipotency into various cell types. Here, we attempted to find the appropriate conditions for neuronal differentiation of tonsil-derived MSCs (TMSCs) and expand the potential application of TMSCs for treating neurological diseases. METHODS: The TMSCs were differentiated in DMEM/F-12 (Dulbecco's Modified Eagle Medium/Nutrient Mixture F-12) supplemented with various neurotrophic factors for 7-28 days to determine the optimal neuronal differentiation condition for the TMSCs. The morphologies as well as the levels of the neural markers and neurotransmitters were assessed to determine neuronal differentiation potentials and the neuronal lineages of the differentiated TMSCs. RESULTS: Our initial study demonstrated that DMEM/F12 supplemented with 50 ng/mL basic fibroblast growth factor with 10 µM forskolin was the optimal condition for neuronal differentiation for the TMSCs. TMSCs had higher protein expression of neuronal markers, including neuron-specific enolase (NSE), GAP43, postsynaptic density protein 95 (PSD95), and synaptosomal-associated protein of 25 kDa (SNAP25) compared to the undifferentiated TMSCs. Immunofluorescence staining also validated the increased mature neuron markers, NeuN and synaptophysin, in the differentiated TMSCs. The expression of glial fibrillar acidic protein and ionized calcium-binding adaptor molecule 1 the markers of astrocytes and microglia, were also slightly increased. Additionally, the differentiated TMSCs released a significantly higher level of acetylcholine, the cholinergic neurotransmitter, as analyzed by the liquid chromatography-tandem mass spectrometry and showed an enhanced choline acetyltransferase immunoreactivity compared to the undifferentiated cells. CONCLUSION: Our study suggests that the optimized condition favors the TMSCs to differentiate into cholinergic neuron-like phenotype, which could be used as a possible therapeutic tool in treating certain neurological disorders such as Alzheimer's disease.

Characterization of Hypothalamic MCH Neuron Development in a 3D Differentiation System of Mouse Embryonic Stem Cells.

Hypothalamic melanin-concentrating hormone (MCH) neurons are important regulators of multiple physiological processes, such as sleep, feeding, and memory. Despite the increasing interest in their neuronal functions, the molecular mechanism underlying MCH neuron development remains poorly understood. We report that a three-dimensional culture of mouse embryonic stem cells (mESCs) can generate hypothalamic-like tissues containing MCH-positive neurons, which reproduce morphologic maturation, neuronal connectivity, and neuropeptide/neurotransmitter phenotype of native MCH neurons. Using this in vitro system, we demonstrate that Hedgehog (Hh) signaling serves to produce major neurochemical subtypes of MCH neurons characterized by the presence or absence of cocaine- and amphetamine-regulated transcript (CART). Without exogenous Hh signals, mESCs initially differentiated into dorsal hypothalamic/prethalamic progenitors and finally into MCH+CART+ neurons through a specific intermediate progenitor state. Conversely, activation of the Hh pathway specified ventral hypothalamic progenitors that generate both MCH+CART- and MCH+CART+ neurons. These results suggest that in vivo MCH neurons may originate from multiple cell lineages that arise through early dorsoventral patterning of the hypothalamus. Additionally, we found that Hh signaling supports the differentiation of mESCs into orexin/hypocretin neurons, a well-defined cell group intermingled with MCH neurons in the lateral hypothalamic area (LHA). The present study highlights and improves the utility of mESC culture in the analysis of the developmental programs of specific hypothalamic cell types.

Chitosan Hydrogel Supplemented with Metformin Promotes Neuron-like Cell Differentiation of Gingival Mesenchymal Stem Cells.

Human gingival mesenchymal stem cells (GMSCs) are derived from migratory neural crest stem cells and have the potential to differentiate into neurons. Metformin can inhibit stem-cell aging and promotes the regeneration and development of neurons. In this study, we investigated the potential of metformin as an enhancer on neuronal differentiation of GMSCs in the growth environment of chitosan hydrogel. The crosslinked chitosan/ß-glycerophosphate hydrogel can form a perforated microporous structure that is suitable for cell growth and channels to transport water and macromolecules. GMSCs have powerful osteogenic, adipogenic and chondrogenic abilities in the induction medium supplemented with metformin. After induction in an induction medium supplemented with metformin, Western blot and immunofluorescence results showed that GMSCs differentiated into neuron-like cells with a significantly enhanced expression of neuro-related markers, including Nestin (NES) and ß-Tubulin (TUJ1). Proteomics was used to construct protein profiles in neural differentiation, and the results showed that chitosan hydrogels containing metformin promoted the upregulation of neural regeneration-related proteins, including ATP5F1, ATP5J, NADH dehydrogenase (ubiquinone) Fe-S protein 3 (NDUFS3), and Glutamate Dehydrogenase 1 (GLUD1). Our results help to promote the clinical application of stem-cell neural regeneration.

Total Ginsenoside Extract from Panax ginseng Enhances Neural Stem Cell Proliferation and Neuronal Differentiation by Inactivating GSK-3ß.

OBJECTIVE: To study the effects of total ginsenosides (TG) extract from Panax ginseng on neural stem cell (NSC) proliferation and differentiation and their underlying mechanisms. METHODS: The migration of NSCs after treatment with various concentrations of TG extract (50, 100, or 200 µ g/mL) were monitored. The proliferation of NSCs was examined by a combination of cell counting kit-8 and neurosphere assays. NSC differentiation mediated by TG extract was evaluated by Western blotting and immunofluorescence staining to monitor the expression of nestin and microtubule associated protein 2 (MAP2). The GSK-3ß/ß-catenin pathway in TG-treated NSCs was examined by Western blot assay. The NSCs with constitutively active GSK-3ß mutant were made by adenovirus-mediated gene transfection, then the proliferation and differentiation of NSCs mediated by TG were further verified. RESULTS: TG treatment significantly enhanced NSC migration (P<0.01 or P<0.05) and increased the proliferation of NSCs (P<0.01 or P<0.05). TG mediation also significantly upregulated MAP2 expression but downregulated nestin expression (P<0.01 or P<0.05). TG extract also significantly induced GSK-3ß phosphorylation at Ser9, leading to GSK-3ß inactivation and, consequently, the activation of the GSK-3ß/ß-catenin pathway (P<0.01 or P<0.05). In addition, constitutive activation of GSK-3ß in NSCs by the transfection of GSK-3ß S9A mutant was found to significantly suppress TG-mediated NSC proliferation and differentiation (P<0.01 or P<0.05). CONCLUSION: TG promoted NSC proliferation and neuronal differentiation by inactivating GSK-3ß.

Regulation of Neurogenesis and Neuronal Differentiation by Natural Compounds.

Neuronal damage or degeneration is the main feature of neurological diseases. Regulation of neurogenesis and neuronal differentiation is important in developing therapies to promote neuronal regeneration or synaptic network reconstruction. Neurogenesis is a multistage process in which neurons are generated and integrated into existing neuronal circuits. Neuronal differentiation is extremely complex because it can occur in different cell types and can be caused by a variety of inducers. Recently, natural compounds that induce neurogenesis and neuronal differentiation have attracted extensive attention. In this paper, the potential neural induction effects of medicinal plant-derived natural compounds on neural stem/progenitor cells (NS/PCs), the cultured neuronal cells, and mesenchymal stem cells (MSCs) are reviewed. The natural compounds that are efficacious in inducing neurogenesis and neuronal differentiation include phenolic acids, polyphenols, flavonoids, glucosides, alkaloids, terpenoids, quinones, coumarins, and others. They exert neural induction effects by regulating signal factors and cellspecific genes involved in the process of neurogenesis and neuronal differentiation, including specific proteins (ß-tubulin III, MAP-2, tau, nestin, neurofilaments, GFAP, GAP-43, NSE), related genes and proteins (STAT3, Hes1, Mash1, NeuroD1, notch, cyclin D1, SIRT1, Reggie-1), transcription factors (CREB, Nkx-2.5, Ngn1), neurotrophins (BDNF, NGF, NT-3), and signaling pathways (JAK/STAT, Wnt/ß-catenin, MAPK, PI3K/Akt, GSK-3ß/ß-catenin, Ca2+/CaMKII/ATF1, Nrf2/HO-1, BMP).The natural compounds with neural induction effects are of great value for neuronal regenerative medicine and provide promising prevention and treatment strategies for neurological diseases.

Total Ginsenoside Extract from Panax ginseng Enhances Neural Stem Cell Proliferation and Neuronal Differentiation by Inactivating GSK-3β / 中国结合医学杂志

OBJECTIVE@#To study the effects of total ginsenosides (TG) extract from Panax ginseng on neural stem cell (NSC) proliferation and differentiation and their underlying mechanisms.@*METHODS@#The migration of NSCs after treatment with various concentrations of TG extract (50, 100, or 200 µ g/mL) were monitored. The proliferation of NSCs was examined by a combination of cell counting kit-8 and neurosphere assays. NSC differentiation mediated by TG extract was evaluated by Western blotting and immunofluorescence staining to monitor the expression of nestin and microtubule associated protein 2 (MAP2). The GSK-3β/β-catenin pathway in TG-treated NSCs was examined by Western blot assay. The NSCs with constitutively active GSK-3β mutant were made by adenovirus-mediated gene transfection, then the proliferation and differentiation of NSCs mediated by TG were further verified.@*RESULTS@#TG treatment significantly enhanced NSC migration (P<0.01 or P<0.05) and increased the proliferation of NSCs (P<0.01 or P<0.05). TG mediation also significantly upregulated MAP2 expression but downregulated nestin expression (P<0.01 or P<0.05). TG extract also significantly induced GSK-3β phosphorylation at Ser9, leading to GSK-3β inactivation and, consequently, the activation of the GSK-3β/β-catenin pathway (P<0.01 or P<0.05). In addition, constitutive activation of GSK-3β in NSCs by the transfection of GSK-3β S9A mutant was found to significantly suppress TG-mediated NSC proliferation and differentiation (P<0.01 or P<0.05).@*CONCLUSION@#TG promoted NSC proliferation and neuronal differentiation by inactivating GSK-3β.

Dendrobium officinale flos increases neurotrophic factor expression in the hippocampus of chronic unpredictable mild stress-exposed mice and in astrocyte primary culture and potentiates NGF-induced neuronal differentiation in PC12 cells.

Dendrobium officinale flos (DOF) is the flower of Dendrobium officinale Kimura et Migo, which is usually regarded as a by-product of Dendrobii Offcinalis Caulis. Based on its use as an alternative medicine, we evaluated the antidepressant-like effect of DOF extracts on chronic, unpredictable, mild stress-induced, depression-like behaviour in mice and tested the effects of DOF on the regulation of neurotrophic factors in mouse astrocyte primary cultures and PC12 cell lines. Oral treatment with DOF ethanol extract (DOF-E) could alleviate depression-like behaviours in stress-exposed mice, as evidenced by increased sucrose consumption and decreased immobile time in a forced swim test. In the hippocampus, DOF extracts increased the expression of NGF and BDNF, both at the transcriptional and protein levels. In astrocytes, DOF-E increased the expression of NGF and BDNF via a cAMP-dependent mechanism and regulated plasminogen and matrix metallopeptidase 9 (MMP-9), which are related to the metabolic regulation of neurotrophic factors. In PC12 cells, DOF-E induced the expression of neurofilaments and potentiated the induction of neurite outgrowth upon treatment with a low dose of NGF. Based on these findings, DOF might be used as a supplement for antidepressant therapy in patients with depression.

Conditional knockout of leptin receptor in neural stem cells leads to obesity in mice and affects neuronal differentiation in the hypothalamus early after birth.

Leptin, secreted by peripheral adipocytes, binds the leptin receptor (Lepr) in the hypothalamus, thereby contributing to the regulation of satiety and body weight. Lepr is expressed in the embryonic brain as early as embryonic day 12.5. However, the function of Lepr in neural precursor cells in the brain has not been resolved. To address this issue, we crossed the Leprflox/flox mice with each of Shh-Cre mice (Shh, sonic hedgehog) and Nestin (Nes)-Cre mice. We found that deletion of Lepr specifically in nestin-expressing cells led to extreme obesity, but the conditional null of Lepr in Shh-expressing cells had no obvious phenotype. Moreover, the level of leptin-activated pSTAT3 decreased in the anterior and central subregions of the arcuate hypothalamus of Shh-Cre; Leprflox/flox mice compared with the controls. By contrast, in Nes-Cre; Leprflox/flox mice, the level of leptin-activated pSTAT3 decreased in all subregions including the anterior, central, and posterior arcuate hypothalamus as well as the dorsomedial, ventromedial, and median eminence of the hypothalamus, revealing that the extensive lack of Lepr in the differentiated neurons of the hypothalamus in the conditional null mice. Notably, conditional deletion of Lepr in nestin-expressing cells enhanced the differentiation of neural precursor cells into neurons and oligodendroglia but inhibited differentiation into astrocytes early in postnatal development of hypothalamus. Our results suggest that Lepr expression in neural precursor cells is essential for maintaining normal body weight as well as the differentiation of neural precursor cells to the neural/glial fate in the hypothalamus shortly after birth.

Effects of Cyperus rotundus Extract on Spatial Memory Impairment and Neuronal Differentiation in Rat Model of Alzheimer's Disease.

BACKGROUND: Alzheimer's disease (AD) is one of the most common neurodegenerative diseases in the older population and characterized by progressive memory and cognitive impairment. Cyperus rotundus, a traditional medicinal herb, has analgesic, sedative, and anti-inflammatory effects and also used to increase memory in Islamic traditional medicine. This study was designed to consider the effects of C. rotundus extract on memory impairment and neurogenesis in the Beta-Amyloid rats' model. MATERIALS AND METHODS: Forty-two male Wistar rats were randomly divided into six groups (n = 7) for the evaluation of baseline training performance in the Morris water maze test. Then, amyloid-beta (Aß1-42) was injected in animal hippocampal CA1 bilaterally in four groups. The first probe trial was performed 21 days after Aß injection. Then, 250, 500, and 750 mg/kg of C. rotundus extract were administered to three Aß-injected groups for 1 month; after that, the second probe trial was performed, and rats were sacrificed after 28 days of the second probe trial. The neurogenesis was detected in the hippocampus, by immunohistochemical staining. RESULTS: This study showed that spatial memory increased in the behavioral test in AD treated group with C. rotundus extract, compared with the AD group (P = 0.02). Immunohistochemical staining revealed that neuronal differentiation has been occurred in the hippocampus in the AD-treated group with C. rotundus extract compared with the AD group (P = 0.01). CONCLUSIONS: This study showed that C. rotundus extract, repaired spatial memory impairment in the Aß rats, through increased neurogenesis in the hippocampus, which could be related to the flavonoid components in the extract.

Complete neural stem cell (NSC) neuronal differentiation requires a branched chain amino acids-induced persistent metabolic shift towards energy metabolism.

Neural stem cell (NSC) neuronal differentiation requires a metabolic shift towards oxidative phosphorylation. We now show that a branched-chain amino acids-driven, persistent metabolic shift toward energy metabolism is required for full neuronal maturation. We increased energy metabolism of differentiating neurons derived both from murine NSCs and human induced pluripotent stem cells (iPSCs) by supplementing the cell culture medium with a mixture composed of branched-chain amino acids, essential amino acids, TCA cycle precursors and co-factors. We found that treated differentiating neuronal cells with enhanced energy metabolism increased: i) total dendritic length; ii) the mean number of branches and iii) the number and maturation of the dendritic spines. Furthermore, neuronal spines in treated neurons appeared more stable with stubby and mushroom phenotype and with increased expression of molecules involved in synapse formation. Treated neurons modified their mitochondrial dynamics increasing the mitochondrial fusion and, consistently with the increase of cellular ATP content, they activated cellular mTORC1 dependent p70S6 K1 anabolism. Global transcriptomic analysis further revealed that treated neurons induce Nrf2 mediated gene expression. This was correlated with a functional increase in the Reactive Oxygen Species (ROS) scavenging mechanisms. In conclusion, persistent branched-chain amino acids-driven metabolic shift toward energy metabolism enhanced neuronal differentiation and antioxidant defences. These findings offer new opportunities to pharmacologically modulate NSC neuronal differentiation and to develop effective strategies for treating neurodegenerative diseases.

Yokukansan, a Kampo medicine, enhances the level of neuronal lineage markers in differentiated P19 embryonic carcinoma cells.

Yokukansan (YKS), a traditional Japanese Kampo medicine, affects neurological and psychiatric disorders. It ameliorates hippocampal neurogenesis in animals. However, its effect on neuronal cell differentiation remains unclear. Therefore, we investigated the effects of YKS on pluripotent P19 embryonic carcinoma cells as neuronal differentiation model cells. Western blotting and immunocytochemistry revealed that 10 µg/mL YKS treatment during embryoid body formation or neuronal differentiation increased the expression of the neuronal stem cell marker, Nestin, by 1.9-fold and 1.7-fold, respectively, and of the mature neuron marker, NeuN, by 1.5-fold and 1.4-fold, respectively. We examined the effect of YKS on intracellular signaling pathways in P19 cells and found significant elevation in phospho-PDK1 and phospho-mTOR expression (1.1-fold and 1.2-fold, respectively). Therefore, we investigated the effect of PDK1 and mTOR inhibitors on the level of neuronal lineage markers. We found that the mTOR inhibitor significantly abolished the YKS effect on the level of neuronal lineage markers. Moreover, to identify the target(s) of YKS, antibody array analysis that simultaneously detects 16 phosphorylated proteins was performed. YKS significantly upregulated 10 phosphorylated proteins including PDK1, Akt, AMPK, PRAS40, mTOR, p70 S6 kinase, GSK-3α, Bad and ERK1/2 under cell proliferation conditions. These results suggest that YKS simultaneously activates multiple signaling pathways. Thus, we concluded that YKS enhances the level of neuronal lineage markers in differentiated P19 cells, however it does not induce neuronal differentiation. Furthermore, mTOR is the predominant mediator of the YKS effect on these cells.

Shexiang Baoxin Pill, a Formulated Chinese Herbal Mixture, Induces Neuronal Differentiation of PC12 Cells: A Signaling Triggered by Activation of Protein Kinase A.

Background: Shexiang Baoxin Pill (SBP) is a well-known composite formula of traditional Chinese medicine (TCM), which is commonly used today in treating cardiovascular diseases. SBP consists of seven materials thereof, including Moschus, extract of Ginseng Radix et Rhizoma, Bovis Calculus Artifactus, Cinnamomi Cortex, Styrax, Bufonis Venenum, and Borneolum Syntheticum. Here, we are investigating the potential roles of SBP in inducing neuron differentiation, i.e., seeking possible application in neurodegenerative diseases. Methods: Water and ethanol extracts of SBP, denoted as SBPwater and SBPEtOH, respectively, as well as its individual herbal materials, were standardized and applied onto cultured rat pheochromocytoma PC12 cells. The potential effect of SBP extracts in neuronal differentiation was suggested by following parameters: (i) induction of neurite outgrowth of PC12 cells, (ii) increase of neurofilament expression, and (iii) activation of transcription of neurofilament. Results: The treatments of SBPwater and SBPEtOH, or extracts from individual herbal materials, with or without low concentration of nerve growth factor (NGF), could potentiate the differentiation of cultured PC12 cells. The differentiation was indicated by increase of neurite outgrowth, as well as expression of neurofilaments. In addition, application of H89, a protein kinase A (PKA) inhibitor, suppressed the SBP-induced neurofilament expressions, as well as the phosphorylation of cAMP-responsive element binding protein (CREB) in cultures. Conclusion: SBP is proposed to possess trophic activity in modulating neuronal differentiation of PC12 cells, and this induction is shown to be mediated partly by a cAMP-PKA signaling pathway. These results indicate the neurite-promoting SBP could be useful in developing potential drug in treating or preventing neurodegenerative diseases.

Anti-aging effects exerted by Tetramethylpyrazine enhances self-renewal and neuronal differentiation of rat bMSCs by suppressing NF-kB signaling.

In order to improve the therapeutic effects of mesenchymal stem cell (MSC)-based therapies for a number of intractable neurological disorders, a more favorable strategy to regulate the outcome of bone marrow MSCs (bMSCs) was examined in the present study. In view of the wide range of neurotrophic and neuroprotective effects, Tetramethylpyrazine (TMP), a biologically active alkaloid isolated from the herbal medicine Ligusticum wallichii, was used. It was revealed that treatment with 30-50 mg/l TMP for 4 days significantly increased cell viability, alleviated senescence by suppressing NF-κB signaling, and promoted bMSC proliferation by regulating the cell cycle. In addition, 40-50 mg/l TMP treatment may facilitate the neuronal differentiation of bMSCs, verified in the present study by presentation of neuronal morphology and expression of neuronal markers: microtubule-associated protein 2 (MAP-2) and neuron-specific enolase (NSE). The quantitative real-time polymerase chain reaction (qRT-PCR) revealed that TMP treatment may promote the expression of neurogenin 1 (Ngn1), neuronal differentiation 1 (NeuroD) and mammalian achaete-scute homolog 1 (Mash1). In conclusion, 4 days of 40-50 mg/l TMP treatment may significantly delay bMSC senescence by suppressing NF-κB signaling, and enhancing the self-renewal ability of bMSCs, and their potential for neuronal differentiation.

Phenotypic Screening Using Mouse and Human Stem Cell-Based Models of Neuroinflammation and Gene Expression Analysis to Study Drug Responses.

High-throughput phenotypic screening enables the identification of new therapeutic targets even when the molecular mechanism underlying the disease is unknown. In the case of neurodegenerative disease, there is a dire need to identify new targets that can ameliorate, halt, or reverse degeneration. Stem cell-based disease models are particularly powerful tools for phenotypic screening because they use the same cell type affected in patients. Here, we describe the expansion of mouse stem cells and human induced pluripotent stem cells as well as the differentiation of these cells into neural lineages that, when exposed to neuroinflammatory stress, can be used for compound screening followed by hit identification, validation, and target deconvolution.

Modulation of mitochondrial respiration underpins neuronal differentiation enhanced by lutein.

Lutein is a dietary carotenoid of particular nutritional interest as it is preferentially taken up by neural tissues. Often linked with beneficial effects on vision, a broader role for lutein in neuronal differentiation has emerged recently, although the underlying mechanisms for these effects are not yet clear. The purpose of this study was to investigate the effect of lutein on neuronal differentiation and explore the associated underpinning mechanisms. We found that lutein treatment enhanced the differentiation of SH-SY5Y cells, specifically increasing neuronal arborization and expression of the neuronal process filament protein microtubule-associated protein 2. This effect was mediated by the intracellular phosphoinositide-3-kinase (PI3K) signaling pathway. While PI3K activity is a known trigger of neuronal differentiation, more recently it has also been shown to modulate the metabolic state of cells. Our analysis of bioenergetics found that lutein treatment increased glucose consumption, rates of glycolysis and enhanced respiratory activity of mitochondrial complexes. Concomitantly, the generation of reactive oxygen species was increased (consistent with previous reports that reactive oxygen species promote neuronal differentiation), as well as the production of the key metabolic intermediate acetyl-CoA, an essential determinant of epigenetic status in the cell. We suggest that lutein-stimulated neuronal differentiation is mediated by PI3K-dependent modulation of mitochondrial respiration and signaling, and that the consequential metabolic shifts initiate epigenetically dependent transcriptomic reprogramming in support of this morphogenesis. These observations support the potential importance of micronutrients supplementation to neurogenesis, both during normal development and in regenerative repair.

Prenatal influenza vaccination rescues impairments of social behavior and lamination in a mouse model of autism.

BACKGROUND: Prenatal infection is a substantial risk factor for neurodevelopmental disorders such as autism in offspring. We have previously reported that influenza vaccination (VAC) during early pregnancy contributes to neurogenesis and behavioral function in offspring. RESULTS: Here, we probe the efficacy of VAC pretreatment on autism-like behaviors in a lipopolysaccharide (LPS)-induced maternal immune activation (MIA) mouse model. We show that VAC improves abnormal fetal brain cytoarchitecture and lamination, an effect associated with promotion of intermediate progenitor cell differentiation in MIA fetal brain. These beneficial effects are sufficient to prevent social deficits in adult MIA offspring. Furthermore, whole-genome analysis suggests a strong interaction between Ikzf1 (IKAROS family zinc-finger 1) and neuronal differentiation. Intriguingly, VAC rescues excessive microglial Ikzf1 expression and attenuates microglial inflammatory responses in the MIA fetal brain. CONCLUSIONS: Our study implies that a preprocessed influenza vaccination prevents maternal bacterial infection from causing neocortical lamination impairments and autism-related behaviors in offspring.