Neuronal Differentiation Rate Research Articles

Transplantation of Wnt5a-modified Bone Marrow Mesenchymal Stem Cells Promotes Recovery After Spinal Cord Injury via the PI3K/AKT Pathway.

Spinal cord injury (SCI) is a severe neurological condition that can lead to paralysis or even death. This study explored the potential benefits of bone marrow mesenchymal stem cell (BMSC) transplantation for repairing SCI. BMSCs also differentiate into astrocytes within damaged spinal cord tissues hindering the cell transplantation efficacy, therefore it is crucial to enhance their neuronal differentiation rate to facilitate spinal cord repair. Wnt5a, an upstream protein in the non-classical Wnt signaling pathway, has been implicated in stem cell migration, differentiation, and neurite formation but its role in the neuronal differentiation of BMSCs remains unclear. Thus, this study investigated the role and underlying mechanisms of Wnt5a in promoting neuronal differentiation of BMSCs both in vivo and in vitro. Wnt5a enhanced neuronal differentiation of BMSCs in vitro while reducing astrocyte differentiation. Additionally, high-throughput RNA sequencing revealed a correlation between Wnt5a and phosphoinositide 3-kinase (PI3K)/protein kinase B(AKT) signaling, which was confirmed by the use of the PI3K inhibitor LY294002 to reverse the effects of Wnt5a on BMSC neuronal differentiation. Furthermore, transplantation of Wnt5a-modified BMSCs into SCI rats effectively improved the histomorphology (Hematoxylin and eosin [H&E], Nissl and Luxol Fast Blue [LFB] staining), motor function scores (Footprint test and Basso-Beattie-Bresnahan [BBB]scores)and promoted neuron production, axonal formation, and remodeling of myelin sheaths (microtubule associated protein-2 [MAP-2], growth-associated protein 43 [GAP43], myelin basic protein [MBP]), while reducing astrocyte production (glial fibrillary acidic protein [GFAP]). Therefore, targeting the Wnt5a/PI3K/AKT pathway could enhance BMSC transplantation for SCI treatment.

IGF1c mimetic peptide-based supramolecular hydrogel microspheres synergize with neural stem cells to promote functional recovery from spinal cord injury

Spinal cord injuries (SCI) often cause severe inflammatory response and neuronal loss, which consequently deteriorates motor function. Although exogenous stem cells are considered to be vital seed cells for cell replacement therapy after SCI, their low survival rate and low efficiency of neuronal differentiation after in vivo transplantation limit their widespread application in clinical practice. To this end, this study proposed a new strategy for dramatically improving the efficiency of insulin-like growth factor 1 (IGF1c) mimetic peptide to activate its receptor by fabricating IGF1c mimetic peptide supramolecular hydrogel microspheres (IGF1c-PHM) using an innovatively designed piezoelectric ceramic-driven thermal electrospray device. IGF1c-PHM displayed an average size of 10 µm and could activate the IGF1R of neural stem cells (NSCs) more effectively than IGF1c peptide block hydrogel (IGF1c-PH). In vitro, IGF1c-PHM exerted strong neuroprotective effects by maintaining the proliferation, inhibiting the apoptosis of NSCs, and promoting the efficiency of differentiation into neurons in presence of myelin extracts. In vivo, IGF1c-PHM improved the inflammatory environment by suppressing the recruitment of the inflammatory cells, altering the subtypes of the macrophages, and promoting the survival rate of endogenous cells and NSCs in the injury area. In particular, IGF1c-PHM enhanced the proliferation ability and neuronal differentiation of NSCs in vivo after SCI. In the presence of IGF1c-PHM, the transplanted NSCs could differentiate into several mature neurons at the injury site, and the axons regenerated in the injury site were remyelinated, all of which closely correlate with the improvement of the electrophysiological indices after SCI. Finally, at 8 weeks post-SCI, the gait analysis and behavioral scores confirmed the significant improvement in motor function in rats treated with IGF1c-PHM and NSCs. Therefore, IGF1c-PHM combined with NSCs is a promising therapy for treating SCI.

Facet-Dependent Activity of CeO2 Nanozymes Regulate the Fate of Human Neural Progenitor Cell via Redox Homeostasis.

Neural progenitor cells (NPCs) therapy, a promising therapeutic strategy for neurodegenerative diseases, has a huge challenge to ensure high survival rate and neuronal differentiation rate. Cerium oxide (CeO2) nanoparticles exhibit multienzyme mimetic activities and have shown the capability of regulating reactive oxygen species (ROS), which is a pivotal mediator for intracellular redox homeostasis in NPCs, regulating biological processes including differentiation, proliferation, and apoptosis. In the present study, the role of facet-dependent CeO2-mediated redox homeostasis in regulating self-renewal and differentiation of NPCs is reported for the first time. The cube-, rod-, and octahedron-shaped CeO2 nanozymes with different facets are prepared. Among the mentioned nanozymes, the cube enclosed by the (100) facet exhibits the highest CAT-like activity, causing it to provide superior protection to NPCs from oxidative stress induced by H2O2; meanwhile, the octahedron enclosed by the (111) facet with the lowest CAT-like activity induces the most ROS production in ReNcell CX cells, which promotes neuronal differentiation by activated AKT/GSK-3β/β-catenin pathways. A further mechanistic study indicated that the electron density of the surface Ce atoms changed continuously with different crystal facets, which led to their different CAT-like activity and modulation of redox homeostasis in NPCs. Altogether, the different surface chemistry and atomic architecture of active sites on CeO2 exert modulation of redox homeostasis and the fate of NPCs.

Neural Stem Cell-Derived Exosomal Netrin1 Contributes to Neuron Differentiation of Mesenchymal Stem Cells in Therapy of Spinal Bifida Aperta.

Spinal bifida aperta (SBA) is a congenital malformation with a high incidence. Bone marrow mesenchymal stem cell (BMSC) transplantation has the potential to repair the structure of damaged tissues and restore their functions. This is an optional treatment that can be used as a supplement to surgery in the treatment of SBA. However, the application of BMSCs is limited, as the neuronal differentiation rate of BMSCs is not satisfactory when used in treating severe SBA. Thus, we aimed to assess the effect of neural stem cell (NSC)-derived exosomes on BMSC neuronal differentiation and observe the therapeutic effect in an ex vivo rat SBA embryo model. We found that NSC-derived exosomes increased the neuronal differentiation rate of BMSCs in vitro and in the SBA embryo model ex vivo. Proteomic analysis showed that NSC-derived exosomes were enriched in Netrin1, which positively regulated neuronal differentiation. Netrin1 increased the neuronal differentiation rate of BMSCs and NSCs and upregulated the expression of the neuronal markers, microtubule-associated protein (Map2), neurofilament, and β3-tubulin. Bioinformatic analysis revealed that Netrin1 treatment increased the expression of the transcription factors Hand2 and Phox2b, related to neuronal differentiation. Furthermore, the Netrin1-induced NSC neuronal differentiation was significantly blocked by Phox2b knockdown. We suggest that NSC-derived exosomal Netrin1 induces neuronal differentiation via the Hand2/Phox2b axis by upregulating the expression of Hand2 and Phox2b. Therefore, NSC-derived exosomes are a critical inducer of BMSC neuronal differentiation and represent a potential treatment agent that can benefit BMSC treatment in SBA.

Life Sciences Discovery and Technology Highlights

Life Sciences Discovery and Technology Highlights

Effects of primary microglia and astrocytes on neural stem cells in in vitro and in vivo models of ischemic stroke.

Transplantation of neural stem cells (NSCs) can protect neurons in animal stroke models; however, their low rates of survival and neuronal differentiation limit their clinical application. Glial niches, an important location of neural stem cells, regulate survival, proliferation and differentiation of neural stem cells. However, the effects of activated glial cells on neural stem cells remain unclear. In the present study, we explored the effects of activated astrocytes and microglia on neural stem cells in vitro stroke models. We also investigated the effects of combined transplantation of neural stem cells and glial cells after stroke in rats. In a Transwell co-culture system, primary cultured astrocytes, microglia or mixed glial cells were exposed to glutamate or H2O2 and then seeded in the upper inserts, while primary neural stem cells were seeded in the lower uncoated wells and cultured for 7 days. Our results showed that microglia were conducive to neurosphere formation and had no effects on apoptosis within neurospheres, while astrocytes and mixed glial cells were conducive to neurosphere differentiation and reduced apoptosis within neurospheres, regardless of their pretreatment. In contrast, microglia and astrocytes induced neuronal differentiation of neural stem cells in differentiation medium, regardless of their pretreatment, with an exception of astrocytes pretreated with H2O2. Rat models of ischemic stroke were established by occlusion of the middle cerebral artery. Three days later, 5 × 105 neural stem cells with microglia or astrocytes were injected into the right lateral ventricle. Neural stem cell/astrocyte-treated rats displayed better improvement of neurological deficits than neural stem cell only-treated rats at 4 days after cell transplantation. Moreover, neural stem cell/microglia-, and neural stem cell/astrocyte-treated rats showed a significant decrease in ischemic volume compared with neural stem cell-treated rats. These findings indicate that microglia and astrocytes exert different effects on neural stem cells, and that co-transplantation of neural stem cells and astrocytes is more conducive to the recovery of neurological impairment in rats with ischemic stroke. The study was approved by the Animal Ethics Committee of Tongji University School of Medicine, China (approval No. 2010-TJAA08220401) in 2010.

Combined Effects of Low Level Laser Therapy and Inducers on the Neural Differentiation of Mesenchymal Stem Cells

Low level laser therapy (LLLT) improves the therapeutic effectiveness of stem cell therapy for neurological injury through its ability to enhance stem differentiation and protect against neuronal apoptosis through its antioxidation effects. However, the specific mechanisms governing these effects are poorly defined. In this study, we investigated the effects of LLLT on stem cell differentiation at the molecular level, oxidative stress balance, and inflammatory factors to provide theoretical support for its clinical application. Cell viability was assessed via MTT assays. Reactive oxygen species (ROS), total superoxide dismutase (SOD), and total antioxidant capacity (TAC) were used to evaluate oxidative stress levels. Western blot analysis was used to quantitatively investigate protein expression. The levels of secreted proteins and tumor necrosis factor- $\alpha $ (TNF- $\alpha$ ) and interleukin- $1\beta $ (IL- $1\beta$ ) expression were measured by ELISA. Compared to LLLT at 808 nm, LLLT at 635 nm enhanced the proliferation of human umbilical cord-derived mesenchymal stem cells (hUCMSCs). The rates of proliferation markedly increased at a power density 20 mW/cm2. LLLT enhanced the antioxidant capacity and caused no inflammation in normal cells. Markers of neural precursors were more highly expressed at 808 nm when combined with inducers for 3 d, compared to the more modest increases observed at 635 nm. The expression of neuN on day 7 also increased, most notably when LLLT at 808 nm was combined with cerebrospinal fluid (CSF)/injured cerebrospinal fluid (iCSF). ELISA assays showed that LLLT at 808 nm with CSF/iCSF also increased the differentiation of hUCMSCs into neurons. LLLT at 808 nm combined with inducers promoted the differentiation into neurons and increased the rate of neuronal differentiation.

Transplantation of Wnt5a-modified NSCs promotes tissue repair and locomotor functional recovery after spinal cord injury

Traditional therapeutic strategies for spinal cord injury (SCI) are insufficient to repair locomotor function because of the failure of axonal reconnection and neuronal regeneration in the injured central nervous system (CNS). Neural stem cell (NSC) transplantation has been considered a potential strategy and is generally feasible for repairing the neural circuit after SCI; however, the most formidable problem is that the neuronal differentiation rate of NSCs is quite limited. Therefore, it is essential to induce the neuronal differentiation of NSCs and improve the differentiation rate of NSCs in spinal cord repair. Our results demonstrate that both Wnt5a and miRNA200b-3p could promote NSC differentiation into neurons and that Wnt5a upregulated miRNA200b-3p expression through MAPK/JNK signaling to promote NSC differentiation into neurons. Wnt5a could reduce RhoA expression by upregulating miRNA200b-3p expression to inhibit activation of the RhoA/Rock signaling pathway, which has been reported to suppress neuronal differentiation. Overexpression of RhoA abolished the neurogenic capacity of Wnt5a and miRNA200b-3p. In vivo, miRNA200b-3p was critical for Wnt5a-induced NSC differentiation into neurons to promote motor functional and histological recovery after SCI by suppressing RhoA/Rock signaling. These findings provide more insight into SCI and help with the identification of novel treatment strategies.

Exploring Sphingolipid Implications in Neurodegeneration.

Over the past decade, it was found that relatively simple sphingolipids, such as ceramide, sphingosine, sphingosine-1-phosphate, and glucosylceramide play important roles in neuronal functions by regulating rates of neuronal growth and differentiation. Homeostasis of membrane sphingolipids in neurons and myelin is essential to prevent the loss of synaptic plasticity, cell death and neurodegeneration. In our review we summarize data about significant brain cell alterations of sphingolipids in different neurodegenerative diseases such as Alzheimer's disease, Parkinson disease, Amyotrophic Lateral Sclerosis, Gaucher's, Farber's diseases, etc. We reported results obtained in brain tissue from both animals in which diseases were induced and humans in autopsy samples. Moreover, attention was paid on sphingolipids in biofluids, liquor and blood, from patients. In Alzheimer's disease sphingolipids are involved in the processing and aggregation of β-amyloid and in the transmission of the cytotoxic signal β-amyloid and TNFα-induced. Recently, the gangliosides metabolism in transgenic animals and the relationship between blood sphingolipids changes and cognitive impairment in Alzheimer's disease patients have been intensively studied. Numerous experiments have highlighted the involvement of ceramide and monohexosylceramide metabolism in the pathophysiology of the sporadic forms of Parkinson's disease. Moreover, gene mutations of the glucocerebrosidase enzyme were considered as responsible for Parkinson's disease via transition of the monomeric form of α-synuclein to an oligomeric, aggregated toxic form. Disturbances in the metabolism of ceramides were also associated with the appearance of Lewy's bodies. Changes in sphingolipid metabolism were found as a manifestation of Amyotrophic Lateral Sclerosis, both sporadic and family forms, and affected the rate of disease development. Currently, fingolimod (FTY720), a sphingosine-1-phosphate receptor modulator, is the only drug undergoing clinical trials of phase II safety for the treatment of Amyotrophic Lateral Sclerosis. The use of sphingolipids as new diagnostic markers and as targets for innovative therapeutic strategies in different neurodegenerative disorders has been included.

Toll-like receptor 4 differentially regulates adult hippocampal neurogenesis in an age- and sex-dependent manner.

Toll-like receptor 4 (TLR4) is primarily responsible for initiating an immune response following pathogen recognition. However, TLR4 is also expressed on neural progenitor cells and has been reported to regulate hippocampal neurogenesis as young male TLR4 knockout mice show increases in cell proliferation and doublecortin positive cells. Whether these effects occur in both sexes and are sustained with normal aging is currently unknown. The present study evaluated whether TLR4 deficiency alters adult hippocampal neurogenesis in young (3-4 months) and aged (18-20 months), male and female, TLR4 deficient (TLR4-/-; B6.B10ScN-Tlr4lps-del/JthJ) and wild type (WT) mice. Additionally, neurogenesis within the dorsal and the ventral hippocampal subdivisions was evaluated to determine if TLR4 has differential effects across the hippocampus. Bromodeoxyuridine (BrdU) was administered to quantify new cell survival as well as cell differentiation. Ki-67 was measured to evaluate cell proliferation. Results show that young TLR4-/- females had higher rates of proliferation and neuronal differentiation in both the dorsal and ventral hippocampus relative to WT females. Young TLR4-/- males show elevated proliferation and neuronal differentiation mainly in the ventral hippocampus. While young TLR4-/- mice show enhanced neurogenesis compared to young WT mice, the increase was not apparent in the aged TLR4-/- mice. Both aged WT and TLR4-/- mice showed a decrease in proliferation, new cell survival, and neuronal differentiation compared to young WT and TLR4-/- mice. The data collectively indicate that TLR4 regulates hippocampal neurogenesis in young adults, but that these effects are region-specific in males and that females show broader changes in neurogenesis throughout the hippocampus.

MiRNA-Based Rapid Differentiation of Purified Neurons from hPSCs Advancestowards Quick Screening for Neuronal Disease Phenotypes In Vitro.

Obtaining differentiated cells with high physiological functions by an efficient, but simple and rapid differentiation method is crucial for modeling neuronal diseases in vitro using human pluripotent stem cells (hPSCs). Currently, methods involving the transient expression of one or a couple of transcription factors have been established as techniques for inducing neuronal differentiation in a rapid, single step. It has also been reported that microRNAs can function as reprogramming effectors for directly reprogramming human dermal fibroblasts to neurons. In this study, we tested the effect of adding neuronal microRNAs, miRNA-9/9*, and miR-124 (miR-9/9*-124), for the neuronal induction method of hPSCs using Tet-On-driven expression of the Neurogenin2 gene (Ngn2), a proneural factor. While it has been established that Ngn2 can facilitate differentiation from pluripotent stem cells into neurons with high purity due to its neurogenic effect, a long or indefinite time is required for neuronal maturation with Ngn2 misexpression alone. With the present method, the cells maintained a high neuronal differentiation rate while exhibiting increased gene expression of neuronal maturation markers, spontaneous calcium oscillation, and high electrical activity with network bursts as assessed by a multipoint electrode system. Moreover, when applying this method to iPSCs from Alzheimer’s disease (AD) patients with presenilin-1 (PS1) or presenilin-2 (PS2) mutations, cellular phenotypes such as increased amount of extracellular secretion of amyloid β42, abnormal oxygen consumption, and increased reactive oxygen species in the cells were observed in a shorter culture period than those previously reported. Therefore, it is strongly anticipated that the induction method combining Ngn2 and miR-9/9*-124 will enable more rapid and simple screening for various types of neuronal disease phenotypes and promote drug discovery.

Draxin-mediated Regulation of Granule Cell Progenitor Differentiation in the Postnatal Hippocampal Dentate Gyrus

The hippocampus is characterized by the presence of life-long neurogenesis. To elucidate the molecular mechanism regulating hippocampal neurogenesis, we studied the functions of the chemorepellent Draxin in neuronal proliferation and differentiation in the postnatal dentate gyrus. The present in vivo cell labeling and fate tracking analyses revealed enhanced differentiation of hippocampal neural stem and progenitor cells (hNSPCs) in the subgranular zone (SGZ) of Draxin-deficient mice. We observed a reduction in the number of BrdU-pulse labeled or Ki-67 immunopositive SGZ cells in the mutant mice. However, Draxin deficiency did not affect cell cycle duration of SGZ cells. In situ hybridization analysis indicated that the receptor component of the canonical Wnt pathway, Lrp6, is expressed in SGZ cells, including Nestin and Sox2 double-positive hNSPCs. Taken together with the previous finding that Draxin interacts physically with Lrp6, we postulate that Draxin plays a pivotal role in the regulation of Wnt-driven hNSPC differentiation to modulate the rate of neuronal differentiation in the progenitor population.

Neural Stem Cells/Neuronal Precursor Cells and Postmitotic Neuroblasts in Constitutive Neurogenesis and After ,Traumatic Injury to the Mesencephalic Tegmentum of Juvenile Chum Salmon, Oncorhynchus keta.

The proliferation of neural stem cells (NSCs)/neuronal precursor cells (NPCs) and the occurrence of postmitotic neuroblasts in the mesencephalic tegmentum of intact juvenile chum salmon, Oncorhynchus keta, and at 3 days after a tegmental injury, were studied by immunohistochemical labeling. BrdU+ constitutive progenitor cells located both in the periventricular matrix zone and in deeper subventricular and parenchymal layers of the brain are revealed in the tegmentum of juvenile chum salmon. As a result of traumatic damage to the tegmentum, the proliferation of resident progenitor cells of the neuroepithelial type increases. Nestin-positive and vimentin-positive NPCs and granules located in the periventricular and subventricular matrix zones, as well as in the parenchymal regions of the tegmentum, are revealed in the mesencephalic tegmentum of juvenile chum salmon, which indicates a high level of constructive metabolism and constitutive neurogenesis. The expression of vimentin and nestin in the extracellular space, as well as additionally in the NSCs and NPCs of the neuroepithelial phenotype, which do not express nestin in the control animals, is enhanced during the traumatic process. As a result of the proliferation of such cells in the post-traumatic period, local Nes+ and Vim+ NPCs clusters are formed and become involved in the reparative response. Along with the primary traumatic lesion, which coincides with the injury zone, additional Nes+ and Vim+ secondary lesions are observed to form in the adjacent subventricular and parenchymal zones of the tegmentum. In the lateral tegmentum, the number of doublecortin-positive cells is higher compared to that in the medial tegmentum, which determines the different intensities and rates of neuronal differentiation in the sensory and motor regions of the tegmentum, respectively. In periventricular regions remote from the injury, the expression of doublecortin in single cells and their groups significantly increases compared to that in the damage zone.

Neuronal differentiation influences progenitor arrangement in the vertebrate neuroepithelium.

ABSTRACTCell division, movement and differentiation contribute to pattern formation in developing tissues. This is the case in the vertebrate neural tube, in which neurons differentiate in a characteristic pattern from a highly dynamic proliferating pseudostratified epithelium. To investigate how progenitor proliferation and differentiation affect cell arrangement and growth of the neural tube, we used experimental measurements to develop a mechanical model of the apical surface of the neuroepithelium that incorporates the effect of interkinetic nuclear movement and spatially varying rates of neuronal differentiation. Simulations predict that tissue growth and the shape of lineage-related clones of cells differ with the rate of differentiation. Growth is isotropic in regions of high differentiation, but dorsoventrally biased in regions of low differentiation. This is consistent with experimental observations. The absence of directional signalling in the simulations indicates that global mechanical constraints are sufficient to explain the observed differences in anisotropy. This provides insight into how the tissue growth rate affects cell dynamics and growth anisotropy and opens up possibilities to study the coupling between mechanics, pattern formation and growth in the neural tube.

Ascl1 Regulates Electric Field-Induced Neuronal Differentiation Through PI3K/Akt Pathway

Directing differentiation of neural stem/progenitor cells (NSCs/NPCs) to produce functional neurons is one of the greatest challenges in regenerative medicine. Our previous paper has confirmed that electrical stimulation has a high efficiency of triggering neuronal differentiation by using isolated filum terminale (FT)-derived NPCs. To further clarify the intrinsic molecular mechanisms, protein–protein interaction (PPI) network analysis was applied to pinpoints novel hubs in electric field (EF)-induced neuronal differentiation. In this study, siRNA transfection of Achaete-scute homolog 1 (Ascl1) in NPCs or NPCs was followed by direct current stimulation at 150 mV/mm. Neuronal differentiation rate and protein expression level were analyzed after 7 or 14 days of electrical stimulation. The data showed that the expression level of Ascl1 was enhanced by electrical stimulation and positively correlated to EF strength. Moreover, we identified that the expression of Ascl1 positively regulated neuronal differentiation of NPCs and can be up-regulated by EF-stimulation through the activation of phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) pathway. Therefore, this study provides new insights into the role of Ascl1 and its relevant PI3K/Akt pathway in regulating of EF-induced neuronal differentiation and pointed out that continuous expression of Ascl1 in NPCs is required for EF-induced neuronal differentiation.

Ferulic Acid exerts concentration-dependent anti-apoptotic and neuronal differentiation-inducing effects in PC12 and mouse neural stem cells

Ferulic Acid (FA) is a phenolic compound with anti-apoptotic and anti-oxidative properties. There are reports regarding its neuro-protective, neuro-proliferative and neuro-differentiative effects. However, effect of FA on neuronal differentiation and its effective neuro-protective and neuro-differentiative concentrations are unknown. Also the role of sirtuin molecules in neuroprotective effects of FA were not reported. We used PC12 and mouse neural stem cells (mNSCs) in our experiments. Intact and apoptotic (H2O2-exposed) cells were treated with different concentrations of FA, and then they were evaluated by MTT, quantitative real-time RT-PCR and immunostaining assays. FA treatment at low concentrations (50 µg/ml) significantly reduced apoptosis in H2O2-treated PC12 cells. Real-time RT-PCR and western blot assays confirmed that FA induced this effect through stabilization and degradation of P53 by increasing the expression rate of SIRT1, SIRT7 and MDM2 but down-regulation of USP7. Beside this anti-apoptotic effect, treatments of PC12 cells and mNSCs with higher concentrations of FA (250–800 µg/ml on PC12 cells and 100–500 µg/ml on mNSCs) increased the rate of neuronal differentiation. Immunocytochemical staining for β-tubulin III and Map2 verified the presence of mature neurons, and western blot assay showed that FA-treated PC12 cells had a stepwise rise of phosphorylated-ERK1/2 with increasing concentrations of FA. Our findings showed that FA at low concentrations has neuroprotective effect through up-regulation of SIRT1, SIRT7 and MDM2, and at higher concentrations can promote neural differentiation and neurite outgrowth.

Maternal Folic Acid Supplementation During Pregnancy Promotes Neurogenesis and Synaptogenesis in Neonatal Rat Offspring.

Maternal folic acid supplementation during pregnancy is associated with improved cognitive performances in offspring. However, the effect of supplementation on offspring's neurogenesis and synaptogenesis is unknown, and whether supplementation should be continued throughout pregnancy is controversial. In present study, 3 groups of female rats were fed a folate-normal diet, folate-deficient diet, or folate-supplemented diet from 1 week before mating until the end of pregnancy. A fourth group fed folate-normal diet from 1 week before mating until mating, then fed folate-supplemented diet for 10 consecutive days, then fed folate-normal diet until the end of pregnancy. Offspring were sacrificed on postnatal day 0 for measurement of neurogenesis and synaptogenesis by immunofluorescence and western blot. Additionally neural stem cells (NSCs) were cultured from offspring's hippocampus for immunocytochemical measurement of their rates of proliferation and neuronal differentiation. The results demonstrated that maternal folic acid supplementation stimulated hippocampal neurogenesis by increasing proliferation and neuronal differentiation of NSCs, and also enhanced synaptogenesis in cerebral cortex of neonatal offspring. Hippocampal neurogenesis was stimulated more when supplementation was continued throughout pregnancy instead of being limited to the periconceptional period. In conclusion, maternal folic acid supplementation, especially if continued throughout pregnancy, improves neurogenesis and synaptogenesis in neonatal offspring.

Mouse maternal protein restriction during preimplantation alone permanently alters brain neuron proportion and adult short-term memory

Maternal protein malnutrition throughout pregnancy and lactation compromises brain development in late gestation and after birth, affecting structural, biochemical, and pathway dynamics with lasting consequences for motor and cognitive function. However, the importance of nutrition during the preimplantation period for brain development is unknown. We have previously shown that maternal low-protein diet (LPD) confined to the preimplantation period (Emb-LPD) in mice, with normal nutrition thereafter, is sufficient to induce cardiometabolic and locomotory behavioral abnormalities in adult offspring. Here, using a range of in vivo and in vitro techniques, we report that Emb-LPD and sustained LPD reduce neural stem cell (NSC) and progenitor cell numbers at E12.5, E14.5, and E17.5 through suppressed proliferation rates in both ganglionic eminences and cortex of the fetal brain. Moreover, Emb-LPD causes remaining NSCs to up-regulate the neuronal differentiation rate beyond control levels, whereas in LPD, apoptosis increases to possibly temper neuron formation. Furthermore, Emb-LPD adult offspring maintain the increase in neuron proportion in the cortex, display increased cortex thickness, and exhibit short-term memory deficit analyzed by the novel-object recognition assay. Last, we identify altered expression of fragile X family genes as a potential molecular mechanism for adverse programming of brain development. Collectively, these data demonstrate that poor maternal nutrition from conception is sufficient to cause abnormal brain development and adult memory loss.

МОЛЕКУЛЯРНЫЕ МЕХАНИЗМЫ ДИФФЕРЕНЦИРОВКИ НЕЙРОНАЛЬНЫХ СТВОЛОВЫХ КЛЕТОК КРЫС ЛИНИИ КРУШИНСКОГО-МОЛОДКИНОЙ

It is well known that epileptogenesis is accompanied with the aberrations in the hippocampal neurogenesis in human as well as in epileptic animals. The Krushinsky--Molodkina (KM) rats genetically prone to audiogenic seizure were recruited in the experiments. Neuronal stem cells (NSC) isolated from the hippocampus of KM rats at the 14--17 days of postnatal development, which corresponds the final stage for the development of the hippocampal neurogenic niche. NSC isolated from the hippocampus of Wistar rats were used as a control. The cells were incubated for 10 days in a medium supplemented with retinoic acid to stimulate neural differentiation. Previously we demonstrated that proliferation level of NSC KM was significantly lower in comparison with the NSC of Wistar rats, but neuronal differentiation rate of NSC KM was significantly higher. This research unravels the underlying molecular mechanisms. Analysis of Akt/GSK3b pathway demonstrated decreased expression of Akt that correlated with attenuation of phosphorylation level of GSK3b at Ser9 and decreased level of b-catenin. It is known that transcriptional factor CREB is one of the main substrates for Akt and on the other hand CREB tightly participates in the regulation of the cell proliferation. Obtained data revealed decreased activity of CREB in NSC KM in comparison to NSC Wistar that could mediate inhibition of cell cycle and potentiation the differentiation. Thus our results demonstrated that glutamatergic differentiation of NSC KM is mediated by AKT/GSK3b/b-catenin/CREB signaling pathway.

Effect of brain traumatic tissue extract on the survival and differentiation of neural stem cells

Objective To simulate the chemical microenvironment after traumatic brain injury (TBI) and to investigate the effect of this microenvironment on the survival and differentiation of neural stem cells (NSCs). Methods The brain tissue homogenate of TBI rat model was harvested to simulate the chemical microenvironment after TBI. The primary NSCs of rat model were isolated and extraction, and then identified the phenotype characteristics with immunofluorescence staining. The experiments were divided into control group, normal brain tissue extract group (BTE group) and traumatic brain injury tissue extract group (TBITE group). The cell growth and morphological changes of each group were observed dynamically. The expression of apoptosis related protein, which includes Bax, Bcl-2, caspase-3 and cleaved caspase-3, were detected by Western Blot 24 h after experiments. The proliferation of NSCs was detected by MTT assay and Western Blot after 3 days. The differentiation level of NSCs to neurons was detected by immunofluorescence staining after 7 days. Results The results of Western Blot showed that compared with the control group, there was no significant change of apoptosis in the BTE group, while the apoptosis in the BTE group was significantly increased, showed a increase of expression levels of Bax (F=18.06, P<0.01) and Cleaved caspase-3 (F=23.86, P<0.01), and a decrease of that of Bcl-2 (F=22.95, P<0.01). The results of MTT assay showed that compared with the BTE group, the proliferation of NSCs in the TBITE group was decreased (F=41.99, P<0.01). The immunofluorescence staining showed that compared with the control group, the neuronal differentiation rate was increased in the BTE group. Further, compared with the BTE group, the neuronal differentiation rate in the TBITE group was decreased (F=66.93, P<0.01). Conclusion The injury microenvironment after TBI can significantly inhibit the survival and differentiation of NSCs, which provides a theoretical basis for clarifying the mechanism of endogenous nerve regeneration after TBI. Key words: Traumatic brain injury; Chemical microenvironment; Neural stem cells; Brain tissue extract