Cell cycle-regulated expression of Fam72a from the |Srgap2–Fam72a| master gene leads to Mis18a downregulation

BackgroundIcariin (ICA), a major ingredient of Epimediumbrevicornum, has various pharmacological activities including central nervous system protective functions such as the improvement of learning and memory function in mice models of Alzheimer’s disease. It has been reported that ICA can promote regeneration of peripheral nerve and functional recovery. The purpose of this study was to investigate the potentiating effect of ICA on the proliferation of rat hippocampal neural stem cells, and explore the possible mechanism involved.MethodsPrimary neural stem cells were prepared from the hippocampus of newly born SD rats, and cells were cultured in special stem cell culture medium. Neural stem cells were confirmed by immunofluorescence detection of nestin, NSE and GFAP expression. The effect of ICA on the growth and proliferation of the neural stem cells was evaluated by 5-ethynyl-2-deoxyuridine (EdU) labeling of proliferating cells, and photomicrographic images of the cultured neural stem cells. Further, the mechanism of ICA-induced cell proliferation of neural stem cells was investigated by analyzing the gene and protein expression of cell cycle related genes cyclin D1 and p21.ResultsThe present study showed that icariin promotes the growth and proliferation of neural stem cells from rat hippocampus in a dose-dependent manner. Incubation of cells with icariin resulted in significant increase in the number of stem cell spheres as well as the increased incorporation of EdU when compared with cells exposed to control vehicle. In addition, it was found that icariin-induced effect on neural stem cells is associated with increased mRNA and protein expression of cell cycle genes cyclin D1 and p21.ConclusionsThis study evidently demonstrates the potentiating effect of ICA on neural stem cell growth and proliferation, which might be mediated through regulation of cell cycle gene and protein expression promoting cell cycle progression.

Patient-derived iPSC-cerebral organoid modeling of the 17q11.2 microdeletion syndrome establishes CRLF3 as a critical regulator of neurogenesis.

The eukaryotic expression vector containing full-length cDNA sequence of rate nerve growth factor (NGF) beta subunit was constructed and its effects on proliferation and differentiation of neural stem cells were observed. By using PCR, full-length cDNA sequence of NGF beta subunit in rats was cloned and ligated into the eukaryotic expression vector pEGFP-N1-NGF. The recombinant plasmid pEGFP-N1-NGF was transfected into the mesencephal neural stem cells of embryonic rats by Lipofectamin and transiently expressed. MTT method was used to determine the effects of NGF on proliferation of neural stem cells, and under phase-contrast microscopy, the effects of NGF on growth of nervous processes following differentiation of neural stem cells were observed. Sequence analysis indicated that the cloned full-length cDNA sequence of rat NGF beta was identical to that of published sequence encoding NGF in gene GeneBank. The transfection of recombinant plasmid pEGFP-N1-NGF into mesencephal neural stem cells of embryonic rats could obviously promote proliferation of neural stem cells and facilitate the growth of neural stem cells-derived nerve cells. It was suggested that neural stem cells could be used as a vehicle of gene transfer, and the expression of NGF beta subunit in the neural stem cells could promote the growth of nerve cells derived from neural stem cells.

Objective To investigate the effect of gene of phosphate and tension homology deleted on chromsome ten (PTEN) inhibitor bpV (pic) on the proliferation and differentiation of neural stem cells (NSCs). Methods Stem cell suspension was prepared from the cerebral cortex of the 16 days of fetal rat. The cells were divided into2 groups: group A (control group): no treatment, group B (experimental group): bpV treatment group (culture medium containing bpV). The number of stem cell spheres was observed by immunofluorescence staining, the proliferation of NSCs was detected by cell counting kit一8 (CCK-8) test, the differentiation of NSCs was detected by immunofluorescence staining, and the expression of PTEN and the mammalian target of rapamycin (mTOR) genes was detected by reverse transcriptase-polymerase chain reaction (RT-PCR) on the fifth day. Results In the experimental group, the number of stem cells per low field (×50) in the experimental group was 60±5, which was significantly higher than that in the control group (40±3, P 0.05). The expression of PTEN in experimental NSCs was 2.13 times higher than that in control NSCs (P<0.01), while the expression of mTOR in experimental NSCs was 3.62 times higher than that in control NSCs (P<0.01). Conclusion PTEN inhibitor bpV (pic) can promote the proliferation of neural stem cells and promote the differentiation of neural stem cells into neurons to a certain extent. Key words: Gene of phosphate and tension homology deleted on chromsome ten; Tumor suppressor gene; Neural stem cells; Proliferation; Differentiation

Neurogenesis involves generation of new neurons through finely tuned multistep processes, such as neural stem cell (NSC) proliferation, migration, differentiation, and integration into existing neuronal circuitry in the dentate gyrus of the hippocampus and subventricular zone. Adult hippocampal neurogenesis is involved in cognitive functions and altered in various neurodegenerative disorders, including Alzheimer disease (AD). Ethosuximide (ETH), an anticonvulsant drug is used for the treatment of epileptic seizures. However, the effects of ETH on adult hippocampal neurogenesis and the underlying cellular and molecular mechanism(s) are yet unexplored. Herein, we studied the effects of ETH on rat multipotent NSC proliferation and neuronal differentiation and adult hippocampal neurogenesis in an amyloid β (Aβ) toxin-induced rat model of AD-like phenotypes. ETH potently induced NSC proliferation and neuronal differentiation in the hippocampus-derived NSC in vitro. ETH enhanced NSC proliferation and neuronal differentiation and reduced Aβ toxin-mediated toxicity and neurodegeneration, leading to behavioral recovery in the rat AD model. ETH inhibited Aβ-mediated suppression of neurogenic and Akt/Wnt/β-catenin pathway gene expression in the hippocampus. ETH activated the PI3K·Akt and Wnt·β-catenin transduction pathways that are known to be involved in the regulation of neurogenesis. Inhibition of the PI3K·Akt and Wnt·β-catenin pathways effectively blocked the mitogenic and neurogenic effects of ETH. In silico molecular target prediction docking studies suggest that ETH interacts with Akt, Dkk-1, and GSK-3β. Our findings suggest that ETH stimulates NSC proliferation and differentiation in vitro and adult hippocampal neurogenesis via the PI3K·Akt and Wnt·β-catenin signaling.

JOURNAL/nrgr/04.03/01300535-202408000-00031/figure1/v/2023-12-16T180322Z/r/image-tiff Proliferation of neural stem cells is crucial for promoting neuronal regeneration and repairing cerebral infarction damage. Transcranial magnetic stimulation (TMS) has recently emerged as a tool for inducing endogenous neural stem cell regeneration, but its underlying mechanisms remain unclear. In this study, we found that repetitive TMS effectively promotes the proliferation of oxygen-glucose deprived neural stem cells. Additionally, repetitive TMS reduced the volume of cerebral infarction in a rat model of ischemic stroke caused by middle cerebral artery occlusion, improved rat cognitive function, and promoted the proliferation of neural stem cells in the ischemic penumbra. RNA-sequencing found that repetitive TMS activated the Wnt signaling pathway in the ischemic penumbra of rats with cerebral ischemia. Furthermore, PCR analysis revealed that repetitive TMS promoted AKT phosphorylation, leading to an increase in mRNA levels of cell cycle-related proteins such as Cdk2 and Cdk4. This effect was also associated with activation of the glycogen synthase kinase 3β/β-catenin signaling pathway, which ultimately promotes the proliferation of neural stem cells. Subsequently, we validated the effect of repetitive TMS on AKT phosphorylation. We found that repetitive TMS promoted Ca2+ influx into neural stem cells by activating the P2 calcium channel/calmodulin pathway, thereby promoting AKT phosphorylation and activating the glycogen synthase kinase 3β/β-catenin pathway. These findings indicate that repetitive TMS can promote the proliferation of endogenous neural stem cells through a Ca2+ influx-dependent phosphorylated AKT/glycogen synthase kinase 3β/β-catenin signaling pathway. This study has produced pioneering results on the intrinsic mechanism of repetitive TMS to promote neural function recovery after ischemic stroke. These results provide a strong scientific foundation for the clinical application of repetitive TMS. Moreover, repetitive TMS treatment may not only be an efficient and potential approach to support neurogenesis for further therapeutic applications, but also provide an effective platform for the expansion of neural stem cells.

The ginkgo biloba extract EGb761 improves memory loss and cognitive impairments in patients with senile dementia. It also promotes proliferation of neural stem cells in the subventricular zone in Parkinson's disease model mice and in the hippocampal zone of young epileptic rats. However, it remains unclear whether EGb761 enhances proliferation of endogenous neural stem cells in the brain of rats with vascular dementia. In this study, a vascular dementia model was established by repeatedly clipping and reperfusing the bilateral common carotid arteries of rats in combination with an intraperitoneal injection of a sodium nitroprusside solution. Seven days after establishing the model, rats were intragastrically given EGb761 at 50 mg/kg per day. Learning and memory abilities were assessed using the Morris water maze and proliferation of endogenous neural stem cells in the subventricular zone and dentate gyrus were labeled by 5-bromo-2-deoxyuridine immunofluorescence in all rats at 15 days, and 1, 2, and 4 months after model establishment. The escape latencies in Morris water maze tests of rats with vascular dementia after EGb761 treatment were significantly shorter than the model group. Immunofluorescence staining showed that the number and proliferation of 5-bromo-2-deoxyuridine-positive cells in the subventricular zone and dentate gyrus of the EGb761-treated group were significantly higher than in the model group. These experimental findings suggest that EGb761 enhances proliferation of neural stem cells in the subventricular zone and dentate gyrus, and significantly improves learning and memory in rats with vascular dementia.

Adult hippocampal neural stem cells can be activated by hippocampal neural activities. When focal cerebral ischemia, known as middle cerebral artery occlusion (MCAO), occurs, neural stem cells are activated to promote their proliferation. However, the mechanism by which these cells are activated is still unclear. Here, we indicate the involvement of metabotropic glutamate receptor 5 (mGluR5) signaling in neural stem cells in their activity-related proliferation after MCAO. We found mGluR5 molecules on neural stem cells by using calcium imaging. We detected the activation of neural stem cells by adding the mGluR5 agonist (RS)-2-chloro-5-hydroxyphenylglycine. On a hippocampal slice, the activation of neural stem cells to promote their proliferation was initiated by theta-burst electrical stimulation at the perforant pathway, and this activation was significantly blocked by an mGluR5 antagonist, 2-methyl-6-(phenylethynyl)pyridine (MPEP). In addition to this, the injection of the blood-brain barrier-permeable mGluR5 agonist 3-cyano-N-(1,3-diphenyl-1H-pyrazol-5-yl)benzamide into live mice promoted the proliferation of neural stem cells. Moreover, in vivo theta-burst electrical stimulation induced proliferation of neural stem cells. A chronic field recording study showed that the activity of the hippocampal formation was elevated after MCAO. Finally, we observed that the mGluR5 antagonist MPEP significantly blocked the stimulated proliferation of neural stem cells induced by MCAO, by blocking mGluR5 signaling. Our results suggest that glutamates released by the elevated neural activities after MCAO may trigger mGluR5 signaling in neural stem cells to promote their proliferation.

Objective To explore the effect of olfactory ensheathing cells (OECs) on the proliferation and differentiation of neural stem cells (NSCs). Methods The cells from the embryonic rat brain were primarily cultured and identified by immunofluorescence and immunocytochemistry. NSCs in the experimental group shared medium with OECs were cultured and induced to differentiate. Simultaneously, NSCs in the control group were cultured alone. The effect of OECs on the proliferation and differentiation of NSCs was observed by immunocytochemistry. Results Nerve growth factor receptor (P75NGFR) was observed in the primarily cultured OECs; nestin was expressed in the primarily cultured neurosphere and the cells differentiated from the neurosphere expressed neurofilament 200 (NF200) and glial fibrillary acidic protein (GFAP). Compared with that in the control group, the number of NSCs in the experimental group was significantly increased (P<0.05). On the 4~(th) and 7~(th) day of differentiation, the percentage of NF200-positive cell was higher in the experimental group than that in the control group (P<0.05), indicating that the appearance of OECs increased the differentiation of NSCs into NF200-positive cells. Conclusion OECs can promote the proliferation of NSCs and induce the differentiation of NSCs into neurons. Key words: Olfactory ensheathing cells; Neural stem cells; Cell proliferation; Cell differentiation

Objective To investigate the effects of functional electrical stimulation (FES) on endogenous proliferation of the neural stem cells within the brain and the behaviors in rats with acute cerebral infarction and explore the FES therapeutic mechanism on improving the neural function after the cerebral infarction. Methods Fifty-four SD adult male rats were randomly allocated into FES treatment group, placebo stimulation group and sham-operated group (n=18). Focal cerebral infarction models were induced by the performent of middle cerebral artery occlusion (MCAO) in rats; the FES treatment group began receiving the FES (10 min/d, once dairy) and the placebo stimulation group did not give any special treatment since the 3rd day of the successful model inducement. The expression of nestin positive cells in the hippocampus subgranular zone and subventricular zone was examined by immunohistochemistry staining and the expression of nestin protein at ischemia side was detected by Western blot analysis on the 3rd, 7th and 14th d after MCAO; meanwhile, the behavior functions of rats at various time points were evaluated. Results The number of nestin positive cells in the hippocampus subgranular zone and subventricular zone and the expression of nestin protein from ischemia side brain in the FES treatment group significantly increased than those in the placebo stimulation group on the 7th and 14th d after MCAO (P<0.05). And statistical difference was noted on the 14th d on the behavior functional evaluation between the FES treatment group and the placebo stimulation group (P<0.05). Conclusion FES may enhance the endogenous proliferation of the neural stem cells within the brain in rats with acute cerebral infarction and improve the behavior function in rats, which may be one of the mechanisms of FES on improving the neural function after cerebral infarction. Key words: Functional electrical stimulation; Neural stem cell; Cerebral infarction; Nestin

The present study comprised a series of experiments to investigate the mechanism underlying the effect of ginsenoside on the self-renewal, proliferation and differentiation of neural stem cells (NSCs) undergoing oxygen-glucose deprivation/reperfusion (OGD/R) in vitro. The NSCs, which were isolated from the hippocampus of embryonic day 17 embryo rats, were subjected to OGD/R to establish an in vitro model of brain ischemia-reperfusion, following which different doses of ginsenoside were administered to the model. The proliferation of the NSCs was determined using MTT colorimetry and nestin/bromodeoxyuridine (BrdU) immunofluorescent double-labeling. The NSCs were identified by measuring the expression of nestin, and the differentiation of NSCs was assessed through the immunofluorescent double-labeling of nestin/vimentin and nestin/neuron-specific class III β-tubulin (tuj-1). The protein levels of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1α (HIF-1α) were detected to investigate the function and mechanism of ginsenoside on ischemic stroke using an enzyme-linked immunosorbent assay. Marked increases in the optical density, area density and numbers of nestin/BrdU-, nestin/vimentin- and nestin/tuj-1-positive cells were found in the ginsenoside-treated group. Compared with the control group, enhanced expression levels of BrdU, tuj-1 and vimentin were found in the ginsenoside-treated group, suggesting that ginsenoside may significantly promote the proliferation and differentiation of NSCs. The results of the present study also showed that ginsenoside significantly increased the protein level of HIF-1α (P<0.05) in the NSCs exposed to OGD/R. These results indicated that ginsenoside may maintain NSC replication, promote NSC proliferation and promote NSC differentiation into neurons and astrocytes. Ginsenoside may initiate the expression of downstream VEGF, which is involved in promoting the survival, self-renewal and differentiation of NSCs.

The effect of the dosage of NT-3/chitosan carriers on the proliferation and differentiation of neural stem cells

CXCL1 promotes the proliferation of neural stem cells by stimulating the generation of reactive oxygen species in APP/PS1 mice

In many recent studies, the inhibitory transmitter gamma-aminobutyric acid has been shown to modulate the proliferation, differentiation and survival of neural stem cells. Most general anesthetics are partial or allosteric gamma-aminobutyric acid A receptor agonists, suggesting that general anesthetics could alter the behavior of neural stem cells. The neuroprotective efficacy of general anesthetics has been recognized for decades, but their effects on the proliferation of neural stem cells have received little attention. This study investigated the potential effect of midazolam, an extensively used general anesthetic and allosteric gamma-aminobutyric acid A receptor agonist, on the proliferation of neural stem cells in vitro and preliminarily explored the underlying mechanism. The proliferation of neural stem cells was tested using both Cell Counting Kit 8 and bromodeoxyuridine incorporation experiments. Cell distribution analysis was performed to describe changes in the cell cycle distribution in response to midazolam. Calcium imaging was employed to explore the molecular signaling pathways activated by midazolam. Midazolam (30–90 μM) decreased the proliferation of neural stem cells in vitro. Pretreatment with the gamma-aminobutyric acid A receptor antagonist bicuculline or Na-K-2Cl cotransport inhibitor furosemide partially rescued this inhibition. In addition, midazolam triggered a calcium influx into neural stem cells. The suppressive effect of midazolam on the proliferation of neural stem cells can be partly attributed to the activation of gamma-aminobutyric acid A receptor. The calcium influx triggered by midazolam may be a trigger factor leading to further downstream events.

SummaryThe consequences of DNA damage generation in mammalian somatic stem cells, including neural stem cells (NSCs), are poorly understood despite their potential relevance for tissue homeostasis. Here, we show that, following ionizing radiation-induced DNA damage, NSCs enter irreversible proliferative arrest with features of cellular senescence. This is characterized by increased cytokine secretion, loss of stem cell markers, and astrocytic differentiation. We demonstrate that BMP2 is necessary to induce expression of the astrocyte marker GFAP in irradiated NSCs via a noncanonical signaling pathway engaging JAK-STAT. This is promoted by ATM and antagonized by p53. Using a SOX2-Cre reporter mouse model for cell-lineage tracing, we demonstrate irradiation-induced NSC differentiation in vivo. Furthermore, glioblastoma assays reveal that irradiation therapy affects the tumorigenic potential of cancer stem cells by ablating self-renewal and inducing astroglial differentiation.