MicroRNA (miRNA) clusters, defined as genomically co-localized miRNAs regulated by a shared promoter and processed from polycistronic transcripts, exhibit synergistic regulatory roles in developmental processes. Among these, the evolutionarily conserved miR-125/let-7 cluster has been identified as a key regulator of neural stem cell (NSC) dynamics. In this study, we used Dicer conditional knockout (cKO) mice to confirm the essential role of miRNAs in mouse neocortical layer formation. The miR-125/let-7 cluster is co-expressed in mice and shows significant enrichment in upper-layer (UL) neurons. Using in utero electroporation (IUE), we found that miR-125b or let-7b overexpression partially rescues cortical phenotypes in Dicer-deficient mice, restoring proper UL organization but failing to rescue laminar fate defects in deep-layer cortical neurons. Our findings demonstrate that the miR-125b/let-7b exhibits a specialized function in regulating UL neuronal fate specification in mice and promotes the differentiation of NSC. Notably, miR-125b and let-7b exhibit both overlapping and distinct regulatory functions. Collectively, these results underscore the cooperative mechanisms by which miRNA clusters orchestrate cortical development.

Hypothalamic neurogenesis is a complex process that plays a crucial role in neuroendocrine homeostasis, making in vivo studies of the hypothalamus particularly challenging. In this study, we present an optimized protocol for isolating and culturing hypothalamic neural stem cells (htNSCs) from neonatal (P1) mice, followed by their directed differentiation in a three-dimensional (3D) Matrigel environment. We successfully established a primary culture system that supports the stability, growth, and distinct characteristics of htNSCs. Notably, we demonstrate that htNSCs can differentiate into GnRH-like neurons within the Matrigel-based 3D culture system. These differentiated neurons exhibit typical neuronal morphology and functional characteristics. Our findings highlight the potential of neonatal htNSCs as an invaluable model for studying hypothalamic function and neurogenesis. Furthermore, this method provides a novel platform for basic research and may serve as important implications for further studying the pathological mechanism of neuroendocrine disorders in hypothalamus.

During cortical development, neural stem/precursor cells (NS/PCs) sequentially produce neurons, astrocytes, and oligodendrocytes. Before producing these cells, human (h) NS/PCs undergo prolonged self-renewal to form a larger cortex than other mammals, although the mechanisms are mostly unknown. Here, we performed a gene knockout screen using the CRISPR/Cas9 system to search for genes involved in hNS/PC self-renewal. We identified RMND5A, encoding an E3 ubiquitin ligase, among the candidate genes. We further demonstrated that knockdown of RMND5A decreased proliferation and promoted neuronal differentiation of hNS/PCs through the activation and suppression of the Wnt and mTOR signaling pathways, respectively. Taken together, our findings suggest that RMND5A participates in the maintenance of hNS/PC self-renewal by modulating the Wnt and mTOR signaling pathways. Impact statement During cortical development, human neural stem/precursor cells (hNS/PCs) undergo prolonged self-renewal to form a larger cortex than other mammals, although the mechanisms are mostly unknown. We identified RMND5A, an E3 ubiquitin ligase, as essential for maintaining self-renewal of hNS/PCs, providing valuable insights into the evolutionary expansion of the human brain.

Aging-related cognitive decline is closely linked to the reduced function of neural progenitor/stem cells (NPSCs), which can be influenced by the neural microenvironment, particularly astrocytes. The aim of this study was to explore how astrocytes affect NPSCs and cognitive function during aging.

H2O2-treated astrocytes were used to mimic the aging phenotype of astrocytes. Proteomic analysis identified altered protein expression, revealing high levels of colony-stimulating factor-1 (CSF-1) in the supernatant of H2O2-treated astrocytes. Primary NPSCs were isolated and cultured in vitro, then stimulated with varying concentrations of recombinant CSF-1 protein to assess its effects on NPSC proliferation, differentiation, and apoptosis. Transcriptome sequencing identified miR-484 related to CSF-1 in H2O2-treated astrocytes, and a dual-luciferase assay verified the interaction between miR-484 and CSF-1. The impact of miR-484 overexpression on NPSC function and cognitive restoration was evaluated both in vitro and in vivo (in 20-month-old rats).

High concentration of CSF-1 inhibited the NPSC proliferation and differentiation into neurons while inducing apoptosis. Overexpression of miR-484 downregulated CSF-1 expression by binding to its 3' untranslated region, thereby promoting the NPSC proliferation and differentiation into neurons. In 20-month-old rats, miR-484 overexpression improved spatial learning and memory in the Morris water maze, increased NPSC proliferation, and reduced apoptosis.

Our findings reveal that miR-484 regulates CSF-1 to influence NPSC proliferation, differentiation into neurons, and apoptosis, consequently improving cognitive function in 20-month-old rats. This study provides a foundation for developing therapeutic strategies targeting age-related hippocampal cognitive impairments.

Glioblastoma is the commonest and deadliest primary brain tumor. Glioblastoma is characterized by significant intra- and inter-tumoral heterogeneity, resistance to treatment and dismal prognoses despite decades of research in understanding its biological underpinnings. Encompassed within this heterogeneity and therapy resistance are severely dysregulated programmed cell death pathways. Glioblastomas recapitulate many neurodevelopmental and neural injury responses; in addition, glioblastoma cells are composed of multiple different transformed versions of CNS cell types. To obtain a greater understanding of the features underlying cell death regulation in glioblastoma, it is important to understand the control of cell death within the healthy CNS during homeostatic and neurodegenerative conditions. Herein, we review apoptotic control within neural stem cells, astrocytes, oligodendrocytes and neurons and compare them to glioblastoma apoptotic control. Specific focus is paid to the Inhibitor of Apoptosis proteins, which play key roles in neuroinflammation, CNS cell survival and gliomagenesis. This review will help in understanding glioblastoma as a transformed version of a heterogeneous organ composed of multiple varied cell types performing different functions and possessing different means of apoptotic control. Further, this review will help in developing more glioblastoma-specific treatment approaches and will better inform treatments looking at more direct brain delivery of therapeutic agents.

In the central nervous system (CNS), cystathionine β-synthase (CBS) is localized in astrocytes. CBS degrades cytotoxic homocysteine and produces cytoprotective hydrogen sulfide; thus the proper expression of CBS is required to maintain CNS functions. CBS expression is very low at the late embryonic stage and increases after birth. This study examined CBS expression in cultured spinal cord cells derived from fetal rats. Treatment of spinal cord cells with epidermal growth factor (EGF) promoted the proliferation and maturation of astrocytes during development. EGF (30 ng/ml, 4 days) increased CBS protein expression and the number of CBS-expressing astrocytes in the culture. A high cell density also increased CBS expression, and EGF was able to increase CBS expression when cellular proliferation was inhibited. The EGF receptor was predominately expressed in neural stem cells rather than astrocytes. These results suggest that EGF acts on neural stem cells, leading to increase in CBS-expressing astrocytes. This effect may reflect the maturation process of astrocytes during embryonic development.

The Drosophila TRIM-NHL RNA-binding protein (RBP), MEI-P26, has previously been shown to suppress tumor formation in the germline. Here we show that, in the Drosophila larval central brain, cell-type-specific expression of MEI-P26 plays a vital role in regulating neural development. MEI-P26 and another TRIM-NHL RBP, Brain tumor (BRAT), have distinct expression patterns in Type I neuroblast (NB) lineages: While both proteins are expressed in NBs, BRAT is expressed in ganglion mother cells (GMCs) but not neurons, whereas MEI-P26 is expressed in neurons but not GMCs. Knockdown of MEI-P26 leads to re-expression of the stem cell marker Deadpan (DPN) and over-production of neurons. In contrast, ectopically expressed MEI-P26 reduces NB lineage size by repressing division of GMCs, resulting in reduced neuron production. We show that MEI-P26 positively regulates expression of Prospero (PROS), a transcription factor that is known to repress cell cycle-related genes. Ectopic expression of PROS phenocopies ectopic expression of MEI-P26. In both cases, Cyclin B (CYCB) expression is downregulated. Importantly, knockdown of PROS in the context of ectopic MEI-P26 rescues the neural lineage. Based on these results, we conclude that MEI-P26 functions to prevent over-production of neurons by promoting production of PROS which, in turn, downregulates cell division.

Female Alzheimer's disease (AD) patients display greater cognitive deficits and worse AD pathology as compared to male AD patients. In this study, we found that conditional knockout (cKO) of Atg5 in female microglia failed to obtain disease-associated microglia (DAM) gene signatures in familiar AD mouse model (5xFAD). Next, we analyzed the maintenance and neurogenesis of neural stem cells (NSCs) in the hippocampus and subventricular zone (SVZ) from 5xFAD mice with Atg5 cKO. Our data indicated that Atg5 cKO reduced the NSC number in hippocampus of female but not male 5xFAD mice. However, in the SVZ, Atg5 cKO only impaired NSCs in male 5xFAD mice. Interestingly, female 5xFAD;Fip200 cKO mice and 5xFAD;Atg14 cKO mice did not show NSC defects. These autophagy genes cKO 5xFAD mice exhibited a higher neurogenesis activity in their SVZ. Together, our data indicate a sex-specific role for microglial Atg5 in postnatal neurogenesis in AD mice.

Human genetic variants of the orphan G protein-coupled receptor GPR50 are suggested risk factors for neuropsychiatric disorders. However, the function of GPR50 in the central nervous system (CNS) and its link to CNS disorders remain poorly defined. Here, we generated GPR50 knockout (GPR50-KO) mice and show that the absence of GPR50 increases neurite outgrowth, cell motility and migration of isolated neural progenitor cells (NPCs) and hypothalamic radial glial cells (tanycytes). These observations were phenocopied in NPCs and tanycytes from wild-type mice treated with neutralizing antibodies the against the prototypical neurite growth inhibitor Nogo-A. Treatment of NPCs and tanycytes from GPR50-KO cells with neutralizing antibodies had no further, additive, effect. Inhibition of neurite growth by GPR50 occurs through activation of the G12/13 protein-RhoA pathway in a manner similar to, but independent of Nogo-A and its receptors. Collectively, we show that GPR50 acts as an inhibitor of neurite growth and cell migration in the brain by activating the G12/13 protein-RhoA pathway.

This review highlights the role of miRNAs in various diseases affecting major organ systems. miRNAs are small, non-coding RNA molecules that regulate numerous genes. Dysregulation of miRNAs is linked to many pathological conditions due to their involvement in gene silencing and cellular pathways. We discuss miRNA expression patterns, their physiological and pathological roles, and how changes in miRNA levels contribute to disease. Notably, miRNAs like miR-499 and miR-21 are implicated in heart failure and atherosclerosis. miRNA dysregulation is also associated with colorectal and gastric cancers, influencing tumorigenesis and chemoresistance. In neurological diseases, miRNAs exhibit diverse profiles that affect neurodevelopment and degeneration. Additionally, miRNAs modulate cell function in reproductive organs, impacting fertility and cancer progression. miRNAs such as miR-192 and miR-204 serve as biomarkers for nephropathy and acute kidney injury. These miRNAs are involved in skeletal muscle diseases, contributing to conditions like osteoporosis and sarcopenia. miRNAs function as oncogenes or tumor suppressors in cancer, highlighting their potential in diagnostics and therapy. Further research is needed to develop miRNA-based diagnostics and treatments.

Nitric oxide (NO) is an essential inorganic signaling molecule produced by constitutive NO synthase (cNOS) in the neurological system. Under pathological conditions, NO rapidly reacts with superoxide (O2•-) to generate peroxynitrite (ONOO¯). Elevated ONOO¯ concentrations induce nitroxidative stress, potentially contributing to numerous pathological processes as observed in neurodegenerative diseases including Alzheimer's disease (AD). Metalloporphyrin nanosensors, (200-300 nm diameter), were applied to quantify the NO/ONOO¯ balance produced by a single human neural progenitor cell (hNPC), in situ. These nanosensors, positioned in proximity of 4-5 ± 1 μm from the hNPCs membrane, enabled real-time measurement of NO and ONOO¯ concentrations following calcium ionophore (CaI) stimulation. The ratio of NO to ONOO¯ concentration ([NO]/[ONOO¯]) was established for the purpose of quantifying nitroxidative stress levels. Normal hNPCs produced a maximum of 107 ± 1 nmol/L of NO and 451 ± 7 nmol/L of ONOO¯, yielding a [NO]/[ONOO¯] ratio of 0.25 ± 0.005. In contrast, the model of the dysfunctional hNPCs, for long-term (48 h) amyloid-beta 42 (Aβ42) exposure significantly altered NO/ONOO¯ production. The NO level decreased to 14 ± 0.1 nmol/L, while ONOO¯ increased to 843 ± 0.8 nmol/L, resulting in a 94 % reduction of the [NO]/[ONOO¯] ratio to 0.016 ± 0.0001. The [NO]/[ONOO¯] ratio is determined by this work as a possible biomarker of nNOS efficiency and hNPC dysfunction, with implications for neurodegenerative disorders such as AD. Promising applications in the early medical diagnosis of neurological illnesses, electrochemical metalloporphyrin nanosensors demonstrate efficacy in real-time nitroxidative stress monitoring.

Neurosphere culture is widely used to expand neural stem and progenitor cells (NSPCs) of the nervous system. Understanding the identity of NSPCs, such as the principals involved in spatiotemporal patterning, will improve our chances of using NSPCs for neurodevelopmental and brain repair studies with the ability to direct NSPCs toward distinct fates. Some reports indicate that aging can affect the nature of NSPCs over time. Therefore, in this study, we aimed to investigate how the initial neural patterning of developing NSPCs changes over time.

In this research, evidence of changing neural patterning potential in the nervous system over time was presented. Thus, the embryonic and adult-derived NSPCs for cardinal characteristics were analyzed, and then, the expression of candidate genes related to neural patterning using real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) was evaluated at various stages of embryonic (E14 and E18), neonatal, and adult brains. Finally, it was assessed the effect of cell attachment and passage on the initial neural patterning of NSPCs.

The analysis of gene expression revealed that although temporal patterning is maintained in vitro, it shows a decrease over time. Embryonic NSPCs exhibited the highest potential for retaining regional identity than neonatal and adult NSPCs. Additionally, it was found that culture conditions, such as cell passaging and attachment status, could affect the initial neural patterning potential, resulting in a decrease over time.

Our study demonstrates that patterning potential decreases over time and aging imposes restrictions on preliminary neural patterning. These results emphasize the significance of patterning in the nervous system and the close relationship between patterning and fate determination, raising questions about the application of aged NSPCs in the treatment of neurodegenerative diseases.

This research introduces innovative tools for studying embryonic neurogenesis in the gecko Paroedura picta. Traditional research methods have been adapted for the gecko's unique biology, including variations of birthdating techniques and the implementation of EdU for tracking neuron generation. We also employ hybridization chain reaction (HCR) to detect specific mRNAs without the need for species-specific antibodies, providing a powerful and flexible tool for studying gene expression patterns in the gecko embryonic brain. However, HCR can also be combined with immunohistochemistry to visualize both RNA and protein distribution. Our work proves the importance of this innovative toolset to understand the evolutionary aspects of nervous system development and the gecko as a valuable model for understanding the ancestral stem amniote.

Neurological diseases are expected to become the leading cause of death in the next decade. Although little is known about it, the interaction between oxidative stress and inflammation is harmful to the nervous system. To find an advanced tool for neural genetics, mouse haploid neural stem cells (haNSCs) from the somite of chimeric mouse embryos at E8.5 is established. The haNSCs present a haploid neural progenitor identity for long-term culture, promising to robustly differentiate into neural subtypes and being able to form cerebral organoids efficiently. Thereafter, haNSC mutants via a high-throughput approach and screened targets of oxidative stress is generated using the specific mutant library. Deletion of Nfkbia (the top hit among the insertion mutants) reduces damage from reactive oxygen species (ROS) in NSCs exposed to H2O2. Transcriptome analysis revealed that Atp2b4 is upregulated significantly in Nfkbia-null NSCs and is probably responsible for the observed resistance. Additionally, overexpression of Atp2b4 itself can increase the survival of NSCs in the presence of H2O2, suggesting that Atp2b4 is closely involved in this resistance. Herein, a powerful haploid system is presented to study functional genetics in neural lineages, shedding light on the screening of critical genes and drugs for neurological diseases.

Neural stem cells (NSCs) reside in discrete regions of the adult mammalian brain where they can differentiate into neurons, astrocytes, and oligodendrocytes. Several studies suggest that mitochondria have a major role in regulating NSC fate. Here, we evaluated mitochondrial properties throughout NSC differentiation and in lineage-specific cells. For this, we used the neurosphere assay model to isolate, expand, and differentiate mouse subventricular zone postnatal NSCs. We found that the levels of proteins involved in mitochondrial fusion (Mitofusin [Mfn] 1 and Mfn 2) increased, whereas proteins involved in fission (dynamin-related protein 1 [DRP1]) decreased along differentiation. Importantly, changes in mitochondrial dynamics correlated with distinct patterns of mitochondrial morphology in each lineage. Particularly, we found that the number of branched and unbranched mitochondria increased during astroglial and neuronal differentiation, whereas the area occupied by mitochondrial structures significantly reduced with oligodendrocyte maturation. In addition, comparing the three lineages, neurons revealed to be the most energetically flexible, whereas astrocytes presented the highest ATP content. Our work identified putative mitochondrial targets to enhance lineage-directed differentiation of mouse subventricular zone-derived NSCs.

Nitric oxide (NO) is an inorganic signaling molecule that plays a crucial role in the regulation of numerous physiological functions. An oxidation product of the cytoprotective NO is cytotoxic peroxynitrite (ONOO-). In biological systems, the concentrations of NO and ONOO- are typically transient, ranging from nanomolar to micromolar, and these increases are normally followed by a swift return to their basal levels due to their short life spans. To understand the vital physiological role of NO and ONOO- in vitro and in vivo, sensitive and selective methods are necessary for direct and continuous NO and ONOO- measurements in real time. Because electrochemical methods can be adjusted for selectivity, sensitivity, and biocompatibility in demanding biological environments, they are suitable for real-time monitoring of NO and ONOO- release. Metalloporphyrin nanosensors, described here, have been designed to measure the concentration of NO and ONOO- produced by a single human neural progenitor cell (hNPC) in real time. These nanosensors (200-300 nm in diameter) can be positioned accurately in the proximity of 4-5 ± 1 μm from an hNPC membrane. The response time of the sensors is better than a millisecond, while detection limits for NO and ONOO- are 1 × 10-9 and 3 × 10-9 mol/L, respectively, with a linear concentration response of up to about 1 μM. The application of these metalloporphyrin nanosensors for the efficient measurement of the concentrations of NO and ONOO- in hNPCs is demonstrated, providing an opportunity to observe in real time the molecular changes of the two signaling molecules in situ.

Neuronal differentiation-the development of neurons from neural stem cells-involves neurite outgrowth and is a key process during the development and regeneration of neural functions. In addition to various chemical signaling mechanisms, it has been suggested that thermal stimuli induce neuronal differentiation. However, the function of physiological subcellular thermogenesis during neuronal differentiation remains unknown. Here we create methods to manipulate and observe local intracellular temperature, and investigate the effects of noninvasive temperature changes on neuronal differentiation using neuron-like PC12 cells. Using quantitative heating with an infrared laser, we find an increase in local temperature (especially in the nucleus) facilitates neurite outgrowth. Intracellular thermometry reveals that neuronal differentiation is accompanied by intracellular thermogenesis associated with transcription and translation. Suppression of intracellular temperature increase during neuronal differentiation inhibits neurite outgrowth. Furthermore, spontaneous intracellular temperature elevation is involved in neurite outgrowth of primary mouse cortical neurons. These results offer a model for understanding neuronal differentiation induced by intracellular thermal signaling.

Mouse postnatal neural stem cells (pNSCs) can be expanded in vitro in the presence of epidermal growth factor and fibroblast growth factor 2 and upon removal of these factors cease proliferation and generate neurons, astrocytes, and oligodendrocytes. The genetic requirements for self-renewal and lineage-commitment of pNSCs are incompletely understood. In this study, we show that the transcription factors NFIA and NFIB, previously shown individually, to be essential for the normal commitment of pNSCs to the astrocytic lineage in vivo, are jointly required for normal self-renewal of pNSCs in vitro and in vivo. Using conditional knockout alleles of Nfia and Nfib, we show that the simultaneous loss of these two genes under self-renewal conditions in vitro reduces the expression of the proliferation markers PCNA and Ki67, eliminates clonogenicity of the cells, reduces the number of cells in S phase, and induces aberrant differentiation primarily into the neuroblast lineage. This phenotype requires the loss of both genes and is not seen upon loss of Nfia or Nfib alone, nor with combined loss of Nfia and Nfix or Nfib and Nfix. These data demonstrate a unique combined requirement for both Nfia and Nfib for pNSC self-renewal.

Retinoic acid (RA), a vitamin A derivative, is an effective cell differentiating factor which plays critical roles in neuronal differentiation induction and the production of neurotransmitters in neurons. However, the specific changes in phosphorylation levels and downstream signalling pathways associated with RA remain unclear. This study employed qualitative and quantitative phosphoproteomics approaches based on mass spectrometry to investigate the phosphorylation changes induced by RA in C17.2 neural stem cells (NSCs). Dimethyl labelling, in conjunction with TiO2 phosphopeptide enrichment, was utilized to profile the phosphoproteome of self-renewing and RA-induced differentiated cells in C17.2 NSCs. The results of our study revealed that, qualitatively, 230 and 14 phosphoproteins were exclusively identified in the self-renewal and RA-induced groups respectively. Quantitatively, we successfully identified and quantified 177 unique phosphoproteins, among which 70 exhibited differential phosphorylation levels. Analysis of conserved phosphorylation motifs demonstrated enrichment of motifs corresponding to cyclin-dependent kinase and MAPK in the RA-induced group. Additionally, through a comprehensive literature and database survey, we found that the differentially expressed proteins were associated with the Wnt/β-catenin and Hippo signalling pathways. This work sheds light on the changes in phosphorylation levels induced by RA in C17.2 NSCs, thereby expanding our understanding of the molecular mechanisms underlying RA-induced neuronal differentiation.

Radial glia (RG) cells generate neurons and glial cells that make up the cerebral cortex. Both in rodents and humans, these stem cells remain for a specific time after birth, named late radial glia (lRG). The knowledge of lRG and molecules that may be involved in their differentiation is based on very limited data. We analyzed whether ascorbic acid (AA) and its transporter SVCT2, are involved in lRG cells differentiation. We demonstrated that lRG cells are highly present between the first and fourth postnatal days. Anatomical characterization of lRG cells, revealed that lRG cells maintained their bipolar morphology and stem-like character. When lRG cells were labeled with adenovirus-eGFP at 1 postnatal day, we detected that some cells display an obvious migratory neuronal phenotype, suggesting that lRG cells continue generating neurons postnatally. Moreover, we demonstrated that SVCT2 was apically polarized in lRG cells. In vitro studies using the transgenic mice SVCT2+/- and SVCT2tg (SVCT2-overexpressing mouse), showed that decreased SVCT2 levels led to accelerated differentiation into astrocytes, whereas both AA treatment and elevated SVCT2 expression maintain the lRG cells in an undifferentiated state. In vivo overexpression of SVCT2 in lRG cells generated cells with a rounded morphology that were migratory and positive for proliferation and neuronal markers. We also examined mediators that can be involved in AA/SVCT2-modulated signaling pathways, determining that GSK3-β through AKT, mTORC2, and PDK1 is active in brains with high levels of SVCT2/AA. Our data provide new insights into the role of AA and SVCT2 in late RG cells.

The central nervous system (CNS) is home to neuronal and glial cells. Traditionally, glia was disregarded as just the structural support across the brain and spinal cord, in striking contrast to neurons, always considered critical players in CNS functioning. In modern times this outdated dogma is continuously repelled by new evidence unravelling the importance of glia in neuronal maintenance and function. Therefore, glia replacement has been considered a potentially powerful therapeutic strategy. Glial progenitors are at the center of this hope, as they are the source of new glial cells. Indeed, sophisticated experimental therapies and exciting clinical trials shed light on the utility of exogenous glia in disease treatment. Therefore, this review article will elaborate on glial-restricted progenitor cells (GRPs), their origin and characteristics, available sources, and adaptation to current therapeutic approaches aimed at various CNS diseases, with particular attention paid to myelin-related disorders with a focus on recent progress and emerging concepts. The landscape of GRP clinical applications is also comprehensively presented, and future perspectives on promising, GRP-based therapeutic strategies for brain and spinal cord diseases are described in detail.

Mutations or dysregulation of nucleoporins (Nups) are strongly associated with neural developmental diseases, yet the underlying mechanisms remain poorly understood. Here, we show that depletion of Nup Seh1 in radial glial progenitors results in defective neural progenitor proliferation and differentiation that ultimately manifests in impaired neurogenesis and microcephaly. This loss of stem cell proliferation is not associated with defects in the nucleocytoplasmic transport. Rather, transcriptome analysis showed that ablation of Seh1 in neural stem cells derepresses the expression of p21, and knockdown of p21 partially restored self-renewal capacity. Mechanistically, Seh1 cooperates with the NuRD transcription repressor complex at the nuclear periphery to regulate p21 expression. Together, these findings identified that Nups regulate brain development by exerting a chromatin-associated role and affecting neural stem cell proliferation.

The interplay between neural progenitors and stem cells (NPSCs), and their extracellular matrix (ECM) is a crucial regulatory mechanism that determines their behavior. Nonetheless, how the ECM dictates the state of NPSCs remains elusive. The hindbrain is valuable to examine this relationship, as cells in the ventricular surface of hindbrain boundaries (HBs), which arise between any two neighboring rhombomeres, express the NPSC marker Sox2, while being surrounded with the membrane-bound ECM molecule chondroitin sulphate proteoglycan (CSPG), in chick and mouse embryos. CSPG expression was used to isolate HB Sox2+ cells for RNA-sequencing, revealing their distinguished molecular properties as typical NPSCs, which express known and newly identified genes relating to stem cells, cancer, the matrisome and cell cycle. In contrast, the CSPG- non-HB cells, displayed clear neural-differentiation transcriptome. To address whether CSPG is significant for hindbrain development, its expression was manipulated in vivo and in vitro. CSPG manipulations shifted the stem versus differentiation state of HB cells, evident by their behavior and altered gene expression. These results provide further understanding of the uniqueness of hindbrain boundaries as repetitive pools of NPSCs in-between the rapidly growing rhombomeres, which rely on their microenvironment to maintain their undifferentiated state during development.

Neural progenitor cells (NPCs) are essential for in vitro drug screening and cell-based therapies for brain-related disorders, necessitating well-defined and reproducible culture systems. Current strategies employing protein growth factors pose challenges in terms of both reproducibility and cost. In this study, we developed a novel DNA-based modulator to regulate FGFR signaling in NPCs, thereby facilitating the long-term maintenance of stemness and promoting neurogenesis. This DNA-based FGFR-agonist effectively stimulated FGFR1 phosphorylation and activated the downstream ERK signaling pathway in human embryonic stem cell (HESC)-derived NPCs. We replaced the basic fibroblast growth factor (bFGF) in the culture medium with our DNA-based FGFR-agonist to artificially modulate FGFR signaling in NPCs. Utilizing a combination of cell experiments and bioinformatics analyses, we showed that our FGFR-agonist could enhance NPC proliferation, direct migration, and promote neurosphere formation, thus mimicking the functions of bFGF. Notably, transcriptomic analysis indicated that the FGFR-agonist could specifically influence the transcriptional program associated with stemness while maintaining the neuronal differentiation program, closely resembling the effects of bFGF. Furthermore, our culture conditions allowed for the successful propagation of NPCs through over 50 passages while retaining their ability to efficiently differentiate into neurons. Collectively, our approach offers a highly effective method for expanding NPCs, thereby providing new avenues for disease-in-dish research and drug screening aimed at combating neural degeneration.

Neural stem cells (NSCs), with the capability of self-renewal, differentiation, and environment modulation, are considered promising for stroke, brain injury therapy, and neuron regeneration. Activation of endogenous NSCs, is attracting increasing research enthusiasm, which avoids immune rejection and ethical issues of exogenous cell transplantation. Yet, how to induce directed growth and differentiation in situ remain a major challenge. In this study, a pure water-driven Ni-Zn micromotor via a self-established electric-chemical field is proposed. The micromotors can be magnetically guided and precisely approach target NSCs. Through the electric-chemical field, bioelectrical signal exchange and communication with endogenous NSCs are allowed, thus allowing for regulated proliferation and directed neuron differentiation in vivo. Therefore, the Ni-Zn micromotor provides a platform for controlling cell fate via a self-established electrochemical field and targeted activation of endogenous NSCs.

Working memory (WM) deficits have frequently been linked to attention deficit hyperactivity disorder (ADHD). Despite previous studies suggested its high heritability, its genetic basis, especially in ADHD, remains unclear. The current study aimed to comprehensively explore the genetic basis of visual-spatial working memory (VSWM) in ADHD using wide-ranging genetic analyses.

The current study recruited a cohort consisted of 802 ADHD individuals, all met DSM-IV ADHD diagnostic criteria. VSWM was assessed by Rey-Osterrieth complex figure test (RCFT), which is a widely used psychological test include four memory indexes: detail delayed (DD), structure delayed (SD), structure immediate (SI), detail immediate (DI). Genetic analyses were conducted at the single nucleotide polymorphism (SNP), gene, pathway, polygenic and protein network levels. Polygenic Risk Scores (PRS) were based on summary statistics of various psychiatric disorders, including ADHD, autism spectrum disorder (ASD), major depressive disorder (MDD), schizophrenia (SCZ), obsessive compulsive disorders (OCD), and substance use disorder (SUD).

Analyses at the single-marker level did not yield significant results (5E-08). However, the potential signals with P values less than E-05 and their mapped genes suggested the regulation of VSWM involved both ocular and neural system related genes, moreover, ADHD-related genes were also involved. The gene-based analysis found RAB11FIP1, whose encoded protein modulates several neurodevelopment processes and visual system, as significantly associated with DD scores (P = 1.96E-06, Padj = 0.036). Candidate pathway enrichment analyses (N = 53) found that forebrain neuron fate commitment significantly enriched in DD (P = 4.78E-04, Padj = 0.025), and dopamine transport enriched in SD (P = 5.90E-04, Padj = 0.031). We also observed a significant negative relationship between DD scores and ADHD PRS scores (P = 0.0025, Empirical P = 0.048).

Our results emphasized the joint contribution of ocular and neural genes in regulating VSWM. The study reveals a shared genetic basis between ADHD and VSWM, with GWAS indicating the involvement of ADHD-related genes in VSWM. Additionally, the PRS analysis identifies a significant relationship between ADHD-PRS and DD scores. Overall, our findings shed light on the genetic basis of VSWM deficits in ADHD, and may have important implications for future research and clinical practice.

Neurodegenerative diseases (ND) are an entire spectrum of clinical conditions that affect the central and peripheral nervous system. There is no cure currently, with treatment focusing mainly on slowing down progression or symptomatic relief. Cellular therapies with various cell types from different sources are being conducted as clinical trials for several ND diseases. They include neural, mesenchymal and hemopoietic stem cells, and neural cells derived from embryonic stem cells and induced pluripotent stem cells. In this review, we present the list of cellular therapies for ND comprising 33 trials that used neural stem progenitors, 8 that used differentiated neural cells ,and 109 trials that involved non-neural cells in the 7 ND. Encouraging results have been shown in a few early-phase clinical trials that require further investigations in a randomized setting. However, such definitive trials may not be possible given the relative cost of the trials, and in the setting of rare diseases.

Spinal cord injury (SCI) is a devastating condition that enormously affects an individual's health and quality of life. Neurogenic lower urinary tract dysfunction (NLUTD) is one of the most important sequelae induced by SCI, causing complications including urinary tract infection, renal function deterioration, urinary incontinence, and voiding dysfunction. Current therapeutic methods for SCI-induced NLUTD mainly target on the urinary bladder, but the outcomes are still far from satisfactory. Stem cell therapy has gained increasing attention for years for its ability to rescue the injured spinal cord directly. Stem cell differentiation and their paracrine effects, including exosomes, are the proposed mechanisms to enhance the recovery from SCI. Several animal studies have demonstrated improvement in bladder function using mesenchymal stem cells (MSCs) and neural stem cells (NSCs). Human clinical trials also provide promising results in urodynamic parameters after MSC therapy. However, there is still uncertainty about the ideal treatment window and application protocol for stem cell therapy. Besides, data on the therapeutic effects regarding NSCs and stem cell-derived exosomes in SCI-related NLUTD are scarce. Therefore, there is a pressing need for further well-designed human clinical trials to translate the stem cell therapy into a formal therapeutic option for SCI-induced NLUTD.

The accuracy of electroencephalogram (EEG) source localization is compromised because of head modelling errors. In this study, we investigated the effect of inaccuracy in the conductivity of head tissues and head model structural deficiencies on the accuracy of EEG source analysis in premature neonates. A series of EEG forward and inverse simulations was performed by introducing structural deficiencies into the reference head models to generate test models, which were then used to investigate head modelling errors caused by cerebrospinal fluid (CSF) exclusion, lack of grey matter (GM)-white matter (WM) distinction, fontanel exclusion and inaccuracy in skull conductivity. The modelling errors were computed between forward and inverse solutions obtained using the reference and test models generated for each deficiency. Our results showed that the exclusion of CSF from the head model had a strong widespread effect on the accuracy of the EEG source localization with position errors lower than 4.17 mm. The GM and WM distinction also caused strong localization errors (up to 3.5 mm). The exclusion of fontanels from the head model also strongly affected the accuracy of the EEG source localization for sources located beneath the fontanels with a maximum localization error of 4.37 mm. Similarly, inaccuracies in the skull conductivity caused errors in EEG forward and inverse modelling in sources beneath cranial bones. Our results indicate that the accuracy of EEG source imaging in premature neonates can be largely improved by using head models, which include not only the brain, skull and scalp but also the CSF, GM, WM and fontanels.

Neural progenitor cells (NPCs) are multipotent neural stem cells (NSCs) capable of self-renewing and differentiating into neurons, astrocytes and oligodendrocytes. In the postnatal/adult brain, NPCs are primarily located in the subventricular zone (SVZ) of the lateral ventricles (LVs) and subgranular zone (SGZ) of the hippocampal dentate gyrus (DG). There is evidence that NPCs are also present in the postnatal/adult hypothalamus, a highly conserved brain region involved in the regulation of core homeostatic processes, such as feeding, metabolism, reproduction, neuroendocrine integration and autonomic output. In the rodent postnatal/adult hypothalamus, NPCs mainly comprise different subtypes of tanycytes lining the wall of the 3^rd ventricle. In the postnatal/adult human hypothalamus, the neurogenic niche is constituted by tanycytes at the floor of the 3^rd ventricle, ependymal cells and ribbon cells (showing a gap-and-ribbon organization similar to that in the SVZ), as well as suprachiasmatic cells. We speculate that in the postnatal/adult human hypothalamus, neurogenesis occurs in a highly complex, exquisitely sophisticated neurogenic niche consisting of at least four subniches; this structure has a key role in the regulation of extrahypothalamic neurogenesis, and hypothalamic and extrahypothalamic neural circuits, partly through the release of neurotransmitters, neuropeptides, extracellular vesicles (EVs) and non-coding RNAs (ncRNAs).

Due to the limited capacity of the adult mammalian brain to self-repair and regenerate, neurological diseases, especially neurodegenerative disorders and stroke, characterized by irreversible cellular damage are often considered as refractory diseases. Neural stem cells (NSCs) play a unique role in the treatment of neurological diseases for their abilities to self-renew and form different neural lineage cells, such as neurons and glial cells. With the increasing understanding of neurodevelopment and advances in stem cell technology, NSCs can be obtained from different sources and directed to differentiate into a specific neural lineage cell phenotype purposefully, making it possible to replace specific cells lost in some neurological diseases, which provides new approaches to treat neurodegenerative diseases as well as stroke. In this review, we outline the advances in generating several neuronal lineage subtypes from different sources of NSCs. We further summarize the therapeutic effects and possible therapeutic mechanisms of these fated specific NSCs in neurological disease models, with special emphasis on Parkinson's disease and ischemic stroke. Finally, from the perspective of clinical translation, we compare the strengths and weaknesses of different sources of NSCs and different methods of directed differentiation, and propose future research directions for directed differentiation of NSCs in regenerative medicine.

Heterozygous loss-of-function variants of FOXP4 are associated with neurodevelopmental disorders (NDDs) that exhibit delayed speech development, intellectual disability, and congenital abnormalities. The etiology of NDDs is unclear. Here we found that FOXP4 and N-cadherin are expressed in the nuclei and apical end-feet of radial glial cells (RGCs), respectively, in the mouse neocortex during early gestation. Knockdown or dominant-negative inhibition of Foxp4 abolishes the apical condensation of N-cadherin in RGCs and the integrity of neuroepithelium in the ventricular zone (VZ). Inhibition of Foxp4 leads to impeded radial migration of cortical neurons and ectopic neurogenesis from the proliferating VZ. The ectopic differentiation and deficient migration disappear when N-cadherin is over-expressed in RGCs. The data indicate that Foxp4 is essential for N-cadherin-based adherens junctions, the loss of which leads to periventricular heterotopias. We hypothesize that FOXP4 variant-associated NDDs may be caused by disruption of the adherens junctions and malformation of the cerebral cortex.

Adult neurogenesis is a process in which the adult neural stem cells produce newborn neurons that are implicated in terms of learning and memory. Methotrexate (MTX) is a chemotherapeutic drug, which has a negative effect on memory and hippocampal neurogenesis in animal models. Metformin is an antidiabetic drug with strong antioxidant capacities. We found that metformin ameliorates MTX induced deteriorations of memory and hippocampal neurogenesis in adult rats. In this study, we focus to investigate neural stem cells, biomarkers of apoptosis, and the protein for synaptogenesis, which involves in the transcription factors of the hippocampus in rats that received metformin and MTX. Male Sprague-Dawley rats were composed of control, MTX, metformin, and MTX+metformin groups. MTX (75 mg/kg, i.v.) was given on days 7 and 14, whereas metformin (200 mg/kg, i.p.) was given for 14 days. Hippocampal neural stem cells in the subgranular zone (SGZ) were quantified using immunofluorescence staining of Sox2 and nestin. Protein expression including PSD95, Casepase-3, Bax, Bcl-2, CREB, and pCREB were determined using Western blotting. MTX-treated rats displayed decreases in Sox2 and nestin-positive cells in the SGZ. Increases in Caspase-3 and Bax levels and decreases in PSD95, Bcl-2, CREB, and pCREB protein expressions in the hippocampus were also detected. However, these negative impacts of MTX were ameliorated by co-treatment with metformin. These consequences postulate that metformin has a potential to increase neural stem cells, synaptic plasticity, decreased apoptotic activities, and transcription factors, resulting in upregulation of hippocampal neurogenesis in MTX-treated rats.

Strokes are a leading cause of morbidity and mortality in adults worldwide. Extensive preclinical studies have shown that neural-stem-cell-based treatments have great therapeutic potential for stroke. Several studies have confirmed that the effective components of traditional Chinese medicine can protect and maintain the survival, proliferation, and differentiation of endogenous neural stem cells through different targets and mechanisms. Therefore, the use of Chinese medicines to activate and promote endogenous nerve regeneration and repair is a potential treatment option for stroke patients. Here, we summarize the current knowledge regarding neural stem cell strategies for ischemic strokes and the potential effects of these Chinese medicines on neuronal regeneration.

Mechanical stimulation modulates neural development and neuronal activity. In a previous study, magnetic "nano-pulling" is proposed as a tool to generate active forces. By loading neural cells with magnetic nanoparticles (MNPs), a precise force vector is remotely generated through static magnetic fields. In the present study, human neural stem cells (NSCs) are subjected to a standard differentiation protocol, in the presence or absence of nano-pulling. Under mechanical stimulation, an increase in the length of the neural processes which showed an enrichment in microtubules, endoplasmic reticulum, and mitochondria is found. A stimulation lasting up to 82 days induces a strong remodeling at the level of synapse density and a re-organization of the neuronal network, halving the time required for the maturation of neural precursors into neurons. The MNP-loaded NSCs are then transplanted into mouse spinal cord organotypic slices, demonstrating that nano-pulling stimulates the elongation of the NSC processes and modulates their orientation even in an ex vivo model. Thus, it is shown that active mechanical stimuli can guide the outgrowth of NSCs transplanted into the spinal cord tissue. The findings suggest that mechanical forces play an important role in neuronal maturation which could be applied in regenerative medicine.

Endogenous neural stem cells and their progeny (together termed neural precursor cells (NPCs)) are promising candidates to facilitate neuroregeneration. Charge-balanced biphasic monopolar stimulation (BPMP) is a clinically relevant approach that can activate NPCs both in vitro and in vivo. Herein, we established a novel ex vivo stimulation system to optimize the efficacy of BPMP electric field (EF) application in activating endogenous NPCs. Using the ex vivo system, we discerned that cathodal amplitude of 200 μA resulted in the greatest NPC pool expansion and enhanced cathodal migration. Application of the same stimulation parameters in vivo resulted in the same NPC activation in the mouse brain. The design and implementation of the novel ex vivo model bridges the gap between in vitro and in vivo systems, enabling a moderate throughput stimulation system to explore and optimize EF parameters that can be applied to clinically relevant brain injury/disease models.

Endogenous neural stem cells (eNSCs) in the adult brain, which have the potential to self-renew and differentiate into functional, tissue-appropriate cell types, have raised new expectations for neurological disease therapy. Low-intensity focused ultrasound (LIFUS)-induced blood-brain barrier modulation has been reported to promote neurogenesis. Although these studies have reported improved behavioral performance and enhanced expression of brain biomarkers after LIFUS, indicating increased neurogenesis, the precise mechanism remains unclear. In this study, we evaluated eNSC activation as a mechanism for neurogenesis after LIFUS-induced blood-brain barrier modulation. We evaluated the specific eNSC markers, Sox-2 and nestin, to confirm the activation of eNSCs. We also performed 3'-deoxy-3'[18F] fluoro-L-thymidine positron emission tomography ([18F] FLT-PET) to evaluate the activation of eNSCs. The expression of Sox-2 and nestin was significantly upregulated 1 week after LIFUS. After 1 week, the upregulated expression decreased sequentially; after 4 weeks, the upregulated expression returned to that of the control group. [18F] FLT-PET images also showed higher stem cell activity after 1 week. The results of this study indicated that LIFUS could activate eNSCs and induce adult neurogenesis. These results show that LIFUS may be useful as an effective treatment for patients with neurological damage or neurological disorders in clinical settings.

Curcumin from turmeric is a natural phenolic compound with a promising potential to regulate fundamental processes involved in neurological diseases, including inflammation, oxidative stress, protein aggregation, and apoptosis at the molecular level. In this regard, employing nanoformulation can improve curcumin efficiency by reducing its limitations, such as low bioavailability. Besides curcumin, growing data suggest that stem cells are a noteworthy candidate for neurodegenerative disorders therapy due to their anti-inflammatory, anti-oxidative, and neuronal-differentiation properties, which result in neuroprotection. Curcumin and stem cells have similar neurogenic features and can be co-administered in a cell-drug delivery system to achieve better combination therapeutic outcomes for neurological diseases. Based on the evidence, curcumin can induce the neuroprotective activity of stem cells by modulating their related signalling pathways. The present review is about the role of curcumin and its nanoformulations in the improvement of neurological diseases alone and through the effect on different categories of stem cells by discussing the underlying mechanisms to provide a roadmap for future investigations.