Search Results (512)

Neuroinflammation is a key response to disturbed CNS homeostasis, largely mediated by activated microglia, and excessive microglia-driven inflammation can negatively impact neurogenesis. ZEB1 plays a crucial role in neurogenesis and brain development by influencing neural stem cell (NSC) maintenance, proliferation, and differentiation. This study aimed to evaluate how the knockdown of ZEB1 influences the behavior of NSCs in inflammatory environments. NSCs were isolated from the subventricular zone of rats, and ZEB1 knockdown was achieved using ZEB1 siRNA. A conditioned medium derived from lipopolysaccharide-activated microglia was utilized to induce inflammatory responses in NSCs. The silencing of ZEB1 in NSCs significantly reduced the expression of ZEB1. Furthermore, ZEB1 knockdown in NSCs resulted in a significant decrease in neurosphere formation, cell migration ability, reactive oxygen species generation, and various cytokine levels under both non-inflammatory and inflammatory conditions. These findings reveal the regulatory role of ZEB1 in the modulation of NSC behavior, suggesting that targeting ZEB1 may provide a potential therapeutic strategy for neuroinflammatory CNS disorders. Full article

The deposition of amyloid beta (Aβ) in the human brain is a hallmark of Alzheimer’s disease (AD). Aβ has been shown to exert a wide range of effects on neurons in cell and animal models. Here, we take advantage of differentiated neurons from iPSC-derived neural stem cells of human donors to examine its effects on human neurons. Specifically, we employed two types of neurons from genetically distinct donors: one male carrying APO E2/E2 (M E2/E2) and one female carrying APO E3/E3 (F E3/E3). Genome-wide RNA-sequencing analysis identified 64 and 44 genes that were induced by Aβ in M E2/E2 and F E3/E3 neurons, respectively. GO and pathway analyses showed that Aβ-induced genes in F E3/E3 neurons do not constitute any statistically significant pathways whereas Aβ-induced genes in M E2/E2 neurons constitute a complex network of activated pathways. These pathways include those promoting inflammatory responses, such as IL1β, IL4, and TNF, and those promoting cell migration and movement, such as chemotaxis, migration of cells, and cell movement. These results strongly suggest that the effects of Aβ on neurons are highly dependent on their genetic background and that Aβ can promote strong responses in inflammation and cell migration in some, but not all, neurons. Full article

Organophosphate (OP) and carbamate pesticides are widely employed in agriculture to facilitate the production of economically viable crops. However, pesticide contamination of food, water, and air leads to undesired human exposure. Neuronal tissue may be particularly vulnerable to pesticide toxicity during periods of neurodevelopment. Hence, this study aimed to investigate the neurotoxicity of three pesticide compounds, namely chlorpyrifos-oxon (CPO), azamethiphos (AZO), and aldicarb, on human neural progenitor cells (hNPCs) and whether toxicity differed between undifferentiated and differentiated stem cells. Undifferentiated and differentiated hNPCs were exposed to these neurotoxicants at concentrations of 0–200 µM for 24 h, and cell viability was evaluated using 3-(4,5 dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assays. The impact of the neurotoxicants on cellular bioenergetics was determined by quantifying cellular ATP levels and the production of reactive oxygen species (ROS) using a 2′,7′-dichlorofluorescein diacetate (DCFDA) assay. Concentration–response curves were also generated to measure their relative inhibition of AChE. The neurotoxicants induced concentration-dependent reductions in cell viability (p < 0.0001), cellular ATP levels (p < 0.0001), and the inhibition of AChE (p < 0.0001). Notably, differentiated neurons displayed higher sensitivity than undifferentiated neural stem cells (NSCs), with a toxicity threshold of ≥1 µM. ROS levels were significantly increased (p < 0.0001) following neurotoxicant exposures, more so in differentiated cells, with levels that correlated with cytotoxicity, cell death, and the induction of oxidatively damaged proteins in surviving cells. These findings suggest a central role of oxidative stress and protein oxidation in mediating the neurotoxic effects of pesticide compounds on NSCs. Furthermore, the heightened susceptibility of NSCs to pesticide toxicity after differentiation is indicative of human vulnerability during periods of neurodevelopment. Full article

N-glycanase 1 (NGLY1) is a cytoplasmic glycoenzyme that removes N-linked glycans from misfolded glycoproteins. It plays an important role in the endoplasmic reticulum-associated degradation (ERAD) pathway in mammalian cells. NGLY1 dysfunction in humans causes NGLY1 deficiency as a rare autosomal recessive disorder that is characterized by neurodevelopmental delay, hypotonia, movement disorders, seizures, and multi-system involvement. In this review, we summarize recent advances in understanding the neural functions of NGLY1 and the neuropathological phenotypes associated with its deficiency. We discuss the molecular basis of NGLY1 deficiency in the central nervous system (CNS) and pathophysiological insights from animal and human induced pluripotent stem cell (iPSC)-based models. We also highlight emerging gene therapy approaches aimed at restoring NGLY1 activity and alleviating neurological symptoms. Full article

Background: Glioblastomas (GBMs) are the most aggressive and common neoplasms that affect glial cells, presenting rapid growth, invasion, and resistance to treatments. Studies have demonstrated the potentially inhibitory effect of flavonoids on glioblastoma cells’ stemness and viability. However, further research is needed to explore sensitivity and the mechanism of action in chemoresistant cells. Methods: In this study, we characterized the impact of apigenin treatment on the viability and differentiation of human GBM cells in vitro and its effects on tumorigenesis and regulation of the inflammatory response in vivo. Results: The flavonoid apigenin reduced the viability of U-251 cells, patient-derived cells TG-1 and OB-1 stem cells in a dose-dependent manner, associated with the induction of acidic vesicle organelles formation and apoptosis. Treatment with apigenin also inhibited migration and induced neural differentiation in the remaining viable cells, characterized by a decrease in the expression of the precursor marker nestin and an increase in the expression of astrocyte and neuron markers, GFAP and β-III tubulin, respectively. The xenotransplantation of apigenin-pretreated U251 cells into rat brains did not lead to tumor formation, unlike untreated cells. The surrounding area of transplanted untreated U251 cells exhibited reactive microglia and astrocytes, along with increased VEGF expression, which was absent in implant sites of apigenin-pretreated GBM cells. Moreover, in this implant area, we observed a significant decrease in the expression of mRNA for inflammatory factors IL-1β, TNF, and NOS2, and the downregulation of IL-10 and IL-4. Conclusions: These results demonstrate that apigenin inhibits the growth of tumoral cells, affecting the viability of tumor stem cells and impairing tumorigenicity, while altering the regulatory profile of immunomodulatory proteins. Therefore, this flavonoid can be considered for further studies to determine its use as an adjuvant to the treatment of human GBMs. Full article

X chromosome inactivation (XCI) is a fundamental epigenetic process that balances X-linked gene expression between females and males by silencing one X chromosome in female cells. Variability or skewing of XCI can influence the clinical presentation of X-linked disorders. Bain type X-linked intellectual disability syndrome (MRXSB), caused by mutations in the X-linked HNRNPH2 gene, is characterized by intellectual disability, developmental delay, and neurological abnormalities. In female patients, XCI heterogeneity complicates disease modeling and therapeutic development. Induced pluripotent stem cells (iPSCs) offer a unique platform to study patient-specific disease mechanisms, but the dynamics of XCI during iPSC reprogramming, maintenance, and differentiation are not fully understood. In this study, we generated 12 iPSC clones from fibroblasts of a female MRXSB patient heterozygous for the HNRNPH2 c.340C > T mutation. Four clones expressed the mutant HNRNPH2 allele and eight expressed the wild-type allele, indicating X chromosome reactivation (XCR) followed by random XCI during reprogramming. Importantly, these XCI patterns remained stable during long-term iPSC propagation and subsequent differentiation into the three germ layers and neural stem cells. Our findings provide new insights into XCI and XCR dynamics in the context of X-linked neurodevelopmental disorders and emphasize the importance of careful clone selection for accurate disease modeling using iPSC-based approaches. Full article

Human induced pluripotent stem cells (hiPSCs) hold great potential for patient-specific therapies. Transplantation of hiPSC-derived neural progenitor cells (NPCs) is a promising reparative strategy for spinal cord injury (SCI), but clinical translation requires efficient differentiation into desired neural lineages and purification before transplantation. Here, differentiated hiPSCs—reprogrammed from human skin fibroblasts using Sendai virus-mediated expression of OCT4, SOX2, KLF4, and C-MYC—into neural rosettes expressing SOX1 and PAX6, followed by neuronal precursors (β-tubulin III⁺/NESTIN⁺) and glial precursors (GFAP+/NESTIN+). Both neuronal and glial precursors expressed the A2B5 surface antigen. A2B5+ NPCs, purified by fluorescence-activated cell sorting (FACS), proliferated in vitro with mitogens, and differentiated into mature neurons and astrocytes under lineage-specific conditions. Then, NOD-SCID mice received a T9 contusion injury followed by transplantation of A2B5+ NPCs, human fibroblasts, or control medium at 8 days post-injury. At two months, grafted NPCs showed robust survival, progressive neuronal maturation (β-tubulin III⁺→doublecortin⁺→NeuN⁺), and astrocytic differentiation (GFAP+), particularly in spared white matter. Transplantation significantly increased spared white matter volume and improved hindlimb locomotor recovery, with no teratoma formation observed. These results demonstrate that hiPSC-derived, FACS-purified A2B5+ NPCs can survive, differentiate into neurons and astrocytes, and enhance functional recovery after SCI. This approach offers a safe and effective candidate cell source for treating SCI and potentially other neurological disorders. Full article

Thyroid hormones (THs) are essential for brain development, and their dysregulation is associated with cognitive deficits and neurodevelopmental disorders. Down syndrome (DS), caused by trisomy 21, is frequently associated with thyroid dysfunction and impaired neurogenesis. Here, we investigated THs signaling dynamics during neural differentiation using human induced pluripotent stem cells (hiPSCs) derived from individuals with DS and controls. We analyzed the gene expression of key THs regulators—deiodinases, transporters, and receptors—and downstream target genes in hiPSCs, hiPSC-derived neural progenitor cells (NPCs), hiPSC-derived astrocytes, and hiPSC-derived neurons. DS-derived hiPSCs, hiPSC-derived NPCs, and hiPSC-derived neurons exhibited 2- to 7-fold increases in the gene expression of DIO2 and 3- to 8-fold reductions in DIO3, alongside 1- to 3-fold downregulation of THRA and THRB isoforms. hiPSC-derived astrocytes showed a 4-fold decrease in the gene expression of DIO2, a 4-fold increase in DIO3, upregulation of SLC16A10 (2-fold), and downregulation of SLC7A5 (0.5-fold) and THs receptors (0.5- to 12-fold). hiPSC-derived neurons exhibited marked downregulation of the gene expression of HOMER1 (0.5-fold), GRIN3A (14-fold), and GRIN3B (4-fold), accompanied by impaired spontaneous activity in multi-electrode array recordings. These findings reveal a robust, cell-type-specific imbalance between THs availability and signaling competence in DS hiPSC-derived neural cells, providing mechanistic insight into THs-related contributions to the function of DS hiPSC-derived neural cells and identifying potential therapeutic targets. Full article

Autism Spectrum Disorder (ASD) is a neurodevelopmental condition that affects communication, social interaction, and behavior. Calcium (Ca²⁺) signaling dysregulation has been frequently highlighted in genetic studies as a contributing factor to aberrant developmental processes in ASD. Herein, we used ASD and control induced pluripotent stem cells (iPSCs) to investigate transcriptomic and functional Ca²⁺ dynamics at various stages of differentiation to cortical neurons. Idiopathic ASD and control iPSC lines underwent the dual SMAD inhibition differentiation protocol to direct their fate toward cortical neurons. Samples from multiple time points along the course of differentiation were processed for bulk RNA sequencing, spanning the following sequential stages: the iPSC stage, neural induction (NI) stage, neurosphere (NSP) stage, and differentiated cortical neuron (Diff) stage. Our transcriptomic analyses suggested that the numbers of Ca²⁺ signaling-relevant differentially expressed genes between ASD and control samples were higher in the iPSC and Diff stages. Accordingly, samples from the iPSC and Diff stages were processed for Ca²⁺ imaging studies. Results revealed that iPSC-stage ASD samples displayed elevated maximum Ca²⁺ levels in response to ATP compared to controls. By contrast, in the Diff stage, ASD neurons showed reduced maximum Ca²⁺ levels in response to ATP but increased maximum Ca²⁺ levels in response to KCl and DHPG relative to controls. Considering the distinct functional signaling contexts of these stimuli, this differential profile of receptor- and ionophore-mediated Ca²⁺ response suggests that aberrant calcium homeostasis underlies the pathophysiology of ASD neurons. Our data provides functional evidence for Ca²⁺ signaling dysregulation during neurogenesis in idiopathic ASD. Full article

Multiple differentiation protocols have emerged in recent years, producing neurons with diverse morphologies, gene and protein expression profiles, and functionality. Many of these differentiation techniques require months of culture and the use of expensive growth factors. Most importantly, the derived neurons usually do not exhibit any electrical activity. This limits the value of the protocol as a tool for engineering and investigating neural networks. Here, we describe an efficacious method for differentiating mouse embryonic stem cells into functional neurons. CGR8 cells were neurally induced via the simultaneous application of retinoic acid and purmorphamine. The derived cells expressed neuronal (TUJ1 and NeuN) and synaptic (GAD2, PSD-95, Synaptophysin, and VGLUT1) markers. During whole-cell recordings, neurons exhibited inward and outward currents, likely caused by fast-inactivating voltage-gated potassium channels. Upon current injection, miniature action potentials were also recorded. The efficient generation of diverse subtypes of functional neurons can be a useful tool in fundamental investigations of neural network activity and translational studies. Full article

Major depressive disorder (MDD) is a prevalent and disabling psychiatric condition with limited treatment options for patients who are resistant to conventional pharmacological and psychotherapeutic interventions. Stem cell (SC)-based therapies have emerged as a promising experimental approach, offering multifaceted mechanisms of action including neurogenesis, immunomodulation, antioxidative protection, and neuromodulation. This narrative review synthesizes current evidence from preclinical studies and early-phase clinical trials on the efficacy of mesenchymal stem cells (MSCs), neural stem cells (NSCs), and induced pluripotent stem cells (iPSCs) in alleviating depressive-like behaviors. Mechanistic insights include enhanced hippocampal neurogenesis, modulation of the brain-derived neurotrophic factor (BDNF)–TrkB pathway, attenuation of neuroinflammation through microglial polarization, and restoration of serotonergic signaling via peripheral-to-central pathways such as via the vagus nerve. In addition, the therapeutic potential of extracellular vesicles (EVs) and intranasal administration as non-invasive delivery strategies is discussed. While animal and first preclinical studies suggest potential benefit, significant translational barriers remain, including issues of scalability, long-term safety, and ethical considerations. Further rigorous studies are needed to validate stem-cell-based therapies as viable treatments for MDD. Full article

CD44, a structurally diverse cell-surface glycoprotein, plays a multifaceted and indispensable role in neural tissue across both physiological and pathological conditions. It orchestrates complex cell–extracellular matrix interactions and intracellular signaling through its variant isoforms and post-translational modifications and is broadly expressed in neural stem/progenitor cells, microglia, astrocytes, and selected neuronal populations. The interactions of CD44 with ligands such as hyaluronan and osteopontin regulate critical cellular functions, including migration, differentiation, inflammation, and synaptic plasticity. In microglia and macrophages, CD44 mediates immune signaling and phagocytic activity, and it is dynamically upregulated in neuroinflammatory diseases, particularly through pathways involving Toll-like receptor 4. CD44 expression in astrocytes is abundant during central nervous system development and in diseases, contributing to glial differentiation, reactive astrogliosis, and scar formation. Though its expression is less prominent in mature neurons, CD44 supports neural plasticity, circuit organization, and injury-induced repair mechanisms. Additionally, its expression at nervous system barriers, such as the blood–brain barrier, underscores its role in regulating vascular permeability during inflammation and ischemia. Collectively, CD44 emerges as a critical integrator of neural cell function and intercellular communication. Although the roles of CD44 in glial cells appear to be similar to those explored in other tissues, the expression of this molecule and its variants on neurons reveals peculiar functions. Elucidating the cell-type-specific roles and regulation of CD44 variants may offer novel therapeutic strategies for diverse neurological disorders. Full article

Tissue engineering has the potential to overcome the limitations of using autografts in nerve gap repair, using cellular biomaterials to bridge the gap and support neuronal regeneration. Various types of therapeutic cells could be considered for use in aligned collagen-based engineered neural tissue (EngNT), including Schwann cells and their precursors, which can be derived from human induced pluripotent stem cells (hiPSCs). Using Schwann cell precursors may have practical advantages over mature Schwann cells as they expand readily in vitro and involve a shorter differentiation period. However, the performance of each cell type needs to be tested in EngNT. By adapting established protocols, hiPSCs were differentiated into Schwann cell precursors and Schwann cells, with distinctive molecular profiles confirmed using immunocytochemistry and RT-qPCR. For the first time, both cell types were incorporated into EngNT using gel aspiration–ejection, a technique used to align and simultaneously stabilise the cellular hydrogels. Both types of cellular constructs supported and guided aligned neurite outgrowth from adult rat dorsal root ganglion neurons in vitro. Initial experiments in a rat model of nerve gap injury demonstrated the extent to which the engrafted cells survived after 2 weeks and indicated that both types of hiPSC-derived cells supported the infiltration of host neurons, Schwann cells and endothelial cells. In summary, we show that human Schwann cell precursors promote infiltrating endogenous axons in a model of peripheral nerve injury to a greater degree than their terminally differentiated Schwann cell counterparts. Full article

Organoids allow to model healthy and diseased human tissues. and have applications in developmental biology, drug discovery, and cell therapy. Traditionally cultured in immersion/suspension, organoids face issues like lack of standardization, fusion, hypoxia-induced necrosis, continuous agitation, and high media volume requirements. To address these issues, we developed an air–liquid interface (ALi) technology for culturing organoids, termed AirLiwell. It uses non-adhesive microwells for generating and maintaining individualized organoids on an air–liquid interface. This method ensures high standardization, prevents organoid fusion, eliminates the need for agitation, simplifies media changes, reduces media volume, and is compatible with Good Manufacturing Practices. We compared the ALi method to standard immersion culture for midbrain organoids, detailing the process from human pluripotent stem cell (hPSC) culture to organoid maturation and analysis. Air–liquid interface organoids (3D-ALi) showed optimized size and shape standardization. RNA sequencing and immunostaining confirmed neural/dopaminergic specification. Single-cell RNA sequencing revealed that immersion organoids (3D-i) contained 16% fibroblast-like, 23% myeloid-like, and 61% neural cells (49% neurons), whereas 3D-ALi organoids comprised 99% neural cells (86% neurons). Functionally, 3D-ALi organoids showed a striking electrophysiological synchronization, unlike the heterogeneous activity of 3D-i organoids. This standardized organoid platform improves reproducibility and scalability, demonstrated here with midbrain organoids. The use of midbrain organoids is particularly relevant for neuroscience and neurodegenerative diseases, such as Parkinson’s disease, due to their high incidence, opening new perspectives in disease modeling and cell therapy. In addition to hPSC-derived organoids, the method’s versatility extends to cancer organoids and 3D cultures from primary human cells. Full article

Central nervous system (CNS) disorders present significant therapeutic challenges due to the limited regenerative capacity of neural tissues, resulting in long-term disability for many patients. Consequently, the development of novel therapeutic strategies is urgently warranted. Stem cell therapies show considerable potential for mitigating brain damage and restoring neural connectivity, owing to their multifaceted properties, including anti-apoptotic, anti-inflammatory, neurogenic, and vasculogenic effects. Recent research has also identified exosomes—small vesicles enclosed by a lipid bilayer, secreted by stem cells—as a key mechanism underlying the therapeutic effects of stem cell therapies, and given their enhanced stability and superior blood–brain barrier permeability compared to the stem cells themselves, exosomes have emerged as a promising alternative treatment for CNS disorders. A key challenge in the application of both stem cell and exosome-based therapies for CNS diseases is the method of delivery. Currently, several routes are being investigated, including intracerebral, intrathecal, intravenous, intranasal, and intra-arterial administration. Intracerebral injection can deliver a substantial quantity of stem cells directly to the brain, but it carries the potential risk of inducing additional brain injury. Conversely, intravenous transplantation is minimally invasive but results in limited delivery of cells and exosomes to the brain, which may compromise the therapeutic efficacy. With advancements in catheter technology, intra-arterial administration of stem cells and exosomes has garnered increasing attention as a promising delivery strategy. This approach offers the advantage of delivering a significant number of stem cells and exosomes to the brain while minimizing the risk of additional brain damage. However, the investigation into the therapeutic potential of intra-arterial transplantation for CNS injury is still in its early stages. In this comprehensive review, we aim to summarize both basic and clinical research exploring the intra-arterial administration of stem cells and exosomes for the treatment of CNS diseases. Additionally, we will elucidate the underlying therapeutic mechanisms and provide insights into the future potential of this approach. Full article