Search Results (280)

Extracellular vesicles (EVs) have emerged as key mediators of intercellular communication in the brain, with glial cell-derived EVs increasingly recognized for their roles in maintaining brain homeostasis and contributing to the progression of neurodegenerative diseases. By transferring a diverse cargo of bioactive molecules, including proteins, RNAs, and organelles, EVs influence recipient cell behavior and overall brain function. In neurodegenerative conditions, glial EVs can either propagate pathogenic signals or deliver neuroprotective and regenerative cues, depending on their cellular origin and molecular composition. This context-dependent heterogeneity highlights the need for physiologically relevant human models to investigate EVs biology. Human induced pluripotent stem cell (iPSC)-derived glial models provide a disease-relevant platform, as they recapitulate key pathological features of Alzheimer’s disease (AD), Parkinson’s disease (PD) and amyotrophic lateral sclerosis (ALS). When further integrated with brain organoid platforms, these iPSC-based systems enable the generation of three-dimensional environments that closely resemble in vivo EVs dynamics. Importantly, glial EVs can modulate cellular pathways involved in neuronal survival and function. Indeed, their potential to interact with and, under specific experimental conditions, traverse the blood–brain barrier (BBB) has contributed to growing interest in their application for biomarker discovery and therapeutic development. Engineered and patient-specific EVs derived from iPSCs are emerging as promising tools for targeted, cell type-specific, therapeutic approaches, although their clinical applicability still requires further validation. This review discusses the emerging evidence supporting the dual role of iPSC-derived glial EVs in health and disease, underscores the translational potential of iPSC-based platforms for mechanistic studies, and outlines their promise as precision medicine tools for diagnostics and therapy. Full article

The intracellular accumulation of amyloid beta 42 (iAβ42) has been proposed as an early pathological indicator of familial Alzheimer’s disease (FAD). DJ-1 is a multifunctional protein sensitive to oxidative stress (OS) that has been associated with neurodegeneration; however, its role in iAβ42 pathology is unclear. In this study, we examined whether oxidized (sulfonic) DJ-1 (Cys¹⁰⁶-SO₃H) drives iAβ42 accumulation using postmortem brain samples and in vitro 3D iPSC-derived cerebral organoids (COs) or 2D induced pluripotent stem cells (iPSC)-derived ChLNs (cholinergic-like neurons) models from a PSEN1 E280A patient and a healthy volunteer (as a control sample). Post-mortem analyses of the temporal and frontal cortices and hippocampus from FAD PSEN1 E280A patients revealed strong intracellular co-localization of sulfonic DJ-1 and iAβ42, which was absent in control samples. To validate these findings, we generated COs from an iPSC PSEN1 E280A FAD patient and a healthy donor. In these organoids, we observed the co-localization of oxidized DJ-1 and Aβ42 in the absence of extracellular fibrils or plaques, as confirmed by BTA-1 staining. To further support these observations, 2D iPSC PSEN1 E280A-derived ChLNs cultures showed that intracellular Aβ42 accumulates progressively in direct correlation with increasing DJ-1 oxidation, as demonstrated by immunofluorescence microscopy and Western blotting analysis. These results indicate that DJ-1 oxidation accompanies the earliest intracellular stages of Aβ42 pathology. Furthermore, complementary in silico molecular docking analysis revealed a higher affinity between Aβ42 and oxidized sulfonic DJ-1 (DJ-1 Cys¹⁰⁶-SO₃H) compared to sulfenic (DJ-1 Cys¹⁰⁶-SOH) or sulfinic acid (DJ-1 Cys¹⁰⁶-SO₂H) forms. Likewise, ELISA tests and seeding assays confirmed that oxidized DJ-1 binds to and decelerates Aβ42 aggregation kinetics. Together, our results identify DJ-1 oxidation as a critical molecular event in the accumulation of iAβ42 in FAD. These findings suggest that oxidized DJ-1 represents not only a potential early biomarker of intracellular pathology but also a pharmacological target. Preventing the oxidation of DJ-1 or its pathological aggregation could provide new biomarkers and therapeutic strategies for reducing the intracellular accumulation of Aβ42 and neurodegeneration in FAD. Full article

Background/Objectives: Magnesium (Mg) is essential for neuronal maturation, yet its role in human cortical development remains poorly defined. Here, we investigated the effects of physiological (1 mM) and elevated (5 mM) concentrations of MgSO₄ and magnesium pidolate (MgPid) on human brain organoids co-cultured with an in vitro blood–brain barrier (BBB) model. Methods: Human brain organoids derived from induced pluripotent stem cells were co-cultured with an in vitro BBB system and treated for 4 days with either MgSO₄ or MgPid at physiological and elevated concentrations. Cortical organization was assessed by transmission electron microscopy and immunofluorescence analysis. Western blotting for neurotransmitter receptors and Mg transporters, quantification of intraorganoid Mg²⁺ levels, ELISA-based measurement of GABA and dopamine, and analysis of glutamate were performed. Results: High Mg exposure enhanced cortical stratification and neuronal organization, as shown by the localization of CTIP2 in the outermost layer and TBR2 in the inner layer, together with ultrastructural features consistent with advanced differentiation. Elevated Mg increased intraorganoid Mg²⁺ levels without altering Mg transporter abundance and selectively modulated neurotransmitter receptor expression: NMDA-R levels were reduced by MgPid, whereas GABA_A-R and GABA_B-R were upregulated, particularly in response to MgPid. Levels of glutamate, GABA, and dopamine remained unchanged. Conclusions: These findings identify Mg, especially in the form of MgPid, as a modulator of cortical architecture and inhibitory–excitatory receptor balance in human organoids, supporting its potential relevance for neurodevelopmental regulation and Mg-based therapeutic strategies. These results also support organoids as human-relevant, animal-free tools for neuroscience and neuropharmacological research. Full article

Brain-on-a-chip (BOC) refers to a miniaturized in vitro platform that integrates living neuronal networks on a micro-engineered chip, enabling the simulation of brain functions, neural activities and physiological responses. BOC technology is an advanced evolution of microphysiological systems (MPS) and Lab-on-a-Chip platforms, providing novel paradigms for in vitro modeling and exploring early-stage biocomputing by interfacing living neural networks with engineered electronics. Microelectrode arrays (MEAs) serve as the critical physical interface for bidirectional communication in these systems. In this review, we systematically examine the technological landscape and engineering requirements of MEAs tailored for BOC applications, evaluating them across electrical characteristics, structural properties, and biocompatibility. Two primary classes of current MEA technologies, including planar arrays for 2D neural cultures and 3D flexible arrays for brain organoids, are discussed in detail. We highlight the transition from passive planar electrodes to high-density active CMOS and TFT-based arrays, and detail how 3D flexible MEAs utilize endogenous integration and exogenous wrapping strategies to overcome tissue-mechanics mismatches. Furthermore, the integration of MEAs with microfluidics, optoelectronics, and electrochemical sensors to enable multimodal monitoring is explored. With the advantages of the various MEAs, the application of MEAs for BOC, particularly in biological computing and network plasticity research, is discussed. Finally, future technological developments in scalability bottlenecks, chronic stability, and the incorporation of artificial intelligence for MEAs of BOC are prospected. Full article

Thyroid hormones (THs) are essential regulators of human brain development, and disrupted TH availability during pregnancy or early life is linked to adverse neurodevelopmental outcomes. Concerns that environmental chemicals interfere with TH signalling have increased the need for human-relevant in vitro systems to identify thyroid hormone system-disrupting chemicals (THSDCs) for risk assessment. Here, we compared two human-induced pluripotent stem cell (hiPSC)-derived brain organoid models for THSDC assessment: (i) human cortical organoids (COs) generated by unguided differentiation, offering higher architectural complexity but lower throughput; and (ii) neural stem cell-derived organoids (NSCOs), designed for scalability with reduced cellular diversity. Both models expressed key TH handling components, including the transporter SLC16A2 (MCT8) and the inactivating enzyme DIO3. Using LC–MS/MS, we show that exogenous T3 is depleted from culture media and metabolized to 3,3′-T2 and 3′-T1 in both models, alongside upregulation of T3-responsive genes (HR, KLF9, DIO3, SEMA3C). Pulse and chronic co-exposures to reference disruptors iopanoic acid (IA, deiodinase inhibitor) and silychristin (SC, MCT8 inhibitor) altered T3 metabolism and modulated T3-responsive transcriptional endpoints. In NSCOs, high-content imaging revealed treatment-associated changes in cell composition, with chronic T3 reducing the SOX2-positive progenitor pool and THSDCs blocking this effect. Together, these findings provide a framework for organoid qualification—linking TH handling, transcriptomic responsiveness, and scalable phenotypic readouts—as a necessary step toward model validation and implementation of brain organoids in THSDC risk assessment pipelines. Full article

Glioblastoma multiforme (GBM) remains the most aggressive and therapeutically intractable primary brain tumor, with many patients experiencing rapid relapse despite maximal surgical resection followed by standard chemoradiation. This persistent failure reflects the convergence of profound tumor-intrinsic genetic heterogeneity and a highly dynamic, spatially structured, and immunosuppressive tumor microenvironment (TME). Together, these forces create strong selective pressures that fuel tumor evolution, intratumoral diversity, phenotype plasticity, diffuse invasion, and robust resistance to therapy. The TME of GBM is orchestrated through a complex interplay between diverse cellular constituents, including tumor-associated macrophages, reactive astrocytes, endothelial cells, pericytes, and GBM stem cells, and non-cellular components such as extracellular matrix remodeling, hypoxia, metabolic and nutrient gradients, and spatially patterned cytokine and chemokine signaling networks. Additionally, heterogeneity in blood–brain barrier (BBB) and blood–tumor barrier (BTB) complicates drug delivery and immune surveillance, reinforcing therapeutic resistance and regional tumor adaptation. Conventional two-dimensional cell cultures and animal models fail to sufficiently capture these multiscale, patient-specific interactions, limiting their translational predictive power. In this narrative review, we synthesize recent advances in GBM organoid technologies as physiologically relevant, three-dimensional platforms that more faithfully recapitulate TME for driving tumor evolution and treatment resistance. We compare complementary organoid strategies, including patient-derived GBM organoids that preserve native cytoarchitecture, cerebral organoid co-culture systems that reconstruct tumor–brain interactions, and advanced platforms incorporating immune and vascular features such as air–liquid interface cultures, microglia-enriched systems, and BBB/BTB-integrated models. Finally, we highlight emerging innovations such as spatial transcriptomics, organoid-on-a-chip systems, live imaging coupled with lineage tracing, genome engineering, and artificial intelligence integration that collectively position GBM organoids at the forefront of precision neuro-oncology, reproducing TME, enabling dynamic mapping of tumor evolution, and accelerating patient-specific therapeutic discovery. Full article

High-density microelectrode arrays (HDMEAs) have become increasingly important tools in neuroscience and biomedical engineering because of their high spatial and temporal resolution for recording and modulating electrical activity across diverse biological systems. Initially developed for in vitro studies of cultured cells, HDMEAs are now being applied to increasingly complex models, including organoids, animal systems, and even human neural systems. These advancements enable a deeper investigation of cellular interactions, network dynamics, and disease mechanisms, as well as providing novel therapeutic and diagnostic tools for neurological disorders. This review explores the evolution of HDMEAs, emphasizing recent innovations in their design, fabrication, and functionalization. We discuss their applications across cellular models, organoid systems, animal studies, and human electrophysiology, and highlight current challenges such as biocompatibility, long-term stability, scalability, and translational deployment. Finally, we outline future directions for advancing HDMEA technologies in both research and clinical settings. Full article

Paraspeckles are subnuclear ribonucleoprotein condensates that regulate host stress responses, including those triggered by viral infection. In vitro studies using non-neuronal cells have shown the involvement of specific paraspeckle components in facilitating the replication of certain viruses, including Herpes Simplex Virus 1 (HSV-1), but these processes have not been investigated in human neuronal cells, which represent a relevant target of the virus. We employed human neural precursor cells (NPCs), neurons, and brain organoids derived from hiPSCs to investigate the previously unexplored dynamics of paraspeckle components in HSV-1-infected human neuronal cells. Our results reveal cell-type-specific differences in the expression of paraspeckle genes in response to HSV-1 infection. Unlike other viruses, HSV-1 orchestrates a previously unreported redistribution of paraspeckle proteins, leading to their accumulation in viral replication compartments (VRCs). Importantly, the expression of the paraspeckle proteins NONO and SFPQ correlates with HSV-1 permissiveness in human neuronal cells and may be required to establish a nuclear environment favoring viral transcription/replication. This enhances our understanding of how stress-response pathways in cells can be exploited by viruses in a cell-type-specific manner. Full article

Background/Objectives: Pediatric cancers are disorders of dysregulated development driven largely by genomic and epigenetic alterations. Precisely modeling these developmental differences is essential for understanding the unique biology of childhood cancers. Patient-derived organoids (PDOs) offer a powerful in vitro platform that recapitulates tumor heterogeneity, plasticity, microenvironment (including immune cells) and disease-relevant features. Methods: Here, we describe a step-by-step protocol for the establishment of PDOs from cells derived from pediatric brain tumors and extracranial solid tumor biopsies and bone marrow aspirates, including tumor processing, organoid culture/subculture, and cryopreservation. Results: Furthermore, we present the use of PDOs for further experimental analysis such as fluorescence imaging, Western blotting, flow cytometry, and immunohistochemistry (IHC) to investigate the underlying pathophysiology of tumorigenesis. Conclusions: Expanding the application of organoids to childhood malignancies holds exceptional promise for elucidating pediatric tumor biology and advancing therapeutic strategies, representing the long-overdue convergence of technology and clinical need. Full article

Tau is a protein associated with microtubules principally expressed in neuronal cells, where it plays a fundamental role in cytoskeleton stabilization and axonal transport. Several diseases collectively named tauopathies, such as Alzheimer’s disease, have been associated with an imbalance in the expression of alternative spliced Tau transcripts and the accumulation of hyperphosphorylated Tau, causing dysfunction and death of neuronal cells. Therefore, understanding the Tau exon splicing mechanisms may contribute to elucidating molecular factors that could underlie the development of neurodegenerative disorders. The aim of this study was to define the role of selected splicing factors in regulating Tau exon expression in cell lines and neuronal organoids. We demonstrated the role of the RNA-binding motif protein 20 (RBM20) splicing factor in regulating Tau exon 6 and exon 10, applying RNA-binding assay and qPCR analyses. Furthermore, we demonstrated that Tau expression was regulated during cerebral organoid differentiation, recapitulating in vivo Tau expression. These results suggest the feasibility of using brain organoid technology to study Tau alternative splicing during neural development, confirming that 3D cellular models could be used to study and characterize pathological processes taking place in Tau-related pathologies. Full article

Peripheral nerves provide a direct connection between the brain and the tumor microenvironment. This connection allows the nervous system to influence processes associated with the development, progression, and metastasis of different tumor types. Therefore, tumor innervation by peripheral nerve fibers is currently emerging as a characteristic that contributes to multiple hallmarks of cancer. Several experimental studies have shown that cancer progression involves actively inducing the ingrowth of autonomic and sensory nerve fibers into tumor tissue. In this process, known as neoaxonogenesis, cancer and other cells in the tumor microenvironment play an important role by synthesizing and releasing neurotrophic factors (e.g., nerve growth factor, brain-derived neurotrophic factor, glial cell line-derived neurotrophic factor), axonal guidance molecules (netrins, semaphorins, ephrins, slits), exosomes (containing microRNA and axonal guidance molecules), and other molecules present in the tumor microenvironment (e.g., granulocyte colony-stimulating factor, leukemia inhibitory factor), which modulate the ingrowth of nerve fibers into the tumor. This results in an increased nerve supply to tumor tissue, which is primarily linked to its growth. However, there are also studies demonstrating the protective effects of increased nerve fiber density against processes associated with cancer progression in certain types of cancer. The findings from these studies contribute to the complexity of neuro-cancer interactions, which is probably based on the type of cancer and the physiological specializations of the nerve fibers in a given organ. Despite contrasting findings, the stimulatory effects of nerve fibers on cancer growth are supported by several studies that described reducing the negative impact of nerve fibers on tumors and thus inhibiting cancer progression. The most significant approaches to reducing neural effects appear to be denervation, the administration of neurotransmitter receptor antagonists, the administration of local anesthetics, and the administration of antibodies against neurotrophic factors. Other significant approaches include methods that improve quality of life, such as psychotherapy and heart rate variability biofeedback. Despite their therapeutic potential, there are several limitations to using approaches that manipulate cancer innervation in clinical practice. These limitations include impaired normal tissue function and nervous system function, as well as the problematic direct application of the therapeutic agent to the tumor site, dosage-dependent, cancer type-dependent, cancer stage-dependent, duration-dependent, and timing-dependent effects. Procedures that modify neoaxonogenesis and nerve fiber signaling appear to be a promising new therapeutic approach in oncology. However, more research is needed to better understand their effects on cancer progression. In the future, the assessment of the presence and density of nerve fibers in tumors, as well as the evaluation of approaches aimed at reducing their negative impact, could be part of personalized anticancer therapy. As part of this therapy, a fresh tumor sample would be collected from the patient to generate patient-derived organoid models to test and consider the possibility of using supportive therapy and to predict its efficacy. Based on these results, it would be possible to evaluate the applicability of nerve-fiber-targeted therapy for a given patient. This review article summarizes and describes the current knowledge concerning the significance of nerve fibers in cancer progression, with a particular emphasis on neoaxonogenesis in tumors and the various factors that influence this process. Full article

Advanced Glycation End Products (AGEs) are reactive compounds formed through the non-enzymatic glycation of proteins, lipids, or nucleic acids due to exposure to reducing sugars. They accumulate through endogenous metabolic dysregulation and exogenous dietary intake, particularly high-fat and high-sugar foods prepared at high temperatures. The interaction between AGEs and their receptor, RAGE (receptor for Advanced Glycation End Products), has been implicated in a range of pathological conditions, including diabetes and metabolic syndrome. However, the impact of AGEs accumulation on neurodevelopment remains poorly understood. In this study, we investigated the effects of AGEs on human-induced pluripotent stem cell (iPSC)-derived cerebral organoids comprising neurons, astrocytes, and microglia. Our findings reveal that AGEs induce RAGE expression, leading to microglial activation, increased deposition of amyloid-beta (Aβ) aggregates, and impaired neurodevelopment. Additionally, elevated levels of AGE-modified proteins, along with altered microglial polarization, were observed in cerebral organoids modeling Western Pacific Amyotrophic Lateral Sclerosis and Parkinsonism–Dementia Complex (ALS-PDC). These findings demonstrate AGEs as active drivers of neurodevelopmental disruption and establish a mechanistic link between metabolic stress and increased susceptibility to neurodegenerative disease. Full article

Brain organoids represent three-dimensional structures that allow for human-specific studies in brain development, pathology and therapeutics. These self-organizing systems, formed through the differentiation of human pluripotent stem cells, can mimic important cellular and molecular events of brain development and therefore serve as a platform for the investigation of neurodevelopmental and neurodegenerative diseases, brain injuries, and tumorigenesis. Although brain organoids show promising perspectives in the study of human physiology, existing brain organoid platforms are hindered by issues of under vascularization, immaturity and protocol variability. Nevertheless, the rapid development of new bioengineering, microfluidic and multi-omics tools and approaches allows us to overcome existing problems and increase the physiological significance of these organoids. Brain organoid transplantation and functional studies further enhance the applications of brain organoids in drug screening, disease modeling and personalized medicine. Here, we provide an overview of recent developments in the field of brain organoid cultures, functional characteristics and translational applications. Full article

Organoid technology is critical for studying human brain development, but existing single-organoid culture systems fail to simulate inter-organ interactions (e.g., optic vesicle–brain) in embryogenesis. This study aimed to establish a more comprehensive in vitro model for early neurodevelopment. We established standardized protocols for cerebral organoids and optic vesicle-containing brain organoids (OVB-organoids), and performed multiscale analyses using immunofluorescence and transcriptomics to compare the two models. Key differences in cellular composition, structure, function and development were found; OVB-organoids better simulated early retinal development, visual-related structures/functions, with specific VSX2 expression and consistent transcriptomic profiles. OVB-organoids are an improved model for early neurodevelopment, providing a reliable basis for exploring eye-brain coordinated development mechanisms and having broad applications in developmental biology, disease research and personalized healthcare. Full article

Alzheimer’s disease is a devastating progressive neurodegenerative disorder characterized by extracellular amyloid-beta plaques and intracellular tau tangles. Despite recent advancements in amyloid-beta-targeting immunotherapies, achieving safe and definitive disease control remains a profound clinical challenge. The CRISPR/Cas9 system has emerged as a powerful technology for precision neurogenetics, offering significant potential to address the fundamental questions behind Alzheimer’s disease. This comprehensive review delineates the trajectory of CRISPR applications in Alzheimer’s disease research and therapeutics. First, we explore the integration of CRISPR in engineering high-fidelity in vitro models, such as isogenic induced pluripotent stem cells and three-dimensional cerebral organoids, alongside advanced in vivo mammalian models. Second, we examine how these platforms facilitate unbiased high-throughput genetic screening to uncover molecular underpinnings regulating tau, lipid metabolism, and neuroinflammation. Third, we critically evaluate precision editing strategies targeting core risk genes (APP, MAPT, APOE, and TREM2), explicitly highlighting the severe physiopathological trade-offs between therapeutic efficacy and loss-of-function toxicity. Finally, we address the ultimate translational bottlenecks impeding clinical application. By dissecting the packaging limits of adeno-associated viral vectors and the physical barricade of the blood–brain barrier, we underscore the necessity of transitioning toward next-generation base editors and non-viral lipid nanoparticles to realize safe and efficacious in vivo clinical gene therapies against Alzheimer’s disease. Full article