Search Results (1,549)

Osteosarcopenia is a widespread geriatric condition resulting from the coexistence of osteoporosis and sarcopenia, where the connection between bone and muscle is, in part, driven by bone–muscle crosstalk. Given the close, reciprocal influence of muscle on nerve, and vice versa, it is not surprising that there are corresponding aging changes in the biochemistry and morphology of the neuromuscular junction (NMJ). Indeed, degeneration of motor neurons and progressive disruption of the neuromuscular connectivity were observed in old age. Extracellular vesicles (EVs) derived from human amniotic fluid stem cells (hAFSC), exhibiting antioxidant properties, which can also explain their anti-aging and cytoprotective effects, can be considered as potential treatment for age-related diseases. To study cell interactions under both healthy and pathological conditions occurring in musculo–skeletal apparatus, we developed a three-culture system exploiting the use of well-known transwell supports. This system allows both myotubes and neurons, eventually treated with EVs, and osteoblasts, induced to osteoporosis, to interact physically and biochemically. Collectively, this method allowed us to understand how the modifications induced in osteoblasts during bone disorders trigger a cascade of detrimental effects in the muscle and neuron parts. Moreover, we demonstrated the efficacy of hAFSC-EVs in preventing NMJ dysfunction, muscle atrophy, and osteoblast impairment. Full article

Rafiq syndrome (RAFQS) is a rare autosomal recessive disorder that is classified as a type II congenital disorder of glycosylation (CDG-II), and caused by MAN1B1 gene mutation. To date, 24 pathogenic MAN1B1 mutations have been reported in association with MAN1B1-CDG. However, the underlying pathogenic mechanisms remain poorly understood. In this study, we recruited a consanguineous family from Pakistan with multiple affected individuals exhibiting mild facial dysmorphism, developmental delay, and intellectual disability. Utilizing exome sequencing and homozygosity mapping, we identified a novel MAN1B1 mutation (c.772_775del) that co-segregated with RAFQS in this family. Analysis of public single-cell transcriptomic data revealed that MAN1B1 is predominantly expressed in dorsal progenitors and intermediate excitatory neurons during human brain development. Knockdown of Man1b1 in primarily cultured mouse excitatory neurons disrupted axon growth, dendrite formation, and spine maturation, and could not be rescued by truncated variants identified in the family. Furthermore, in utero, electroporation experiments revealed that Man1b1 knockdown in the murine cortex impaired neural stem cells’ proliferation and differentiation, as well as cortical neuron migration. Collectively, these findings elucidate a critical role for MAN1B1 in the etiology of RAFQS and demonstrate that loss-of-function mutation in MAN1B1 disrupt neuro-developmental processes, providing mechanistic insights into the pathogenesis of this disorder. Full article

5-HT4 receptors play an important role in the regulation of synaptic plasticity. However, the effect of 5-HT4Rs on neural network activity and intercellular calcium signaling remains enigmatic. Using calcium imaging and original software, we determined the network-level characteristics of calcium dynamics within primary hippocampal cultures. We found that the single activation of 5-HT4 receptors by BIMU8 significantly reduced the correlation of activity within neuron–glial networks of primary cultures, without altering the proportion of active cells or the frequency of calcium events. In contrast, chronic stimulation of 5-HT4Rs promoted greater cell involvement in Ca²⁺ signal generation and increased the frequency of calcium events, while maintaining the connectivity level of the neuron–glial network. Moreover, our immunocytochemical labeling results indicated that chronic stimulation of 5-HT4Rs increased the size of both presynaptic and postsynaptic terminals. The acute blockade of 5-HT4Rs by RS23597-190 exerted a marked inhibitory effect on calcium activity in primary hippocampal cultures. Network connectivity and correlation of calcium activity were disrupted, and the number of functional connections among cells sharply declined. Our study showed that 5-HT4 receptors exhibit diverse effects based on the type and duration of activation, mediating several key functions in regulating neural network calcium activity. 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

The rhythm of epileptiform activity occurs in various brain injuries (ischemia, stroke, concussion, mechanical damage, AD, PD). The epileptiform rhythm is accompanied by periodic Ca²⁺ pulses, which are necessary for the neurotransmitter release, the repair of damaged connections between neurons, and the growth of new projections. The duration and amplitude of these pulses depend on intracellular calcium-binding proteins. The effect of the synthetic fast calcium buffer BAPTA on the epileptiform activity of neurons induced by the GABA(A)-receptor inhibitor, bicuculline, was investigated in a 14-DIV rat hippocampal culture. In the epileptiform activity mode, neurons periodically synchronously generate action potential (AP) bursts in the form of paroxysmal depolarization shift (PDS) clusters and their corresponding high-amplitude Ca²⁺ pulses. Changes in the paroxysmal activity and Ca²⁺ pulses were recorded continuously for 10–11 min as BAPTA accumulated. It was shown that during BAPTA accumulation, transformation of neuronal patch activity occurs. Moreover, GABAergic and glutamatergic neurons respond differently to the presence of calcium buffer. Experiments were performed on two populations of neurons: a population of GABAergic neurons that responded selectively to ATPA, a calcium-permeable GluK1 kainate receptor agonist, and a population of glutamatergic neurons with a large amplitude of cluster depolarization (greater than −20 mV). These neurons made up the majority of neurons. In the population of GABAergic neurons, during BAPTA accumulation, the amplitude of PDS clusters decreases, which leads to a switch from the PDS mode to the classical burst mode with an increase in the electrical activity of the neuron. In glutamatergic neurons, the duration of PDS clusters decreased during BAPTA accumulation. However, the amplitude changed little. The data obtained showed that endogenous calcium-binding proteins play a significant role in switching the epileptiform rhythm to the recovery rhythm and perform a neuroprotective function by reducing the duration of impulses in excitatory neurons and the amplitude of impulses in inhibitory neurons. Full article

Forty per cent of major depression patients are resistant to antidepressant medication. Thus, it is necessary to search for alternative treatments. Melatonin (N-acetyl-5-hydroxytryptamine) enhances neurogenesis and neuronal survival in the adult mouse hippocampal dentate gyrus. Additionally, melatonin stimulates the activity of Ca²⁺/Calmodulin-dependent Kinase II (CaMKII), promoting dendrite formation and neurogenic processes in human olfactory neuronal precursors and rat organotypic cultures. Similarly, ketamine, an N-methyl-D-aspartate receptor (NMDAR) antagonist, modulates CaMKII activity. Importantly, co-treatment of low doses of ketamine (10⁻⁷ M) in combination with melatonin (10⁻⁷ M) produces additive effects on neurogenic responses in olfactory neuronal precursors. Importantly, enhanced neurogenic responses are produced by conventional antidepressants like ISSRs. The goal of this study was to investigate whether hippocampal CaMKII participates in the signaling pathway elicited by combining doses of melatonin with ketamine acutely administered to mice, 30 min before being subjected to the forced swimming test. The results showed that melatonin, in conjunction with ketamine, significantly enhances CaMKII activation and changes its subcellular distribution in the dentate gyrus of the hippocampus. Remarkably, melatonin causes nuclear translocation of the active form of CaMKII. Luzindole, a non-selective MT₁ and MT₂ receptor antagonist, abolished these effects, suggesting that CaMKII is downstream of the melatonin receptor pathway that causes the antidepressant-like effects. These findings provide molecular insights into the combined effects of melatonin and ketamine on neuronal plasticity-related signaling pathways and pave the way for combating depression using combination therapy. Full article

Background/Objectives: Epilepsy is a brain condition that affects millions of people worldwide. Although there are many antiepileptic drugs with different mechanisms of action, many patients still fail to control their agonizing symptoms, a situation that highlights the need for more strategies to address this issue. In this in vitro study, we elucidated and characterized the alterations in intracellular Ca²⁺ levels in cell cultures where diazepam and repetitive transcranial magnetic stimulation were implemented, alone or in combination. Methods: Using the differentiated human-derived neuroblastoma cell line SH-SY5Y, we measured the alterations in intracellular Ca²⁺ levels under the impact of either low-frequency repetitive transcranial magnetic stimulation (1 Hz), diazepam (14 μM), or their combination. We used the Ca²⁺-sensitive fluorescent indicator Fluo-4 acetoxymethyl ester for calcium imaging, while neuronal excitation was achieved with 50 mM KCl. Results: The highest median fluorescence intensity increase (%ΔF/F = 24.80) was observed in control cell cultures, followed by rTMS cultures (%ΔF/F = 16.96) and diazepam cultures (%ΔF/F = 11.46). The lowest median fluorescence intensity value (%ΔF/F =−0.44) was observed when diazepam was used concomitantly with repetitive transcranial magnetic stimulation. Post hoc analysis assessed pairwise differences, showing statistically significant differentiation between the control group and all other groups. Additionally, statistically significant results were observed between repetitive transcranial magnetic stimulation or diazepam and their combination, but not between them. Conclusions: The combination of diazepam and repetitive transcranial magnetic stimulation resulted in the most significant reduction in intracellular Ca²⁺ levels, as indicated by the lowest fluorescence values compared with the control group. Individually, each treatment produced a notable but less pronounced effect. We conclude that both diazepam and low-frequency repetitive transcranial magnetic stimulation can control epileptiform activity in vitro, while their combination is the most effective treatment. Full article

The thalamus is an important sensory relay station. It integrates all somatic sensory pathways (excluding olfaction) and transmits information through thalamic relay neurons before projecting to the cerebral cortex via thalamocortical axons (TCAs). Emerging evidence has shown that FGF3, a member of the morphogen family, is an axon guidance molecule that repels TCAs away from the hypothalamus and into the internal capsule so that they subsequently reach different regions of the cortex. However, current studies on FGF-mediated axon guidance predominantly focus on phenomenological observations, with limited exploration of the underlying molecular mechanisms. To address this gap, we investigated both direct and indirect downstream signaling pathways mediating FGF3-dependent chemorepulsion of TCAs at later developmental stages. Firstly, we used pharmacological inhibitors to identify the signaling cascade(s) responsible for FGF3-triggered direct chemorepulsion of TCAs, in vitro and in vivo. Our results demonstrate that the PC-PLC pathway is required for FGF3 to directly stimulate the asymmetrical repellent growth of developing TCAs. Then, we found the FGF3-mediated repulsion can be indirectly induced by Slit1 because the addition of FGF3 in the culture media induced an increase in Slit1 expression in the diencephalon. Furthermore, by using downstream inhibitors, we found that the indirect repulsive effect of FGF3 is mediated through the PI3K downstream pathway of FGFR1. Full article

Oxidative stress, driven by impaired antioxidant defence systems, is a major contributor to cognitive decline and neurodegenerative processes in brain ageing. This study investigates the neuroprotective effects of a natural compound mixture—composed of Hericium erinaceus, Palmitoylethanolamide, Bilberry extract, and Centella asiatica—using a multi-step in vitro strategy. An initial evaluation in a 3D intestinal epithelial model demonstrated that the formulation preserves barrier integrity and may be bioaccessible, as evidenced by transepithelial electrical resistance (TEER) and the expression of tight junctions. Subsequent analysis in an integrated gut–brain axis model under oxidative stress conditions revealed that the formulation significantly reduces inflammatory markers (NF-κB, TNF-α, IL-1β, and IL-6; about 1.5-fold vs. H₂O₂), reactive oxygen species (about 2-fold vs. H₂O₂), and nitric oxide levels (about 1.2-fold vs. H₂O₂). Additionally, it enhances mitochondrial activity while also improving antioxidant responses. In a co-culture of neuronal and astrocytic cells, the combination upregulates neurotrophic factors such as BDNF and NGF (about 2.3-fold and 1.9-fold vs. H₂O₂). Crucially, the formulation also modulates key biomarkers associated with cognitive decline, reducing APP and phosphorylated tau levels (about 98% and 1.6-fold vs. H₂O₂) while increasing Sirtuin 1 and Nrf2 expression (about 3.6-fold and 3-fold vs. H₂O₂). These findings suggest that this nutraceutical combination may support the cellular pathways involved in neuronal resilience and healthy brain ageing, offering potential as a functional food ingredient or dietary supplement. Full article

As the global population continues to age, the incidence of neurodegenerative diseases and neural injuries is increasing, presenting major challenges for healthcare systems. Due to the brain’s limited regenerative capacity, there is an urgent need for strategies that promote neuronal repair and functional integration. Brain-derived neurotrophic factor (BDNF) is a key regulator of synaptic plasticity and neuronal development. In this study, we investigated whether constitutive BDNF expression in human induced pluripotent stem cell (iPSC)-derived neural progenitor cells (NPCs) enhances their neurogenic and integrative potential in vitro. We found that NPCs engineered to overexpress BDNF produced neuronal cultures with increased numbers of mature and spontaneously active neurons, without altering the overall structure or organization of functional networks. Furthermore, BDNF-expressing neurons exhibited significantly greater axonal outgrowth, including directed axon extension in a compartmentalized microfluidic system, suggesting a chemoattractive effect of localized BDNF secretion. These effects were comparable to those observed with the early supplementation of recombinant BDNF. Our results demonstrate that sustained BDNF expression enhances neuronal maturation and axonal projection without disrupting network integrity. These findings support the use of BDNF not only as a therapeutic agent to improve cell therapy outcomes but also as a tool to accelerate the development of functional neural networks in vitro. Full article

Parishin C (PaC) is an active ingredient in Gastrodia elata Bl. that has neuroprotective effects. However, research on its role in oxidative stress and neuroinflammation is still limited. This study used LPS–stimulated HT22 cells to investigate the antioxidant properties of PaC. Through the co–culture system of HT22 and BV2 cells, the effect of PaC on neuroinflammation was explored. The current results indicated that PaC can inhibit the levels of reactive oxygen species and peroxides in LPS–stimulated HT22 cells and increase the levels of antioxidant factors. Meanwhile, PaC can also inhibit neuronal ferroptosis and the levels of pro–inflammatory cytokines in BV2 cells. Importantly, the antioxidant and anti–inflammatory effects of PaC are achieved by activating the Nrf2 signaling pathway. The WB and IF results indicated that PaC can promote nuclear translocation of Nrf2, activate downstream antioxidant factors, and thereby regulate inflammatory responses. Inhibition of Nrf2 can significantly inhibit the regulation of PaC on the Nrf2 signaling pathway. These results indicated that PaC can activate the Nrf2 signaling pathway to inhibit oxidative stress and inflammation. Full article

Background: Accelerated demographic aging in Hungary and across Europe presents significant public health and socioeconomic challenges, particularly in preserving cognitive function and preventing neurodegenerative diseases. Modifiable lifestyle factors—especially dietary habits—play a critical role in brain aging and cognitive decline. Objective: This narrative review explores the mechanisms by which Western dietary patterns contribute to cognitive impairment and neurovascular aging, with specific attention to their relevance in the Hungarian context. It also outlines the rationale and design of the Semmelweis Study and its workplace-based health promotion program targeting lifestyle-related risk factors. Methods: A review of peer-reviewed literature was conducted focusing on Western diet, cognitive decline, cerebrovascular health, and dietary interventions. Emphasis was placed on mechanistic pathways involving systemic inflammation, oxidative stress, endothelial dysfunction, and decreased neurotrophic support. Key findings: Western dietary patterns—characterized by high intakes of saturated fats, refined sugars, ultra-processed foods, and linoleic acid—are associated with elevated levels of 4-hydroxynonenal (4-HNE), a lipid peroxidation product linked to neuronal injury and accelerated cognitive aging. In contrast, adherence to Mediterranean dietary patterns—particularly those rich in polyphenols from extra virgin olive oil and moderate red wine consumption—supports neurovascular integrity and promotes brain-derived neurotrophic factor (BDNF) and nerve growth factor (NGF) activity. The concept of “cognitive frailty” is introduced as a modifiable, intermediate state between healthy aging and dementia. Application: The Semmelweis Study is a prospective cohort study involving employees of Semmelweis University aged ≥25 years, collecting longitudinal data on dietary, psychosocial, and metabolic determinants of aging. The Semmelweis–EUniWell Workplace Health Promotion Model translates these findings into practical interventions targeting diet, physical activity, and cardiovascular risk factors in the workplace setting. Conclusions: Improving our understanding of the diet–brain health relationship through population-specific longitudinal research is crucial for developing culturally tailored preventive strategies. The Semmelweis Study offers a scalable, evidence-based model for reducing cognitive decline and supporting healthy aging across diverse populations. Full article

Heparan sulfate proteoglycans (HSPGs) within the neuronal niche are expressed during brain development, contributing to multiple aspects of neurogenesis, yet their roles in glial lineage commitment remain elusive. This study utilised three human cell models expanded under basal culture conditions followed by media-induced lineage induction to identify a reproducible and robust model of gliogenesis. SH-SY5Y human neuroblastoma cells (neuronal control), ReNcell CX human neural progenitor cells (astrocyte inductive) and ReNcell VM human neural progenitor (mixed neural induction) models were examined. The cultures were characterised during basal and inductive states via Q-PCR, Western Blotting, immunocytochemistry (ICC) and calcium signalling activity analyses. While the ReNcell lines did not produce fully mature or homogeneous astrocyte cultures, the ReNcell CX cultures most closely resembled an astrocytic phenotype with ReNcell VM cells treated with platelet-derived growth factor (PDGF) biased toward an oligodendrocyte lineage. The glycated variant of surface-bound glypican-2 (GPC2) was found to be associated with lineage commitment, with GPC6 and 6-O HS sulfation upregulated in astrocyte lineage cultures. Syndecan-3 (SDC3) emerged as a lineage-sensitive proteoglycan, with its cytoplasmic domain enriched in progenitor-like states and lost upon differentiation, supporting a role in maintaining neural plasticity. Conversely, the persistence of transmembrane-bound SDC3 in astrocyte cultures suggest continued involvement in extracellular signalling and proteoglycan secretion, demonstrated by increased membrane-bound HS aggregates. This data supports HSPGs and HS GAGs as human neural lineage differentiation and specification markers that may enable better isolation of human neural lineage-specific cell populations and improve our understanding of human neurogenesis. Full article

Fumonisin B1, deoxynivalenol (DON), and zearalenone (ZEA) are toxic secondary metabolites produced by Fusarium molds. These mycotoxins are common food and feed pollutants and represent a risk to human and animal health. Although the mycotoxins produced by this genus can cross the blood–brain barrier in many species, their effect on neuronal function remains unclear. We investigated the cell viability effects of these toxins on specified neural cell types, including mouse primary neuronal, astroglial, and mixed-cell cultures 24 or 48 h after mycotoxin administration. DON decreased cell viability in a dose-dependent manner, independent of the culture type. Fumonisin B1 was toxic in pure neuronal cultures only at high doses, but toxicity was attenuated in mixed and pure astroglial cultures. ZEA had significant effects on all culture types in 10 nM by increasing cell viability and network activity, as revealed by multi-electrode array measurements. Since ZEA is a mycoestrogen, we analyzed the effects of ZEA on the expression of estrogen receptor isotypes ERα and ERβ and the mitochondrial voltage-dependent anion channel via qRT-PCR. In neuronal and mixed cultures, ZEA administration decreased ERα expression, while in astroglial cultures, it induced the opposite effect. Thus, our results emphasize that Fusarium mycotoxins act in a cell-specific manner. Full article

In familial Alzheimer’s disease (FAD), presenilin 1 (PSEN1) E280A cholinergic-like neurons (ChLNs) induce aberrant secretion of extracellular amyloid beta (eAβ). How PSEN1 E280A ChLNs-eAβ affects microglial activity is still unknown. We obtained induced microglia-like cells (iMG) from human peripheral blood cells (hPBCs) in a 15-day differentiation process to investigate the effect of bolus addition of Aβ42, PSEN1 E280A cholinergic-like neuron (ChLN)-derived culture supernatants, and PSEN1 E280A ChLNs on wild type (WT) iMG, PSEN1 E280A iMG, and sporadic Alzheimer’s disease (SAD) iMG. We found that WT iMG cells, when challenged with non-cellular (e.g., lipopolysaccharide, LPS) or cellular (e.g., Aβ42, PSEN1 E280A ChLN-derived culture supernatants) microenvironments, closely resemble primary human microglia in terms of morphology (resembling an “amoeboid-like phenotype”), expression of surface markers (Ionized calcium-binding adapter molecule 1, IBA-1; transmembrane protein 119, TMEM119), phagocytic ability (high pHrodo™ Red E. coli BioParticles™ phagocytic activity), immune metabolism (i.e., high generation of reactive oxygen species, ROS), increase in mitochondrial membrane potential (ΔΨm), response to ATP-induced transient intracellular Ca²⁺ influx, cell polarization (cluster of differentiation 68 (CD68)/CD206 ratio: M1 phenotype), cell migration activity according to the scratch wound assay, and especially in their inflammatory response (secretion of cytokine interleukin-6, IL-6; Tumor necrosis factor alpha, TNF-α). We also found that PSEN1 E280A and SAD iMG are physiologically unresponsive to ATP-induced Ca²⁺ influx, have reduced phagocytic activity, and diminished expression of Triggering Receptor Expressed on Myeloid Cells 2 (TREM2) protein, but when co-cultured with PSEN1 E280A ChLNs, iMG shows an increase in pro-inflammatory phenotype (M1) and secretes high levels of cytokines IL-6 and TNF-α. As a result, PSEN1 E280A and SAD iMG induce apoptosis in PSEN1 E280A ChLNs as evidenced by abnormal phosphorylation of protein TAU at residue T205 and cleaved caspase 3 (CC3). Taken together, these results suggest that PSEN1 E280A ChLNs initiate a vicious cycle between damaged neurons and M1 phenotype microglia, resulting in excessive ChLN death. Our findings provide a suitable platform for the exploration of novel therapeutic approaches for the fight against FAD. Full article