Search Results (224)

Parkinson’s disease (PD) is recognized as one of the most common neurodegenerative disorders globally. The primary factor contributing to this condition is the loss of dopaminergic neurons, which results in both motor and nonmotor symptoms. The etiology of neurodegeneration remains unclear. However, it is characterized by the elevated production of reactive oxygen species, which subsequently leads to oxidative stress, lipid peroxidation, mitochondrial dysfunction, and inflammation. The investigation of the applicability of natural compounds and their derivatives to various diseases is becoming increasingly important. The possible role of curcumin from Curcuma longa L. and its derivatives in the treatment of PD has been partially investigated, but there are no data on the action of synthetic cyclic C₅-curcuminoids and chalcones tested in a Parkinson’s model. Two chalcones and five synthetic cyclic C₅-curcuminoids with potential antioxidant properties were investigated in an in vitro model of 6-hydroxydopamine (6-OHDA)-induced neurodegeneration in differentiated SH-SY5Y cells. Reactive oxygen species (ROS) production, total antioxidant capacity, antioxidant enzyme activity, thiol and ATP levels, caspase-3 activity, and cytokine release were examined after treatment with the test compounds. Based on these results, one cyclic chalcone (compound 5) and three synthetic cyclic C₅-curcuminoids (compounds 9, 12, and 13) decreased oxidative stress and apoptosis in our in vitro model of neurodegeneration. Compounds 5 and 9 were also successful in decreasing the production of pro-inflammatory cytokines (IL-6, IL-8, and TNF-α), while promoting the release of anti-inflammatory cytokines (IL-4 and IL-10). These findings indicate that these two compounds exhibit potential antioxidant, anti-apoptotic, and anti-inflammatory properties, rendering them promising candidates for drug development. Full article

Sulforaphane (SFN) is a bioactive compound belonging to the isothiocyanate family, known for its neuroprotective properties. While transcriptomic studies have highlighted SFN’s role in regulating gene expression, its impact on alternative splicing (AS), a key regulatory mechanism in neuronal metabolism, remains underexplored. In this study, we investigated whether SFN pre-treatment influences mRNA splicing patterns in an in vitro neuronal model using retinoic acid (RA)-differentiated SH-SY5Y cells. Using a dedicated RNA-seq-based splicing analysis pipeline, we identified 194 differential alternative splicing events (DASEs) associated with SFN treatment. Gene Ontology enrichment revealed significant over-representation of DNA repair processes. To better understand the functional implications, we integrated in silico predictions of premature stop codons, DASE/miRNA hybridizations, and DASE/RNA-binding protein (RBP) motif occurrences. Our findings suggest that SFN may modulate splicing of key DNA repair genes, contributing to protecting neurons against DNA damage. These preliminary results underscore a novel layer of SFN’s molecular effects and propose it as a valuable adjuvant in physiological conditions to enhance cellular health. Further studies are warranted to dissect the mechanistic underpinnings of SFN-mediated AS and its relevance in DNA-damage-related disorders. Full article

Background: Alzheimer’s disease is a progressive neurodegenerative disorder characterized by cognitive decline and memory loss. Gamma (γ) oscillations are closely linked to learning and memory, and recent interest has grown around Gamma ENtrainment Using Sensory stimulation (GENUS) as a non-invasive neuromodulation strategy. However, the therapeutic impact of vibrotactile gamma stimulation under varying physical parameters such as acceleration remains underexplored. Methods: Differentiated SH-SY5Y cells were treated with amyloid-β (Aβ) and exposed to vibrotactile stimulation at 2.2 or 4.0 m/s². In vivo, male C57BL/6N mice (7 weeks old, 35 g) were administered scopolamine to induce neurotoxicity and randomly assigned to sham, scopolamine, donepezil, or vibrotactile stimulation groups (n = 10 each). Behavioral tests, biochemical assays, Western blotting, and immunohistochemistry were performed to evaluate cognitive function, oxidative stress, cholinergic activity, synaptic plasticity, and neuroinflammation. Results: In vitro, SH-SY5Y cells exposed to amyloid-beta (Aβ) were treated with vibrotactile stimulation, resulting in enhanced neuronal marker expression at 2.2 m/s². In vivo, mice receiving stimulation at 2.2 m/s² showed improved cognitive performance, reduced oxidative stress, restored cholinergic function, suppressed neuroinflammation, and enhanced synaptic plasticity. Mechanistically, these effects were associated with activation of the AKT/GSK3β/β-catenin pathway. Conclusions: Our findings demonstrate that vibrotactile gamma stimulation at 2.2 m/s² exerts greater therapeutic efficacy than higher acceleration, highlighting the importance of optimizing stimulation parameters. This work supports the potential of acceleration-tuned, non-invasive GENUS-based therapies as effective strategies for cognitive recovery in neurodegenerative conditions. Full article

Background: Chemotherapy-induced neuropathic pain is a major side effect of antineoplastic treatment. This study investigated the neuroprotective potential of Acmella oleracea L. extracts containing the N-alkylamide spilanthol, phenolic acids, and glycosylated flavonoids. Methods: Hydroalcoholic extracts of aerial parts (AP) and roots (R) of in vitro seedlings were quali-quantitatively characterized by HPLC-DAD-MS and by ¹H-NMR. Different concentrations (15–150 µg/mL) of AP and R were tested in SH-SY5Y cells differentiated into neurons exposed to oxaliplatin (10 µM), assessing cell viability (MTT), cytotoxicity (LDH), SOD activity, and expression of ATF-3, Ire1α, and Nf-H genes. To evaluate the impact on neuropathic pain, CD-1 mice were treated intraperitoneally with oxaliplatin (2.4 mg/kg), the effect of AP and R extracts (200–1200 mg/kg) were measured by the cold plate test. Results: AP extract was rich in phenols and alkylamides, whereas R extract showed higher phenolic levels but lower alkylamides content. Both extracts significantly reduced mortality and cytotoxicity and counteracted oxidative imbalance by enhancing SOD activity. Gene expression analysis confirmed their neuroprotective effects. In vivo, oxaliplatin induced a 50% reduction in pain threshold, while acute treatment with AP and R extracts dose-dependently alleviated neuropathic pain. Despite the lower spilanthol content in R extract, its efficacy was comparable to AP, suggesting an important role of phenolic compounds. Conclusions: Extracts from both aerial parts and roots of A. oleracea show promise in alleviating chemotherapy-induced neuropathy through mechanisms not solely related to spilanthol. Further studies to elucidate the contribution of phenolic components are desirable. Full article

Cardiovascular disease (CVD) and dementia may share common pathogenic factors such as atherosclerosis and hyperlipoproteinemia. Dyslipidemia-induced oxidative stress contributes to dementia comorbidity in CVD. Abelmoschus esculentus (AE, okra) potentiates in alleviating hyperlipidemia and diabetes-related cognitive impairment. This study evaluated the effects of AE in hyperlipidemic ApoE^−/− mice treated with streptozotocin (50 mg/kg) and fed a high-fat diet (17% lard oil, 1.2% cholesterol). AE fractions F1 or F2 (0.65 mg/kg) were administered for 8 weeks. AE significantly reduced serum LDL-C, HDL-C, triglycerides, and glucose, improved cognitive and memory function, and protected hippocampal neurons. AE also lowered oxidative stress markers (8-hydroxy-2′-deoxyguanosine, 8-OHdG) and modulated neuronal nuclei (NeuN) and doublecortin (DCX) expression. In vitro, AE promoted neurite outgrowth and neuronal differentiation in retinoic acid (RA)-differentiated human SH-SY5Y cells under metabolic stress (glucose and palmitate), alongside the upregulation of heme oxygenase-1 (HO-1), Nuclear factor-erythroid 2-related factor 2 (Nrf2), and brain-derived neurotrophic factor (BDNF). These findings suggest AE may counter cognitive decline via oxidative stress regulation and the enhancement of neuronal differentiation. Full article

Aizoon africanum (L.) Klak (Synonym Galenia africana L.) is traditionally used for a variety of medicinal purposes; however, it has been reported to cause liver damage and severe ascites, particularly in sheep and Angora goats in the arid regions of the Western Cape. This study explores its cytotoxic properties to identify potential cytotoxic compound(s) in the plant. The methanolic extract of A. africanum was re-investigated and subjected to various chromatographic techniques, including preparative HPLC, resulting in the isolation of eight compounds (1–8). Structural elucidation was primarily based on NMR data. Among the isolated compounds, four were flavanones, one was a flavonone, and three were chalcones. Notably, compound 8 was identified as a new chalcone, while compounds 2 and 3 were reported for the first time from this plant. The toxicity of these isolated compounds was evaluated against the HepG2 and SH-SY5Y cancer cell lines using the MTT assay. We further investigated markers of cell death using spectrophotometric and luminometric methods. Among the isolated compounds, 7 and 8 exhibited cytotoxic activities within the range of 3.0–20.0 µg/mL. Notably, the compounds demonstrated greater cytotoxicity towards liver-derived HepG2 cells compared to the neuronal SH-SY5Y cell line. Compound 7 (2′,4′-dihydroxychalcone) was identified as inducing apoptosis through the intrinsic pathway without causing overt necrosis. The findings indicate that the phytochemicals derived from A. africanum exhibit differential cytotoxic effects based on cell type, suggesting potential for developing novel anticancer agents, particularly compound 7. Additionally, the identification of compound 8 provides insight into the liver toxicity of this plant observed in sheep in South Africa. 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

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

Modified oligonucleotides (oligos) are widely used as convenient tools in many scientific fields, including biomedical applications and therapies. In particular, oligos with lipophilic groups attached to the backbone ensure penetration of the cell membrane without the need for transfection. This study examines the interaction between amphiphilic DNA duplexes, in which one of the chains contains a lipophilic substituent, and several DNA repair proteins, particularly DNA-damage-dependent PARPs, using various biochemical approaches. DNA with a lipophilic substituent (LS-DNA) demonstrates more efficient binding with DNA damage activated poly(AD-ribose) polymerases 1-3 (PARP1, PARP2, PARP3) and DNA polymerase β. Chemically reactive LS-DNA derivatives containing a photoactivatable nucleotide (photo-LS-DNAs) or a 5′ deoxyribose phosphate (dRP) group in the vicinity of double-strand breaks (DSBs) are used for the affinity labelling of PARPs and other proteins in several whole-cell extracts of human cells. In particular, photo-LS-DNAs are used to track the level of Ku antigen in the extracts of neuron-like differentiated SH-SY5Y, undifferentiated SH-SY5Y, and olfactory epithelial cells. In vitro, PARP1–PARP3 are shown to be able to slowly excise the 5′ dRP group at DSBs. LS-DNAs can activate PARP1 and PARP2 for autoPARylation, albeit less effectively than regular DNA duplexes. Full article

Lead (Pb) exposure poses a significant public health concern due to its neurotoxic effects. While mitochondrial dysfunction is implicated in lead neurotoxicity, the precise molecular mechanisms, particularly the role of non-coding RNA-mediated competing endogenous RNA networks, remain underexplored. SH-SY5Y neuroblastoma cells were treated with 10 μM lead acetate. Cell viability was assessed by Cell Counting Kit-8 (CCK-8). Mitochondrial ultrastructure and quantity were analyzed via transmission electron microscopy (TEM). Key mitochondrial dynamics proteins were examined by Western blot. Comprehensive transcriptome sequencing, including long non-coding RNAs (lncRNAs), circular RNAs (circRNAs), microRNAs (miRNAs) and mRNAs, was performed followed by functional enrichment and ceRNA network construction. Selected RNAs and hub genes were validated using quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). Lead exposure significantly reduced SH-SY5Y cell viability and induced mitochondrial damage (decreased quantity, swelling, fragmentation). Western blot confirmed an imbalance in mitochondrial dynamics, as indicated by decreased mitofusin 2 (MFN2), increased total and phosphorylated dynamin-related protein 1 (DRP1). Transcriptomic analysis revealed widespread differential expression of lncRNAs, circRNAs, miRNAs, and mRNAs. Enrichment analysis highlighted mitochondrial function and oxidative stress pathways. A ceRNA network identified five key hub genes: SLC7A11, FOS, HMOX1, HGF, and NR4A1. All validated RNA and hub gene expression patterns were consistent with sequencing results. Our study demonstrates that lead exposure significantly impairs mitochondrial quantity and morphology in SH-SY5Y cells, likely via disrupted mitochondrial dynamics. We reveal the potential regulatory mechanisms of lead-induced neurotoxicity involving ceRNA networks, identifying hub genes crucial for cellular stress response. This research provides a foundational framework for developing therapeutic strategies against lead-induced neurotoxicity. Full article

Neuroinflammation is a key process in Alzheimer’s disease (AD). We aimed to examine the development and evaluation of a comprehensive in vitro model that captures the complex interplay between neurons and immune cell types. Retinoic acid-differentiated SH-SY5Y neuroblastoma cells exposed to LPS-conditioned media (CM) from RAW264.7 macrophages, BV2 microglia, and HL60 promyelocytic cells differentiated into neutrophil- or monocyte-like phenotypes were analyzed. The effects of CM containing inflammatory factors on neuronal viability and function were systematically evaluated. Neuronal oxidative stress, mitochondrial function, autophagy and protein aggregates were analyzed. The involvement of insulin resistance was studied by assaying glucose uptake and determining its IC₅₀ values for cell viability improvement and GSK3β phosphorylation. After short-term exposure (3 h), most inflammatory CMs induced peroxide production in neurons, with the strongest effect observed in media from DMSO- or RA-differentiated HL60 cells. Mitochondrial membrane potential was markedly reduced by LPS-stimulated BV2 and HL60-derived CMs. Prolonged exposure (72 h) revealed partial normalization of oxidative stress and mitochondrial membrane potential. Glucose uptake was significantly impaired in cells treated with LPS-activated RAW264.7, BV2, and DMSO-differentiated HL60 cell media, while insulin partially rescued this effect, except for the CM of BV2 cells. Notably, insulin IC₅₀ increased dramatically under LPS-treated BV2 cells induced inflammation (35 vs. 198 pM), confirming the development of insulin resistance. Immune cell-specific inflammation causes distinct effects on neuronal oxidative stress, mitochondrial function, protein aggregation, insulin signaling and viability. LPS-activated BV2-derived CM best recapitulates AD-related pathology, offering a relevant in vitro model for further studies. Full article

Calpain-5/CAPN5 is a calcium-activated, non-lysosomal cysteine (thiol) protease. The substrate repertoire of CAPN5 is not known. Calpains catalyze limited proteolysis of their substrates, generating neo-N-termini that correspond to internal residues of their nascent substrate proteins. To identify such neo-N-termini generated by CAPN5, we employed an N-terminomics approach called TAILS (Terminal amine isotopic labeling of substrates) to quantitatively compare the N-terminal peptides detected in parental and CAPN5-deficient SH-SY5Y neuroblastoma cells. Thirty neo-N-termini corresponding to 29 protein groups and 24 unique proteins were detected to be depleted in the CAPN5^−/− cells. A subset of the identified putative substrates was further studied with CAPN5 co-immunoprecipitation, in vitro calcium-induced CAPN5 proteolysis assay, and their cellular fragmentation patterns were compared in parental and CAPN5-deficient SH-SY5Y cells. Here, we provide evidence for CAPN5-mediated proteolysis of the synaptic proteins DLGAP4, IQSEC1 and MPDZ, the neurodegeneration-related EWS, hnRNPU, TFG and UGP2, the DNA replication regulator MCM3, and the neuronal differentiation regulator LMTK1. Our data provide new relevance for neovascular inflammatory vitreoretinopathy (NIV), a progressive eye disease caused by pathogenic mutations in CAPN5. Data are available via ProteomeXchange with identifier PXD064313. Full article

The nervous system maintains homeostasis and coordinated behavior through complex neuronal and glial cells. Traditional models, such as primary rodent neurons and human-induced pluripotent stem cell (hIPSC)-derived neurons, have advanced our understanding of neuronal function and neurotoxic damage; however, they are costly and labor-intensive. SH-SY5Y cells, an immortalized human neuroblastoma cell line, provide a more accessible alternative for studying neuronal processes and neurotoxicity. However, their limited capacity to differentiate into specific neuronal phenotypes remains a challenge. To address this limitation, differentiation protocols using neuronal factors and vitamins have been developed, primarily in two-dimensional (2D) cultures, which reduces physiological relevance. Here, we present a novel three-dimensional (3D) SH-SY5Y model incorporating 2D differentiation protocols to generate cholinergic neurons (ChAT+). This model enhances neurotoxicity studies related to pesticides and mycotoxins. Our protocol produces homogeneous spheroids differentiated into cholinergic neurons using serum restriction and specific factors, maintaining viability and circularity for up to 22 days. Differentiation was validated by immunofluorescence and Western blot by Choline acetyltransferase (ChAT) expression. This scalable and reproducible 3D model provides a valuable in vitro tool for neurotoxicological research, improving physiological relevance and enabling the study of cholinergic neuron differentiation and function. Full article

This study investigated the potential neuroprotective mechanisms of porcine brain enzyme hydrolysate (PBEH) against Alzheimer’s disease pathology using differentiated SH-SY5Y cells. Differentiated neuronal cells were treated with 40 μM amyloid-β(1-42; Aβ) to induce neurotoxicity, followed by PBEH treatment (12.5–400 μg/mL), Com-A (peptide-based neuroprotective supplement; 200 μg/mL) treatment, and Com-B (herbal extract known for improving memory function; 100 μg/mL) treatment. Key assessments included cell viability, Aβ aggregation in adding 10 μM Aβ, amyloidogenic proteins (APP, BACE), synaptic markers (BDNF, ERK), apoptotic markers (BAX/BCL-2, caspase-3), oxidative stress (reactive oxygen species (ROS)), cholinergic function (ChAT, AChE), MAPK signaling (JNK, p38), and neuroinflammation (IL-1β). PBEH contained high concentrations of amino acids, including L-lysine (32.3 mg/g), L-leucine (42.4 mg/g), L-phenylalanine (30.0 mg/g) and the PSIS peptide (86.9 μg/g). Treatment up to 400 μg/mL showed no cytotoxicity and had cognitive protection effects up to 152% under Aβ stress (p < 0.05). PBEH significantly attenuated Aβ aggregation, decreased APP (28%) and BACE (51%) expression, enhanced synaptic function through increased BDNF, and restored ERK phosphorylation (p < 0.05). Anti-apoptotic effects included a 76% reduction in the BAX/BCL-2 ratio, a 47% decrease in caspase-3, and a 56% reduction in ROS levels. Cholinergic function showed restoration via increased ChAT activity (p < 0.01) and decreased AChE activity (p < 0.05). PBEH reduced IL-1β levels by 70% and suppressed JNK/p38 phosphorylation (p < 0.05). While Com-A enhanced BDNF and Com-B showed anti-inflammatory effects, PBEH demonstrated activity across multiple pathway markers. In conclusion, these findings suggest that PBEH may enable neuronal preservation through multi-pathway modulation, establishing foundational evidence for further mechanistic investigation in cognitive enhancement applications. Full article

Seliciclib, a cyclin-dependent kinase 5 (CDK5) inhibitor, has demonstrated neuroprotective potential. However, its therapeutic application is limited by poor permeability across the blood–brain barrier (BBB). In this study, polymeric nanoparticles (NPs) modified with a BBB-targeting peptide ligand (His-Ala-Ile-Tyr-Pro-Arg-His) were employed to encapsulate seliciclib. In vitro transport studies showed that the peptide-modified NPs exhibited significantly greater translocation across a bEnd.3 cell monolayer compared to unmodified NPs. Furthermore, in vivo biodistribution analysis revealed that the brain accumulation of peptide-modified NPs was 3.38-fold higher than that of unmodified NPs. Notably, the peptide-conjugated, seliciclib-loaded NPs demonstrated a significant neuroprotective effect against the neurotoxin 1-methyl-4-phenylpyridinium (MPP⁺) in differentiated SH-SY5Y cells. Full article