Human Neuroblastoma SH-SY5Y Cells Research Articles

The impact of Aureobasidium melanogenum cells and extracellular vesicles on human cell lines

Aureobasidium melanogenum is a black yeast-like fungus that occurs frequently both in nature and in domestic environments. It is becoming increasingly important as an opportunistic pathogen. Nevertheless, its effect on human cells has not yet been studied. In this study, we investigated the effect of A. melanogenum cells and extracellular vesicles (EVs) on human cell lines A549 (human lung cells), HDFa (human dermal fibroblasts), and SH-SY5Y (human neuroblastoma cells). Scanning electron microscopy (SEM) showed no direct interaction between A. melanogenum cells and human cell lines, but there were some changes in HDFa cells. As a possible cause for this change, we tested the cytotoxic effect of EVs from A. melanogenum on the same cell lines. We isolated EVs from the fungus and prepared three different pools: a non-melanin pool (containing mainly EVs), a melanin pool (containing mainly melanin nanoparticles), and a total pool (containing both EVs and melanin nanoparticles). All three pools were characterized and then added to human cell lines to test their cytotoxicity. Unlike in some other fungal opportunistic pathogens, no effects of fungal EVs on human cell viability were observed. Therefore, the opportunistic potential of A. melanogenum remains only partially understood.

Investigating the neuroprotective properties of Sargassum wightii extract against MPP+-induced apoptosis in SH-SY5Y human neuroblastoma cells.

This study aims to assess the neuroprotective effects of the methanolic extract of Sargassum wightii against oxidative stress and cell death induced by neurotoxins MPP + in SH-SY5Y cells. Briefly, the methanolic extract of S.wightii decreased the cytotoxicity of MPP + in SH-SY5Y cells. Treatment with S.wightii extract at a concentration of 400µg/ml resulted in a notable decrease in cell death, particularly in MPP + -induced cells. Flow cytometry analysis with annexin V/PI staining reveals apoptosis and necrosis in SH-SY5Y cell lines upon exposure to 1mM of MPP + . However, 100-400µg/ml concentrations of S.wightii extract effectively decrease apoptosis in SH-SY5Y cells. Furthermore, S.wightii inhibits caspase-3 activity, effectively shielding neuronal cells against MPP + -induced cell death. Mitochondrial membrane potential (MMP) assay using a JC-1 fluorescent probe indicates that the methanolic extract of S.wightii exhibits protective effects against MPP + -induced cell death and maintains mitochondrial membrane potential. Our results conclude that exposing SH-SY5Y cells to a methanolic extract of S.wightii could potentially increase the likelihood of inhibiting the cascade mechanism, stopping MPP+-induced apoptosis, and preventing the rupture of the mitochondrial membrane. However, the lack of low solubility and poor bioavailability reduce the therapeutic efficacy of S.wightii. Liposome-based drug delivery systems can improve the bioavailability and stability of bioactive compounds, enhancing their therapeutic potential. Hence, S.wightii may hold promise as an innovative treatment for neurological ailments.

Cytoprotective effect of melatonin against MPP+ toxicity in SH-SY5Y cells: Role sharing of two types of antioxidative activities of melatonin

Melatonin is a neurohormone that is not only a regulator of circadian cycles, but also a potent antioxidant. Parkinson’s disease (PD) is a major neurodegenerative disease that may result from oxidative stress as a part of its pathogenic cascade. Therefore, antioxidants, including melatonin, have attracted attention as potential candidates for neuroprotection against PD-related neurotoxicity. In this study, we report that melatonin has 2 types of antioxidant mechanisms of neuroprotection in an experimental cellular PD model using 1-Methyl-4-phenylpyridinium ion (MPP+) in human neuroblastoma SH-SY5Y cells. The first mechanism is a classical antioxidative mechanism through the direct action of melatonin, which reduces lipid hydroperoxide and 8-OHdG. The second mechanism is an indirect antioxidative effect via the melatonin receptor (Mel-R)/PI3K/Akt/Nrf2 cascade. Melatonin and Mel-R agonist activated PI3K/Akt signaling and Nrf2. Both Mel-R antagonist and the PI3K inhibitor blocked transcription induced by Nrf2 and the cytoprotective effect of melatonin. Interestingly, the antioxidative effect due to the Nrf2-related mechanism contributed mainly to a decrease of protein carbonyl, but not to lipid hydroperoxide and 8-OHdG. Mel-R agonist also showed a similar effect. Our results elucidate the mechanism of melatonin's powerful antioxidative effect and suggest the application of melatonin therapy to PD-related cytotoxicity.

Bioactive compounds from Crataegus pinnatifida Bge. leaves: potential health benefits.

The leaves of Crataegus pinnatifida Bge., a medicinal plant and a source of brewed tea, are often overlooked as a byproduct of fruit consumption. Despite this, these leaves have demonstrated significant cardiovascular protective and lipid-lowering properties. To further investigate their potential applications, nine novel compounds (1-9) and twelve known flavonoids were isolated from C. pinnatifida leaves. Their structures were elucidated through comprehensive spectroscopic analysis, including experimental and calculated electronic circular dichroism. The antioxidant capacities of these 21 compounds were assessed using DPPH˙ and ABTS˙+ assays. Notably, the newly identified biphenyl compounds 7 and 8 exhibited potent antioxidant activities, surpassing that of vitamin C in both assays. Additionally, the ABTS˙+ assay revealed that flavonoid O-glycosides generally possess stronger antioxidant activity compared to flavonoid C-glycosides. To evaluate their cytoprotective potential, compounds 12-14 and 18-20 were tested against H2O2-induced neurotoxicity in human neuroblastoma SH-SY5Y cells. At 100 μM, these compounds demonstrated significant protective effects. Furthermore, flavonoid C-glycosides exhibited stronger protective activity against alcohol-induced injury in BRL-3A hepatocytes than flavonoid O-glycosides. In conclusion, this study provides compelling evidence that C. pinnatifida leaves contain valuable bioactive compounds with potent antioxidant and cytoprotective properties. These findings further validate the traditional use of this plant and offer promising avenues for future research and potential therapeutic applications.

Static Magnetic Field Exposure Causes Small Cell Cycle Disruptions and Changes in Reactive Oxygen Species Levels in Ionizing Radiation Exposed Human Neuroblastoma Cells.

Although static magnetic fields (SMFs) have been reported to induce only minimal biological effects, it has been proposed that they may alter the effects of other agents, such as ionizing radiation. We sham-exposed or exposed human SH-SY5Y neuroblastoma cells to 0.5-, 1.5-, 2.5-, or 3.5-mT SMFs for 24 h either before or after irradiation at 0, 0.4 or 2.0 Gy. After the exposures, cell cycle distribution (subG1 for apoptosis), reactive oxygen species (ROS) levels, caspase-3 activity, and clonogenic survival were assayed. Increase of G0/G1 and decrease of S phase cells was observed in samples exposed to a 3.5-mT SMF after irradiation. The same exposure schedule with a 1.5-mT SMF was associated with an increase of S phase cells, and an increase in ROS levels. Conversely, a decrease in ROS levels was observed in cells exposed to a 2.5-mT SMF before ionizing radiation. No cell cycle changes were observed with SMF exposures before irradiation. Caspase-3 activity or clonogenic survival was not affected by SMF exposures, irrespective of the exposure schedule. In conclusion, small changes in cell cycle distribution and ROS levels were observed in SH-SY5Y cells exposed to SMFs, with more prominent effects observed when SMF exposure was applied after irradiation. Our results suggest that SMF-induced effects show no linear dependency on magnetic flux density below 5 mT. Notably, SMF exposures did not significantly potentiate the effects of ionizing radiation but rather caused an independent additive effect. Bioelectromagnetics. 00:00-00, 2024.

Effects of Pelargonium Sidoides Extract on Apoptosis and Oxidative Stress in Human Neuroblastoma Cells.

Background and Objectives: Neuroblastoma is the most common extracranial solid tumor in children, often presenting challenges in treatment due to its clinical and genetic heterogeneity. This study investigated the anticancer potential of Pelargonium sidoides root extract on the human neuroblastoma cell line (SH-SY5Y). Using XTT assays, ELISA-based oxidative stress markers, and RT-PCR analysis of apoptotic genes, the study explored the extract's effects on cell proliferation, oxidative stress, and apoptosis. Materials and Methods: For the cell culture, SH-SY5Y human neuroblastoma cells were thawed, cultured, and maintained under appropriate conditions for experiments. The dose- and time-dependent activity of Pelorgonium sidoides extract on SH-SY5Y neuroblastoma cells was investigated by XTT assay. The change in the oxidative stress marker 8-Hydroxy-2'-deoxyguanosine (8-OhDG) level was determined by ELISA for the doses applied to the control group root extract at a concentration of 25 μg/mL. Total antioxidant status (TAS) and total oxidant status (TOS) were measured from the cells in the study group with the help of a commercial kit. The oxidative stress index (OSI) was calculated by dividing the TAS by the TOS and multiplying by 100. In order to evaluate the expression levels of apoptosis-related Bax, Bcl-2, Caspase-3, Caspase-8, and Caspase-9 genes at the mRNA level in control and dose group cells, RNA isolation was performed from the SH-SY5Y control and dose group cells (IC50 value). Results: It is observed that the P. sidoides substance inhibits proliferation in cells at 24 h (p < 0.05). As the dose increases, cell proliferation decreases (p < 0.05). The IC50 value was calculated to be 113.83 μg/mL at 24 h. The concentration of 8-OhDG increased in neuroblastoma cells as a result of P. sidoides extract treatment (p < 0.05). TOS levels increased in neuroblastoma cells treated with P. sidoides extract (p < 0.01). OSI levels increased in cells treated with P. sidoides extract (p < 0.001). BAX and Caspase-8 expression increased are statistically significant in the P. sidoides dose group (p < 0.05). Conclusions: P. sidoides extract induces apoptosis in neuroblastoma cells through oxidative stress and mitochondrial- and death receptor-mediated pathways. This study highlights the potential of P. sidoides as a complementary therapeutic agent for neuroblastoma, warranting further in vivo and clinical investigations to assess its safety and efficacy.

Lactuca racemosa Willd., Source of Antioxidants with Diverse Chemical Structures.

Ethanolic extracts from the roots and aerial parts of the hitherto chemically uninvestigated lettuce species Lactuca racemosa Willd. (Cichorieae, Asteraceae) were chromatographically separated to obtain eight sesquiterpenoids, two apocarotenoids (loliolide and (6S,9S) roseoside), and three phenolic glucosides (apigenin 7-O-glucoside, eugenyl-4-O-β-glucopyranoside, and 5-methoxyeugenyl-4-O-β-glucopyranoside). Four of the isolated sesquiterpene lactones (8-α-angeloyloxyleucodin, matricarin, 15-deoxylactucin, and deacetylmatricarin 8-β-glucopyranoside) have not previously been found either in Lactuca spp. or in Cicerbita spp. In addition, HPLC-PAD chromatographic methods were used to estimate the deacetylmatricarin derivatives, luteolin 7-O-glucoside, and caffeic acid derivatives contents in the analyzed plant material. The aerial parts contained c. 3.0% dry weight of chicoric acid and equal amounts (0.4%) of caftaric acid and luteolin 7-O-glucoside. The roots contained fewer phenolic metabolites but were rich in deacetylmatricarin glucoside (c. 1.3%). The aglycone of the most abundant sesquiterpene lactone was evaluated with respect to its neuroprotective effect in H2O2- and 6-OHDA-treated human neuroblastoma SH-SY5Y cells. This compound, at concentrations of 10 and 50 μM, provided partial protection of undifferentiated cells, and at a concentration of 50 μM, it provided partial protection of retinoic acid-differentiated cells from H2O2-induced damage. In a model of 6-OHDA-evoked cytotoxicity, the sesquiterpenoid was less effective. Our findings may support the inclusion of this plant into the human diet.

Protective effects of naringenin against methamphetamine-induced cell death in dopaminergic SH-SY5Y cells

ABSTRACT Background: Methamphetamine is a psychoactive substance that competes with the dopamine transporter, disrupting its flow and storage. This can trigger oxidative stress, finally resulting in neural cell death. Due to the increasing prevalence of methamphetamine use, extensive research has been devoted to finding treatments that ameliorate its detrimental effects. Naringenin, a dietary flavonoid found in citrus fruits, has shown several neuroprotective and pharmacological properties. Objectives: This study was aimed to assess the protective effects of naringenin against methamphetamine-induced dopaminergic cell death. Methods: Before exposure to methamphetamine, human neuroblastomaSH-SY5Y cells were either pretreated or not treated (controls) with naringenin. Cell viability, level of oxidative stress markers, and expression of some genes involved in apoptosis and autophagy processes were then assessed using MTT, ROS, and MMP assays, and qRT-PCR and Western blotting techniques. Results: Naringenin pretreatment significantly enhanced cell viability following methamphetamine exposure (p < .01). It significantly decreased ROS levels (p < .001), preserved mitochondrial membrane potential, and moderated upregulation of apoptotic (CytC, Casp3, and Bax) and autophagic genes (Beclin-1, and LC-3) and down-regulation of Bcl-2 as an anti-apoptotic gene. Similar naringenin-mediated patterns were observed for cytochrome C and caspase 3 proteins. Conclusion: Naringenin administration can be considered for treating the neurotoxic effects of methamphetamine.

Filbertone-Induced Nrf2 Activation Ameliorates Neuronal Damage via Increasing BDNF Expression

Neurotrophic factors are endogenous proteins that promote the survival of various neuronal cells. Increasing evidence has suggested a key role for brain-derived neurotrophic factor (BDNF) in the dopaminergic neurotoxicity associated with Parkinson’s Disease (PD). This study explores the therapeutic potential of filbertone, a bioactive compound found in hazelnuts, in neurodegeneration, focusing on its effects on neurotrophic factors and the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway. In our study, filbertone markedly elevated the expression of neurotrophic factors, including BDNF, Glial cell line-Derived Neurotrophic Factor (GDNF), and Nerve Growth Factor (NGF), in human neuroblastoma SH-SY5Y cells, mouse astrocyte C8-D1A cells, and mouse hypothalamus mHypoE-N1 cells. Moreover, filbertone effectively countered neuroinflammation and reversed the decline in neurotrophic factors and Nrf2 activation induced by a high-fat diet (HFD) in neurodegeneration models. The neuroprotective effects of filbertone were further validated in models of neurotoxicity induced by palmitic acid (PA) and the neurotoxin MPTP/MPP+, where it was observed to counteract PA and MPTP/MPP+-induced decreases in cell viability and neuroinflammation, primarily through the activation of Nrf2 and the subsequent upregulation of BDNF and heme oxygenase-1 expression. Nrf2 deficiency negated the neuroprotective effects of filbertone in MPTP-treated mice. Consequently, our finding suggests that filbertone is a novel therapeutic agent for neurodegenerative diseases, enhancing neuronal resilience through the Nrf2 signaling pathway and upregulation of neurotrophic factors.Graphical s

Protective effect of astragaloside IV against zinc oxide nanoparticles induced human neuroblastoma SH-SY5Y cell death: a focus on mitochondrial quality control.

Occupational and unintentional exposure of zinc oxide nanoparticles (ZnONPs) raises concerns regarding their neurotoxic potential and there is an urgent need for the development of effective agents to protect against the toxic effects of ZnONPs. Astragalus memeranaceus (AM), a famous Traditional Chinese Medicine, as well as its bioactive components, showing a potential neuroprotective function. This study aims to investigate the neuroprotective effects of bioactive components of AM against ZnONPs-induced toxicity in human neuroblastoma SH-SY5Y cells and its underlying mechanisms. The cell apoptosis, ROS generation, MMP changes, mitochondrial fission/fusion, biogenesis, and mitophagy were assessed. In this study, AM treatment inhibited ZnONPs-induced cell apoptosis and ROS overproduction in SH-SY5Y cells. And astragaloside IV (ASIV) played a dominant role in the attenuation of cytotoxicity after ZnONPs exposure, rather than flavonoids and polysaccharides. ASIV treatment significantly reduced ROS generation and MMP collapse in ZnONPs-exposed cells. Furthermore, the protein expressions of mitochondrial biogenesis (PGC-1α), fusion (Mfn1 and Mfn2), and fission (Drp1) were markedly increased. Meanwhile, the PINK1/Parkin-mediated mitophagy was activated after ASIV administration, which ameliorated ZnONPs-induced SH-SY5Y cell death. Collectively, ASIV administration mitigated ZnONPs-induced cytotoxicity in SH-SY5Y cells through restoring mitochondrial quality control process, which hinted the protective role of ASIV in ZnONPs-induced neurotoxicity.

Characterization of Walnut Oil and Evaluation of Its Neuroprotective Effects in an In Vitro Model of Parkinson's Disease.

Parkinson's disease (PD) is a common neurodegenerative disorder marked by the degeneration of dopaminergic neurons and the buildup of α-synuclein aggregates. The current treatments focus on symptom relief, with no drugs available to halt disease progression. This has prompted interest in plant-based extracts as alternative therapies. This study examines the neuroprotective and antioxidant effects of walnut oil (WO), extracted from Juglans regia L., in an in vitro PD model using the neurotoxin rotenone (ROT). WO, rich in polyunsaturated fatty acids (PUFAs), including linoleic acid (LA) and α-linolenic acid (ALA), together with minor bioactive components, is known for its neuroprotective properties. Using human HMC3 microglial and SH-SY5Y neuroblastoma cells, we tested WO's effects on ROT-induced toxicity. The experiments were performed at different time points. The results showed that the co-administration of WO with ROT significantly improved cell viability and reduced reactive oxygen species (ROS) levels. Additionally, conditioned media from WO-treated HMC3 cells enhanced SH-SY5Y cell survival, indicating positive microglia-neuron interactions. Cell viability appeared to be concentration- and time-dependent. These findings highlight WO's potential, mainly due to its PUFA content, as a promising candidate for preventing neurodegenerative diseases like PD; they underscore the potential of WO content in food for the prevention of neurodegenerative diseases such as PD.

T-2 toxin induces senescence in human neuroblastoma SH-SY5Y cells by regulating the HIF-1α/p53/p21 pathway

The mechanisms by which T-2 toxin induces senescence in neurons, along with the involvement of hypoxia-inducible factor-1α (HIF-1α), remain poorly understood. Using human neuroblastoma SH-SY5Y cells as a model, T-2 toxin (6 nM, 1–48 h) induced senescence in SH-SY5Y cells, leading to cell cycle arrest. This was mediated by the upregulation of proteins involved in cell cycle regulation and stimulation of IL-6 and NF-κB expression. T-2 toxin rapidly triggered the expression of senescence-associated β-galactosidase, disrupted mitochondrial function, and altered the levels of Alzheimer’s disease-associated proteins (Tau, p-Tau, and APP). T-2 toxin also transiently elevated HIF-1α levels. Notably, HIF-1α regulated the expression of cell cycle inhibitory proteins (p16, p21, p53) and senescence-associated secretory phenotype factors (IL-8, IL-6, CCL2), along with the expression of senescence-associated β-galactosidase, thereby exacerbating T-2 toxin-induced cellular senescence. These findings underscore the vital role of HIF-1α in T-2 toxin-induced senescence in SH-SY5Y cells.

Enhanced brain delivery and antiproliferative activity of resveratrol using milk-derived exosomes

The polyphenol resveratrol (RSV) undergoes extensive phase II metabolism, which limits its bioavailability and bioactivity, including anticancer effects. Growing preclinical evidence reinforces milk-derived exosomes (EXOs) as promising nanocarrier drugs and bioactives delivery systems. Herein, to overcome the mentioned RSV’s drawbacks, EXOs were loaded with RSV (EXO-RSV) to evaluate its brain delivery in Sprague-Dawley rats and its in vitro antiproliferative effects against human neuroblastoma SH-SY5Y and glioblastoma U-87MG cancer cells compared with non-encapsulated RSV. Pharmacokinetic analyses in perfused brains showed RSV detection only after EXO-RSV administration, not after non-encapsulated RSV administration. Encapsulation also resulted in lower concentrations of phase II-derived RSV metabolites in circulation. Additionally, blood-brain barrier (BBB) transport assays using human brain microvascular endothelial cells (HBMECs) corroborated that encapsulation enhanced RSV transport efficiency and protected it from cellular metabolism. In vitro and in silico analyses of phase-II conjugates, primarily RSV 3-glucuronide, showed lower capacity than RSV to cross the BBB. Finally, EXO-RSV (47−750 nM) showed an increased dose-dependent antiproliferative efficacy on both cancer cell lines compared with the non-encapsulated RSV (10 μM), which was ineffective after 72 h of treatment. Our findings suggest that EXOs protect RSV from phase II metabolism and enhance its brain delivery and antiproliferative activity.

Impact of a Withania somnifera and Bacopa monnieri Formulation on SH-SY5Y Human Neuroblastoma Cells Metabolism Through NMR Metabolomic.

Objectives: This study investigates the effectiveness of an herbal formulation, STRESSLESS (ST-65), which combines ashwagandha (Withania somnifera) and bacopa (Bacopa monnieri), on SH-SY5Y human neuroblastoma cells. Given the rising interest in natural compounds for neuroprotection and stress alleviation, we aimed to explore the cellular and molecular effects of this formulation. Methods: Utilizing a nuclear magnetic resonance (NMR) metabolomic approach and ultra-high-performance liquid chromatography-high-resolution mass spectrometry (UHPLC-HRMS), we identified key bioactive compounds in ST-65, including withanolides from ashwagandha and bacosides from bacopa. Results: Our findings indicate that ST-65 treatment significantly alters the metabolic profile of SH-SY5Y cells. Key changes included increased levels of metabolites linked to neuroprotection, energy metabolism, and antioxidant defense. Notable enhancements were observed in specific amino acids and neuroprotective compounds, suggesting activation of neuroprotective mechanisms and mitigation of stress-induced damage. Conclusions: The study reveals a complex phyto-chemical profile of ST-65 and underscores its potential as a natural active agent for addressing stress-related neurodegenerative conditions. These insights into neuronal mechanisms provide a foundation for further exploration of herbal formulations in neuroprotection.

Detoxification of paraoxon-ethyl by Lysiphyllum strychnifolium extracts in undifferentiated human neuroblastoma SH-SY5Y cells

Context Lysiphyllum strychnifolium (Craib) A. Schmitz. (Fabaceae) is a Thai traditional medicine used to remove food and alcohol toxins from the body. Objective This study investigates the molecular mechanism of L. strychnifolium extracts against paraoxon-ethyl-induced apoptosis in SH-SY5Y cells. Materials and methods The ethanol and water extracts of leaves and stems of L. strychnifolium were prepared at various concentrations (0–100 μg/mL) and co-treated to the cells with 0.375 mM paraoxon-ethyl for 24 and 48 h. Cell viability was performed using the PrestoBlue assay. ROS and caspase activity were detected using 2′,7′-dichlorodihydrofluorecein diacetate and caspase-Glo® 3/7, 8, and 9 assay kits. Apoptotic and ER stress-related gene expression were determined by real-time PCR, and nuclear and mitochondrial morphology were observed using Hoechst 33342 and MitoTracker® Deep Red FM staining. Results The most effective concentrations of each extract against paraoxon-ethyl-induced cell death were 25 μg/mL of leaf ethanol, 12.5 μg/mL of stem ethanol, 100 μg/mL of leaf water, and 25 μg/mL of stem water extracts. The leaf ethanol extract was the most effective at detoxifying, while stem extracts were highly toxic in high doses. The detoxifying L. strychnifolium extracts against paraoxon-ethyl-induced oxidative stress decreased p53, BiP/GRP78, and CHOP gene expression and minimized caspase 9 and caspase 3, protecting cells from apoptosis. The extracts could also restore mitochondrial membrane potential and reduce the swollen globule mitochondrial shape. Discussion and conclusion These findings could potentially protect neuron cells from neurodegenerative issues due to oxidative damage, apoptosis, and other potential consequences.

Screening of a Fraction with Higher Amyloid β Aggregation Inhibitory Activity from a Library Containing 210 Mushroom Extracts Using a Microliter-Scale High-Throughput Screening System with Quantum Dot Imaging.

Alzheimer's disease (AD) is a highly prevalent neurodegenerative disease hallmarked by amyloid plaques and neurofibrillary tangles. Amyloid plaques are formed by the amyloid β (Aβ) aggregation, so substances that inhibit this aggregation are useful for preventing and treating AD. Mushrooms are widely used medicinal fungi with high edible and nutritional value. Mushrooms have a variety of biologically active ingredients, and studies have shown that they have certain effects in anti-bacterial, anti-oxidation, anti-inflammatory, anti-tumor, and immune regulation. Previously, we developed a microliter-scale high-throughput screening (MSHTS) system using quantum dot (QD) nanoprobes to screen Aβ aggregation inhibitors. In this study, we appraised the Aβ aggregation inhibitory activity of 210 natural mushrooms from Hokkaido (Japan) and found 11 samples with high activity. We then selected Elfvingia applanata and Fuscoporia obliqua for extraction and purification as these samples were able to suppress Aβ-induced neurocytotoxicity and were readily available in large quantities. We found that the ethyl acetate (EtOAc) extract of E. applanata has high Aβ aggregation inhibitory activity, so we performed silica gel column chromatography fractionation and found that fraction 5 (f5) of the EtOAc extract displayed the highest Aβ aggregation inhibitory activity among all mushroom samples. The half-maximal effective concentration (EC50) value was 2.30 µg/mL, higher than the EC50 of 10.7 µg/mL for rosmarinic acid, a well-known Aβ aggregation inhibitor. This inhibitory activity decreased with further purification, suggesting that some compounds act synergistically. The f5 fraction also inhibited the deposition of Aβ aggregates on the cell surface of human neuroblastoma SH-SY5Y cells. Our expectation is that f5, with additional tests, may eventually prove to be an inhibitor for the prevention of AD.

Understanding Aβ25-35 peptide altered exosomal proteome and associated pathways linked with the Alzheimer's disease pathogenesis using human neuroblastoma SH-SY5Y Cells.

The central nervous system (CNS) involves a complex interplay of communications between the neurons and various glial cells, which is crucial for brain functions. The major interactomes are exosomes that transmit sundry molecules (DNA, miRNAs, and proteins) between the cells and thus alter cell physiology. Exosomes can act as neuroprotective or neurodegenerative agents depending on the microenvironment of cells secreting them. Therefore, revealing exosome proteome becomes important to understand donor cells' physiology and its effect on the recipient cell. In this study, oxidative stress was induced by Aβ25-35 in the human neuroblastoma SH-SY5Y cells and the protective effects of phytochemical ferulic acid (FA) were evaluated alone and in combination with Aβ25-35 (pre-treated for 3h before Aβ25-35 exposure) and proteome of their secreted exosomes was analyzed, which was carried out via a high-resolution LC-MS Triple-ToF and further network-based analysis has been carried out using various bioinformatics tools. The proteomic profiling enlightened the multiple roles of exosomes as proteins associated with the various pathways advocate that exosomes can mediate a wide range of effects, from normal physiological processes like synaptic plasticity, neuronal metabolic support, nerve regeneration, DNA repair, axon guidance, and long-term potentiation (LTP) to abnormal pathological processes like inflammatory responses, oxidative stress, apoptosis, and formation of neutrophil extracellular traps (NETs). On comparison, treatment with Aβ25-35 resulted in a significant modulation of the exosomal proteome, promoting pathways associated with neurodegeneration. Conversely, the phytochemical FA displayed a protective effect by effectively countering Aβ25-35-induced oxidative stress responses linked with neurodegeneration, as seen in Alzheimer's disease (AD). Taken together, this study highlights the dual role of exosomes in physiological and pathophysiological neurodegenerative AD, which intricately depend on the particular cellular milieu.

Differentiated and mature neurons are more responsive to neurotoxicant exposure at both transcriptional and translational levels

SH-SY5Y human neuroblastoma cells have been extensively used as an in vitro model system in a diverse range of studies involving neurodevelopment, neurotoxicity, neurodegeneration, and neuronal ageing. Both naïve and differentiated phenotypes of SH-SY5Y cells are utilized to model human neurons under in vitro conditions. The process of differentiation causes extensive remodeling of neuronal cells at multiple omic levels, including the epigenome and proteome. In the present investigation, the miRNAome and proteome profiles of arsenic-treated naïve and differentiated SH-SY5Y cells were generated using the miRNA OpenArray technology and high-resolution mass spectrometry. Our findings demonstrated that differentiation dramatically affected the response of SH-SY5Y cells to toxicant exposure, as indicated by increased tolerance of differentiated cells against arsenic exposure compared to naïve cells in cell viability assay. Arsenic-exposed naïve and differentiated SH-SY5Y cells possess distinct miRNA and protein profiles with few similarities. Compared to naïve cells, differentiated cells have undergone higher deregulation in the expression of brain-enriched miRNAs and proteins and have shown a more drastic decrease in oxygen consumption rate, which is a measure of mitochondrial respiration after exposure to arsenic. Proteins identified in arsenic-treated differentiated SH-SY5Y cells were more enriched in pathways underlying multifactorial neurotoxic events. Additionally, more functional regulatory modules have been identified between the miRNAs and proteins differentially expressed in arsenic-treated differentiated SH-SY5Y cells relative to naïve cells. Collectively, our studies have shown that differentiated SH-SY5Y cells displayed alterations in the expression of a greater number of miRNAs and proteins following neurotoxicant exposure, indicating their higher responsivity.

Selective Noradrenergic Activation of BDNF Translation by Mirtazapine.

Antidepressants are known for their neurotrophic effects, particularly through the regulation of brain-derived neurotrophic factor (BDNF) expression. Mirtazapine, a tetracyclic noradrenergic and specific serotonergic antidepressant (NaSSA) has been observed to upregulate BDNF, though its underlying mechanism remains unclear. In this study, we used the human neuroblastoma SH-SY5Y cell line to investigate whether mirtazapine could enhance BDNF translation by modulating serotonin and/or norepinephrine and their receptors. A 1-h stimulation with 1 or 10µM mirtazapine led to downregulation of serotonergic receptors 5HT1A, while increasing ADRA2A and ADRB2 receptors. Mirtazapine at 10 µM upregulated endogenous BDNF after 3h, but not 1h stimulation. To investigate the translation of major BDNF transcripts, we used chimeric BDNF-luciferase constructs with the untranslated 5'UTR exons I, IIc, IV, or VI, and the long version of the 3'UTR. Luciferase assays and Western blotting revealed that mirtazapine selectively enhanced exon-IIc-BDNF-long3'UTR-Luciferase translation. This increase was associated with norepinephrine release and was inhibited by blocking ADRA2A or ADRB2 adrenoceptors for the exon-IIc-BDNF-long3'UTR-Luciferase, and ADR2B for endogenous BDNF. These findings provide a new perspective on the critical role of the noradrenergic system in mediating mirtazapine's effects on BDNF translation. We propose a novel mechanism of action in which mirtazapine promotes norepinephrine release and noradrenergic responses by upregulating ADRA2A and ADRB2 while downregulating serotonergic receptors.

Protective effect of Apelin-13 on D-glutamic acid-induced excitotoxicity in SH-SY5Y cell line: An in-vitro study

Excitotoxicity, resulting from excessive accumulation of glutamate in the extracellular space, leads to neuronal cell death. This study investigates the protective effects of Apelin-13 on D-Glutamic acid-induced excitotoxicity in SH-SY5Y human neuroblastoma cells, an in-vitro model for neurodegenerative diseases. Unlike the commonly studied L-glutamic acid, this research focuses on D-Glutamic acid to understand its specific impacts. SH-SY5Y cells were treated with varying concentrations of D-Glutamic acid and Apelin-13, followed by analyses at 12 and 24 h to evaluate cell viability, oxidative stress markers, and inflammatory cytokine levels. Cell viability assays revealed significant cytotoxic effects of D-Glutamic acid at doses of 10 mM and 20 mM, reducing viability by over 50 %. However, Apelin-13 treatment mitigated these effects, especially at 2 μg/ml, enhancing cell viability and reducing inflammatory cytokine levels (IL-1β and TNF-α). Apelin-13 also increased anti-inflammatory cytokine levels (IL-10 and TGF-β1) and brain-derived neurotrophic factor (BDNF), indicating its neuroprotective role. Oxidative stress markers, including ROS, AGE, AOPP, DT, T-SH, were significantly elevated by D-Glutamic acid but effectively reduced by Apelin-13. The neuroprotective mechanisms of Apelin-13 involve modulation of cAMP/PKA and MAPK signaling pathways, enhancing BDNF synthesis and suppressing oxidative stress and inflammatory responses. This study is the first to demonstrate the effects of D-Glutamic acid on SH-SY5Y cells. It highlights Apelin-13's potential as a therapeutic agent against excitotoxicity-induced neuronal damage, emphasizing its ability to modulate key molecular pathways involved in inflammation and oxidative stress. Further in-vivo studies are warranted to explore the long-term neuroprotective effects of Apelin-13 in treating neurodegenerative diseases.