Investigating the molecular mechanisms of Fuzheng Yiliu Shenji prescription in SH-SY5Y neuroblastoma cells.

Background: Neuroblastoma is the most common extracranial solid tumor in childhood. Fuzheng Yiliu Shenji Prescription (FYSP) has shown potential in treating malignant pediatric tumors in clinical settings. This study aims to explore the molecular mechanisms behind its effects, specifically in the context of neuroblastoma cell lines. Objective: To elucidate the active compounds in FYSP and their mechanisms of action in inhibiting neuroblastoma cell viability, inducing apoptosis, and affecting the cell cycle in SH-SY5Y cells through network pharmacology and empirical validation. Materials and methods: We identified the major compounds in FYSP and their predicted targets, constructing a protein-protein interaction (PPI) network and performing GO and KEGG pathway analyses. The effects of FYSP were empirically validated through assays on cell viability, cell cycle, apoptosis, and protein expression in SH-SY5Y cells. Results: The study identified 172 active chemical components in FYSP, with 188 common targets related to neuroblastoma. Network analysis highlighted the PI3K-Akt pathway as a significant target. Experimental validation in SH-SY5Y cells confirmed that FYSP could inhibit cell viability, induce G2/M cell cycle arrest, and promote apoptosis through modulation of the PI3K-Akt pathway, specifically upregulating caspase-3 and downregulating Bcl-2/Bax expression. Conclusion: The study elucidates the molecular basis of FYSP's effects on neuroblastoma cells in vitro, demonstrating its ability to modulate key pathways involved in cell cycle and apoptosis. While these findings suggest a potential therapeutic role for FYSP, they are limited to in vitro observations, and further research, including in vivo studies, is necessary to explore its clinical applicability.

In vitro modelling of Parkinson's disease using 6-OHDA is associated with increased NQO2 activity.

The pathogenesis of Parkinson's disease (PD) involves abnormalities in the metabolism of catecholamines. The enzyme quinone reductase 2 (NQO2) reduces quinone derivatives of catecholamines, which promotes the formation of reactive oxygen species (ROS), suggesting a role for NQO2 in the development of cellular damage typical of PD. In the present study, we investigated the relationship between 6-hydroxydophamine (6-OHDA) induced cellular damage and NQO2 activity and its levels in SH-SY5Y cell culture to establish an experimental model to evaluate the pharmacological properties of NQO2 inhibitors. Cellular damage was evaluated using the MTT and comet assays. It was shown that oxidative damage of SH-SY5Y cells upon incubation with 6-OHDA for 6, 12 and 24 h was accompanied by an increase in NQO2 activity. The increase in NQO2 protein level in SH-SY5Y cells was observed 24 h after incubation with 6-OHDA at concentrations of 50 and 100 µM. Oxidative damage of SH-SY5Y cells upon 1 h incubation with 6-OHDA is increased in the presence of the selective enzyme co-substrate 1-benzyl-1,4-dihydronicotinamide (BNAH), but is not accompanied by changes in NQO2 activity and protein levels. The data obtained demonstrate the contribution of NQO2 to the cytotoxic mechanism of 6-OHDA action.

LncRNA BASP1-AS1 is a positive regulator of stemness and pluripotency in human SH-SY5Y neuroblastoma cells.

Neuroblastoma is the most common extra-cranial solid tumor diagnosed mostly in children below the age of five years and comprises of about 15 % of all paediatric cancer deaths. Tumor initiating cancer stem cells (CSCs) can be targeted for better treatment approaches. BASP1-AS1 is a long non coding (Lnc) RNA that is a divergent LncRNA for its coding gene brain abundant membrane attached signal protein 1 (BASP1). We had earlier demonstrated it to be expressed in foetus derived human neural progenitor cells (hNPCs), where it was a positive regulator of BASP1 and was critical for neural differentiation. In this study, we have investigated the role of BASP1-AS1 in CSCs derived from the human neuroblastoma cell line SH-SY5Y. We cultured SH-SY5Y cells on Poly-d-Lysine coated flasks in serum free media supplemented with growth factors, which led to the enrichment of CSCs as determined by marker expression. When grown on ultra-low attachment flasks, these cells formed CSCs enriched neurospheres. We examined the effects of BASP1-AS1 siRNA mediated knockdown on CSCs enriched SH-SY5Y cells and SH-SY5Y derived neurospheres. BASP1-AS1 knockdown decreased the levels of the corresponding gene BASP1 and the rate of cell proliferation of CSCs enriched cells along with low expression of Ki67. It also reduced the mRNA levels of stem cell and pluripotency gene markers (CD133, CD44, c-KIT, SOX2, OCT4 and NANOG), as also Wnt 2 and the Wnt pathway effector ß catenin. It also abrogated the formation of neurospheres in ultra-low attachment flasks. A similar effect on proliferation and stemness related properties was seen on BASP1 knockdown. BASP1-AS1 and its related pathways may provide a point of intervention for the CSCs population in neuroblastoma.

Effect of lycopene on TiO2 nanoforms induced oxidative stress and neuroinflammation in SH-SY5Y cells: an in vitro study.

Due to its antioxidant action, the carotenoid lycopene has been demonstrated to have a protective effect in several disease models; however, its effect on the nanoform of titanium oxide (nano-TiO2)-induced neurotoxicity has not yet been determined. The purpose of this study was to evaluate how lycopene affects neuronal damage brought on by nano-TiO2 and the mechanisms involved. SH-SY5Y cells were treated with different concentrations of nano-TiO2 for 48 hours, the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) test was used after that to evaluate cell viability. IC50 of nano-TiO2 was determined and the results revealed that IC50 is equal 40 µM/mL, lycopene (10 µM) was applied to SH-SY5Y human neuroblastoma cells an hour before exposure to 40 µM nano-TiO2. Reactive oxygen species, lipid peroxidation, nitric oxide, glutathione, superoxide dismutase, and catalase, tumor necrosis factor-alpha, interleukin 1 beta, nuclear factor kappa B, and apoptotic markers (Bcl2, Bax, and caspase-3), were measured to determine the anti-oxidant effect of lycopene. In SH-SY5Y neuroblastoma cells, pretreatment with 10 µM lycopene significantly reduced the toxicity brought on by exposure to nano-TiO2, according to MTT assay findings and lactate dehydrogenase (LDH) cytotoxicity assessment. In cells exposed to nano-TiO2, lycopene pretreatment significantly boosted the activity of antioxidative enzymes and reduced oxidative stress. Furthermore, when SH-SY5Y cells were subjected to nano-TiO2, lycopene pretreatment stopped neuroinflammation and apoptosis. The findings of this study suggest that lycopene may be an effective neuroprotective against oxidative stress and neuroinflammation and may be used to stop neuronal death or injury in a variety of neurological illnesses.

The epigenetic signatures of opioid addiction and physical dependence are prevented by D-cysteine ethyl ester and betaine.

We have reported that D,L-thiol esters, including D-cysteine ethyl ester (D-CYSee), are effective at overcoming opioid-induced respiratory depression (OIRD) in rats. Our on-going studies reveal that co-injections of D-CYSee with multi-day morphine injections markedly diminish spontaneous withdrawal that usually occurs after cessation of multiple injections of morphine in rats. Chronically administered opioids are known (1) to alter cellular redox status, thus inducing an oxidative state, and (2) for an overall decrease in DNA methylation, therefore resulting in the transcriptional activation of previously silenced long interspersed elements (LINE-1) retrotransposon genes. The first objective of the present study was to determine whether D-CYSee and the one carbon metabolism with the methyl donor, betaine, would maintain redox control and normal DNA methylation levels in human neuroblastoma cell cultures (SH-SY5Y) under overnight challenge with morphine (100 nM). The second objective was to determine whether D-CYSee and/or betaine could diminish the degree of physical dependence to morphine in male Sprague Dawley rats. Our data showed that overnight treatment with morphine reduced cellular GSH levels, induced mitochondrial damage, decreased global DNA methylation, and increased LINE-1 mRNA expression. These adverse effects by morphine, which diminished the reducing capacity and compromised the maintenance of the membrane potential of SH-SY5Y cells, was prevented by concurrent application of D-CYSee (100 µM) or betaine (300 µM). Furthermore, our data demonstrated that co-injections of D-CYSee (250 µmol/kg, IV) and to a lesser extent, betaine (250 µmol/kg, IV), markedly diminished the development of physical dependence induced by multi-day morphine injections (escalating daily doses of 10-30 mg/kg, IV), as assessed by the lesser number of withdrawal phenomena elicited by the injection of the opioid receptor antagonist, naloxone (1.5 mg/kg, IV). These findings provide evidence that D-CYSee and betaine prevent the appearance of redox alterations and epigenetic signatures commonly seen in neural cells involved in opioid physical dependence/addiction, and lessen development of physical dependence to morphine.

Traditionally used edible medicinal plants protect against rotenone induced toxicity in SH-SY5Y cells-a prospect for the development of herbal nutraceuticals.

Plants are good sources of pharmacologically active compounds. The present study aimed to examine the neuroprotective potentials of the methanol extracts of Salix tetrasperma Roxb. leaf (STME) and Plantago asiatica L. (PAME), two edibles medicinal plants of Manipur, India against neurotoxicity induced by rotenone in SH-SY5Y cells. Free radical quenching activities were evaluated by ABTS and DPPH assays. The cytotoxicity of rotenone and the neuronal survival were assessed by MTT assay and MAP2 expression analysis. DCF-DA, Rhodamine 123 (Rh-123), and DAPI measured the intracellular reactive oxygen species (ROS) levels, mitochondrial membrane potential (MMP), and apoptotic nuclei, respectively. Superoxide dismutase (SOD), glutathione peroxidase (GPx), and catalase (CAT) activities were also assessed. LC-QTOF-MS analysis was performed for the identification of the compounds present in STME and PAME. The study showed that both the plant extracts (STME and PAME) showed antioxidant and neuroprotective capabilities in rotenone-induced neurotoxicity by preventing oxidative stress through the reduction of intracellular ROS levels and reversing the activities of GPx, SOD, and CAT caused by rotenone. Further, both plants prevented apoptotic cell death by normalizing the steady state of MMP and protecting nuclear DNA condensation. LC-QTOF-MS analysis shows the presence of known neuroprotective compounds like uridine and gabapentin in STME and PAME respectively. The two plants might be an important source of natural antioxidants and nutraceuticals with neuroprotective abilities. This could be investigated further to formulate herbal nutraceuticals for the treatment of neurodegenerative disease like Parkinson's disease.

A Novel and Robust Protocol for Differentiation of SH-SY5Y Neuroblastoma Cells into Neuron Like Cells.

Introduction: Human neuroblastoma cell line SH-SY5Y is a frequently used experimental cellular model in a variety of neuropsychiatric and neurodegenerative disorders. It is crucial to use a culture protocol that supports the fully differentiation of SH-SY5Y into neuron-like phenotype for the consistency of the results with neurons in vivo. However, a standardized neuronal differentiation protocol for SH-SY5Y cells still does not exist. Numerous differentiation methods have been proposed in the literature, yet SH-SY5Y cells with stronger neuronal characteristics and a more favorable environment for these differentiated cells are required in order to best representation of neurons. Therefore, in the study, we aimed to establish a more successful differentiation protocol for SH-SY5Y cells based on the primary neuron culture technique, which neuronal maturation is very well defined. Methods: In the study, we rearranged previous SH-SY5Y differentiation protocols, combined them with our primary neuron culture protocol and created a robust and reproducible protocol for differentiation of SH-SY5Y. Results: Our proposed "retinoic acid+brain-derived neurotrophic factor (RA+BDNF)-induced 7 days differentiation (conalbumin- on day 4) protocol provided well developed neurites, adequate expression and localization of neuronal and synaptic markers resembling mature neurons. Conclusion: The differentiation protocol we present can enable researchers to obtain satisfactory and properly differentiated SH-SY5Y cells in each independent experiment, achieving the closest possible in vivo results.

Lactoferrin Protects Against Rotenone-Induced Toxicity in Dopaminergic SH-SY5Y Cells through the Modulation of Apoptotic-Associated Pathways.

Parkinson's disease (PD) is a common motor neurodegenerative disease that still lacks effective therapeutic options. Previous studies have reported that lactoferrin exhibited neuroprotective effects in cellular and animal models of PD, typically induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) or 1-methyl-4-phenylpyridinium (MPP+) synthetic toxin. However, the neuroprotective capacity of lactoferrin in the rotenone-induced cellular model of PD remains relatively less established. Unlike MPTP/MPP+, rotenone is a naturally occurring environmental toxin known to induce chronic toxicity and increase the risk of PD in humans. In this study, we constructed a cellular model of PD by differentiating SH-SY5Y neuroblastoma cells with retinoic acid into mature dopaminergic neurons with increased ß-tubulin III and tyrosine hydroxylase expression, followed by 24 h of rotenone exposure. Using this cellular model of PD, we showed that lactoferrin (1-10 µg/ml) pre-treatment for 48 h decreased loss of cell viability, mitochondrial membrane potential impairment, reactive oxygen species generation and pro-apoptotic activities (pan-caspase activation and nuclear condensation) in cells exposed to rotenone (1 and 5 µM) using biochemical assays, Hoechst 33342 staining and immunocytochemical techniques. We further demonstrated that 48 h of lactoferrin (10 µg/ml) pre-treatment decreased Bax:Bcl2 ratio and p42/44 mitogen-activated protein kinase expression but increased pAkt expression in 5 µM rotenone-exposed cells. Our study demonstrates that lactoferrin neuroprotective capacity is present in the rotenone-induced cellular model of PD, further supporting lactoferrin as a potential PD therapeutic that warrants further studies.

α-Lipoic acid: a potential regulator of copper metabolism in Alzheimer's disease.

Alzheimer's disease (AD) is characterized by classic hallmarks such as amyloid plaques and neurofibrillary tangles, however, intensive research has broadened its scope to explore additional underlying mechanisms. Notably, disruptions in metal homeostasis, particularly involving copper, have gained significant attention. In AD pathology, an imbalance is evident: there is an excess of extracellular copper alongside a deficiency in intracellular copper in brain tissue. Our previous work demonstrated that α-lipoic acid (LA) can effectively shift copper from the extracellular space to the intracellular environment in a neuronal cell model. However, the precise mechanism of action and role of LA in copper metabolism remained elusive. In this study, we compared the cellular effects of LA with those of different synthetic copper-binding ligands: diethyldithiocarbamate (DETC), clioquinol (CQ), D-penicillamine (D-PA) and elesclomol (ES). Using differentiated SH-SY5Y cell culture as a neuronal model, we found that, unlike other synthetic compounds, natural ligand LA is not toxic in the presence of extracellular copper, even at high doses. LA gradually increased intracellular copper levels over 24 h. In contrast, DETC, CQ, and ES acted as fast copper ionophores, potentially explaining their higher toxicity compared to LA. D-PA did not facilitate copper uptake into cells. We demonstrated that a slow increase of LA inside the cells is enhanced in the presence of copper. Furthermore, the ability of LA to modulate the equilibrium of extra- and intracellular copper was evident when we added copper isotope 65Cu. The ratio of copper isotopes changed rapidly, reflecting the impact of LA on the equilibrium of copper distribution without affecting the copper transport network. Our results provide compelling evidence that α-lipoic acid holds promise as a non-toxic agent capable of normalizing copper metabolism in Alzheimer's disease.

Randomly methylated ß-cyclodextrin improves water - solubility, cellular protection and mucosa permeability of idebenone.

Neurodegenerative diseases such as Alzheimer's are very common today. Idebenone (IDE) is a potent antioxidant with good potential for restoring cerebral efficiency in cases of these and other medical conditions, but a serious drawback for the clinical use of IDE in neurological disorders lies in its scarce water solubility, which greatly inhibits its bioavailability. In this work, we prepared the inclusion complex of IDE with randomly methylated ß-cyclodextrin (RAMEB), resulting in improved water solubility of the included drug; then its in vitro biological activity and ex vivo permeability was evalutated. The solid complex was characterized through FT-IR spectroscopy, Thermogravimetric analysis (TGA) and Differential Scanning Calorimetry (DSC). A 78-fold improvement of the solubility of IDE in water resulted, together with a strong 1:1 host-guest interaction (association constant of 12630 M-1), and dissolution of the complex within 15 min, all evidenced during the in-solution studies. Biological in vitro studies were then performed on differentiated human neuroblastoma cells (SH-SY5Y) subjected to oxidative stress. Pretreatment with IDE/RAMEB positively affected cell viability, promoted the nuclear translocation of Nrf2, and increased the levels of GSH as well as those of the endogenous antioxidant enzymes Mn-SOD and HO-1. Lastly, the complexation significantly improved the permeation of IDE through isolated rat nasal mucosa.

The role of neuron-like cell lines and primary neuron cell models in unraveling the complexity of neurodegenerative diseases: a comprehensive review.

Neurodegenerative diseases (NDs) are characterized by the progressive loss of neurons. As to developing effective therapeutic interventions, it is crucial to understand the underlying mechanisms of NDs. Cellular models have become invaluable tools for studying the complex pathogenesis of NDs, offering insights into disease mechanisms, determining potential therapeutic targets, and aiding in drug discovery. This review provides a comprehensive overview of various cellular models used in ND research, focusing on Alzheimer's disease, Parkinson's disease, Huntington's disease, and amyotrophic lateral sclerosis. Cell lines, such as SH-SY5Y and PC12 cells, have emerged as valuable tools due to their ease of use, reproducibility, and scalability. Additionally, co-culture models, involving the growth of distinct cell types like neurons and astrocytes together, are highlighted for simulating brain interactions and microenvironment. While cell lines cannot fully replicate the complexity of the human brain, they provide a scalable method for examining important aspects of neurodegenerative diseases. Advancements in cell line technologies, including the incorporation of patient-specific genetic variants and improved co-culture models, hold promise for enhancing our understanding and expediting the development of effective treatments. Integrating multiple cellular models and advanced technologies offers the potential for significant progress in unraveling the intricacies of these debilitating diseases and improving patient outcomes.

Synthesis of novel hydrophilic celastrol nanoformulation by entrapment within calcium phosphate nanoparticle and study of its antioxidant activity against neurotoxin-induced damage in human neuroblastoma cells.

Celastrol, a pentacyclic triterpenoid found in Chinese herb Tripterygium wilfordii, is considered as one of the top-five natural medicinal compounds with high antioxidant property. However, celastrol has poor aqueous solubility and thereby low bioavailability, restricting its clinical application as drug. To overcome this problem, we nanonized celastrol by entrapping it within hydrophilic nanocarrier - calcium phosphate nanoparticle. The synthesized calcium phosphate celastrol nanoparticle (CPCN) had average size of 35 nm, spherical shape, significant stability with (-) 37 mV zeta potential, celastrol entrapment efficiency around 75 % and low celastrol release kinetics spanning over 7 days, as measured by different techniques like FESEM, AFM, DLS, and spectrophotometry. Studies on the antioxidant potency of CPCN by flow cytometry and fluorescence microscopy depicted that the toxicity developed in human neuroblastoma cells SH-SY5Y by treatment with the selective neurotoxin MPP+ iodide (N-Methyl-4-phenylpyridinium iodide) got reduced by pretreatment of the cells with CPCN. Determination of cellular ROS content, depolarization level of mitochondrial membrane potential, cell cycle analysis and nuclear damage in MPP+-exposed cells demonstrated that CPCN had about 65 % more antioxidant efficacy over that of bulk celastrol. Thus, the nanonization process transformed hydrophobic celastrol into hydrophilic CPCN, having high potentiality to be developed as an effective antioxidant drug.

Design, synthesis, molecular modeling, in vitro evaluation of novel piperidine-containing hydrazone derivatives as cholinesterase inhibitors.

In an effort to develop new and effective therapeutic agents for Alzheimer's disease, a series of hydrazone derivatives bearing piperidine rings have been designed and synthesized. The chemical structures of the compounds were characterized by various spectroscopic techniques. In vitro antioxidant and cholinesterase activities of the compounds were evaluated. Among the compounds, N12 exhibited the most antioxidant activity in all methods (CUPRAC, FRAP, DPPH, ABTS). In vitro acetylcholinesterase (AChE) activity results of the compounds showed good IC50 values between 14.124 ± 0.084 and 49.680 ± 0.110 µM were obtained (IC50 = 38.842 ± 0.053 µM for Donepezil). Among the compounds, N7 and N6 are much more effective derivatives than the standard compound donepezil with IC50 values of 14.124 ± 0.084 and 17.968 ± 0.072 µM, respectively. In vitro, butyrylcholinesterase (BChE) inhibition values of the compounds were between 13.505 ± 0.025 and 52.230 ± 0.027 µm. Among the compounds, N6 has the highest BChE inhibition with an IC50 value of 13.505 µm in the series. The cytotoxicity and AChE inhibitory activity of the compounds on SH-SY5Y cell lines were also evaluated. Kinetic studies were also performed to determine the behavior of the compounds as competitive or noncompetitive inhibitors. The binding modes of N6, which was determined to be highly effective according to in vitro analyses, with AChE and BChE were investigated using molecular docking studies, and the stability of the complexes was determined by molecular dynamics simulations. These findings indicated that AChE and BChE enzymes maintained their overall structural stability and compactness during interactions with compound N6.

Examination of Akt and GSK3ß in BDNF-mediated reductions in BACE1 activity in neuronal cells.

Brain-derived neurotrophic factor (BDNF) content and signaling has been identified as one potential regulator of amyloid precursor protein (APP) processing. Recently published work has demonstrated that BDNF reduces BACE1 activity while also elevating the inhibition of GSK3ß in the prefrontal cortex of male C57BL/6J mice. These results provide evidence that BDNF alters APP processing by reducing BACE1 activity, which may act through GSK3ß inhibition. The purpose of this study was to further explore the role of GSK3ß in BDNF-induced regulation on BACE1 activity. We utilized a cell culture and an in vitro activity assay model to pharmacologically target BDNF and GSK3ß signaling to confirm its involvement in the BDNF response. Treatment of differentiated SH-SY5Y neuronal cells with 75 ng/mL BDNF resulted in elevated pTrkB content, pAkt content, pGSK3ß content, and reduced BACE1 activity. An in vitro BACE1 activity assay utilizing mouse prefrontal cortex (n = 6/group) supplemented with BDNF, BDNF + ANA12 (Trkb antagonist), or BDNF + wortmannin (Akt inhibitor) demonstrated that BDNF reduced BACE1 activity; however, in the presence of TrkB or Akt inhibition, this effect was abolished. An in vitro ADAM10 activity assay utilizing mouse prefrontal cortex (n = 6/group) supplemented with BDNF, BDNF + ANA12 (Trkb antagonist), or BDNF + wortmannin (Akt inhibitor) demonstrated that BDNF did not alter ADAM10 activity. However, inhibiting BDNF signaling reduced ADAM10 activity. Collectively these studies suggest that GSK3ß inhibition may be necessary for BDNF-induced reductions in BACE1 activity. These findings will allow for the optimization of future therapeutic strategies by selectively targeting TrkB activation and GSK3ß inhibition.

Dopamine and Citicoline-Co-Loaded Solid Lipid Nanoparticles as Multifunctional Nanomedicines for Parkinson's Disease Treatment by Intranasal Administration.

This work aimed to evaluate the potential of the nanosystems constituted by dopamine (DA) and the antioxidant Citicoline (CIT) co-loaded in solid lipid nanoparticles (SLNs) for intranasal administration in the treatment of Parkinson disease (PD). Such nanosystems, denoted as DA-CIT-SLNs, were designed according to the concept of multifunctional nanomedicine where multiple biological roles are combined into a single nanocarrier and prepared by the melt emulsification method employing the self-emulsifying Gelucire® 50/13 as lipid matrix. The resulting DA-CIT-SLNs were characterized regarding particle size, surface charge, encapsulation efficiency, morphology, and physical stability. Differential scanning calorimetry, FT-IR, and X ray diffraction studies were carried out to gain information on solid-state features, and in vitro release tests in simulated nasal fluid (SNF) were performed. Monitoring the particle size at two temperatures (4 °C and 37 °C), the size enlargement observed over the time at 37 °C was lower than that observed at 4 °C, even though at higher temperature, color changes occurred, indicative of possible neurotransmitter decomposition. Solid-state studies indicated a reduction in the crystallinity when DA and CIT are co-encapsulated in DA-CIT-SLNs. Interestingly, in vitro release studies in SNF indicated a sustained release of DA. Furthermore, DA-CIT SLNs displayed high cytocompatibility with both human nasal RPMI 2650 and neuronal SH-SY5Y cells. Furthermore, OxyBlot assay demonstrated considerable potential to assess the protective effect of antioxidant agents against oxidative cellular damage. Thus, such protective effect was shown by DA-CIT-SLNs, which constitute a promising formulation for PD application.

Tannic acid protects neuroblastoma cells against hydrogen peroxide - triggered oxidative stress by suppressing oxidative stress and apoptosis.

Recent investigations indicate that tannic acid is associated with a decrease in oxidative damage. Growing evidence supports the protective effects of tannic acid on the central nervous system (CNS). However, uncertainties persist regarding its influence on hydrogen peroxide (H2O2)-triggered oxidative impairment in nerve cells and its interaction with apoptosis. Hence, the objective of this work was to examine the neuroprotective impact of tannic acid on SH-SY5Y cell impairment following H2O2-induced oxidative stress, particularly concerning apoptotic pathways. The control group received no treatment, while the H2O2 group underwent treatment with 0.5 mM H2O2 for a duration of 24 h. The tannic acid group received treatment with different concentrations of tannic acid for a duration of 24 h. Meanwhile, the tannic acid + H2O2 group underwent pre-treatment with tannic acid for one hour and was subsequently subjected to 0.5 mM H2O2 for one day. Within the tannic acid + H2O2 group, the cell viability in SH-SY5Y cells was notably enhanced by tannic acid at concentrations of 2.5, 5, and 10 µM. It also resulted in a considerable rise in TAS (Total Antioxidant Status) levels and a concurrent decline in TOS (Total Oxidant Status) levels, serving as indicators of reduced oxidative stress. Additionally, tannic acid treatment resulted in decreased levels of apoptotic markers (Bax, cleaved PARP, and cleaved caspase 3) and oxidative DNA damage marker (8-oxo-dG), while increasing the anti-apoptotic marker Bcl-2. The findings from flow cytometry also revealed a significant reduction in the apoptosis rate following pretreatment with tannic acid. In summary, tannic acid demonstrates protective effects on SH-SY5Y cells in the face of H2O2-triggered oxidative damage by suppressing both oxidative stress and apoptosis. Nevertheless, additional research is warranted to assess the neuroprotective potential of tannic acid.

Distinct In Vitro Differentiation Protocols Differentially Affect Cytotoxicity Induced by Heavy Metals in Human Neuroblastoma SH-SY5Y Cells.

The SH-SY5Y cell line is widely used in neurotoxicity studies. However, the effects of inducing cell differentiation on the cytotoxic effects of heavy metals are unclear. Therefore, we investigated the effects of mercuric chloride (HgCl2), cadmium chloride (CdCl2), arsenic trioxide (As2O3), and methylmercury (MeHg) on SH-SY5Y cells differentiated in the presence of insulin-like growth factor-I (IGF-I) or all-trans retinoic acid (ATRA). Neurite outgrowth with distinct changes in neuronal marker expression, phenotype, and cell cycle was induced in SH-SY5Y cells by IGF-I treatment for 1 day or ATRA treatment for up to 7 days. The cytotoxic effects of HgCl2 decreased at lower concentrations and increased at higher concentrations in both IGF-I- and ATRA-differentiated cells compared with those in undifferentiated cells. Differentiation with IGF-I, but not with ATRA, increased the cytotoxic effects of CdCl2. Decreased cytotoxic effects of As2O3 and MeHg were observed at lower concentrations in IGF-I-differentiated cells, whereas increased cytotoxic effects of As2O3 and MeHg were observed at higher concentrations in ATRA-differentiated cells. Changes in the cytotoxic effects of heavy metals were observed even after 1 day of ATRA exposure in SH-SY5Y cells. Our results demonstrate that the differentiation of SH-SY5Y cells by IGF-I and ATRA induces different cellular characteristics, resulting in diverse changes in sensitivity to heavy metals, which depend not only on the differentiation agents and treatment time but also on the heavy metal species and concentration.

Putative Adverse Outcome Pathway for Parkinson's Disease-like Symptoms Induced by Silicon Quantum Dots based on In Vivo/Vitro Approaches.

Given the commercial proliferation of silicon quantum dots (SiQDs) and their inevitable environmental dispersal, this study critically examines their biological and public health implications, specifically regarding Parkinson's disease. The study investigated the toxicological impact of SiQDs on the onset and development of PD-like symptoms through the induction of ferroptosis, utilizing both in vivo [Caenorhabditis elegans (C. elegans)] and in vitro (SH-SY5Y neuroblastoma cell line) models. Our findings demonstrated that SiQDs, characterized by their stable and water-soluble physicochemical properties, tended to accumulate in neuronal tissues. This accumulation precipitated dopaminergic neurodegeneration, manifested as diminished dopamine-dependent behaviors, and escalated the expression of PD-specific genes in C. elegans. Importantly, the results revealed that SiQDs induced ferritinophagy, a selective autophagy pathway that triggered ferroptosis and resulted in PD-like symptoms, even exacerbating disease progression in biological models. These insights were incorporated into a putatively qualitative and quantitative adverse outcome pathway framework, highlighting the serious neurodegenerative risks posed by SiQDs through ferroptosis pathways. This study provides a multidisciplinary analysis critical for informing policy on the regulation of SiQDs exposure to safeguard susceptible populations and guiding the responsible development of nanotechnologies impacting environmental and public health.

Imidacloprid affects human cells through mitochondrial dysfunction and oxidative stress.

Given their relatively low persistence and mammalian toxicity, neonicotinoid pesticides have been extensively used worldwide and are omnipresent in the environment. Recent studies have shown that neonicotinoids may pose adverse effects on non-target organisms other than the known neurotoxicity, raising emerging concerns that these insecticides might pose human health risk through additional toxicity pathways. In the present study, the mitochondria function, oxidative stress, DNA damages, and genes transcription levels were examined in the human neuroblastoma SH-SY5Y cells after 48-h exposure to imidacloprid at concentrations from 0.05 to 200 µmol/L. Results showed that imidacloprid induced mitochondrial dysfunction with the degradation of adenosine triphosphate (ATP) and mitochondrial membrane potential (MMP) levels. In addition, imidacloprid caused oxidative stress by stimulating the generation of reactive oxygen species (ROS) and hydrogen peroxide (H2O2) via the disruption of calcium ion level and mitochondrial function. Ultimately, the oxidative stress continued to produce DNA damage and apoptosis in SH-SY5Y cells at imidacloprid concentrations above 47.6 µmol/L. Among the evaluated endpoints, ATP was the most sensitive, with a median activity concentration of 0.74 µmol/L. The 5 % hazard concentration of imidacloprid was estimated to be 0.69 µmol/L, which can be used as a threshold for human health risk assessment for imidacloprid. Collectively, our results provide an important support for further research on potential toxicity of neonicotinoids related to mitochondrial toxicity in humans.

Ecklonia cava Polyphenols Have a Preventive Effect on Parkinson's Disease through the Activation of the Nrf2-ARE Pathway.

Parkinson's disease (PD) is a degenerative neurological disorder defined by the deterioration and loss of dopamine-producing neurons in the substantia nigra, leading to a range of motor impairments and non-motor symptoms. The underlying mechanism of this neurodegeneration remains unclear. This research examined the neuroprotective properties of Ecklonia cava polyphenols (ECPs) in mitigating neuronal damage induced by rotenone via the activation of the nuclear factor erythroid 2-related factor 2 (Nrf2)-antioxidant response element (ARE) pathway. Using human neuroblastoma SH-SY5Y cells and PD model mice, we found that ECP, rich in the antioxidant polyphenol phlorotannin, boosted the gene expression and functionality of the antioxidant enzyme NAD(P)H quinone oxidoreductase-1. ECP also promoted Nrf2 nuclear translocation and increased p62 expression, suggesting that p62 helps sustain Nrf2 activation via a positive feedback loop. The neuroprotective effect of ECP was significantly reduced by Compound C (CC), an AMP-activated protein kinase (AMPK) inhibitor, which also suppressed Nrf2 nuclear translocation. In PD model mice, ECPs improved motor functions impaired by rotenone, as assessed by the pole test and wire-hanging test, and restored intestinal motor function and colon tissue morphology. Additionally, ECPs increased tyrosine hydroxylase expression in the substantia nigra, indicating a protective effect on dopaminergic neurons. These findings suggest that ECP has a preventative effect on PD.