Activity In SH-SY5Y Cells Research Articles

Genkwanin suppresses MPP+-induced cytotoxicity by inhibiting TLR4/MyD88/NLRP3 inflammasome pathway in a cellular model of Parkinson’s disease

Parkinson’s disease (PD) is a complicated multifactorial neurodegenerative disorder. Oxidative stress, neuroinflammatory response, and activation of apoptosis have been proposed to be tightly involved in the pathogenesis of PD. Genkwanin is a typical bioactive non-glycosylated flavonoid with anti-inflammatory and anti-oxidant activities. However, the effect of genkwanin on PD remains unclear. Cell viability, lactate dehydrogenase (LDH) release, caspase-3/7 activity, and apoptosis was evaluated by MTT, LDH release assay, caspase-3/7 activity assay, and TUNEL assay, respectively. The secretion of prostaglandin E2 (PGE2), tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6 were measured by respective commercial ELISA kits. The mRNA expression of TNF-α, IL-1β, and IL-6 was detected by qRT-PCR. The protein levels of cycloxygenase-2 (COX-2), toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88), and NOD-like receptor (NLR) protein: 3 (NLRP3) were determined by western blot analysis. Genkwanin at concentrations less than 40 μM had no impact on cell viability and LDH release. Genkwanin suppressed MPP+-induced neuroinflammation in SH-SY5Y cells. MPP+ treatment inhibited cell viability, increased LDH release, apoptosis, and ROS generation, and reduced superoxide dismutase (SOD) activity in SH-SY5Y cells, which were abolished by genkwanin treatment. Genkwanin suppressed MPP+-induced activation of TLR4/MyD88/NLRP3 inflammasome pathway in SH-SY5Y cells. TLR4 overexpression weakened the anti-inflammatory and anti-neurotoxicity of genkwanin in SH-SY5Y cells. In conclusion, genkwanin attenuated neuroinflammation and neurotoxicity by inhibiting TLR4/MyD88/NLRP3 inflammasome pathway in MPP+-induced cellular model of PD.

Tris (2,3-Dibromopropyl) Isocyanurate (TDBP-TAZTO or TBC) Shows Different Toxicity Depending on the Degree of Differentiation of the Human Neuroblastoma (SH-SY5Y) Cell Line

Tris (2,3-dibromopropyl) isocyanurate (TDBP-TAZTO or TBC) is a heterocyclic hexabromated flame retardant. It is widely used during the production of many synthetic compounds. High concentrations of TDBP-TAZTO were found in river water, surface sediments, soil, earthworms, and carp tissues. Moreover, it has been shown that this compound can cross the blood–brain barrier and accumulate in the gut and brain of carp. The aryl hydrocarbon receptor (AhR) has been characterized as a multifunctional intracellular sensor and receptor. AhR is an activator of cytochrome P450 1A1 and 1A2, which metabolize various toxic compounds. The aim of the study was to explain how/whether TDBP-TAZTO increases the expression and/or activity of the CYP1A1 enzyme and the AhR and TUBB3 expression during SH-SY5Y cell differentiation. SH-SY5Y cells were differentiated for 7 and 14 days using retinoic acid. Cell viability, ethoxyresorufin-O-deethylase (EROD) activity, and mRNA expression of CYP1A1, AhR, and TUBB3 were assessed. Our experiment showed that, during the differentiation process, the ability of TDBP-TAZTO to induce EROD activity in SH-SY5Y cells subsequently decreased, which may have been an effect of cell differentiation into neurons. Moreover, the results suggest that TDBP-TAZTO can affect the differentiation process. Since no CYP2B6 mRNA expression was detected, the CAR receptor may not be involved in the TDBP-TAZTO mechanism of action. However, more research is needed in this field to elucidate this mechanism precisely.

Neuroprotective Activities of Crossyne flava Bulbs and Amaryllidaceae Alkaloids: Implications for Parkinson's Disease.

Parkinson’s disease (PD) is one of the most common neurodegenerative diseases and affects approximately 6.3 million people worldwide. To date, the treatment of PD remains a challenge, as available treatment options are known to be associated with serious side effects; hence, the search for new treatment strategies is critical. Extracts from the Amaryllidaceae plant family as well as their alkaloids have been reported to have neuroprotective potentials. This study, therefore, investigated the biological activities of Crossyne flava and its isolated alkaloids in an in vitro MPP+ (1-methyl-4-phenylpyridinium) PD model using SH-SY5Y cells. The effects of the total extract as well as the four compounds isolated from Crossyne flava (i.e., pancratinine B (1), bufanidrine (2), buphanisine (3), and epibuphanisine (4)) were evaluated for cell viability, neuroprotection, levels of reactive oxygen species (ROS), adenosine triphosphate activity (ATP), and caspase 3/7 activity in SH-SY5Y cells. The results obtained showed that pre-treatment with both the extract and the isolated compounds was effective in protecting the SH-SY5Y cells from MPP+-induced neurotoxicity and inhibited ROS generation, ATP depletion as well as apoptosis induction in the SH-SY5Y cells. The results of this study show that the Amaryllidaceae plant family may be a source of novel compounds for the treatment of neurodegenerative diseases, which validates the reported traditional uses.

Design, synthesis, and neuroprotective effects of novel hybrid compounds containing edaravone analogue and 3-n-butylphthalide ring-opened derivatives

To develop anti-ischemic stroke drugs with higher blood-brain barrier (BBB) penetrating capability and neuroprotective activity, a series of hybrid compounds containing edaravone analogue and 3-n-butylphthalide (NBP) ring-opened derivatives were synthesized and biologically evaluated. Among them, compound 10a displayed the highest protective activity in SH-SY5Y cells against oxygen and glucose deprivation (OGD) and H2O2 insults. Experiment results indicated that 10a could inhibit platelet aggregation via the synergistic action of the edaravone analogue and NBP, and its oral administration protected the rats against ischemia/reperfusion-induced brain injury. Moreover, 10a effectively inhibited apoptosis and reduced oxidative stress in OGD-exposed cells. Further analysis suggested that 10a might alleviate oxidative damage in SH-SY5Y cells via the modulation of the Nrf2 pathway. Collectively, these findings demonstrate that 10a can emerge as a potential candidate drug for the treatment of ischemic stroke.

Novel pentacyclic triterpenes exhibiting strong neuroprotective activity in SH-SY5Y cells in salsolinol- and glutamate-induced neurodegeneration models.

Novel triterpene derivatives were prepared and evaluated in salsolinol (SAL)- and glutamate (Glu)-induced models of neurodegeneration in neuron-like SH-SY5Y cells. Among the tested compounds, betulin triazole 4 bearing a tetraacetyl-β-d-glucose substituent showed a highly potent neuroprotective effect. Further studies revealed that removal of tetraacetyl-β-d-glucose part (free triazole derivative 10) resulted in strong neuroprotection in the SAL model at 1μM, but this derivative suffered from cytotoxicity at higher concentrations. Both compounds modulated oxidative stress and caspase-3,7 activity, but 10 showed a superior effect comparable to the Ac-DEVD-CHO inhibitor. Interestingly, while both 4 and 10 outperformed the positive controls in blocking mitochondrial permeability transition pore opening, only 4 demonstrated potent restoration of the mitochondrial membrane potential (MMP) in the model. Derivatives 4 and 10 also showed neuroprotection in the Glu model, with 10 exhibiting the strongest oxidative stress reducing effect among the tested compounds, while the neuroprotective activity of 4 was probably due recovery of the MMP.

Long noncoding RNA SRY-box transcription factor 2 overlapping transcript participates in Parkinson’s disease by regulating the microRNA-942-5p/nuclear apoptosis-inducing factor 1 axis

ABSTRACT Parkinson’s disease (PD) is a neurodegenerative disorder. Studies have shown that long noncoding RNA SRY-box transcription factor 2 overlapping transcript (lncRNA SOX2-OT) is highly expressed in PD patients, but its specific functions and mechanisms require further research. To address this gap, this study utilized an in vitro PD cell model induced by 1-methyl-4-phenylpyridinium (MPP+). Cell viability, apoptosis, lactate dehydrogenase (LDH) activity, inflammatory factor secretion, and oxidative stress indicators were determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-dipheyltetrazolium bromide assay, LDH assay, flow cytometry, enzyme linked immunosorbent assay (ELISA), and corresponding kits, respectively. Gene and protein expression were measured using quantitative real-time-PCR and western blotting, respectively. The results indicated that microRNA-942-5p (miR-942-5p) was a direct target of lncRNA SOX2-OT and nuclear apoptosis-inducing factor 1 (NAIF1) was a direct target of miR-942-5p. The expression levels of lncRNA SOX2-OT and NAIF1 were increased, and miR-942-5p expression was decreased in SH-SY5Y cells following MPP+ treatment. In addition, MPP+ treatment reduced SH-SY5Y cell viability, increased apoptosis; increased cleaved caspase-3 protein expression and cleaved caspase-3/caspase-3 ratio; enhanced lactate dehydrogenase viability; increased tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and reactive oxygen species, and decreased superoxide dismutase activity in SH-SY5Y cells were inhibited by SOX2-OT-siRNA, and these inhibitions were reversed by miR-942-5p inhibitor. Moreover, the protective role of miR-942-5p mimic in MPP+-induced SH-SY5Y cells was eliminated by the NAIF1 plasmid. Overall, lncRNA SOX2-OT-mediated regulation of oxidative stress, inflammation, and neuronal apoptosis were directly controlled by the miR-942-5p/NAIF1 signal axis in MPP+-induced SH-SY5Y cells.

MicroRNA-155 influences cell damage in ischemic stroke via TLR4/MYD88 signaling pathway

ABSTRACT Cerebral ischemic stroke (CIS) is extremely harmful, and its treatment should be underpinned by understanding its pathogenic mechanism. This study was designed to determine the involvement of miR-155 in CIS development via the TLR4/MyD88 signaling pathway. First, we quantified serum miR-155 in patients with CIS and healthy individuals, and found high expression of miR-155 in such patients and a decrease in it in the patients after therapy (P < 0.05). Serum miR-155 demonstrated a favorable function in predicting the development and prognosis of CIS (P < 0.001). We also conducted a mouse assay, and found that knocking out miR-155 can improve the neurological function of mice and suppress protein TLR4 and MyD88 (all P < 0.05). Finally, we carried out a cell assay, and found enhancement in the activity of SH-SY5Y cells, decrease in their apoptosis, and protein TLR4 and MyD88 in them after suppression of miR-155 (all P < 0.05). Furthermore, we also found complete reverse by TLR4/MyD88 pathway inhibitor on the influence of increasing miR-155 on cells (P > 0.05). Therefore, with an increase in cases with CIS, miR-155 takes a part in the development of cell damage by activating TLR4/MyD88, and it is probably the key to diagnosing and treating CIS.

Treatment of Neuroblastoma Cells with Inhibitors of Protein Disulfide Isomerase Upregulates NQO1 Activity.

Hallmarks of neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease include oxidative stress, accumulation of unfolded proteins, and neuronal cell death. One key player in maintaining redox homeostasis and oxidative protein folding is the protein disulfide isomerase (PDI). PDI has been the focus of drug discovery studies in neurodegenerative diseases, which have reported, paradoxically, that PDI inhibition is neuroprotective in cellular disease models. This study investigated the molecular implications of PDI inhibition by examining the effect of the PDI inhibitors securinine and 16F16 on the gene expression profile of SH-SY5Y neuroblastoma cells. Microarray analysis identified 36 genes that were differentially expressed in both inhibitor treatments. Computational approaches revealed that these differentially expressed genes are involved in apoptosis and cell death and that they are part of a protein-protein interaction network. Among the 36 identified genes, NAD(P)H quinone dehydrogenase 1 (NQO1) displayed the highest average expression change. As a central player in the cellular oxidative stress response, NQO1 was the focus of further investigation. Immunoblotting confirmed the increased expression level of NQO1, and activity assays demonstrated substantial increases in NQO1 activity in SH-SY5Y cells after treatment with PDI inhibitors. In summary, this study suggests a novel link between PDI inhibition and NQO1 activity, providing insights into the dynamic interplay between protein folding, oxidative stress, and cell death in neurodegenerative diseases, which can be exploited for drug development in the future.

Comparative Effects of Alpha- and Gamma-Tocopherol on Mitochondrial Functions in Alzheimer\u2019s Disease In Vitro Model

Vitamin E acts as an antioxidant and reduces the level of reactive oxygen species (ROS) in Alzheimer’s disease (AD). Alpha-tocopherol (ATF) is the most widely studied form of vitamin E besides gamma-tocopherol (GTF) which also shows beneficial effects in AD. The levels of amyloid-beta (Aβ) and amyloid precursor protein (APP) increased in the brains of AD patients, and mutations in the APP gene are known to enhance the production of Aβ. Mitochondrial function was shown to be affected by the increased level of Aβ and may induce cell death. Here, we aimed to compare the effects of ATF and GTF on their ability to reduce Aβ level, modulate mitochondrial function and reduce the apoptosis marker in SH-SY5Y cells stably transfected with the wild-type or mutant form of the APP gene. The Aβ level was measured by ELISA, the mitochondrial ROS and ATP level were quantified by fluorescence and luciferase assay respectively whereas the complex V enzyme activity was measured by spectrophotometry. The expressions of genes involved in the regulation of mitochondrial membrane permeability such as voltage dependent anion channel (VDAC1), adenine nucleotide translocase (ANT), and cyclophilin D (CYPD) were determined by quantitative real-time polymerase chain reaction (qRT-PCR), while the expressions of cyclophilin D (CypD), cytochrome c, Bcl2 associated X (BAX), B cell lymphoma-2 (Bcl-2), and pro-caspase-3 were determined by western blot. Our results showed that mitochondrial ROS level was elevated accompanied by decreased ATP level and complex V enzyme activity in SH-SY5Y cells expressing the mutant APP gene (p < 0.05). Treatment with both ATF and GTF reduced the mitochondrial ROS level with maximum reduction was observed in the cells treated with high concentrations of ATF and GTF (p < 0.05). However, only GTF at 80 µM significantly increase the ATP level and complex V enzyme activity (p < 0.05). VDAC1 and CYPD were downregulated and CypD protein was significantly overexpressed in cells transfected with the wild-type (WT) and mutant APP gene (p < 0.05). Cytochrome c release, the ratio of BAX/Bcl-2, and pro-caspase-3 expression increased in cells expressing mutated APP gene (p < 0.05). The expression of CypD and pro-caspase 3 protein, and the ratio of BAX/Bcl-2 were increased in the following order; SH-SY5Y-APP-WT < SH-SY5Y-APP Swe <SH-SY5Y-APP Swe/Ind. Treatment with both ATF and GTF reduced the release of cytochrome c and the ratio of BAX/Bcl-2. However, only GTF significantly reduced the expression of CypD and pro-caspase-3, suggestive of its unique role in AD. In conclusion, GTF has an effect that was not shown by ATF and thus suggest its potential role in the development of therapeutic agents for AD.

Chinese Veterinary Medicine B307 Promotes Cardiac Performance and Skeletal Muscle Contraction via Enhancing Intracellular Calcium Levels and Neural Electrical Activity in Animal and Cell Models

The study mainly investigated the effects of Chinese veterinary medicine B307 in cardiac and motor functions in animal models of pigeons and mice. Related cellular mechanisms were also studied in the neuroblastoma cell model of SH-SY5Y. Cardiac functions of pigeons and mice were examined by using moorFLPI Laser color Doppler imager and M-mode echocardiography, and motor functions were examined by using muscle electrical stimulation and force recording in the isolated breast muscle. Intracellular calcium levels and electrical activity of SH-SY5Y cells were examined by using Fura 2-AM fluorescence and MED64 system separately. Our results in vivo found that those pigeons under oral B307 treatment obviously enhanced subcutaneous microcirculation and contractile force and prolonged fatigue time in their breast muscles. Those mice under oral B307 treatment obviously elevated ejection fraction and cardiac output in their hearts. Our results in vitro showed that those SH-SY5Y cells under B307 treatment obviously increased intracellular calcium mobilization and electrical activities. These results revealed that improvement of cardiac and motor functions under B307 treatments may be caused by increasing electrical activities and intracellular calcium levels in neuromuscular cells and a similar mechanism may also occur in muscle cells. Thus, we suggested that B307 can be a functional Chinese veterinary medicine for flying pigeons.

Butein attenuates the cytotoxic effects of LPS-stimulated microglia on the SH-SY5Y neuronal cell line

Neuroinflammation is involved in brain aging and neuronal cell death in neurodegenerative diseases such as Alzheimer's disease (AD). Butein has been suggested to have anti-inflammatory, anti-apoptotic, and anti-cancer effects. However, few studies have been done to evaluate whether butein exerts protective effects on neurons, and the potential mechanism for this effect has not been studied. Here, we examined the effect of butein on SH-SY5Y neuroblastoma cells grown with conditioned medium from BV2 microglia cells that had been activated by lipopolysaccharide (LPS) as a neuroinflammation model. We found butein pretreatment significantly increased SH-SY5Y cell viability in a dose-dependent manner by inhibiting the apoptosis normally induced by microglia-conditioned medium. SH-SY5Y cells treated with microglia-conditioned medium showed upregulated ERK signaling pathway-related mRNA expression and protein phosphorylation, which was dose-dependently reversed by butein. Immunocytochemistry and Western blot results showed that BV2-LPS conditioned medium-induced Nuclear factor kappaB (NF-κB) transactivational activity in SH-SY5Y cells, but this was attenuated by butein treatment of the BV2 cells prior to their exposure to LPS. Collectively, our results indicate that butein effectively mitigates inflammatory injury caused by LPS-conditioned medium from microglia, possibly due to reductions in the transactivational activity of NF-κB p65 and ERK signaling pathway activation, and provide evidence for a neuroprotective role of butein through blocking negative consequences of microglial activation.

Synergistic Effects of Curcumin and Piperine as Potent Acetylcholine and Amyloidogenic Inhibitors With Significant Neuroprotective Activity in SH-SY5Y Cells via Computational Molecular Modeling and in vitro Assay.

Hallmarks of Alzheimer’s disease (AD) pathology include acetylcholine (ACh) deficiency and plaque deposition. Emerging studies suggest that acetylcholinesterase (AChE) may interact with amyloid β (Aβ) to promote aggregation of insoluble Aβ plaques in brains of patients. Current therapeutic options available for AD patients, such as AChE inhibitors, provide only symptomatic relief. In this study, we screened four natural compounds believed to harbor cognitive benefits—curcumin, piperine, bacoside A, and chebulinic acid. In the first section, preliminary screening through computational molecular docking simulations gauged the suitability of the compounds as novel AChE inhibitors. From here, only compounds that met the in silico selection criteria were selected for the second section through in vitro investigations, including AChE enzyme inhibition assay, 3-(4,5-dimenthylthiazol-2-yl)-2,5-dimethyltetrazolium bromide (MTT) assay, Thioflavin T (ThT) assay, and biochemical analysis via a neuronal cell line model. Of the four compounds screened, only curcumin (−9.6 kcal/mol) and piperine (−10.5 kcal/mol) showed favorable binding affinities and interactions towards AChE and were hence selected. In vitro AChE inhibition demonstrated that combination of curcumin and piperine showed greater AChE inhibition with an IC50 of 62.81 ± 0.01 μg/ml as compared to individual compounds, i.e., IC50 of curcumin at 134.5 ± 0.06 μg/ml and IC50 of piperine at 76.6 ± 0.08 μg/ml. In the SH-SY5Y cell model, this combination preserved cell viability up to 85%, indicating that the compounds protect against Aβ-induced neuronal damage (p < 0.01). Interestingly, our results also showed that curcumin and piperine achieved a synergistic effect at 35 μM with an synergism quotient (SQ) value of 1.824. Synergistic behavior indicates that the combination of these two compounds at lower concentrations may provide a better outcome than singularly used for Aβ proteins. Combined curcumin and piperine managed to inhibit aggregation (reduced ThT intensity at 0.432 a.u.; p < 0.01) as well as disaggregation (reduced ThT intensity at 0.532 a.u.; p < 0.01) of fibrillar Aβ42. Furthermore, combined curcumin and piperine reversed the Aβ-induced up-regulation of neuronal oxidative stress (p < 0.01). In conclusion, curcumin and piperine demonstrated promising neuroprotective effects, whereas bacoside A and chebulinic acid may not be suitable lead compounds. These results are hoped to advance the field of natural products research as potentially therapeutic and curative AD agents.

Dipropargyl substituted diphenylpyrimidines as dual inhibitors of monoamine oxidase and acetylcholinesterase

Alzheimer's disease (AD) is a multifactorial neurological disorder involving complex pathogenesis. Single target directed drugs proved ineffective and since last few years' different pharmacological strategies including multi-targeting agents are being explored for the effective drug development for AD. A total of 19 dipropargyl substituted diphenylpyrimidines have been synthesized and evaluated for the monoamine oxidase (MAO) and acetylcholinesterase (AChE) inhibition potential. All the compounds were found to be selective and reversible inhibitors of MAO-B isoform. These compounds also displayed good AChE inhibition potential with IC50 values in low micromolar range. AVB4 was found to be the most potent MAO-B inhibitor with IC50 value of 1.49 ± 0.09 μM and AVB1 was found to be the most potent AChE inhibitor with IC50 value of 1.35 ± 0.03 μM. In the ROS protection inhibition studies, AVB1 and AVB4 displayed weak but interesting activity in SH-SY5Y cells. In the cytotoxicity studies involving SH-SY5Y cells, both AVB1 and AVB4 were found to be non-toxic to the tissue cells. In the molecular dynamic simulation studies of 30 ns, the potent compounds were found to be quite stable in the active site of MAO-B and AChE. The results suggested that AVB1 and AVB4 are promising dual inhibitors and have the potential to be developed as anti-Alzheimer's drug.

Glycyrrhizic Acid Alleviates 6-Hydroxydopamine and Corticosterone-Induced Neurotoxicity in SH-SY5Y Cells Through Modulating Autophagy.

Recent researches have shown that autophagy is associated with the pathogenesis of neurodegenerative disorders, but there is no paper to investigate the effects of autophagy modulation on Parkinson's disease depression (PDD). In addition, glycyrrhizic acid (GA), the major bioactive ingredient of Radix glycyrrhizae, can induce autophagy and ease rotenone-induced Parkinson's disease (PD). However, there is also no paper to study the action and molecular mechanisms of GA on PDD. In this research, we built the injury model of SH-SY5Y cells through 6-hydroxydopamine (6-OHDA) and corticosterone (CORT). Then, our results showed that GA markedly increased the viability and decreased the apoptosis in SH-SY5Y cells after pre-treating with 6-OHDA and CORT. Moreover, GA notably decreased the expressions of α-Syn and p-S1292-LRRK2 proteins, and significantly increased the levels of CREB and BDNF proteins. Previous papers have suggested that CORT contributed to dopaminergic neurodegeneration via the glucocorticoid (GC)/glucocorticoid receptor (GR) interaction, and our results showed that GA reduced GC level and hypothalamic-pituitary-adrenal (HPA) activity in SH-SY5Y cells by regulating GR signaling pathway. Furthermore, mechanism investigations also showed that GA had the ability to up-regulate the conversion of LC3B II/I and the expression of Beclin-1, and induce autophagy in SH-SY5Y cells, which were reversed by the autophagy inhibitor 3-methyladenine (3-MA). Collectively, these findings proved that GA exerted efficient activity against neurotoxicity in SH-SY5Y cells induced by 6-OHDA and CORT via activation of autophagy, which should be developed as an efficient candidate for treating PDD in the future.

Comparative Therapeutic Effects of Plant-Extract Synthesized and Traditionally Synthesized Gold Nanoparticles on Alcohol-Induced Inflammatory Activity in SH-SY5Y Cells In Vitro.

The present study describes potential beneficial and adverse effects of plant-extract synthesized gold nanoparticles (AuNPs) on ethanol toxicity in SH-SY5Y cells. Although kudzu root extract (K), edible-gum extract (G), alone or in combination (KG), reduced Au3+ into AuNPs, the extract’s composition and the reaction temperature determined their size (AuNPKG(90<50<37) << AuNPK (90,50<37) < AuNPG (90<50); the subscript KG, K, or G is extract identification and numerical vales are reaction temperature in Celsius) and biological properties (AuNPKG (90,50>37) << AuNPK (90,50>37) < AuNPG (90,50)). The surface of each AuNP contained the extract’s active ingredients, that were analyzed and confirmed using laser desorption ionization (LDI)) and low-matrix laser desorption-ionization (LMALDI). AuNPKG-50 was (i) least toxic to SH-SY5Y cells, but most effective in suppressing the adverse effects of ethanol on SH-SY5Y cells, and (ii) more effective than a combination of free kudzu and gum extracts. The beneficial and adverse effects of AuNPs may have been modified by the formation of proteins corona. This study provides a proof of concept for possible application of plant-extract synthesized AuNPs in mitigating ethanol toxicity.

SLM, a novel carbazole-based fluorophore attenuates okadaic acid-induced tau hyperphosphorylation via down-regulating GSK-3β activity in SH-SY5Y cells

Phosphorylated tau dissociates from microtubules and aggregates to form neurofibrillary tangles resulting in neuronal toxicity and cognitive deficits. Attenuating tau hyperphosphorylation is considered as an effective therapeutic approach for Alzheimer's disease (AD). From our previous study, SLM, a carbazole-based fluorophore prevents Aβ aggregation, reduced glycogen synthase kinase-3β (GSK-3β) activity and tau hyperphosphorylation in triple transgenic mouse model of AD. However, the mechanism by which SLM attenuates tau hyperphosphorylation warrants further investigation. In the current study, we intend to evaluate the effects of SLM against okadaic acid (OA)-induced tau hyperphosphorylation and microtubules instability in human neuroblastoma (SH-SY5Y) cells. The results showed that, SLM reduced the OA-induced cell neurotoxicity and tau hyperphosphorylation in SH-SY5Y cells. SLM treatment down-regulated GSK-3β activity. However, in the presence of GSK-3β inhibitor (SB216763, 10μM), SLM treatment could not reduce GSK-3β activity and tau hyperphosphorylation as compared with SB216763 treatment alone. Furthermore, SLM treatment also ameliorated OA-induced microtubules instability and cytoskeleton damage. Collectively, SLM attenuated OA-induced tau hyperphosphorylation via down-regulating GSK-3β activity in SH-SY5Y cells. Therefore, this study supports SLM as a potential compound for AD and other tau pathology-related neurodegenerative disorders.

The effect of UV-filters on the viability of neuroblastoma (SH-SY5Y) cell line

Topical application of cosmetic products, containing ultraviolet filters (UV filters) are recommended as a protection against sunburns and in order to reduce the risk of skin cancer. However, some UV filters can be absorbed through skin and by consuming contaminated food. Among the chemical UV filters, benzophenone-3 (BP-3), 3-(4-methylbenzylidene)camphor (4-MBC) and 2-ethylhexyl-4-methoxycinnamate (OMC) are absorbed through the skin to the greatest extent. So far, these lipophilic compounds were demonstrated to influence the gonadal and thyroid hormone function, but their effect on central nervous system cells has not been investigated, yet.In the present study, we investigated the effect of some UV filters on cell viability and caspase-3 activity in SH-SY5Y cells. It has been found that benzophenone-2 (BP-2), BP-3, 4-methylbenzophenone (4-MBP) and OMC present in the culture medium for 72h in high concentration (10−5 and 10−4M) and 4-MBC only 10−4M produced a significant cytotoxic effect, as determined both by the MTT reduction test and LDH release assay. In contrast to necrotic changes, all tested UV filters increased caspase-3 activity in much lower concentrations (from 10−8 to 10−7M). Proapoptotic properties of the test compounds were positively verified by Hoechst staining.The obtained results indicated that UV filters adversely affected the viability of nerve cells, most likely by enhancing the process of apoptosis. The most potent effect was exerted by BP-3 and 4-MBC and at concentrations that may be reached in vivo. Since human exposure to UV filters is significant these compound should be taken into consideration as one of the possible factors involved in pathogenesis of neurodegenerative diseases.

Nicotinamide N-methyltransferase increases complex I activity in SH-SY5Y cells via sirtuin 3

Nicotinamide N-methyltransferase (NNMT, E.C. 2.1.1.1) N-methylates nicotinamide to 1-methylnicotinamide. We have previously shown that NNMT is significantly overexpressed in the brains of patients who have died of Parkinson's disease, and others have shown that NNMT is significantly overexpressed in a variety of diseases ranging from cancer to hepatic cirrhosis. In vitro overexpression has revealed many cytoprotective effects of NNMT, in particular increased complex I activity and ATP synthesis. Although this appears to be mediated by an increase in 1-methylnicotinamide production, the molecular mechanisms involved remain unclear. In the present study, we have investigated the role that sirtuins 1, 2 and 3, class III DNA deacetylase enzymes known to regulate mitochondrial energy production and cell cycle, have in mediating the effects of NNMT upon complex I activity. Expression of NNMT in SH-SY5Y human neuroblastoma cells, which have no endogenous expression of NNMT, significantly increased the expression of all three sirtuins. siRNA-mediated silencing of sirtuin 3 expression decreased complex I activity in NNMT-expressing SH-SY5Y cells to that observed in wild-type SH-SY5Y, and significantly reduced cellular ATP content also. These results demonstrate that sirtuin 3 is a key mediator of NNMT-induced complex I activity and ATP synthesis. These results further reinforce a central role for NNMT in the regulation of energy homeostasis and provide further mechanistic insight into the consequences of enhanced NNMT expression.

Involvement of diacylglycerol produced by phospholipase D activation in Aβ-induced reduction of sAPPα secretion in SH-SY5Y neuroblastoma cells

We previously reported that the thiol proteinase inhibitor, E-64-d, ameliorated amyloid β (Aβ)-induced reduction of soluble amyloid precursor protein α (sAPPα) secretion by reversing ceramide-induced protein kinase C down-regulation in SH-SY5Y neuroblastoma cells. In the present study, we showed that Aβ (1–42) peptide enhanced diacylglycerol (DAG) production by phospholipase D (PLD) activation in these cells. We subsequently examined whether PLD was involved in Aβ-induced reduction of sAPPα secretion and showed that 2μM CAY10593, which selectively inhibits PLD2, ameliorated reduction of sAPPα secretion, whereas 50nM CAY10593, which selectively inhibits PLD1, did not. Moreover, 50µM propranolol, a phosphatidic acid phosphohydrolase inhibitor, also ameliorated Aβ-induced reduction of sAPPα secretion, suggesting that DAG may be responsible for Aβ-induced reduction of sAPPα. We subsequently examined whether DAG affects sAPPα secretion and showed that a DAG analog reduced sAPPα secretion in SH-SY5Y cells. In addition, DAG enhanced ceramide production by stimulating neutral sphingomyelinase (N-SMase) activity. We previously demonstrated that Aβ stimulates N-SMase activity in SH-SY5Y cells. Here, we showed that inhibition of PLD2 by 2μM CAY10593 suppressed Aβ-induced N-SMase activation. Taken together, the results suggest that DAG produced through the PLD pathway is involved in Aβ-induced reduction of sAPPα secretion in SH-SY5Y cells.

Anti-oxidative and anti-inflammatory property of ethanol extracts of Chungyul medicines in human neuroblastoma cells, SH-SY5Y

Previous studies demonstrated that Prunella vulgaris, Scutellaria baicalensis and Lonicera japonica have Chungyul activity, anti-inflammatory activity, anti-oxidant activity, and anti-cancer activity. They are also known to have the effect on degenerative cerebropathia of cranial nerve system, cerebrovascular diseases and brain tumors. We selected those herbs (Chungyul medicines herbs) and performed experiments to investigate whether they have anti-oxidant and anti-inflammatory effects in human neuroblastoma cells, SH-SY5Y. We found that Prunella vulgaris has the highest total phenolic contents and DPPH radical scavenging activity suggesting that DPPH scavenging activity is associated with total phenolic level of the extracts. The highest superoxide radical scavenging activity was also shown in Prunella vulgaris. We also observed that Prunella vulgaris, Scutellaria baicalensis, and Lonicera japonica suppressed 6-hydroxydopamine (6-OHDA)-induced TBARS increase suggesting that Chungyul medicines herbs have antioxidant activity. Additionally, catalase (CAT) level which was reduced by 6-OHDA was restored by Chungyul medicines. The herbs reduced NO production induced by 6-OHDA and Prunella vulgaris decreased cell viability in a dose-dependent manner. Prunella vulgaris suppressed 6-OHDA-induced expression of iNOS and COX-2. Our study demonstrates that Chungyul medicines herbs have anti-oxidant and anti-inflammatory activity in SH-SY5Y cells and Prunella vulgaris seems to be the most effective herb for the treatment of inflammatory diseases.