Pathway In SH-SY5Y Cells Research Articles

Schizophrenia risk candidate EGR3 is a novel transcriptional regulator of RELN and regulates neurite outgrowth via the Reelin signal pathway in vitro.

Schizophrenia is a severe psychiatric disorder with a strong hereditary component that affects approximately 1% of the world's population. The disease is most likely caused by the altered expression of a number of genes that function at the level of biological pathways or gene networks. Transcription factors (TF) are indispensable regulators of gene expression. EGR3 is a TF associated with schizophrenia. In the current study, DNA microarray and ingenuity pathway analyses (IPA) demonstrated that EGR3 regulates Reelin signaling pathway in SH-SY5Y cells. ChIP and luciferase reporter studies confirmed that EGR3 directly binds to the promoter region of RELN thereby activating RELN expression. The expression of both EGR3 and RELN was decreased during neuronal differentiation induced by retinoic acid (RA) in SH-SY5Y cells, and EGR3 over-expression reduced neurite outgrowth which could be partially reversed by the knockdown of RELN. The expression levels of EGR3 and RELN in peripheral blood of subjects with schizophrenia were found to be down-regulated (compared with healthy controls), and were positively correlated. Furthermore, data mining from public databases revealed that the expression levels of EGR3 and RELN were presented a positive correlation in post-mortem brain tissue of subjects with schizophrenia. Taken together, this study suggests that EGR3 is a novel TF of the RELN gene and regulates neurite outgrowth via the Reelin signaling pathway. Our findings contribute to the understanding of the regulatory role of EGR3 in the pathophysiology and molecular mechanisms of schizophrenia, and potentially to the development of new therapies and diagnostic biomarkers for the disorder.

Protective effects of Buyinqianzheng Formula on mitochondrial morphology by PINK1/Parkin pathway in SH-SY5Y cells induced by MPP+

Buyinqianzheng Formula (BYQZF) is clinically employed in traditional Chinese medicine to treat Parkinson’s disease (PD) by improving mitochondrial dysfunction. However, the underlying mechanisms by which BYQZF affects mitochondrial morphology remain unknown. Therefore, we observed the effects of BYQZF on mitochondria from the perspective of the PINK1/Parkin pathway. Cell survival rates were assessed by Cell Counting Kit-8 assay. Expression levels of PINK1 and Parkin mRNA were examined by qRT-PCR. Protein expression levels of PINK1, PINK1-Ser228, Parkin, Parkin-Ser65, Drp1, and Drp1-Ser637 were examined by western blotting. PINK1, Parkin, and MitoTracker® Red CMXRos (MTR) were stained by triple-labeled immunofluorescence, and observed under laser confocal microscopy. Cell survival rate, mitochondrial form factor, mean length and number of mitochondrial network branches, mitochondrial activity, mRNA expression levels of PINK1 and Parkin, and protein expression levels of PINK1, Parkin, and Drp1-Ser637 were reduced after 1-methyl-4-phenylpyridinium (MPP + ) intervention. In contrast, Pearson’s correlation coefficients between PINK1 and Parkin, and between Parkin and MTR, as well as protein expression levels of PINK1-Ser228, Parkin-Ser65, and Drp1 increased significantly after MPP + intervention. Treatment with BYQZF increased cell survival rate, mitochondrial form factor, mean length and number of mitochondrial network branches, mitochondrial activity, mRNA expression levels of PINK1 and Parkin , and expression of PINK1, Parkin, and Drp1-Ser637 proteins. Pearson’s correlation coefficients between PINK1 and Parkin, and between Parkin and MTR, as well as protein expression levels of PINK1 -Ser228, Parkin -Ser65, and Drp1 decreased after BYQZF treatment. These results demonstrate that BYQZF has a protective effect on mitochondrial molecular mechanisms in the PD cell model, and the mechanism is related to the PINK1/Parkin pathway.

Speedy/RINGO protein interacts with ERK/MAPK and PI3K/AKT pathways in SH-SY5Y neuroblastoma cells.

Abnormal activity of ERK/MAPK and PI3K/AKT pathways is one of the most important factors for the development of many cancer types including neuroblastoma cancer. Apart from these two pathways, some cell cycle regulators such as Speedy/RINGO also contribute to neuroblastoma development. There is data reinforcing the possible communication of the components of ERK/MAPK and PI3K/AKT pathways in carcinogenic process. In addition to this, there are studies about the direct/indirect interaction of Speedy/RINGO with these pathways in different cell types other than neuroblastoma. However, there is not any study available showing the interaction of Speedy/RINGO with both pathways in neuroblastoma cells. Therefore, the aim of this study is to determine the possible effect of Speedy/RINGO on PI3K/AKT and ERK/MAPK pathways in SH-SY5Y neuroblastoma cells. For this aim, Speedy/RINGO was silenced by siRNA technique to analyze the effects of direct inhibition of Speedy/RINGO on these pathways. Results showed that Speedy/RINGO silencing caused a significant decrease in MEK1/2 expression and AKT phosphorylation. Afterward, MEK1/2 was inhibited using a specific inhibitor U0126. Data reveal a corresponding decrease in the Speedy/RINGO expression and AKT phosphorylation indicating a reciprocal interaction between ERK/MAPK and Speedy/RINGO. In addition, MTS analysis showed that both ERK/MAPK inhibition and Speedy/RINGO silencing significantly reduced the viability of SH-SY5Y cells. This study provides information about a possible interaction of Speedy/RINGO with PI3K/AKT and ERK/MAPK pathways in SH-SY5Y cells for the first time. It will not only help to better understand the cancer-prone interactions of these pathways but also enable us to identify the appropriate molecular targets for developing efficient treatment strategies.

HIF1α-dependent mitophagy facilitates cardiomyoblast differentiation.

Mitophagy is thought to play a key role in eliminating damaged mitochondria, with diseases such as cancer and neurodegeneration exhibiting defects in this process. Mitophagy is also involved in cell differentiation and maturation, potentially through modulating mitochondrial metabolic reprogramming. Here we examined mitophagy that is induced upon iron chelation and found that the transcriptional activity of HIF1α, in part through upregulation of BNIP3 and NIX, is an essential mediator of this pathway in SH-SY5Y cells. In contrast, HIF1α is dispensable for mitophagy occurring upon mitochondrial depolarisation. To examine the role of this pathway in a metabolic reprogramming and differentiation context, we utilised the H9c2 cell line model of cardiomyocyte maturation. During differentiation of these cardiomyoblasts, mitophagy increased and required HIF1α-dependent upregulation of NIX. Though HIF1α was essential for expression of key cardiomyocyte markers, mitophagy was not directly required. However, enhancing mitophagy through NIX overexpression, accelerated marker gene expression. Taken together, our findings provide a molecular link between mitophagy signalling and cardiomyocyte differentiation and suggest that although mitophagy may not be essential per se, it plays a critical role in maintaining mitochondrial integrity during this energy demanding process.

Neuroprotective Activity of Mentha Species on Hydrogen Peroxide-Induced Apoptosis in SH-SY5Y Cells.

Alzheimer’s disease (AD) is a progressive neurodegenerative disorder with an unclear cause. It appears that multiple factors participate in the process of neuronal damage including oxidative stress and accumulation of the protein amyloid β (Aβ) in the brain. The search for a treatment for this disorder is essential as current medications are limited to alleviating symptoms and palliative effects. The aim of this study is to investigate the effects of mint extracts on selected mechanisms implicated in the development of AD. To enable a thorough investigation of mechanisms, including effects on β-secretase (the enzyme that leads to the formation of Aβ), on Aβ aggregation, and on oxidative stress and apoptosis pathways, a neuronal cell model, SH-SY5Y cells, was selected. Six Mentha taxa were investigated for their in vitro β-secretase (BACE) and Aβ-aggregation inhibition activities. Moreover, their neuroprotective effects on H2O2-induced oxidative stress and apoptosis in SH-SY5Y cells were evaluated through caspase activity. Real-time PCR and Western blot analysis were carried out for the two most promising extracts to determine their effects on signalling pathways in SH-SY5Y cells. All mint extracts had strong BACE inhibition activity. M. requienii extracts showed excellent inhibition of Aβ-aggregation, while other extracts showed moderate inhibition. M. diemenica and M. requienii extracts lowered caspase activity. Exposure of SH-SY5Y cells to M. diemenica extracts resulted in a decrease in the expression of pro-apoptotic protein, Bax, and an elevation in the anti-apoptotic protein, Bcl-xL, potentially mediated by down-regulation of the ASK1-JNK pathway. These results indicate that mint extracts could prevent the formation of Aβ and also could prevent their aggregation if they had already formed. M. diemenica and M. requienii extracts have potential to suppress apoptosis at the cellular level. Hence, mint extracts could provide a source of efficacious compounds for a therapeutic approach for AD.

Pyrroloquinoline Quinine and LY294002 Changed Cell Cycle and Apoptosis by Regulating PI3K-AKT-GSK3β Pathway in SH-SY5Y Cells

To verify the role of PI3K-AKT-GSK3β pathway during manganese (Mn)-induced cell death, apoptosis, related indicators were investigated. SH-SY5Y cells were directly exposed to different concentrations of MnCl2. Then, cell viability, apoptosis, necrosis rate, and cell cycle were detected by MTT, FITC Annexin V Apoptosis Detection Kit with PI and PI staining. Then, in two intervention groups, cells were preconditioned with agonist (PQQ) and suppressant (LY294002). The cell viability decreased with a dose-response relationship (p < 0.05), while apoptosis and necrosis increased (p < 0.05). The ratio of G0/G1 and G2/M also decreased, but the percentage of S phase increased (p < 0.05). During above process, PI3K-AKT-GSK3β pathway was involved by regulating the expression of PI3K, AKT, p-AKT, and GSK3β (p < 0.05). For further research, cell cycle and apoptosis were detected pretreatment with PQQ and LY294002 before Mn exposure. The result showed cell ability, apoptosis, and necrosis rate changed obviously compared with non-pretreated group (p < 0.05). The variance of G0/G1 and G2/M ratio and percentage of S phase were also different, especially in 2.0mM (p < 0.05). Mn can cause apoptosis and necrosis, varying cell cycle of SH-SY5Y cells, which could be changed by PQQ and LY294002 by regulating PI3K-AKT-GSK3β pathway.

MiR-18b attenuates cerebral ischemia/reperfusion injury through regulation of ANXA3 and PI3K/Akt signaling pathway

Recent, research has displayed that the disorders of miR-18b are related to ischemic stroke. Here, we aimed to investigate the underlying neuroprotective mechanism of miR-18b in cerebral ischemia/reperfusion (I/R) injury. Oxygen-glucose deprivation/reperfusion (OGDR) model in vitro and middle cerebral artery occlusion (MCAO) model in vivo were established to simulate cerebral I/R injury. RT-PCR, western blotting, CCK-8, TUNEL, and TTC staining assays were applied in this study to explore the effect of miR-18b on cerebral I/R injury. Results displayed that miR-18b expression was reduced after cerebral I/R injury. Besides, miR-18b showed neuroprotective effects on cerebral I/R injury both in vitro and in vivo, These neuroprotective effects included promoting cell viability, decreasing cell apoptosis, reducing the production of inflammatory cytokines in SH-SY 5Y cells after OGDR and depressing MCAO-induced infarct size, neurological deficits and apoptotic cells in mice. Moreover, miR-18b negatively regulated ANXA3 expression, and its neuroprotection on cerebral I/R injury was overturned by ANXA3. Additionally, increasing miR-18b or decreasing ANXA3 promoted the activation of the PI3K/Akt signaling pathway in SH-SY 5Y cells after cerebral I/R injury. In conclusion, these data indicate that miR-18b protects against cerebral I/R injury by inhibiting ANXA3 and activating PI3K/Akt pathway, which provides a promising therapeutic target for ischemic stroke therapy.

Ozone alleviates ischemia/reperfusion injury by inhibiting mitochondrion-mediated apoptosis pathway in SH-SY5Y cells.

Cerebral ischemia/reperfusion (I/R) injuries are common and often cause severe complications. Ozone has been applied for protecting I/R injury in animal models of several organs including cerebra, but the detailed mechanism remains unclear. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and lactate dehydrogenase measurement were used to determine the influence of ozone on cell activity and damage of SH-SY5Y cells. Some redox items such as catalase (CAT), malondialdehyde (MDA), glutathione peroxidase (GSH-Px), and superoxide dismutase (SOD) were measured by enzyme-linked immunosorbent assay. The mitochondrial membrane potential (ΔΨm ) was determined by JC-1 assay. Cytochrome-c (cyt-c) level in the cytoplasm and mitochondrion was measured by western blotting. Apoptosis was determined by flow cytometry, and some apoptosis-related molecules were detected by quantitative real-time polymerase chain reaction and western blotting. Ozone alleviated oxidative damage by increasing GSH-Px, SOD, CAT, and decreasing MDA. Ozone decreased mitochondrial damage caused by I/R injury and inhibited the release of cyt-c from mitochondrion to cytoplasm in SH-SY5Y cells. The cell apoptosis caused by I/R was inhibited by ozone, and ozone could decrease apoptosis by increasing the ratio of Bcl-2/Bax and inhibiting caspase signaling pathway in SH-SY5Y cells. Ozone has the ability of maintaining redox homeostasis, decreasing mitochondrion damage, and inhibiting neurocytes apoptosis induced by I/R. Therefore, ozone may be a promising protective strategy against cerebral I/R injury.

Berberine alleviates rotenone-induced cytotoxicity by antioxidation and activation of PI3K/Akt signaling pathway in SH-SY5Y cells.

Berberine, an isoquinoline alkaloid isolated from traditional Chinese medicine, has been widely studied for its efficacy in the treatment of neurodegenerative diseases. However, berberine-mediated neuroprotection in the pathogenesis of Parkinson's disease is still uncertain. In this study, the effects of berberine on rotenone-induced neurotoxicity in SH-SY5Y cells were investigated. The results showed that berberine treatment significantly alleviated rotenone-induced decrease in the cell viability in SH-SY5Y cells. Further studies demonstrated that berberine suppressed the production of intracellular reactive oxygen species, restored the mitochondrial transmembrane potential, increased Bcl-2/Bax ratio, and decreased caspase-3 activation that induced by rotenone. Furthermore, berberine also restored the phosphorylation of Akt, which was downregulated by rotenone in SH-SY5Y cells. These results suggest that berberine protects rotenone-treated SH-SY5Y cells by antioxidation and activation of PI3K/Akt signaling pathway.

SNHG1 promotes MPP+-induced cytotoxicity by regulating PTEN/AKT/mTOR signaling pathway in SH-SY5Y cells via sponging miR-153-3p

BackgroundLong non-coding RNA small molecule RNA host gene 1 (SNHG1) was previously identified to be relevant with Parkinson’s disease (PD) pathogenesis. This work aims to further elucidate the regulatory networks of SNHG1 involved in PD.Methods1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-hydrochloride (MPTP)-induced mice and 1-methyl-4-phenylpyridinium (MPP+)-treated SH-SY5Y cells were respectively constructed as the in vivo and in vitro PD models. Expression levels of SNHG1 and miR-153-3p were detected by qRT-PCR. Protein expression levels of phosphate and tension homology deleted on chromosome ten (PTEN) were measured by western blotting assay. Cell viability and apoptosis were determined by MTT and flow cytometry assays. The interactions among SNHG1, miR-153-3p and PTEN were identified by luciferase reporter assay, RNA immunoprecipitation, and/or RNA pull-down analysis.ResultsIncreased SNHG1 expression was found in midbrain of MPTP-induced PD mice and MPP+-treated SH-SY5Y cells. Overexpression of SNHG1 lowered viability and enhanced apoptosis in MPP+-treated SH-SY5Y cells. Moreover, SNHG1 acted as a molecular sponge to inhibit the expression of miR-153-3p. Furthermore, miR-153-3p-mediated suppression of MPP+-induced cytotoxicity was abated following SNHG1 up-regulation. Additionally, PTEN was identified as a direct target of miR-153-3p, and SNHG1 could serve as a competing endogenous RNA (ceRNA) of miR-153-3p to improve the expression of PTEN. Besides, enforced expression of PTEN displayed the similar functions as SNHG1 overexpression in regulating the viability and apoptosis of MPP+-treated SH-SY5Y cells. Finally, SNHG1 was found to activate PTEN/AKT/mTOR signaling pathway in SH-SY5Y cells by targeting miR-153-3p.ConclusionSNHG1 aggravates MPP+-induced cellular toxicity in SH-SY5Y cells by regulating PTEN/AKT/mTOR signaling via sponging miR-153-3p, indicating the potential of SNHG1 as a promising therapeutic target for PD.

Schizandrol A protects against Aβ1-42-induced autophagy via activation of PI3K/AKT/mTOR pathway in SH-SY5Y cells and primary hippocampal neurons.

Autophagy, a lysosomal degradative pathway, is crucial for the pathogenesis of Alzheimer's disease (AD). Schizandrol A (SchA) shows multiple pharmacological effects. However, the potential effects and mechanisms of SchA on amyloid-β (Aβ)-induced autophagy remain unclear. In this study, differentiated SH-SY5Y cells or primary hippocampal neurons were pretreated with SchA (2μg/ml) for 1h before subjected to Aβ1-42 (10μM) for 24h to test its effects on cell viability, apoptosis, oxidative stress, and autophagy. Then an mTOR inhibitor (rapamycin) and a PI3K inhibitor (LY294002) were employed to explore the role of PI3K/AKT/mTOR pathway. The results showed that SchA significantly inhibited Aβ1-42-triggered reduction of viable cells, increases of apoptotic cell number and pro-apoptotic protein expressions, as well as alterations of oxidative stress markers. In addition, the increases of LC3-II/LC3-I and Beclin-1 and decrease of p62 were suppressed by SchA. At the molecular level, we found that the inactivation of PI3K/AKT/mTOR pathway was ameliorated by SchA. Inhibition of PI3K/AKT/mTOR pathway deteriorated the protective effects of SchA against Aβ1-42-induced autophagy activation, cell death, and apoptosis. In conclusion, we demonstrate that SchA attenuates Aβ1-42-induced autophagy through activating PI3K/AKT/mTOR signaling pathway. SchA may be a novel drug for the prevention and treatment of AD.

Tetramethylpyrazine Attenuated Sevoflurane-Induced Neurotoxicity by Enhancing Autophagy through GPR50/CREB Pathway in SH-SY5Y Cells.

Tetramethylpyrazine has shown neuroprotective and axonal outgrowth-promoting effects and can improve cognitive deficit in a rat model of chronic hypoperfusion. However, the role of tetramethylpyrazine in sevoflurane-induced neurotoxicity is still vague. Therefore, this study was designed to investigate the effects and mechanisms of tetramethylpyrazine on sevoflurane-induced autophagy, apoptosis, and the expression of BACE1 and A[Formula: see text] in SH-SY5Y cells. We measured the expression levels of the apoptosis protein markers Bax and Bcl-2, autophagy protein markers Atg5 and LC3-II, BACE1, and A[Formula: see text] in SH-SY5Y cells after sevoflurane treatment and determined the effects of tetramethylpyrazine on sevoflurane-induced expression of these proteins after silencing GPR50 or Atg5 with siRNA in vitro. We found that exposure to 3.4% sevoflurane for 6 h decreased the expression of autophagy protein markers and increased the expression of the apoptosis protein markers, BACE1, and A[Formula: see text] in SH-SY5Y cells. The number of red puncta (autolysosomes) and yellow puncta (autophagosomes) in each SH-SY5Y cell decreased after transient transfection with the mRFP-GFP-LC3 expression plasmid. Silencing of GPR50 decreased the expression of pCREB, Atg5, and LC3-II, while silencing of Atg5 increased the expression of BACE1 and A[Formula: see text] in SH-SY5Y cells. Our results demonstrate that tetramethylpyrazine attenuated sevoflurane-induced neurotoxicity by enhancing autophagy through the GPR50/CREB pathway in SH-SY5Y cells.

Withdrawal: Cai, H., Tao, X., Zheng, L., Huang, L., Peng, Y., Liao, R. and Zhu, Y. (2019), Ozone alleviates ischemia-reperfusion injury by inhibiting mitochondrion-mediated apoptosis pathway in SH-SY5Y cells. Cell Biol Int. Accepted Author Manuscript. https://doi.org/10.1002/cbin.11087.

The above article, published online on 20 December 2018 in Wiley Online Library (https://doi.org/10.1002/cbin.11087), has been withdrawn by agreement with the journal Editor, Sergio Schenkman, and John Wiley & Sons Ltd. The withdrawal has been agreed because the authors have not provided a signed Copyright Transfer Agreement.

Tanshinone IIA attenuates A\u03b2-induced neurotoxicity by down-regulating COX-2 expression and PGE2 synthesis via inactivation of NF-\u03baB pathway in SH-SY5Y cells

Amyloid-β (Aβ)-induced neurotoxicity is a major pathological mechanism of Alzheimer’s disease (AD). Tanshinone IIA (Tan IIA), extracted from traditional Chinese herb Radix salvia miltiorrhiza, possesses anti-oxidant and anti-inflammatory actions, as well as neuroprotective effects. The present study aims to explore the possible mechanism by which Tan IIA attenuated Aβ-induced neurotoxicity. Exposure of SH-SY5Y cells to different concentrations of Aβ led to neurotoxicity by reducing cell viability, inducing cell apoptosis and increasing neuroinflammation in a dose-dependent manner. Moreover, Aβ treatment promoted cyclooxygenase-2 (COX-2) expression and Prostaglandin E2 (PGE2) secretion, and activated nuclear transcription factor kappa (NF-κB) pathway in SH-SY5Y cells. However, pretreatment of SH-SY5Y cells with Tan IIA prior to Aβ prevented these Aβ-induced cellular events noticeably. These data suggested that Tan IIA exerted its neuroprotective action by alleviating Aβ-induced increase in COX-2 expression and PGE2 secretion via inactivation of NF-κB pathway.

Neuroprotection of miR-214 against isoflurane-induced neurotoxicity involves the PTEN/PI3K/Akt pathway in human neuroblastoma cell line SH-SY5Y

Isoflurane, one of the commonly used inhalation anesthetics worldwide in clinical practice, may generate substantial risks of neurotoxicity in the developing brains. The present study aimed to illustrate the effects and underlying mechanisms of miR-214 on isoflurane-induced neurotoxicity in human neuroblastoma cell line SH-SY5Y. SH-SY5Y cells were transfected with miR-214 or miR-con alone or in combination with pcDNA empty vector or pcDNA-PTEN in the presence of 3% isoflurane and incubated for 48 h. Cell viability, lactate dehydrogenase (LDH) release, apoptosis, and caspase-3/7 activity were evaluated using CCK-8, LDH release assay, flow cytometry analysis, and caspase-3/7 activity assay, respectively. The superoxide dismutase (SOD), glutathione (GSH), and malondialdehyde (MDA) activities were measured using commercial kits. miR-214 expression and alterations of the phosphatase and tensin homolog (PTEN)/phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) pathway were detected by qRT-PCR and Western blot, respectively. The interaction between miR-214 and PTEN was explored by luciferase reporter assay. We found that isoflurane exposure induced neurotoxicity in SH-SY5Y cells, as evidenced by the reduced cell viability, increased LDH release, apoptotic rate, caspase-3/7 activity, and oxidative stress levels. Moreover, isoflurane exposure decreased the expression of miR-214 and affected the PTEN/PI3K/Akt pathway in SH-SY5Y cells. miR-214 overexpression significantly suppressed isoflurane-induced viability reduction, LDH release, apoptosis and oxidative stress, as well as inactivation of the PI3K/Akt pathway in SH-SY5Y cells. Interestingly, PTEN was identified as a target of miR-214. Moreover, PTEN upregulation blocked the effects of miR-214 on isoflurane-induced neurotoxicity in SH-SY5Y cells. In conclusion, miR-214 protected against isoflurane-induced neurotoxicity in SH-SY5Y cells via regulation of PI3K/Akt pathway by targeting PTEN, contributing to better understanding the underlying mechanisms of anesthetics-induce neurotoxicity.

Manganese activates autophagy to alleviate endoplasmic reticulum stress–induced apoptosis via PERK pathway

Overexposure to manganese (Mn) is neurotoxic. Our previous research has demonstrated that the interaction of endoplasmic reticulum (ER) stress and autophagy participates in the early stage of Mn‐mediated neurotoxicity in mouse. However, the mechanisms of ER stress signalling pathways in the initiation of autophagy remain confused. In the current study, we first validated that ER stress–mediated cell apoptosis is accompanied by autophagy in SH‐SY5Y cells. Then, we found that inhibiting ER stress with 4‐phenylbutyrate (4‐PBA) decreased ER stress–related protein expression and reduced cell apoptosis, whereas blocking autophagy with 3‐methyladenine (3‐MA) increased cell apoptosis. These data indicate that protective autophagy was activated to alleviate ER stress–mediated apoptosis. Knockdown of the protein kinase RNA‐like ER kinase (PERK) gene inhibited Mn‐induced autophagy and weakened the interaction between ATF4 and the LC3 promoter. Our results reveal a novel molecular mechanism in which ER stress may regulate autophagy via the PERK/eIF2α/ATF4 signalling pathway. Additionally, Mn may activate protective autophagy to alleviate ER stress–mediated apoptosis via the PERK/eIF2α/ATF4 signalling pathway in SH‐SY5Y cells.

Curcumin inhibits BACE1 expression through the interaction between ERβ and NFκB signaling pathway in SH-SY5Y cells.

Alzheimer's disease (AD) is the leading cause of dementia, which characterized by toxic senile plaques is composed of amyloid-β (Aβ). β-site amyloid precursor protein-cleaving enzyme 1 (BACE1) is the rate-limiting protease in Aβ generation. Therefore, pharmacology BACE1 inhibition is one of the prime targets for potential treatment of AD. Curcumin, a yellow polyphenol derived from the rhizomes of the plant Curcuma longa Linn, has been reported to cross the blood-brain barrier and prevent Aβ aggregation in AD models. However, its neuroprotective mechanism is still unclear. In the present study, we find that curcumin markedly reduces Aβ levels in HEK293-APPswe cells. Our results show that curcumin inhibits BACE1 gene expression in SH-SY5Y cells at transcriptional and translational levels. Furthermore, we reveal that nuclear factor kappa B (NFκB) signaling is involved in the regulation of curcumin on BACE1. Interestingly, the estrogenicity of curcumin is found to partially contribute to its protective action. Our data show that curcumin activates estrogen receptor β (ERβ) selectively and the activation of ERβ directly effects on the upstream factors of the NFκB signaling pathway. The above results indicate that curcumin reduces BACE1 expression through ERβ and NFκB pathway, providing a novel mechanism for curcumin as a candidate for AD therapy.

Merging memantine and ferulic acid to probe connections between NMDA receptors, oxidative stress and amyloid-β peptide in Alzheimer's disease

N-methyl-d-aspartate receptors (NMDAR) are critically involved in the pathogenesis of Alzheimer's disease (AD). Acting as an open-channel blocker, the anti-AD drug memantine preferentially targets NMDAR overactivation, which has been proposed to trigger neurotoxic events mediated by amyloid β peptide (Aβ) and oxidative stress. In this study, we applied a multifunctional approach by conjugating memantine to ferulic acid, which is known to protect the brain from Aβ neurotoxicity and neuronal death caused by ROS. The most interesting compound (7) behaved, like memantine, as a voltage-dependent antagonist of NMDAR (IC50 = 6.9 μM). In addition, at 10 μM concentration, 7 exerted antioxidant properties both directly and indirectly through the activation of the Nrf-2 pathway in SH-SY5Y cells. At the same concentration, differently from the parent compounds memantine and ferulic acid alone, it was able to modulate Aβ production, as revealed by the observed increase of the non-amyloidogenic sAPPα in H4-SW cells. These findings suggest that compound 7 may represent a promising tool for investigating NMDAR-mediated neurotoxic events involving Aβ burden and oxidative damage.

Artemisinin Attenuated Hydrogen Peroxide (H2O2)-Induced Oxidative Injury in SH-SY5Y and Hippocampal Neurons via the Activation of AMPK Pathway.

Oxidative stress is believed to be one of the main causes of neurodegenerative diseases such as Alzheimer’s disease (AD). The pathogenesis of AD is still not elucidated clearly but oxidative stress is one of the key hypotheses. Here, we found that artemisinin, an anti-malarial Chinese medicine, possesses neuroprotective effects. However, the antioxidative effects of artemisinin remain to be explored. In this study, we found that artemisinin rescued SH-SY5Y and hippocampal neuronal cells from hydrogen peroxide (H2O2)-induced cell death at clinically relevant doses in a concentration-dependent manner. Further studies showed that artemisinin significantly restored the nuclear morphology, improved the abnormal changes in intracellular reactive oxygen species (ROS), reduced the mitochondrial membrane potential, and caspase-3 activation, thereby attenuating apoptosis. Artemisinin also stimulated the phosphorylation of the adenosine monophosphate -activated protein kinase (AMPK) pathway in SH-SY5Y cells in a time- and concentration-dependent manner. Inhibition of the AMPK pathway attenuated the protective effect of artemisinin. These data put together suggested that artemisinin has the potential to protect neuronal cells. Similar results were obtained in primary cultured hippocampal neurons. Cumulatively, these results indicated that artemisinin protected neuronal cells from oxidative damage, at least in part through the activation of AMPK. Our findings support the role of artemisinin as a potential therapeutic agent for neurodegenerative diseases.

Brain-derived neurotrophic factor induces thioredoxin-1 expression through TrkB/Akt/CREB pathway in SH-SY5Y cells

Brain-derived neurotrophic factor (BDNF) is one of the neurotrophic factors that are vital to the survival and proliferation of neuron. Thioredoxin-1 (Trx-1) is a redox regulating protein and plays various roles in regulating transcript factors and inhibiting apoptosis. It has been reported that Trx-1 is required for nerve growth factor-mediated signal transduction and neurite outgrowth, and is involved in synaptic protein expression induced by BDNF. However, the molecular mechanism on BDNF inducing Trx-1 expression has not been fully verified. The present study investigated the expression of Trx-1 after treatment with BDNF in SH-SY5Y cells. We first demonstrated that cell viability and the expression of Trx-1 were increased by BDNF in SH-SY5Y cells, which were inhibited by the tyrosine kinase B (TrkB) inhibitor, K252a, and the phosphatidylinositol 3-kinase (PI3-K) inhibitor, LY294002. Moreover, BDNF increased the activity of cAMP response element-binding protein (CREB) through TrkB/PI3-K/Akt pathway. Whereas the expression of Trx-1 induced by BDNF was suppressed by CREB siRNA. Thus, our data suggest that BDNF induces the expression of Trx-1 through the TrkB/Akt/CREB pathway.