Human Neuroblastoma Cell Line Research Articles

Serine Hydroxyl-Methyltransferase 2 Inhibition Induces Differentiation and Impairs Tumorigenesis in High-Risk Neuroblastoma Cells

INTRODUCTION: High-risk neuroblastoma (NB) is associated with SHMT2 (serine hydroxyl-methyltransferase 2) expression, which is upregulated in several cancers and is associated with tumor aggressiveness. We have previously shown that SHMT2 silencing impairs cellular proliferation, migration, and colony formation, and treatment with a SHMT2 inhibitor induces differentiation in vitro. The stem cell marker genes CD133, CKIT, and NOTCH1 have been shown to downregulate differentiation in NB. We hypothesize that treatment with SHIN2, a novel SHMT2 inhibitor, will induce differentiation and impair tumorigenesis via downregulation of CD133, CKIT, and NOTCH1. METHODS: Human NB cell lines, BE(2)-C and SK-N-SH, were used for the study. Cells were treated with SHIN2 for 24 hours and quantitative reverse transcription polymerase chain reaction and immunoblotting were performed. Cellular proliferation was assessed using CCK-8 assays at 24 and 72 hours after SHIN2 treatment. Wound healing assays were performed to assess cellular migration at 24 hours after SHIN2 treatment. RESULTS: SHIN2 decreased CD133, CKIT, and NOTCH1 mRNA and protein expression by 1.2- to 2.5-fold and 1.1- to 1.4-fold, respectively, in both cell lines. SHIN2 significantly impaired cellular proliferation in both cell lines at 24 and 72 hours. SHIN2 treatment significantly increased wound gap and closure rate in both cell lines (Figs. 1A–1C).Figure 1CONCLUSION: SHMT2 plays an important role in NB tumorigenesis. Inhibition of SHMT2 with SHIN2 impairs cellular proliferation and cellular migration, and it induces differentiation via downregulation of stem cell genes CD133, CKIT, and NOTCH1, in vitro. The use of SHIN2 may serve as a novel therapeutic strategy in high-risk NB.

Exosome mimetics derived from bone marrow mesenchymal stem cells ablate neuroblastoma tumor in vitro and in vivo

PurposeTo develop exosome-mimetics derived from bone marrow mesenchymal stem cells (EM) as a novel nanoscale drug delivery system(nanoDDS) with improved tumor targeting activity, therapeutic effect, and biosafety, and to evaluate the therapeutic effect of doxorubicin loaded EM (EM-Dox) on neuroblastoma (NB) in vitro and in vivo. MethodsEM was prepared by serial extrusion of bone marrow mesenchymal stem cells (BMSCs), ammonium sulfate gradient method was used to promote the active loading of doxorubicin, and EM-Dox was obtained after removal of free doxorubicin by dialysis. The obtained EM and EM-Dox were characterized by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), Western Blot assay(WB), and the yield of exosomes and EM was further compared. Confocal fluorescent microscopy was used to verify the uptake of EM-Dox and free doxorubicin (Free-Dox) by NB cells. CCK-8 assay, cell cycle assay, and cell apoptosis assay were used to evaluate the antitumor effect of EM-Dox on NB cells in vitro. In addition, the targeted therapeutic effect and biosafety of EM-Dox against NB were evaluated in tumor-bearing nude mice. ResultsTEM, NTA, and WB verified that both EM and EM-Dox feature highly similar morphology, size and marker protein expression in comparison with naturally occurred exosomes, but the particle size of EM-Dox increased slightly after loading doxorubicin. The protein yield and particle yield of EM-Dox were 16.8 and 26.3-folds higher than those of exosomes, respectively. Confocal fluorescent microscopy showed that EM and doxorubicin had a definite co-localization. EM-Dox was readily internalized in two well-established human NB cell lines. The intracellular content of doxorubicin in cells treated with EM-Dox was significantly higher than that treated with Free-Dox. CCK-8 assay and flow cytometry confirmed that EM-Dox could inhibit NB cell proliferation, induce G2/M phase cell cycle arrest, and promote NB cell apoptosis in vitro. In vivo bioluminescence imaging results demonstrated that EM-Dox effectively targets NB tumors in vivo. Compared with Free-Dox, EM-Dox had a significantly increased inhibitory effect against NB tumor proliferation and progression in vivo, without inducing any myocardial injury. ConclusionsEM-Dox showed significantly increased anti-tumor activity in comparison with free doxorubicin in vitro and in vivo, and scalable EMs may represent a new class of NanoDDS that can potentially replace naturally occurred exosomes in preclinical or clinical translations.

Combination bromo- and extraterminal domain and poly (ADP-ribose) polymerase inhibition synergistically enhances DNA damage and inhibits neuroblastoma tumorigenesis

PurposeJQ1 is a bromo- and extraterminal (BET) domain inhibitor that downregulates MYC expression and impairs the DNA damage response. Poly (ADP-ribose) polymerase (PARP) inhibitors prevent DNA damage sensing and repair. We hypothesized that JQ1 would promote a DNA repair-deficient phenotype that sensitizes neuroblastoma cells to PARP inhibition.MethodsFour human neuroblastoma cell lines were examined: two MYCN-amplified (BE(2)-C and IMR-32), and two non-MYCN-amplified (SK-N-SH and SH-SY5Y). Cells were treated with JQ1 (BET inhibitor), Olaparib (PARP inhibitor), or in combination to assess for therapeutic synergy of JQ1 and Olaparib. Treated cells were harvested and analyzed. Quantitative assessment of combination treatment synergy was performed using the median effect principle of Chou and Talalay.ResultsCombination treatment with Olaparib decreased the IC50 of JQ1 by 19.9-fold, 2.0-fold, 12.1-fold, and 2.0-fold in the BE(2)-C, IMR-32, SK-N-SH, and SH-SY5Y cell lines, respectively. In the MYCN-amplified cell lines, BE(2)-C and IMR-32, combination treatment decreased gene expression of MYCN relative to single-drug treatment alone or control. Combination treatment decreased protein expression of DNA repair proteins Ku80 and RAD51, led to accumulation of DNA damage marker phospho-histone H2A.X, and increased caspase activity. In the non-MYCN-amplified cell lines, SK-N-SH and SH-SY5Y, combination treatment induced G0/G1 cell cycle arrest.ConclusionsCombination BET and PARP inhibition synergistically inhibited neuroblastoma tumorigenesis in vitro. In MYCN-amplified neuroblastoma cells, this effect may be induced by downregulation of MYCN transcription, defects in DNA repair, accumulation of DNA damage, and apoptosis. In non-MYCN-amplified cell lines, combination treatment induced cell cycle arrest.

The Therapeutic Potential of Naturally Occurring Peptides in Counteracting SH-SY5Y Cells Injury.

Peptides have revealed a large range of biological activities with high selectivity and efficiency for the development of new drugs, including neuroprotective agents. Therefore, this work investigates the neuroprotective properties of naturally occurring peptides, endomorphin-1 (EM-1), endomorphin-2 (EM-2), rubiscolin-5 (R-5), and rubiscolin-6 (R-6). We aimed at answering the question of whether well-known opioid peptides can counteract cell injury in a common in vitro model of Parkinson’s disease (PD). Antioxidant activity of these four peptides was evaluated by the 2-diphenyl-1-picrylhydrazyl radical (DPPH) scavenging activity, oxygen radical absorbance capacity (ORAC), and ferric-reducing antioxidant power (FRAP) assays, while neuroprotective effects were assessed in a neurotoxic model induced by 6-hydroxydopamine (6-OHDA) in a human neuroblastoma cell line (SH-SY5Y). The mechanisms associated with neuroprotection were investigated by the determination of mitochondrial membrane potential (MMP), reactive oxygen species (ROS) production, and Caspase-3 activity. Among the tested peptides, endomorphins significantly prevented neuronal death induced by 6-OHDA treatment, decreasing MMP (EM-1) or Caspase-3 activity (EM-2). Meanwhile, R-6 showed antioxidant potential by FRAP assay and exhibited the highest capacity to recover the neurotoxicity induced by 6-OHDA via attenuation of ROS levels and mitochondrial dysfunction. Generally, we hypothesize that peptides’ ability to suppress the toxic effect induced by 6-OHDA may be mediated by different cellular mechanisms. The protective effect caused by endomorphins results in an antiapoptotic effect (mitochondrial protection and decrease in Caspase-3 activity), while R-6 potency to increase a cell’s viability seems to be mediated by reducing oxidative stress. Our results may provide new insight into neurodegeneration and support the short peptides as a potent drug candidate to treat PD. However, further studies should be conducted on the detailed mechanisms of how tested peptides could suppress neuronal injuries.

Design and synthesis of new indole drug candidates to treat Alzheimer’s disease and targeting neuro-inflammation using a multi-target-directed ligand (MTDL) strategy

A novel series of indole-based compounds was designed, synthesised, and evaluated as anti-Alzheimer’s and anti-neuroinflammatory agents. The designed compounds were in vitro evaluated for their AChE and BuChE inhibitory activities. The obtained results revealed that compound 3c had higher selectivity for AChE than BuChE, while, 4a, 4b, and 4d showed selectivity for BuChE over AChE. Compounds 5b, 6b, 7c, and 10b exerted dual AChE/BuChE inhibitory activities at nanomolar range. Compounds 5b and 6b had the ability to inhibit the self-induced Aβ amyloid aggregation. Different anti-inflammatory mediators (NO, COX-2, IL-1β, and TNF-α) were assessed for compounds 5b and 6b. Cytotoxic effect of 5b and 6b against human neuroblastoma (SH-SY5Y) and normal hepatic (THLE2) cell lines was screened in vitro. Molecular docking study inside rhAChE and hBuChE active sites, drug-likeness, and ADMET prediction were performed.

The GBA variant E326K is associated with alpha-synuclein aggregation and lipid droplet accumulation in human cell lines.

Sequence variants or mutations in the GBA gene are numerically the most important risk factor for Parkinson disease (PD). The GBA gene encodes for the lysosomal hydrolase enzyme, glucocerebrosidase (GCase). GBA mutations often reduce GCase activity and lead to the impairment of the autophagy-lysosomal pathway, which is important in the turnover of alpha-synuclein, accumulation of which is a key pathological hallmark of PD. Although the E326K variant is one of the most common GBA variants associated with PD, there is limited understanding of its biochemical effects. We have characterized homozygous and heterozygous E326K variants in human fibroblasts. We found that E326K variants did not cause a significant loss of GCase protein or activity, endoplasmic reticulum (ER) retention or ER stress, in contrast to the L444P GBA mutation. This was confirmed in human dopaminergic SH-SY5Y neuroblastoma cell lines overexpressing GCase with either E326K or L444P protein. Despite no loss of the GCase activity, a significant increase in insoluble alpha-synuclein aggregates in E326K and L444P mutants was observed. Notably, SH-SY5Y overexpressing E326K demonstrated a significant increase in the lipid droplet number under basal conditions, which was exacerbated following treatment with the fatty acid oleic acid. Similarly, a significant increase in lipid droplet formation following lipid loading was observed in heterozygous and homozygous E326K fibroblasts. In conclusion, the work presented here demonstrates that the E326K mutation behaves differently to the common loss of function GBA mutations; however, lipid dyshomeostasis and alpha-synuclein pathology are still evident.

Mechanisms underlying citrinin-induced toxicity via oxidative stress and apoptosis-mediated by mitochondrial-dependent pathway in SH-SY5Y cells

Citrinin (CIT) is a mycotoxin produced as a secondary product by the genera Aspergillus, Penicillium, Monascus, and other strains. CIT has the potential for contaminating animal feed and human food such as maize, wheat, rye, barley, oats, rice, cheese, and sake. Although CIT is primarily known as a nephrotoxic mycotoxin, it also affects other organs, including the liver and bone marrow, and its mechanisms of toxicity have not been clearly elucidated. There is a further lack of studies investigating the potential for CIT-induced neurotoxicity and its mechanisms. In the current study, SH-SY5Y human neuroblastoma cell line was treated with CIT for 24 h to evaluate various toxicological endpoints, such as reactive oxygen species (ROS) production and apoptosis induction. Results indicate that CIT has an IC50 value of 250.90 μM and cell proliferation decreased significantly at 50 and 100 μM CIT concentrations. These same concentrations also caused elevated ROS production (≥34.76%), apoptosis (≥9.43-fold) and calcium ion mobilization (≥36.52%) in the cells. Results show a significant decrease in the mitochondrial membrane potential (≥86.8%). We also found that CIT significantly upregulated the expression of some genes related to oxidative stress and apoptosis, while downregulating others. These results suggest that apoptosis and oxidative stress may be involved in the mechanisms underlying CIT-induced neurotoxicity.

Neuroblastoma SH-SY5Y Cell Differentiation to Mature Neuron by AM580 Treatment

Neuroblastoma is a type of developmental childhood cancer that arises from the neural crest. It is the most common pediatric solid tumor in the world. AM580 is a powerful cyto-differentiating molecule on acute promyelocytic leukemia cells and induced pluripotent stem cells, but its effect on neuroblastoma is still unknown. In this study, the neuronal differentiation impact of AM580 was investigated using the human neuroblastoma cell line SH-SY5Y as a model. AM580 successfully stimulated the SH-SY5Y cells to develop into neuron-like cells. Functional enrichment analysis of RNAseq data revealed that differentially expressed genes (DEGs) were substantially enriched for GO keywords and KEGG pathways linked to neuron development. Some potassium ion channel genes associated with neuronal excitation, such as KCNT1, were shown to be upregulated. Through the MEA tests, we found the AM580-induced neurons possessed electrical spikes as mature neurons. AM580 also induced the neuronal marker β-tubulin III and mature neurons marker Neurofilament H. Our study proved that AM580 can promote the differentiation of neurons and has the potential to treat neuroblastoma, neurodevelopmental and neurodegenerative diseases.

3D engineered neural co-culture model and neurovascular effects of marine fungi-derived citreohybridonol

Marine-based biomolecules are emerging metabolites that have gained attention for developing novel biomaterials, drugs, and pharmaceutical in vitro platforms. Here, we developed a 3D engineered neural co-culture model via a 3D prototyped sliding frame-platform for multi-step UV lithography and investigated the neurovascular potential of citreohybridonol in neuroblastoma treatment. Citreohybridonol was isolated from a sponge-derived fungus Penicillium atrovenetum. The model was characterized by Fourier-transform infrared spectroscopy and scanning electron microscopy analysis. Human umbilical cord vein endothelial cells (HUVECs) and neuroblastoma (SH-SY5Y) cell lines were encapsulated in gelatin methacrylate (GelMA) with and without citreohybridonol. The effect of citreohybridonol on the proliferation capacity of cells was assessed via cell viability and immunostaining assays. GelMA and 3D culture characterization indicated that the cells were successfully encapsulated as axenic and mixed with/without citreohybridonol. The cytotoxic test confirmed that the 3D microenvironment was non-toxic for cultural experiments, and it showed the inhibitory effects of citreohybridonol on SH-SY5Y cells and induced the proliferation of HUVECs. Finally, immunohistochemical staining demonstrated that citreohybridonol suppressed SH-SY5Y cells and induced vascularization of HUVECs in mixed 3D cell culture.

Bioactivity of Ionic Liquids Based on Valproate in SH-SY5Y Human Neuroblastoma Cell Line

The search for alternative and effective therapies to fight cancer is one of the main goals of the pharmaceutical industry. Recently, ionic liquids (ILs) have emerged as potential therapeutic agents with antitumor properties. The goal of this study was to synthesize and evaluate the bioactivity of different ILs coupled with the active pharmaceutical ingredient (API) valproate (VPA) as an antitumor agent. The toxicity of the prepared ionic liquids was evaluated by the MTT cell metabolic assay in human neuroblastoma SH-SY5Y and human primary Gingival Fibroblast (GF) cell lines, in which they showed inhibitory effects during the study period. In addition, low cytotoxicity against GF cell lines was observed, suggesting that these compounds are not toxic to human cell lines. [C2OHDMiM][VPA] demonstrated an outstanding antitumor activity against SH-SY5Y and lower activity against the non-neoplastic GF line. The herein assessed compounds played an important role in the modulation of the signaling pathways involved in the cellular behavior. This work also highlights the potential of these ILs-API as possible antitumor agents.

In Vitro Effects of Acitretin on Human Neuronal SH-SY5Y Cells

Acitretin is an oral drug approved by the Food and Drug Administration that is commonly used to treat psoriasis. In recent years, acitretin has been identified as a candidate drug for the treatment of Alzheimer’s disease, but its role in neuronal development is still unclear. In this study, the human neuroblastoma cell line SH-SY5Y was used as a model to study neuronal differentiation. We found that acitretin effectively promoted the differentiation of SH-SY5Y cells into neuronal cells and upregulated the expression of the neuronal marker β-III tubulin and the mature neuronal marker NFH. Differentially expressed genes were identified by RNA sequencing and analyzed by bioinformatics approaches. The results showed that genes associated with neuron development-related pathways, such as SSPO and KCNT1, had significant changes in expression. Analysis showed that PRKCA and CAMK2B may play important roles in the process by which acitretin promotes neurodevelopment. Through whole-cell patch clamping and a microelectrode array assay, we found that acitretin-treated neurons generated electrical spikes similar to those generated by mature neurons. This study provided evidence to support an accessible and safe model of neuron-like cells and verified that acitretin can promote the differentiation of neurons and has the potential to treat brain tumors and neurodevelopmental and neurodegenerative diseases.

Syzysamalactone, an Unusual 11-Carbon δ-Lactone Derivative from the Fresh Ripe Fruits of Syzygium samarangense (Wax Apple).

To study the chemical constituents from the ripe fresh fruits of Syzygium samarangense (wax apple) and their potential health effects, a phytochemical investigation was undertaken. A new δ-lactone derivative, syzysamalactone (1), along with a known biogenetically related δ-lactone derivative, 6-pentyl-α-pyrone (2), were isolated from the fresh ripe fruits of S. samarangense. Syzysamalactone (1) is an unusual 11-carbon δ-lactone derivative, and its chemical structure and absolute configuration were elucidated by spectroscopic data analysis. A plausible biogenetic pathway for 1 was also proposed. Furthermore, the potential neuroprotective effects of compounds 1 and 2 were assessed. As a result, compounds 1 and 2 displayed notable neuroprotective effects with EC50 values of 0.29 ± 0.03 and 1.28 ± 0.06 μM, respectively, using the SH-SY5Y human neuroblastoma cell line. This is the first report of δ-lactone derivatives showing significant neuroprotective activities.

Calcium channel antagonists interfere with the mechanism of action of elastin-derived peptide VGVAPG in mouse cortical astrocytes in vitro

Elastin-derived peptides (EDPs) contain replications of the Val-Gly-Val-Ala-Pro-Gly (VGVAPG) hexapeptide. It has been described that the VGVAPG peptide induces reactive oxygen species (ROS) production in murine monocytes and astrocytes, human fibroblasts, and the human neuroblastoma (SH-SY5Y) cell line. To date, there is growing evidence that calcium channel blockers (CCBs) reduce oxidative stress and development of inflammation in the nervous system. Therefore, the aim of the present study was to evaluate the impact of such CCBs as Nifedipine, Verapamil, and MK-801 on the expression of peroxisome proliferator-activated receptor (Pparγ), i.e. ROS-related and inflammation-related proteins, in mouse astrocytes exposed in vitro to the VGVAPG peptide. The experiments showed that Nifedipine or MK-801 used in co-treatment with the VGVAPG peptide potentiated the effect of this peptide on the Pparγ level after the 24-h and 48-h treatment. Moreover, all studied compounds decreased the VGVAPG-induced caspase-1 activity in both time intervals. The data also showed that the VGVAPG peptide decreased the interleukin 1 beta (IL-1β) level in both studied time intervals. Upon a short-time exposure, the use of CCBs intensified the decrease in IL-1β stimulated by the VGVAPG peptide, opposite to the longer treatment. Moreover, the VGVAPG peptide decreased the IL-1βR1 level in both studied time intervals. After 24 h, Nifedipine and Verapamil potentiated the effect of the VGVAPG peptide. The VGVAPG peptide decreased the catalase (Cat) protein expression only after 24 h, whereas CCBs did not affect the expression of Cat induced by the VGVAPG peptide. The VGVAPG peptide increased the expression of the superoxide dismutase 1 (Sod1) protein. After 24 h of exposure, Nifedipine and Verapamil potentiated the increase in the Sod1 protein expression. Finally, our data showed that VGVAPG did not change the level of estradiol (E2) in the astrocytes. Interestingly, Nifedipine and Verapamil in co-treatment with VGVAPG increased the E2 level. Summarizing, it can be assumed that increased amounts of the VGVAPG during lifetime can play a certain role in calcium channel functioning in neurodegenerative diseases.

Antiproliferative Activity of Aminobenzylnaphthols Deriving from the Betti Reaction

Two aminobenzylnaphthols, which are representative items of the family of compounds synthesized with the Betti reaction, were investigated as antiproliferative agents against adenocarcinoma human colorectal (Caco-2) and human neuroblastoma (SH-SY5Y) cell lines, using cisplatin as a positive control. A better antiproliferative activity was recorded after 24 h of incubation for the first tested molecule, whereas the other one was more effective after 72 h of incubation. These results support the hypothesis that both of the tested aminobenzylnaphthols could potentially be endowed with a biological activity.

N-trans-feruloyltyramine Protects Human Neuroblastoma SK-N-SH Cell Line Against H2O2-Induced Cytotoxicity

Excessive accumulation of reactive oxygen species (ROS) has been regarded as a major contributor of pathogenesis in neurodegenerative diseases. N- trans-feruloyltyramine (NTF), an alkaloid isolated from several plants, has demonstrated an ability to be a potent antioxidant. In this study, the antioxidative and anti-apoptotic properties of NTF extracted from the stems of Polyalthia suberosa were investigated in the human neuroblastoma cell line SK-N-SH. NTF at concentrations ranging from 10 µM to 500 µM were not toxic to cells and reduced intracellular ROS levels significantly. Furthermore, pre-treatment of NTF significantly decreased H2O2-induced ROS generation and attenuated H2O2-mediated cytotoxicity. An increase in the expression of Bax and activated caspase-3 and reduction of Bcl-2 mediated by H2O2 was reversed by pre-treating the cells with 100 µM NTF. Likewise, NTF suppressed the increase of caspase-3 activity induced by H2O2. In conclusion, the findings reveal that NTF improves H2O2-induced intracellular ROS generation and decreases apoptosis. These protective effects of NTF could be useful for oxidative stress-related neurodegenerative conditions.

Alpha-Amylase Inhibits Cell Proliferation and Glucose Uptake in Human Neuroblastoma Cell Lines

The present article describes a study of the effects of alpha-amylase (α-amylase) on the human neuroblastoma (NB) cell lines SH-SY5Y, IMR-32, and LA-N-1. NB is the most common malignancy diagnosed in infants younger than 12 months. Some clinical observations revealed an inverse association between the risk of NB development and breastfeeding. α-Amylase which is present in breast milk was shown to have anticancer properties already in the beginning of the 20th century. Data presented here show that pancreatic α-amylase inhibits cell proliferation and has a direct impact on glucose uptake in the human NB cell lines. Our results point out the importance of further research which could elucidate the α-amylase mode of action and justify the presence of this enzyme in breast milk as a possible inhibitor of NB development. α-Amylase can be thus recognized as a potential safe and natural mild/host anticancer agent minimizing chemotherapy-related toxicity in the treatment of NB.

In vitro Evaluation of Selective Cytotoxic Activity of Chaerophyllum macropodum Boiss. on Cultured Human SH-SY5Y Neuroblastoma Cells.

Neuroblastoma is the most common solid tumor in children. New treatment approaches are needed because of the harmful side effects and costs of the methods used in the treatment of neuroblastoma. Medicinal and aromatic plants are important for new treatment approaches due to their minimal side effects and economic advantages. Therefore, the present study was carried out to examine the cytotoxic effect of Chaerophyllum macropodum extract on human neuroblastoma (SH-SY5Y) and fibroblast (HDFa) cell lines. 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and lactate dehydrogenase release (LDH) assays were used to determine the cytotoxic effect of C. macropodum. The extracts were analyzed for their phenolic content by HPLC-PDA. Major components were determined as 63.600% o-coumaric acid, 15.606% catechine hydrate, 8.713% rosmarinic acid, 4.376% clorogenic acid, and 3.972% salicylic acid. The obtained results from cytotoxicity testing revealed that C. macropodum exerted a significant cytotoxic effect on human neuroblastoma cells at all tested concentrations (p < 0.05). But it did not lead to any cytotoxic potential on human fibroblasts. As a result, the obtained data clearly revealed C. macropodum exerted a selective cytotoxic action on neuroblastoma cells for the first time.

Effect of Sublethal Copper Overload on Cholesterol De Novo Synthesis in Undifferentiated Neuronal Cells.

Although copper (Cu) is an essential trace metal for cells, it can induce harmful effects as it participates in the Fenton reaction. Involuntary exposure to Cu overload is much more common than expected and has been linked with neurodegeneration, particularly with Alzheimer’s disease (AD) evidenced by a positive correlation between free Cu in plasma and the severity of the disease. It has been suggested that Cu imbalance alters cholesterol (Chol) homeostasis and that high membrane Chol promotes the amyloidogenic processing of the amyloid precursor protein (APP) secreting the β-amyloid (Aβ) peptide. Despite the wide knowledge on the effects of Cu in mature brain metabolism, the consequence of its overload on immature neurons remains unknown. Therefore, we used an undifferentiated human neuroblastoma cell line (SH-SY5Y) to analyze the effect of sublethal concentrations of Cu on 1— de novo Chol synthesis and membrane distribution; 2—APP levels in cells and its distribution in membrane rafts; 3—the levels of Aβ in the culture medium. Our results demonstrated that Cu increases reactive oxygen species (ROS) and favors Chol de novo synthesis in both ROS-dependent and independent manners. Also, at least part of these effects was due to the activation of 3-hydroxy-3-methyl glutaryl CoA reductase (HMGCR). In addition, Cu increases the Chol/PL ratio in the cellular membranes, specifically Chol content in membrane rafts. We found no changes in total APP cell levels; however, its presence in membrane rafts increases with the consequent increase of Aβ in the culture medium. We conclude that Cu overload favors Chol de novo synthesis in both ROS-dependent and independent manners, being at least in part, responsible for the high Chol levels found in the cell membrane and membrane rafts. These may promote the redistribution of APP into the rafts, favoring the amyloidogenic processing of this protein and increasing the levels of Aβ.

Structural, optical characteristics and Anti-Cancer effect of Cd0.99Ni0.01O nanoparticles on human neuroblastoma and cervical cancer cell lines

Cd0.99Ni0.01O nanoparticles are outstanding nanomaterials used in different applications. So the structural, morphological, and optical properties of these nanoparticles are employed in anticancer applications. X-ray diffraction data shows the single phase with polycrystalline nature. The structural parameters were calculated from the MAUD program. Also, the TEM confirmed the size distribution of Cd0.99Ni0.01O nanoparticles obtained from the co-precipitated method. The optical properties show the different optical characteristics of Cd0.99Ni0.01O nanoparticles with 2.06 eV energy gab. The Cd0.99Ni0.01O nanoparticles are employed for some medicinal and pharmaceutical applications. Cytotoxicity of Cd0.99Ni0.01O against human neuroblastoma IMR-32 and cervical cancer HeLa cell lines was studied. The cytotoxicity of Cd0.99Ni0.01O on the previous cancer cell lines resulted in inhibition of cell growth estimated by MTT assay. The exposure of IMR-32 and HeLa cancer cells to Cd0.99Ni0.01O for 48hrs verified a potential apoptotic activity by significant up-regulation of p53 gene expression. Furthermore, anticancer activity was tested by down-regulation of platelet-derived growth factor receptor beta (PDGFR-β) gene expression.

Protective effects of cyclosporine A on neurodegeneration and motor impairment in rotenone-induced experimental models of Parkinson's disease

The development of neuroprotective drugs targeting mitochondria could be an important strategy in combating the progressive clinical course of Parkinson's disease. In the current study, we demonstrated that in SH-SY5Y cells (human dopaminergic neuroblastoma cell line), rotenone caused a dose-dependent (0.25–1 μM) and time-dependent (up to 48 h) loss of cell viability and a loss of cellular ATP content with mitochondrial membrane depolarization and an increased formation of reactive oxygen species; all these processes were markedly prevented by the mitochondrial permeability transition pore blocker cyclosporine A, which did not affect complex I inhibition by rotenone. The nuclear morphology of rotenone-treated cells for 48 h indicated the presence of both necrosis and apoptosis. We then examined the effects of cyclosporine A on the rotenone-induced model of Parkinson's disease in Wistar rats. Cyclosporine A significantly improved the motor deficits and prevented the loss of nigral dopaminergic neurons projecting into the striatum in rotenone-treated rats. Being a marketed immuno-suppressive drug, cyclosporine A should be further evaluated for its putative neuroprotective action in Parkinson's disease.