Anti-neuroblastoma cell line antibodies in inflammatory demyelinating polyneuropathy: Inhibition in vitro and in vivo by IV immunoglobulin

Microglia, the main immune cells in the brain, participate in the innate immune response in the central nervous system (CNS). Studies have shown that microglia can be polarized into pro-inflammatory M1 and anti-inflammatory M2 phenotypes. Accumulated evidence suggests that over-activated M1 microglia release pro-inflammatory mediators that damage neurons and lead to Parkinson’s disease (PD). In contrast, M2 microglia release neuroprotective factors and exert the effects of neuroprotection. Camptothecin (CPT), an extract of the plant Camptotheca acuminate, has been reported to have anti-inflammation and antitumor effects. However, the effect of CPT on microglia polarization and microglia-mediated inflammation responses has not been reported. In our study we found that CPT improved motor performance of mice and reduced the loss of neurons in the substantia nigra (SN) of the midbrain in LPS-injected mice. In the mechanism study, we found that CPT inhibited M1 polarization of microglia and promotes M2 polarization via the AKT/Nrf2/HO-1 and NF-κB signals. Furthermore, CPT protected the neuroblastoma cell line SH-SY5Y and dopaminergic neuron cell line MN9D from damage mediated by microglia activation. In conclusion, our results demonstrate that CPT regulates the microglia polarization phenotype via activating AKT/Nrf2/HO-1 and inhibiting NF-κB pathways, inhibits neuro-inflammatory responses, and exerts neuroprotective effects in vivo and in vitro.

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A human neuroblastoma cell line with a stable ornithine decarboxylase in vivo and in vitro

Biochemical and Biophysical Research Communications ◽

10.1016/0006-291x(85)90576-5 ◽

1985 ◽

Vol 126 (1) ◽

pp. 96-102 ◽

Cited By ~ 8

Author(s):

Eija Karvonen ◽

Leif C. Andersson ◽

Hannu Pösö

Keyword(s):

Cell Line ◽

Ornithine Decarboxylase ◽

Neuroblastoma Cell ◽

Neuroblastoma Cell Line ◽

Human Neuroblastoma Cell ◽

Human Neuroblastoma ◽

Human Neuroblastoma Cell Line

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A novel in vitro assay model developed to measure both extracellular and intracellular acetylcholine levels for screening cholinergic agents

PLoS ONE ◽

10.1371/journal.pone.0258420 ◽

2021 ◽

Vol 16 (10) ◽

pp. e0258420

Author(s):

Ryohei Tanaka-Kanegae ◽

Koichiro Hamada

Keyword(s):

Cell Line ◽

Neuroblastoma Cell ◽

Cholinergic Neurons ◽

Mechanisms Of Action ◽

Neuroblastoma Cell Line ◽

Ache Inhibitor ◽

Vitro Assay ◽

Food Ingredients ◽

High Doses

Background Cholinergic neurons utilize choline (Ch) to synthetize acetylcholine (ACh) and contain a high-affinity Ch transporter, Ch acetyltransferase (ChAT), ACh receptors, and acetylcholinesterase (AChE). As the depletion or malfunction of each component of the cholinergic system has been reported in patients with dementia, many studies have sought to evaluate whether treatment candidates affect each of the cholinergic components. The associated changes in the cholinergic components may be reflected by intra- or extra-cellular ACh levels, with an increase in extracellular ACh levels occurring following AChE inhibition. We hypothesized that increases in intracellular ACh levels can be more sensitively detected than those in extracellular ACh levels, thereby capturing subtle effects in the cholinergic components other than AChE. The objective of this study was to test this hypothesis. Methods We developed an in vitro model to measure both extracellular and intracellular ACh levels using the human cholinergic neuroblastoma cell line, LA-N-2, which have been reported to express Ch transporter, ChAT, muscarinic ACh receptor (mAChR), and AChE. With this model, we evaluated several drug compounds and food constituents reported to improve cholinergic function through various mechanisms. In addition, we conducted western blotting to identify the subtype of mAChR that is expressed on the cell line. Results Our cell-based assay system was capable of detecting increases in extracellular ACh levels induced by an AChE inhibitor at relatively high doses, as well as increases in intracellular ACh levels following the administration of lower AChE-inhibitor doses and an mAChR agonist. Moreover, increases in intracellular ACh levels were observed even after treatment with food constituents that have different mechanisms of action, such as Ch provision and ChAT activation. In addition, we revealed that LA-N-2 cells expressed mAChR M2. Conclusion The findings support our hypothesis and indicate that the developed assay model can broadly screen compounds from drugs to food ingredients, with varying strengths and mechanisms of action, to develop treatments for ACh-relevant phenomena, including dementia and aging-related cognitive decline.

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Hexarelin exerts neuroprotective and antioxidant effects against hydrogen peroxide-induced toxicity through the modulation of MAPK and PI3K/Akt patways in Neuro-2A cells

10.1101/2020.11.16.384321 ◽

2020 ◽

Author(s):

Ramona Meanti ◽

Laura Rizzi ◽

Elena Bresciani ◽

Laura Molteni ◽

Vittorio Locatelli ◽

...

Keyword(s):

Oxidative Stress ◽

Hydrogen Peroxide ◽

Apoptotic Cell ◽

Neuroblastoma Cell ◽

Neuroblastoma Cell Line ◽

Inhibitory Influence ◽

No Production ◽

Mrna Levels

AbstractHexarelin, a synthetic hexapeptide, protects cardiac and skeletal muscles by inhibiting apoptosis, both in vitro and in vivo. Moreover, evidence suggests that hexarelin could have important neuroprotective bioactivity.Oxidative stress and the generation of free radicals has been implicated in the etiologies of several neurodegenerative diseases, including amyotrophic lateral sclerosis, Parkinson’s disease, Alzheimer’s disease, Huntington’s disease and multiple sclerosis. In addition to direct oxidative stress, exogenous hydrogen peroxide (H2O2) can penetrate biological membranes and enhance the formation of other reactive oxygen species.The aim of this study was to examine the inhibitory influence of hexarelin on H2O2-induced apoptosis in Neuro-2A cells, a mouse neuroblastoma cell line. Our results indicate that H2O2 reduced the viability of Neuro-2A cells in a dose-related fashion. Furthermore, H2O2 induced significant changes in the morphology of Neuro-2A cells, reflected in the formation of apoptotic cell bodies, and an increase of nitric oxide (NO) production. Hexarelin effectively antagonized H2O2 oxidative damage to Neuro-2A cells as indicated by improved cell viability, normal morphology and reduced nitrite (NO2−) release. Hexarelin treatment of Neuro-2A cells also reduced mRNA levels of caspases−3 and −7 and those of the pro-apoptotic molecule Bax; by contrast, hexarelin treatment increased anti-apoptotic Bcl-2 mRNA levels. Hexarelin also reduced MAPKs phosphorylation induced by H2O2 and concurrently increased p-Akt protein expression.In conclusion, our results identify several neuroprotective and anti-apoptotic effects of hexarelin. These properties suggest that further investigation of hexarelin as a neuroprotective agent in an investigational and therapeutic context are merited.

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Cell Plasticity during in Vitro Differentiation of a Human Neuroblastoma Cell Line

Plasticity and Regeneration of the Nervous System - Advances in Experimental Medicine and Biology ◽

10.1007/978-1-4684-8047-4_10 ◽

1991 ◽

pp. 91-102

Author(s):

F. Clementi ◽

C. Gotti ◽

E. Sher ◽

A. Zanini

Keyword(s):

Cell Line ◽

Neuroblastoma Cell ◽

Neuroblastoma Cell Line ◽

Human Neuroblastoma Cell ◽

In Vitro Differentiation ◽

Cell Plasticity ◽

Human Neuroblastoma ◽

Human Neuroblastoma Cell Line

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Silencing of AURKA augments the antitumor efficacy of the AURKA inhibitor MLN8237 on neuroblastoma cells

Cancer Cell International ◽

10.1186/s12935-019-1072-y ◽

2020 ◽

Vol 20 (1) ◽

Cited By ~ 1

Author(s):

Yan Yang ◽

Lili Ding ◽

Qi Zhou ◽

Li Fen ◽

Yuhua Cao ◽

...

Keyword(s):

Small Interfering Rna ◽

Neuroblastoma Cell ◽

Neuroblastoma Cells ◽

Inhibitory Effect ◽

Neuroblastoma Cell Line ◽

Inhibitory Effects ◽

Rna Transfection ◽

Interfering Rna

Abstract Background Aurora kinase A (AURKA) has been implicated in the regulation of cell cycle progression, mitosis and a key number of oncogenic signaling pathways in various malignancies including neuroblastoma. Small molecule inhibitors of AURKA have shown potential, but still not as good as expected effects in clinical trials. Little is known about this underlying mechanism. Here, we evaluated the inhibitory effects of AURKA inhibitor MLN8237 on neuroblastoma cells to understand the potential mechanisms responsible for tumor therapy. Methods MLN8237 treatment on neuroblastoma cell line IMR32 was done and in vivo inhibitory effects were investigated using tumor xenograft model. Cellular senescence was evaluated by senescence-associated β-gal Staining assay. Flow cytometry was used to tested cell cycle arrest and cell apoptosis. Senescence-associated signal pathways were detected by western blot. CD133 microbeads and microsphere formation were used to separate and enrich CD133+ cells. AURKA small interfering RNA transfection was carried to downregulate AURKA level. Finally, the combination of MLN8237 treatment with AURKA small interfering RNA transfection were adopted to evaluate the inhibitory effect on neuroblastoma cells. Results We demonstrate that MLN8237, an inhibitor of AURKA, induces the neuroblastoma cell line IMR32 into cellular senescence and G2/M cell phase arrest. Inactivation of AURKA results in MYCN destabilization and inhibits cell growth in vitro and in a mouse model. Although MLN8237 inhibits AURKA kinase activity, it has almost no inhibitory effect on the AURKA protein level. By contrast, MLN8237 treatment leads to abnormal high expression of AURKA in vitro and in vivo. Knockdown of AURKA reduces cell survival. The combination of MLN8237 with AURKA small interfering RNA results in more profound inhibitory effects on neuroblastoma cell growth. Moreover, MLN8237 treatment followed by AURKA siRNA forces senescent cells into apoptosis via suppression of the Akt/Stat3 pathway. Conclusions The effect of AURKA-targeted inhibition of tumor growth plays roles in both the inactivation of AURKA activity and the decrease in the AURKA protein expression level.

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Functional Changes in Potassium Conductances of the Human Neuroblastoma Cell Line SH-SY5Y During In Vitro Differentiation

Journal of Neurophysiology ◽

10.1152/jn.1998.79.2.648 ◽

1998 ◽

Vol 79 (2) ◽

pp. 648-658 ◽

Cited By ~ 43

Author(s):

Patrizia Tosetti ◽

Vanni Taglietti ◽

Mauro Toselli

Keyword(s):

Steady State ◽

Cell Line ◽

Neuroblastoma Cell ◽

Neuroblastoma Cell Line ◽

Human Neuroblastoma Cell ◽

Human Neuroblastoma ◽

Human Neuroblastoma Cell Line ◽

Differentiated Cells ◽

Voltage Dependent

Tosetti, Patrizia, Vanni Taglietti, and Mauro Toselli. Functional changes in potassium conductances of the human neuroblastoma cell line SH-SY5Y during in vitro differentiation. J. Neurophysiol. 79: 648–658, 1998. The electrophysiological properties of voltage-dependent outward currents were investigated under voltage-clamp conditions in the human neuroblastoma cell line SH-SY5Y before and after in vitro differentiation with retinoic acid, by using the whole cell variant of the patch-clamp technique. Voltage steps to depolarizing potentials from a holding level of −90 mV elicited, in both undifferentiated and differentiated cells, outward potassium currents that were blocked by tetraethylammonium, but were unaffected by 4-aminopyridine, cadmium, and by shifts of the holding potentials to −40 mV. These currents activated rapidly and inactivated slowly in a voltage-dependent manner. In undifferentiated cells the threshold for current activation was about −30 mV, with a steady-state half activation potential of 19.5 mV. Maximum conductance was 4.3 nS and mean conductance density was 0.34 mS/cm2. Steady-state half inactivation potential was −13.8 mV and ∼10% of the current was resistant to inactivation. Both activation and inactivation kinetics were voltage dependent. In differentiated cells the threshold for current activation was about −20 mV, with a half potential for steady-state activation of 37.0 mV. Maximum conductance was 15.2 nS and mean conductance density was 0.78 mS/cm2. Steady-state half inactivation potential was −9.7 mV and ∼37% of the current was resistant to inactivation. Both activation and inactivation kinetics were voltage dependent. This diversity in potassium channel properties observed between undifferentiated and differentiated cells was related to differences in cell excitability. Under current-clamp conditions, the action potential repolarization rate in differentiated cells was about threefold faster than that of the abortive action potentials elicitable in undifferentiated cells. Furthermore, during prolonged stimulation, trains of spikes could be generated in some differentiated cells but not in undifferentiated cells.

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A cullin-RING ubiquitin ligase targets exogenous α-synuclein and inhibits Lewy body–like pathology

Science Translational Medicine ◽

10.1126/scitranslmed.aau6722 ◽

2019 ◽

Vol 11 (495) ◽

pp. eaau6722 ◽

Cited By ~ 8

Author(s):

Juan A. Gerez ◽

Natalia C. Prymaczok ◽

Edward Rockenstein ◽

Uli S. Herrmann ◽

Petra Schwarz ◽

...

Keyword(s):

Lewy Body ◽

Cultured Cells ◽

Neuroblastoma Cell ◽

Lewy Bodies ◽

Neuroblastoma Cell Line ◽

Loss Of Function ◽

Intracerebral Administration ◽

F Box Protein

Parkinson’s disease (PD) is a neurological disorder characterized by the progressive accumulation of neuronal α-synuclein (αSyn) inclusions called Lewy bodies. It is believed that Lewy bodies spread throughout the nervous system due to the cell-to-cell propagation of αSyn via cycles of secretion and uptake. Here, we investigated the internalization and intracellular accumulation of exogenous αSyn, two key steps of Lewy body pathogenesis, amplification and spreading. We found that stable αSyn fibrils substantially accumulate in different cell lines upon internalization, whereas αSyn monomers, oligomers, and dissociable fibrils do not. Our data indicate that the uptake-mediated accumulation of αSyn in a human-derived neuroblastoma cell line triggered an adaptive response that involved proteins linked to ubiquitin ligases of the S-phase kinase-associated protein 1 (SKP1), cullin-1 (Cul1), and F-box domain–containing protein (SCF) family. We found that SKP1, Cul1, and the F-box/LRR repeat protein 5 (FBXL5) colocalized and physically interacted with internalized αSyn in cultured cells. Moreover, the SCF containing the F-box protein FBXL5 (SCFFBXL5) catalyzed αSyn ubiquitination in reconstitution experiments in vitro using recombinant proteins and in cultured cells. In the human brain, SKP1 and Cul1 were recruited into Lewy bodies from brainstem and neocortex of patients with PD and related neurological disorders. In both transgenic and nontransgenic mice, intracerebral administration of exogenous αSyn fibrils triggered a Lewy body–like pathology, which was amplified by SKP1 or FBXL5 loss of function. Our data thus indicate that SCFFXBL5 regulates αSyn in vivo and that SCF ligases may constitute targets for the treatment of PD and other α-synucleinopathies.

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