Restricted growth of attenuated poliovirus strains in cultured cells of a human neuroblastoma.

It has been reported that overproduction of reactive oxygen species occurs after brain injury and mediates neuronal cells degeneration. In the present study, we examined the role of Ras signaling on hydrogen peroxide-induced neuronal cells degeneration in dopaminergic neuroblastoma SH-SY5Y cells. Hydrogen peroxide significantly reduced cell viability in SH-SY5Y cultured cells. An inhibitor of the enzyme that catalyzes the farnesylation of Ras proteins, FTI-277, and a competitive inhibitor of GTP-binding proteins, GDP-beta-S significantly decreased hydrogen peroxide-induced reduction in cell viability in SH-SY5Y cultured cells. The results of this study might indicate that a Ras-dependent signaling pathway plays a role in hydrogen peroxide-induced toxicity in neuronal cells.

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Melatonin reduces induction of Bax, caspase and cell death in methamphetamine-treated human neuroblastoma SH-SY5Y cultured cells

Neuroscience Research ◽

10.1016/j.neures.2009.09.846 ◽

2009 ◽

Vol 65 ◽

pp. S163

Author(s):

Wilaiwan Wisessmith ◽

Pansiri Phansuwan-Pujito ◽

Piyarat Govitrapong ◽

Banthit Chetsawang

Keyword(s):

Cell Death ◽

Cultured Cells ◽

Human Neuroblastoma

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Prolonged Amphetamine Treatments Cause Long-Term Decrease of Dopamine Uptake in Cultured Cells

Neurochemical Research ◽

10.1007/s11064-019-02938-7 ◽

2019 ◽

Vol 45 (6) ◽

pp. 1399-1409

Author(s):

Nafisa Ferdous ◽

Sirisha Kudumala ◽

Serena Sossi ◽

Lucia Carvelli

Keyword(s):

Cultured Cells ◽

Neuroblastoma Cell ◽

Neuroblastoma Cell Line ◽

Human Neuroblastoma Cell ◽

Long Term Effects ◽

Human Neuroblastoma ◽

Daughter Cells ◽

Cell Divisions

AbstractAmphetamine (AMPH) is a systemic stimulant used to treat a variety of diseases including Attention Deficit Hyperactive Disorder, narcolepsy and obesity. Previous data showed that by binding to catecholamine transporters, AMPH prevents the reuptake of the neurotransmitters dopamine (DA) and norepinephrine (NE). Because AMPH, either used therapeutically at final concentrations of 1–10 µM or abused as recreational drug (50–200 µM), is taken over long periods of time, we investigated the prolonged effects of this drug on the uptake of DA. We found that, in LLC-PK1 cells stably expressing the human DA transporter (hDAT), pretreatments with 1 or 50 µM AMPH caused significant reduction in DA uptake right after the 15-h pretreatment. Remarkably, after 50 but not 1 µM AMPH pretreatment, we observed a significant reduction in DA uptake also after one, two or three cell divisions. To test whether these long-term effects induced by AMPH where conserved in a model comparable to primordial neuronal cells and native neurons, we used the human neuroblastoma cell line SH-SY5Y cells, which were reported to endogenously express both hDAT and the NE transporter. Pretreatments with 50 µM AMPH caused a significant reduction of DA uptake both right after 15 h and 3 cell divisions followed by neuro-differentiation with retinoic acid (RA) for 5 days. Under these same conditions, AMPH did not change the intracellular concentrations of ATP, ROS and cell viability suggesting, therefore, that the reduction in DA uptake was not cause by AMPH-induced toxicity. Interestingly, while 1 µM AMPH did not cause long-term effects in the LLC-PK1 cells, in the SH-SY5Y cells, it decreased the DA uptake after one, two, but not three, cell divisions and 5-day RA differentiation. These data show that besides the well-known acute effects, AMPH can also produce long-term effects in vitro that are maintained during cell division and transmitted to the daughter cells.

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Predominance of the acylation route in the metabolic processing of exogenous sphingosine in neural and extraneural cells in culture

Biochemical Journal ◽

10.1042/bj3380147 ◽

1999 ◽

Vol 338 (1) ◽

pp. 147-151 ◽

Cited By ~ 17

Author(s):

Laura RIBONI ◽

Rosaria BASSI ◽

Alessandro PRINETTI ◽

Paola VIANI ◽

Guido TETTAMANTI

Keyword(s):

Cultured Cells ◽

Primary Cultures ◽

Calf Serum ◽

Neuroblastoma Cells ◽

Cell Types ◽

Total Radioactivity ◽

Metabolic Fate ◽

Pulse Period ◽

Human Neuroblastoma ◽

Sphingosine 1 Phosphate

The metabolic fate of exogenous [3H]sphingosine was investigated in five types of cultured cells: primary cultures of neurons and astrocytes, murine and human neuroblastoma cells and human skin fibroblasts. After administration of 40 nM [3-3H]sphingosine into a cell-conditioned medium containing fetal calf serum, all cell types rapidly and efficiently incorporated the long-chain base in a time-dependent fashion. In all cases, after a 120 min pulse, the amount of radioactivity taken up was in the range of the endogenous sphingosine content. However, unchanged [3H]sphingosine represented only a very minor portion of the label incorporated into cells throughout the pulse period (10–120 min), indicating rapid and efficient sphingosine metabolism in these cells. Most of the [3H]sphingosine taken up was metabolically processed, either by degradation (assessed as 3H2O release into the culture medium) or by N-acylation (mainly to radioactive ceramide, sphingomyelin, neutral glycolipids and gangliosides). [3H]Sphingosine 1-phosphate accounted for less than 2% of the total radioactivity incorporated in all cases. Throughout the pulse period and in all cell types, 3H-labelled organic metabolites largely prevailed over 3H2O, indicating that N-acylation is the major metabolic fate of sphingosine in these cells under apparently physiological conditions. These results are consistent with the notion that sphingosine has a rapid turnover in the cells studied, and indicate that regulation of the basal level of this bioactive molecule occurs mainly through N-acylation.

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The protective effect of melatonin on methamphetamine-induced calpain-dependent death pathway in human neuroblastoma SH-SY5Y cultured cells

Journal of Pineal Research ◽

10.1111/j.1600-079x.2009.00731.x ◽

2010 ◽

Vol 48 (2) ◽

pp. 94-101 ◽

Cited By ~ 32

Author(s):

Wilasinee Suwanjang ◽

Pansiri Phansuwan-Pujito ◽

Piyarat Govitrapong ◽

Banthit Chetsawang

Keyword(s):

Protective Effect ◽

Cultured Cells ◽

Human Neuroblastoma

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Identification of a biologically active fragment of ALK and LTK-Ligand 2 (augmentor-α)

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1807881115 ◽

2018 ◽

Vol 115 (33) ◽

pp. 8340-8345 ◽

Cited By ~ 4

Author(s):

Andrey V. Reshetnyak ◽

Jyotidarsini Mohanty ◽

Francisco Tomé ◽

David E. Puleo ◽

Alexander N. Plotnikov ◽

...

Keyword(s):

Tyrosine Kinases ◽

Deletion Mutant ◽

Biochemical Characterization ◽

Cultured Cells ◽

Variable Region ◽

Biologically Active ◽

Local Concentration ◽

Dependent Manner ◽

Human Neuroblastoma ◽

Anaplastic Lymphoma

Elucidating the physiological roles and modes of action of the recently discovered ligands (designated ALKAL1,2 or AUG-α,β) of the receptor tyrosine kinases Anaplastic Lymphoma Kinase (ALK) and Leukocyte Tyrosine Kinase (LTK) has been limited by difficulties in producing sufficient amounts of the two ligands and their poor stability. Here we describe procedures for expression and purification of AUG-α and a deletion mutant lacking the N-terminal variable region. Detailed biochemical characterization of AUG-α by mass spectrometry shows that the four conserved cysteines located in the augmentor domain (AD) form two intramolecular disulfide bridges while a fifth, primate-specific cysteine located in the N-terminal variable region mediates dimerization through formation of a disulfide bridge between two AUG-α molecules. In contrast to AUG-α, the capacity of AUG-α AD to undergo dimerization is strongly compromised. However, full-length AUG-α and the AUG-α AD deletion mutant stimulate similar tyrosine phosphorylation of cells expressing either ALK or LTK. Both AUG-α and AUG-α AD also stimulate a similar profile of MAP kinase response in L6 cells and colony formation in soft agar by autocrine stimulation of NIH 3T3 cells expressing ALK. Moreover, both AUG-α and AUG-α AD stimulate neuronal differentiation of human neuroblastoma NB1 and PC12 cells in a similar dose-dependent manner. Taken together, these experiments show that deletion of the N-terminal variable region minimally affects the activity of AUG-α toward LTK or ALK stimulation in cultured cells. Reduced dimerization might be compensated by high local concentration of AUG-α AD bound to ALK at the cell membrane and by potential ligand-induced receptor–receptor interactions.

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Melatonin reduces induction of Bax, caspase and cell death in methamphetamine-treated human neuroblastoma SH-SY5Y cultured cells

Journal of Pineal Research ◽

10.1111/j.1600-079x.2009.00680.x ◽

2009 ◽

Vol 46 (4) ◽

pp. 433-440 ◽

Cited By ~ 35

Author(s):

Wilaiwan Wisessmith ◽

Pansiri Phansuwan-Pujito ◽

Piyarat Govitrapong ◽

Banthit Chetsawang

Keyword(s):

Cell Death ◽

Cultured Cells ◽

Human Neuroblastoma

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Mitotic mechanisms in Alzheimer's disease?

The Journal of Cell Biology ◽

10.1083/jcb.132.3.413 ◽

1996 ◽

Vol 132 (3) ◽

pp. 413-425 ◽

Cited By ~ 248

Author(s):

I Vincent ◽

M Rosado ◽

P Davies

Keyword(s):

Alzheimer’S Disease ◽

Alzheimer's Disease ◽

Human Brain ◽

Cultured Cells ◽

Neuroblastoma Cells ◽

Human Brain Tissue ◽

Human Neuroblastoma ◽

Mitotic Kinase ◽

M Phase ◽

Mature Neurons

The mechanism(s) leading to widespread hyper-phosphorylation of proteins in Alzheimer's disease (AD) are unknown. We have characterized seven new monoclonal antibodies recognizing independent phospho-epitopes in the paired helical filament proteins (PHF) found in AD brain. These antibodies show pronounced immunoreactivity with cultured human neuroblastoma cells that are in the M phase of cell division, but have no discernible reactivity with interphase cells. Immunoreactivity with these antibodies does not localize to the microtubule spindles or chromosomes in M phase, but is confined to the surrounding cytoplasm. Similar staining in M phase is observed with cultured cells of various tissue types and species. Cells arrested in M phase with the microtubule depolymerizing agent, nocodazole, show marked increases in immunoreactivity with the antibodies by immunofluorescence staining, ELISA, and immunoblotting. In neuroblastoma cells, the appearance of the TG/MC phospho-epitopes coincides with activation of mitotic protein kinases, but not with the activity of the neuronal specific cyclin-dependent kinase, cdk5. These data suggest that the TG/MC epitopes are conserved mitotic phospho-epitopes produced as a result of increased mitotic kinase activity. To investigate this possibility in AD, we examined the staining of human brain tissue with MPM-2, a marker antibody for mitotic phospho-epitopes. It was found that MPM-2 reacts strongly with neurofibrillary tangles, neuritic processes, and neurons in AD but has no staining in normal human brain. Our data suggest that accumulation of phospho-epitopes in AD may result from activation of mitotic posttranslational mechanisms which do not normally operate in mature neurons of brain.

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Cornel Iridoid Glycoside Attenuates Tau Hyperphosphorylation by Inhibition of PP2A Demethylation

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2013/108486 ◽

2013 ◽

Vol 2013 ◽

pp. 1-9 ◽

Cited By ~ 12

Author(s):

Cui-cui Yang ◽

Xue-xian Kuai ◽

Ya-li Li ◽

Li Zhang ◽

Jian-chun Yu ◽

...

Keyword(s):

Cultured Cells ◽

Glycogen Synthase ◽

Iridoid Glycoside ◽

Tau Hyperphosphorylation ◽

Human Neuroblastoma ◽

Microtubular Cytoskeleton ◽

Gf 109203X ◽

Phosphatase 2A ◽

Cornel Iridoid Glycoside ◽

Underlying Mechanisms

Aim. The aim of the present study was to investigate the effect of cornel iridoid glycoside (CIG) on tau hyperphosphorylation induced by wortmannin (WT) and GF-109203X (GFX) and the underlying mechanisms.Methods. Human neuroblastoma SK-N-SH cells were preincubated with CIG (50, 100, and 200 µg/ml, resp.) for 24 h and then exposed to 10 µM WT and 10 µM GFX for 3 h after washing out CIG. Immunohistochemistry was used to observe the microtubular cytoskeleton of the cultured cells. Western blotting was used to measure the phosphorylation level of tau protein, glycogen synthase kinase 3β(GSK-3β), and protein phosphatase 2A (PP2A). The activity of PP2A was detected by a biochemical assay.Results. Preincubation of CIG significantly attenuated the WT/GFX-induced tau hyperphosphorylation at the sites of Thr205, Thr212, Ser214, Thr217, Ser396, and PHF-1 and improved the damage of morphology and microtubular cytoskeleton of the cells. CIG did not prevent the decrease in p-AKT-ser473 and p-GSK-3β-ser9 induced by WT/GFX. However, CIG significantly elevated the activity of PP2A by reducing the demethylation of PP2A catalytic subunit (PP2Ac) at Leu309 and the ratio of PME-1/LCMT in the WT/GFX-treated cells. The results suggest that CIG may be beneficial to the treatment of AD.

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DNA Polymerase-Associated Lectin (DPAL) and Its Binding to the Galactose-Containing Glycoconjugate of the Replication Complex

Bioscience Reports ◽

10.1023/a:1020268407342 ◽

1999 ◽

Vol 19 (5) ◽

pp. 433-447

Author(s):

Thomas J. Kelley ◽

Tara St. Amand ◽

Jeremy M. Groll ◽

Satyajit Ray ◽

Subhash Basu

Keyword(s):

Molecular Weight ◽

Monoclonal Antibody ◽

Dna Polymerase ◽

Recombinant Dna ◽

Cultured Cells ◽

Polyclonal Antibodies ◽

Immunoblot Analysis ◽

Post Translational Modification ◽

Human Neuroblastoma ◽

Dna Polymerase Α

The highly purified DNA Pol-α from rat prostate tumor (PA-3) and human neuroblastoma (IMR-32) cells appeared to be inhibited by Ricin (RCA-II), and Con-A. Loss of activity (40 to 60%) of a specific form of DNA polymerase from IMR-32 was observed when the cells were treated with tunicamycin [Bhattacharya, P. and Basu, S. (1982) Proc. Natl. Acad. Sci., USA79:1488–1492]. Binding of ConA and RCA to human recombinant DNA polymerase-α showed a specific labile site in the N-terminus [Hsi et al.. (1990) Nucleic Acid Res.18:6231–6237]. The catalytic polypeptide, DNA polymerase-α of eukaryotic origin, was isolated from developing tissues or cultured cells as a family of 180 to 120 kDa polypeptides, perhaps derived from a single primary structure. Immunoblot analysis with a monoclonal antibody (SJK-237-71) indicated that the lower molecular weight polypeptides resulted from either proteolytic cleavage of post-translational modification after specific cleavages. Present results suggest DNA polymerase-α from embryonic chicken brain (ECB) contains an α-galactose-binding subunit which may be involved in developmental regulation of the enzyme. It was shown before that the catalytic subunit of DNA polymerase-α reduces from 186 kDa in 11-day-old ECB to 120 kDa in 19-day-old ECB [Ray, S. et al. Cell Growth and Differentiation2:567–573] by the treatment with methyl-α-galactose. The low molecular weight DNA polymerase activity (120 kDa) can be reconstituted to high molecular weight (Mr = 186 kDa) with an α-galactose binding, 56 kDa lectin-like protein. Polyclonal antibodies raised against the purified lectin were able to precipitate DNA. Pol-α as determined by immunostaining with the polymerase-α-specific monoclonal antibody SJK 132-20, suggesting this is a DNA polymerase associated-lectin (DPAL). RCA-II and GS-I-Sepharose 4B chromatographies resulted in significant purification of DNA-α and a complete separation of polymerase complex and primase.

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