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Query: UMLS:C0027819 (neuroblastoma)
 27,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two clonal immortalized neurons designated CL8c4.7 and CL8a5.2 were established by somatic cell fusion between a hypoxanthine phosphoribosyltransferase-(HPRT-) deficient neuroblastoma N18TG2 and newborn mouse cerebellar/brain stem neurons. In the serum-containing medium without extra differentiating agents, both clones exhibited a morphology of differentiated neurons. They contained high levels of glutamate but no gamma-aminobutyric acid (GABA). The CL8a5.2 clone synthesized choline acetyltransferase and serotonin. In immunocytochemical studies, both clones expressed 200 kD neurofilament protein, neuron-specific enolase, microtubule-associated protein 2 (MAP2), tau protein, neuronal cell adhesion molecule (N-CAM), HNK-1, Thy-1.2, saxitoxin-binding sodium channel protein, and glutamate. Synaptophysin immunoreactivity was identified in the neuritic terminals of CL8c4.7 cells. Most of these antigens were barely detectable on N18TG2 cells. Electrophysiologically, both clones generated action potentials in response to electrical stimuli. The hybrid clones that express characteristics of differentiated neurons derived from the cerebellar and brain stem regions might be invaluable for the study of the molecular basis of neuronal differentiation and degeneration in these regions.
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PMID:Establishment of mouse-immortalized hybrid clones expressing characteristics of differentiated neurons derived from the cerebellar and brain stem regions. 135 6

The use of a panel of monoclonal antibodies (mAbs) directed against different determinants of microtubule-associated protein 2 (MAP2) enabled us to identify two distinct high-molecular-mass MAP2 species (270 and 250 kDa) and a substantial amount of MAP2c (70 kDa) in human neuroblastoma cells. The 250-kDa MAP2 species appears to be confined to the human neuroblastoma cells and was not observed in microtubules (MTs) from bovine and rat brain, mouse neuroblastoma, or MTs from human cerebellum. A new overlay method was developed, which demonstrates binding of tubulin to human neuroblastoma high-molecular-mass MAP2 by exposing nitrocellulose-bound MT proteins under polymerization conditions to tubulin. Bound tubulin was detected with a mAb directed against beta-tubulin. The binding of tubulin to MAP2 could be abolished by a peptide homologous to positions 426-445 of the C-terminal region of beta-tubulin. Immunological cross-reactivity with several mAbs directed against bovine brain MAP2, taxol-promoted coassembly into MTs, and immunocytochemical visualization within cells were further criteria utilized to characterize these proteins as true MAPs. Indirect immunofluorescence with anti-MAP2 and anti-beta-tubulin mAbs demonstrated that there is a change in the spatial organization of MTs during induced cell differentiation, as indicated by the appearance of MT bundles and the redistribution of MAP2.
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PMID:Characterization and intracellular distribution of microtubule-associated protein 2 in differentiating human neuroblastoma cells. 169 15

A pool of ten monoclonal antibodies to SDS-insoluble epitopes of Alzheimer neurofibrillary tangles (NFT) was used to screen an adult human brain cDNA expression library. Fourteen clones were isolated, two of which are described. The largest of the clones encodes 80 kD, or approximately 600 amino acids, of microtubule-associated protein 2 (MAP 2). The MAP 2 region encoded by the clone shares at least two epitopes with human tau, another microtubule-associated protein which cross-reacts with NFT. In rat brain mRNA, the MAP 2 cDNA hybridizes to a single transcript of 9.5 kb. In human neuroblastoma mRNA, the MAP 2 cDNA hybridizes, at high stringency, to two transcripts of 9.5 kb and 6 kb. The 6-kb transcript comigrates with the transcript for tau, as detected by a human tau cDNA. The properties of the MAP 2 cDNA suggest that, in humans, MAP 2 and tau have a common domain which may play a role in NFT formation. Another clone isolated with the anti-NFT antibodies shares epitopes, but not nucleic acid homology, with the MAP 2 cDNA. This clone detects a single abundant transcript of 1 kb present in RNA from human neuroblastoma and from several non-neuronal human cell lines. The properties of this cDNA suggest that it encodes a protein other than those previously reported to cross-react with NFT.
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PMID:Isolation and characterization of cDNA clones encoding epitopes shared with Alzheimer neurofibrillary tangles. 244 45

A type II calcium/calmodulin-dependent protein kinase (CaM kinase II) was purified approximately 300-fold from cultured neuroblastoma/glioma (NG108) cell homogenate. The purification of the kinase, which used a combination of differential centrifugation and chromatography on cation-exchange, calmodulin-affinity, and gel-filtration resins, was monitored by the ability of the kinase to phosphorylate the high-molecular-weight microtubule-associated protein 2 (MAP-2). The kinase was compared with authentic CaM kinase II purified from rat brain cytosol. Based upon holoenzyme molecular weight, subunit composition and molecular weight, calcium-dependent calmodulin-binding to subunits, calcium/calmodulin-dependent autophosphorylation of subunits, substrate specificity, apparent km's for ATP and calmodulin, phosphopeptide maps of subunits, time course, and heat lability, the kinase was identified as a type II calcium/calmodulin-dependent protein kinase. When cellular differentiation was induced under specific conditions of cell culture, a significant increase in the apparent activity and amount of the kinase per mg protein was observed relative to control cells. These studies suggest that there is an increase in CaM kinase II expression during cellular differentiation, which may relate to the concurrent development of electrical excitability, synaptogenesis, and elaboration of cytoskeletal elements. Thus, the NG108 cell should provide a useful model to study the physiological functions of CaM kinase II.
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PMID:Differentiation increases type II calmodulin-dependent protein kinase in the neuroblastoma/glioma cell line 108CC15 (NG108-15). 253 90

We examined the expression and distribution of microtubule-associated protein 2 (MAP2) during the differentiation in culture of both mouse NB2a neuroblastoma and primary embryonic rat neurons. The differentiation of NB2a cells was induced with retinoic acid (RA) which stimulated the extension of a highly branched neuritic network and dibutyryl cAMP which stimulated the outgrowth of long bipolar or monopolar processes. We found that although monoclonal antibodies to MAP2 stained the cell bodies of control and differentiated cells, only the RA-induced neurites were positive for this antigen. These data support our ultrastructural studies indicating that the RA-induced neurites were dendrite-like and that the dibutyryl cAMP-induced processes were axon-like. Studies on the biosynthesis of MAP2 indicated that RA induced a 2-3-fold increase in MAP2 synthesis in 24 h; however, this effect was transient, with the synthesis of MAP2 in RA-treated cells returning to control level by 72 h. Although biosynthetic studies suggested the synthesis of species at 250-300 kdalton, the major molecular weight form in the neuroblastoma cells was 230 kdalton. Immunocytochemical analysis of primary neurons showed staining of neuronal cell bodies and of short processes, but virtually no staining of the long axon-like processes. The staining of neuronal cell bodies and processes was evident at all stages of cell differentiation. This finding was corroborated by immunoblots which showed significant amounts of MAP2 throughout cell development. The molecular weight of the immunoreactive material was ca. 300 kdalton in both primary neurons and rat brain. Immunoblots also revealed that embryonic neurons expressed only MAP2B as they differentiated in culture for 14 days. Biosynthesis studies suggested that early in culture there was a modest increase in MAP2 synthesis, but no detectable change was observed thereafter. We concluded therefore that both neuroblastoma cells and primary neurons can differentiate neuritic processes, which show dendritic properties in terms of morphology and preferential distribution of MAP2.
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PMID:Expression and distribution of microtubule-associated protein 2 (MAP2) in neuroblastoma and primary neuronal cells. 351 42

Monoclonal antibodies against microtubule-associated protein 2 (MAP2) were prepared and their specificity was verified by visualization of the antigens using the antibody overlay technique and by radioimmunoassay. MAP2 was cleaved by alpha-chymotrypsin to generate a series of high-molecular-mass fragments ranging between 270 and 140 kDa. The precursor-product relationship of these fragments was suggested from the rate of their appearance and from the analysis of the tryptic peptide map of each fragment. A group of monoclonal antibodies was found to react predominantly with the intact 270-kDa MAP2 molecule and a fragment having a mass of 240 kDa and to some extent with a 215-kDa fragment. Another group of monoclonal antibodies reacted with an antigenic determinant which was located on the 270-kDa molecule as well as on fragments as small as 140 kDa. None of the two groups of monoclonal antibodies reacted with the microtubule-binding domain of MAP2. These results suggest that one group of antibodies reacts with sites located at or dependent upon a terminal 60-kDa domain(s) distal to the microtubule-binding site of MAP2. The second group of antibodies, which can still bind to smaller proteolytic products, appear to be associated with the central region of the MAP2 molecule. Indirect immunofluorescence experiments with the antibody preparations indicated that at least some of the antigenic determinants are exposed when MAP2 is associated with microtubules in the cell body and neurite outgrowths of differentiated rat brain neuroblastoma B104 cells.
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PMID:Mapping of distinct structural domains on microtubule-associated protein 2 by monoclonal antibodies. 618 36

The human neuroblastoma cell line, SH-SY5Y, differentiates into a neuronal, sympathetic phenotype in the presence of phorbol ester and serum. Growth cones prepared from differentiating SH-SY5Y cells have characteristics similar to those of growth cones from embryonic rat brain. In addition, SH-SY5Y growth cones contain ribosomes. In this study we show, by metabolic labeling of isolated growth cones, that local protein synthesis occurred in these structures. The pattern of labeled proteins was very similar to that of the corresponding cell body fraction. RNA was shown to be transported to the growth cone compartment, and by in situ hybridization. beta-actin mRNA could be visualized in intact neuritic growth cones. Comparison by Northern blot hybridizations of RNA prepared from growth cones and cell bodies, respectively, showed that mRNAs coding for growth-associated protein 43, microtubule-associated protein 2, actin, neuropeptide tyrosine, and glyceraldehyde-3-phosphate dehydrogenase were present in both fractions. In contrast, mRNAs coding for the nuclear proteins c-jun and N-myc were virtually absent in the growth cone, but readily detectable in the cell body preparation. The selective distribution of mRNAs to the growth cones was not restricted to stable, abundant mRNA species, since mRNA coding for the insulin-like growth factor I receptor was stable, but not present in growth cones. Thus, differentiating SH-SY5Y cells can sort and transport RNA to the growth cone compartment, suggesting that this system of clonal cells could be useful to unravel mechanisms involved in the compartmentalization of mRNA.
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PMID:Protein synthesis and mRNA in isolated growth cones from differentiating SH-SY5Y neuroblastoma cells. 817 54

In this study, the effects of oxidative stress on calpain-mediated proteolysis and calpain I autolysis in situ were examined. Calpain activity was stimulated in SH-SY5Y human neuroblastoma cells with the calcium ionophore, ionomycin. Calpain-mediated proteolysis of the membrane-permeable fluorescent substrate N-succinyl-L-leucyl-L-leucyl-L-valyl-L-tyrosine-7-amido-4-methylcouma rin, as well as the endogenous protein substrates microtubule-associated protein 2, tau and spectrin, was measured. Oxidative stress, induced by addition of either doxorubicin or 2-mercaptopyridine N-oxide, resulted in a significant decrease in the extent of ionophore-stimulated calpain activity of both the fluorescent compound and the endogenous substrates compared with control, normoxic conditions. Addition of glutathione ethyl ester, as well as other antioxidants, resulted in the retention/recovery of calpain activity, indicating that oxidation-induced calpain inactivation was preventable/reversible. The rate of autolytic conversion of the large subunit of calpain I from 80 to 78 to 76 kDa was decreased during oxidative stress; however, the extent of calpain autolysis was not altered. These data indicate that oxidative stress may reversibly inactivate calpain I in vivo.
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PMID:Oxidative stress inhibits calpain activity in situ. 958 80

alpha-Synuclein is presynaptic nerve terminal protein and its immunoreactivity has been observed in such neurodegenerative structures as senile plaques of Alzheimer's disease or Lewy bodies of Parkinson's disease. The physiological role of alpha-synuclein is still unknown. It is speculated that alpha-synuclein may be expressed in brain tumors, especially in those showing neuronal differentiation. We examined the immunohistochemical localization of alpha-synuclein in 77 human brain tumors. alpha-Synuclein was widely distributed in the brain tumors showing neuronal differentiation. As a result, positive immunostaining for alpha-synuclein was observed in ganglioglioma, medulloblastoma, neuroblastoma, primitive neuroectodermal tumor, pineocytoma/pineoblastoma, and central neurocytoma. Compared with other neuronal markers, the positive ratio of alpha-synuclein was not as high as synaptophysin, microtubule-associated protein 2, neuron-specific enolase and tau, but it was higher than neurofilament and chromogranin A. The expression of synaptophysin was diffusely observed in the cytoplasm, cellular processes and nucleus in tumors showing neuronal differentiation; however, the expression of alpha-synuclein was predominantly observed in the cytoplasm of the tumors as well as in the cellular processes. On the other hand, non-neuronal brain tumors such as astrocytic tumors or meningiomas were totally negative for alpha-synuclein. In conclusion, the appearance of an alpha-synuclein-positive structure was not limited to neurodegenerative diseases, but could also be detected in neoplastic cells showing neuronal differentiation.
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PMID:alpha-Synuclein is expressed in a variety of brain tumors showing neuronal differentiation. 1067 22

A rapid and simple procedure is presented to obtain nearly pure populations of human neuron-like cells from the SH-SY5Y neuroblastoma cell line. Sequential exposure of SH-SY5Y cells to retinoic acid and brain-derived neurotrophic factor in serum-free medium yields homogeneous populations of cells with neuronal morphology, avoiding the presence of other neural crest derivatives that would normally arise from those cells. Cells are withdrawn from the cell cycle, as shown by 5-bromo-2'-deoxyuridine uptake and retinoblastoma hypophosphorylation. Cell survival is dependent on the continuous presence of brain-derived neurotrophic factor, and removal of this neurotrophin causes apoptotic cell death accompanied by an attempt to reenter the cell cycle. Differentiated cells express neuronal markers, including neurofilaments, neuron-specific enolase, and growth-associated protein-43 as well as neuronal polarity markers such as tau and microtubule-associated protein 2. Moreover, differentiated cultures do not contain glial cells, as could be evidenced after the negative staining for glial fibrillary acidic protein. In conclusion, the protocol presented herein yields homogeneous populations of human neuronal differentiated cells that present many of the characteristics of primary cultures of neurons. This model may be useful to perform large-scale biochemical and molecular studies due to its susceptibility to genetic manipulation and the availability of an unlimited amount of cells.
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PMID:Sequential treatment of SH-SY5Y cells with retinoic acid and brain-derived neurotrophic factor gives rise to fully differentiated, neurotrophic factor-dependent, human neuron-like cells. 1093 80


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