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

Astroglial-conditioned medium (GCM) induced two distinct, but intimately related, phases of neuritogenesis in NB2a/d1 neuroblastoma cells--a "rapid-outgrowth," unstable phase, and a delayed, relatively stable phase, which are apparently regulated by glial-derived protease inhibitors and laminin, respectively. The initial rapid outgrowth (less than 4 hr) may be mediated by inhibition of a thrombin-like protease, present as a serum component and/or adsorbed to the outer neuronal surface, since (1) a similar effect was obtained by serum removal or by adding the specific thrombin inhibitor, hirudin; (2) exogenous thrombin inhibited the rapid outgrowth of neurites by GCM; and (3) cell-free enzyme assays confirmed the presence of thrombin-inhibitory activity in GCM. Although neurites induced by removal of serum removal or hirudin addition are rapidly resorbed following serum replenishment or hirudin depletion, GCM-induced neurites continued to elongate after GCM removal, indicating that GCM contained additional neurite-promoting factors. Anti-laminin antiserum did not inhibit the initial elaboration of neurites by GCM but prevented their continued elongation. Anti-laminin antiserum had no affect on neurite outgrowth induced by serum deprivation. The more protracted, second phase of neurite outgrowth could also be achieved by the addition of soluble purified laminin to undifferentiated cells. Unlike neurites at 4 hr, neurites at 24 hr were no longer dependent on the protease inhibitors in GCM, since exogenous thrombin no longer caused them to retract. Simultaneous addition of thrombin and anti-laminin antiserum with GCM had identical inhibitory effects on continued neurite elaboration at 24 hr as did anti-laminin antiserum without thrombin.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Sequential effects of astroglial-derived factors on neurite outgrowth: initiation by protease inhibitors and potentiation by extracellular matrix components. 157 81

The initial outgrowth of neuritogenesis in mouse NB2a/d1 neuroblastoma cells may be regulated by thrombin or a thrombin-like protease, present either in serum or adsorbed to the plasma membrane, since neuritogenesis is induced by serum deprivation and treatment with the specific thrombin inhibitor, hirudin (Shea et al., 1991, J. Neurochem., 56:842). Cultured astroglial cells secrete factors that promote neuritogenesis, including protease inhibitors active against thrombin, leading to suggestions that the inhibition of specific neuronal surface proteases by the surrounding glial environment may represent an initial step in axonal outgrowth in situ. To examine the relative importance of glial-derived protease inhibitory activities on neurine outgrowth, we tested the neurite promoting effect of glial-conditioned medium (GCM) on NB2a/d1 cells. Like serum deprivation and hirudin treatment, GCM induced neurite outgrowth within 4 hr. Exogenous thrombin inhibited the effect of GCM, and cell-free enzyme assays confirmed the presence of thrombin-inhibitory activity in GCM, suggesting that GCM induces neuritogenesis by inhibition of a thrombin-like protease. Unlike neurites induced by serum removal or hirudin addition, which are rapidly resorbed following serum replenishment or hirudin depletion, however, GCM-induced neurites continued to elongate after GCM removal. Furthermore, cultures treated simultaneously with GCM and thrombin exhibited delayed outgrowth of neurites following GCM removal which were insensitive to further thrombin treatment. These findings indicate that the initial elaboration of neurites can be mediated by glial-derived protease inhibitor(s) active against a thrombin-like protease, but indicate the requirement of additional glial-derived factors for the maintenance and continued elaboration of these neurites.
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PMID:Transient neuritogenesis in NB2a/d1 neuroblastoma cells induced by glial-derived protease inhibitors. 191 50

Recent studies have suggested important roles for certain proteases and protease inhibitors in the growth and development of the CNS. In the present studies, inhibitors of urokinase or thrombin in cultured neural cells and serum-free medium from the cells were identified by screening for components that formed sodium dodecyl sulfate-stable complexes with 125I-urokinase or 125I-thrombin. Rinsed glioblastoma possessed two components that complexed 125I-urokinase. One was type 1 plasminogen activator inhibitor (PAI-1), because the 125I-urokinase-containing complexes were immunoprecipitated with anti-PAI-1 antibodies. The other component formed complexes with 125I-urokinase that were not recognized by antibodies to PAI-1 or protease nexin-1 (PN-1). Its identity is unknown. In addition to these cell-bound components, the glioblastoma cells also secreted two inhibitors that formed complexes with 125I-urokinase; one was PAI-1, and the other was PN-1. The secreted PN-1 also formed complexes with 125I-thrombin. It was the only thrombin inhibitor detected in these studies. Human neuroblastoma cells did not contain components that formed detectable complexes with either 125I-urokinase or 125I-thrombin. However, human neuroblastoma cells did contain very low levels of PN-1 mRNA and PN-1 protein. Added PN-1 bound to the surface of both glioblastoma and neuroblastoma cells. This interaction accelerated the inhibition of thrombin by PN-1 and blocked the ability of PN-1 to form complexes with 125I-urokinase. Thus, cell-bound PN-1 was a specific thrombin inhibitor.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Inhibitors of urokinase and thrombin in cultured neural cells. 198 20

Protease nexin-1 (PN-1) is a protein proteinase inhibitor recently shown to be identical with the glial-derived neurite-promoting factor or glial-derived nexin. It has been shown to promote neurite outgrowth in neuroblastoma cells and in sympathetic neurons. The present experiments were designed to further test the hypothesis that this activity on neuroblastoma cells is due to its ability to complex and inhibit thrombin. It has been suggested that PN-1:thrombin complexes might mediate the neurite outgrowth activity of PN-1. However, the present studies showed that such complexes, unlike free PN-1, did not promote neurite outgrowth. The neurite outgrowth activity of PN-1 was only detected in the presence of thrombin or serum (which contains thrombin). PN-1 did not affect the rate or extent of neurite outgrowth that occurred when neuroblastoma cells were placed in serum-free medium. Retraction of neurites by thrombin was indistinguishable in cells whose neurites had been extended in the presence or absence of PN-1. The neurite-promoting activity of PN-1 was inhibited by an anti-PN-1 monoclonal antibody, which blocks its capacity to complex serine proteinases. The plasma thrombin inhibitor, antithrombin III, stimulated neurite outgrowth but only when its thrombin inhibitory activity was accelerated by heparin. The neurite outgrowth activity of both antithrombin III and PN-1 corresponded to their inhibition of thrombin. Together, these observations show that PN-1 promotes neurite outgrowth from neuroblastoma cells by inhibiting thrombin and suggest that this depends on the ability of thrombin to retract neurites.
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PMID:Neurite outgrowth activity of protease nexin-1 on neuroblastoma cells requires thrombin inhibition. 229 18

Glia-derived nexin (GDN) is a serine protease inhibitor which promotes the outgrowth of neurites from neuroblastoma cells and from chick sympathetic neurons. However, it has not been demonstrated that this effect is independent of its protease inhibitory activity. We report here that, 48 h after the addition of GDN to astrocyte-free cultures of rat hippocampal cells, there was a significant increase in axon length, although dendrite length and the total number of neurites were unaffected. Nerve growth factor added alone had no significant effect nor was there any additional effect when it was added together with GDN. However, hirudin, a thrombin inhibitor purified from leeches, was found to mimic the GDN effect at similar molar concentrations of active protein. This suggests that the protease inhibitory activity is crucial for the neurite-promoting effect of GDN on hippocampal neurons.
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PMID:Glia-derived nexin potentiates neurite extension in hippocampal pyramidal cells in vitro. 233 38

Thrombin has been reported to inhibit neurite outgrowth from neuroblastoma cells grown in serum-containing medium after switching to serum-free medium. A test of the serum and substrate-dependence of this inhibition became possible with the development of Neurobasal/B27 serum-free medium. Inhibition of sprouting of Nb2a neuroblastoma cells by thrombin occurred from the substrate where it was bound to material adsorbed from serum. Neuritogenesis of primary hippocampal neurons was unaffected by exogenous thrombin on polylysine substrates with or without serum treatment. However, sprouting of hippocampal neurons was stimulated by treating the substrate with hirudin, a highly specific thrombin inhibitor. This suggests that hippocampal neurons are not directly responsive to added thrombin, perhaps because they produce their own thrombin.
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PMID:Exogenous thrombin inhibits neuritogenesis in cultured neuroblastoma cells but not in rat hippocampal neurons. 755 63

We examined the interdependence of calpain and protein kinase C (PKC) activities on neurite outgrowth in SH-SY-5Y human neuroblastoma cells. SH-SY-5Y cells elaborated neurites when deprived of serum or after a specific thrombin inhibitor, hirudin, was added to serum-containing medium. The extent of neurite outgrowth under these conditions was enhanced by treatment of cells with the cell-permeant cysteine protease inhibitors N-acetyl-leucyl-leucyl-norleucinal ("C1") and calpeptin or by the phospholipid-mediated intracellular delivery of either a recombinant peptide corresponding to a conserved inhibitory sequence of human calpastatin or a neutralizing anti-calpain antisera. Calpain inhibition in intact cells was confirmed by immunoblot analysis showing inhibition of calpain autolysis and reduced proteolysis of the known calpain substrates fodrin and microtubule-associated protein 1. The above inhibitory peptides and antiserum did not induce neurites in medium containing serum but lacking hirudin, suggesting that increased surface protein adhesiveness is a prerequisite for enhancement of neurite outgrowth by calpain inhibition. Treatment of cells with the PKC inhibitor H7, staurosporine, or sphingosine induced neurite outgrowth independently of serum concentration. Because calpain is thought to regulate PKC activity, we examined this potential interrelationship during neurite outgrowth. Simultaneous treatment with calpain and PKC inhibitors did not produce additive or synergistic effects on neurite outgrowth. PKC activation by 2-O-tetradecanoylphorbol 13-acetate (TPA) prevented and reversed both neurite initiation by serum deprivation and its enhancement by calpain inhibitors. Treatment of cells with the calpain inhibitor C1 retarded PKC down-regulation following TPA treatment. Cell-free analyses demonstrated the relative specificity of various protease and kinase inhibitors for calpain and PKC and confirmed the ability of millimolar calcium-requiring calpain to cleave the SH-SY-5Y PKC regulatory subunit from the catalytic subunit, yielding a free catalytic subunit (protein kinase M). These findings suggest that the influence of PKC on neurite outgrowth is downstream from that of surface adhesiveness and calpain activity.
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PMID:Enhancement of neurite outgrowth following calpain inhibition is mediated by protein kinase C. 761 5

We have tested the hypothesis that hippocampal neurons respond to thrombin via a neuronal thrombin receptor. A human neuroblastoma cell line, SK-N-SH, known to be thrombin responsive morphologically, responded both to thrombin and thrombin receptor agonist peptide (TRAP 42-55) with elevation of intracellular calcium. In Western blots of membranes from SK-N-SH cells and cultured rat hippocampal neurons using an antibody against the N-terminal peptide of the human thrombin receptor, putative receptor proteins of 66 and 47 kDa were detected in both cells. Neurons were treated with thrombin and TRAP 42-55 (TRAP-14) to determine their effects on intracellular levels of calcium and cAMP. Only 10% of the neurons showed a rapid response to thrombin, but most responded rapidly to agonist peptide with a prolonged elevation of intracellular free calcium. Neuronal cAMP levels were decreased by 40% after 24 h thrombin treatment. This decrease in cAMP level could be blocked by both the Gi-protein inhibitor, pertussis toxin, and the thrombin inhibitor, hirudin, suggesting a possible involvement of Gi-protein-coupled receptor activation. Furthermore, rapid calcium and cAMP responses were apparently induced by pre-treatment of neurons with thrombin for 24 h and subsequent washout. In summary, these data indicate that rat primary hippocampal neurons have thrombin receptors whose responses to thrombin apparently are up-regulated by 24 h thrombin pre-treatment. These results may have implications for synaptic remodeling, learning and memory.
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PMID:Thrombin receptor on rat primary hippocampal neurons: coupled calcium and cAMP responses. 924 61

Thrombin is a multifunctional protease. Recent studies on cultured neuronal cells have suggested a function for thrombin in the development and maintenance of the nervous system. Thrombin has been found to induce neurite retraction and reverse stellation in neuroblastoma cell lines and rat astrocytes, respectively. The major focus of our study was to investigate the potential role of thrombin in peripheral nervous system development using the rat embryonic dorsal root ganglion model. We found a dose dependent inhibition of neurite outgrowth from explant dorsal root ganglion cultures upon exposure to 2 to 200 nM thrombin. This effect was reversed by the specific thrombin inhibitor, hirudin. A synthetic peptide that imitates the fully active receptor, thrombin receptor activating peptide, was also found to inhibit neurite outgrowth from dorsal root ganglia. bis-Benzimide stained neuronal cultures did not show any evidence of cell death after exposure to thrombin or thrombin receptor activating peptides. Immunohistochemical studies revealed specific staining of the thrombin receptor on neurons, with intense labeling along neurites. Enriched neuronal cultures exposed to thrombin and thrombin receptor activating peptides revealed rapid activation of phospholipase Cgamma-1, a second messenger associated with the thrombin receptor. These findings are the first to describe the localization of the thrombin receptor to dorsal root ganglion neurons. We propose that receptor activation is associated with thrombin induced inhibition of neurite outgrowth.
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PMID:Thrombin induced inhibition of neurite outgrowth from dorsal root ganglion neurons. 966 59

Increasing evidence indicates several roles for thrombin-like serine proteases and their cognate inhibitors (serpins) in normal development and/or pathology of the nervous system. In addition to its prominent role in thrombosis and/or hemostasis, thrombin inhibits neurite outgrowth in neuroblastoma and primary neuronal cells in vitro, prevents stellation of glial cells, and induces cell death in glial and neuronal cell cultures. Thrombin is known to act via a cell surface protease-activated receptor (PAR-1), and recent evidence suggests that rodent neurons express PAR-1. Previously, we have shown that the thrombin inhibitor, protease nexin-1, significantly prevents neuronal cell death both in vitro and in vivo. Here we have examined the effects of human alpha-thrombin and the presence and/or activation of PAR-1 on the survival and differentiation of highly enriched cultures of embryonic chick spinal motoneurons. We show that thrombin significantly decreased the mean neurite length, prevented neurite branching, and induced motoneuron death by an apoptosis-like mechanism in a dose-dependent manner. These effects were prevented by cotreatment with hirudin, a specific thrombin inhibitor. Treatment of the cultures with a synthetic thrombin receptor-activating peptide (SFLLRNP) mimicked the deleterious effects of thrombin on motoneurons. Furthermore, cotreatment of the cultures with inhibitors of caspase activities completely prevented the death of motoneurons induced by either thrombin or SFLLRNP. These findings indicate that (1) embryonic avian spinal motoneurons express functional PAR-1 and (2) activation of this receptor induces neuronal cell degeneration and death via stimulation of caspases. Together with previous reports, our results suggest that thrombin, its receptor(s), and endogenous thrombin inhibitors may be important regulators of neuronal cell fate during development, after injury, and in pathology of the nervous system.
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PMID:Thrombin perturbs neurite outgrowth and induces apoptotic cell death in enriched chick spinal motoneuron cultures through caspase activation. 971 58


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