Cell line neuroblastoma

Most recently, a phase-I-study defined a dose of 13-ris-retinoic acid that was tolerable in patients after myeloablative therapy, and a phase-III-trial showed that postconsolidation therapy with 13-cis-retinoic acid improved EFS for patients with high-risk neuroblastoma [7]. Preclinical studies in neuroblastoma indicate that ATRA or 13-cw-RA can antagonize cytotoxic chemotherapy and radiation, such that use of 13-cis-RA in neuroblastoma is limited to maintenance after completion of cytotoxic chemotherapy and radiation. It is likely that recurrent disease seen during or after 13-cis-RA therapy in neuroblastoma is due to tumor cell resistance to retinoid-mediated differentiation induction. Studies in neuroblastoma cell lines resistant to 13-cw-RA and ATRA have shown that they can be sensitive, and in some cases collaterally hypersensitive, to the cytotoxic retinoid fenretinide. Here, fenretinide induces tumor cell cytotoxicity rather than differentiation, acts independently from RA receptors, and in initial phase-I-trials has been well tolerated. Clinical trials of fenretinide, alone and in combination with ceramide modulators, are in development. 

Bottenstein, J.E. Sato, G.H. (1979). Growth of a rat neuroblastoma cell line in senan-ffee supplemented medium. Proc. Natl. Acad. Sci. USA 76,514-517. 

Tissue Culture Assay. Kogure et al. (48) report a novel tissue culture assay for detecting several types of sodium channel blockers. The mouse neuroblastoma cell line ATCC CCL 131 is grown in RPMI 1640 supplemented with 13.5% fetal bovine serum and 100 pg/ml gentamycin, in an atmosphere of 5% C0 95% air at 37 C. Ninety-six well plates are seeded with 1 x 10 cells in 200 pi of medium containing 1 mM ouabain and 0.075 mM veratridine. Veratridine and ouabain cause neuroblastoma cells to round-up and die. In the presence of sodium channel blockers (e.g., TTXs or STXs), the lethal action of veratridine is obviated and cells retain normal morphology and viability. An important feature of this assay is that a positive test for sodium channel blockers results in normal cell viability. Since bacterial extracts can contain cytotoxic components, this assay offers an advantage over tests that use cell death as an endpoint. The minimum detectable level of TTX is approximately 3 nM, or approximately 1/1000 mouse unit. 

In addition to the pre-synaptic effects, CX3CL1 modulates the functional properties of ligand-gated channels at post-synaptic sites. In SK-N-SH cells, a human neuroblastoma cell line, CX3CL1 reduces the amplitude of NMDA-induced calcium transients (Deiva et al. 2004). In these cells, CX3CL1 application causes a PTX insensitive transient increase in the intracellular Ca " concentration dependent on... 

Besides the synthetic inhibitors, a variety of natural compounds is known to inhibit the CP. One of these natural inhibitors, lactacystin, was discovered by its ability to induce neurite outgrowth in a murine neuroblastoma cell line. Incubation of cells in the presence of radioactive lactacystin leads to the labelling of the yS5 subunit (Fenteany et al. 1995) and to irreversible inhibition of the CP. As shown by X-ray analysis, the inhibitor is covalently attached to subunit fS5 by an ester bond with the N-terminal ThrlO (Groll et al. 1997) (see Figure 10.7A). The subunit selectivity of lactacystin can be attributed to its dimethyl group, which mimics a valine or a leucine side chain and closely interacts with Met45 in the hydrophobic SI pocket of subunit j85. 

Newton RA, Phipps SL, Flanigan TP, Newberry NR, Carey JE, Kumar C, McDonald B, Chen C, Elliott JM. (1996). Characterisation of human 5-hydroxytryptamine2A and 5-hydroxytryptamine2C receptors expressed in the human neuroblastoma cell line SH-SY5Y comparative stimulation by hallucinogenic drugs. J Neurochem. 67(6) 2521-31. 

The implementation of animal test protocols in the 1980s has been accompanied by the development of a host of alternative methods to study adverse effects of chemicals on reproductive and developmental parameters. For example, rat whole embryo culture stems from the seventies (16), as does the rat limb bud organ culture (17) and rat limb bud and brain micromass was developed in the eighties (18). An elegant nonvertebrate alternative model used regeneration of polyps of Hydra atUnuata from dissociated cells (19). Animal-free in vitro alternatives include those employing the proliferation of a human embryonic palatal mesenchymal cell line (20), the attachment of a mouse ovarian tumor cell line (21), and the differentiation of a neuroblastoma cell line (22) and a embryonal carcinoma cell line (23). Various overviews of methods have been published over the years (24). The predictability of... 

In a human neuroblastoma cell line, Cova et al. (1992) found acrylonitrile to be highly toxic, showing an EC50 of 72.5 nM for cytotoxicity. The cytotoxic potency of potassium cyanide was 2.5 J,M, thus acrylonitrile toxicity in these cells cannot be attributed to its metabolism to cyanide. 

Cova, D., Fumagalli, P. Santagostino, A. (1992) Toxicity of acrylonitrile in a human neuroblastoma cell line and its effect on glutathione and glutathione-S-transferase. Bull, environ. Contam. Toxicol., 49, 886-891... 

Before implantation several in vitro tests were performed. For evaluation of a possible toxic reaction, we investigated the material and the whole devices in vitro with cell culture methods. Direct contact and extraction tests with a mouse fibroblasts cell line (L 929) and a neuroblastoma cell line (neuro-2-a) were performed according to the international standard ISO 10993 ( Biological Evaluation of Medical Devices ). The materials and devices showed no toxicity, i.e. no significant differences in membrane integrity of the cell membranes, mitochondrial activity and DNA synthesis rate. The neuro-2-a cell line is so sensitive that even small changes in process technology are detectable. The flexible polyimide structures proved to be non toxic. 

Fountoulakis M, Schlaeger EJ. The mitochondrial proteins of the neuroblastoma cell line IMR-32. Electrophoresis 2003 24 260-275. 

Tsuji, S., Arita, M., and Nagai, Y. (1983). GQlb, a bioactive ganglioside that exhibits novel nerve growth factor (NGF)-like activities in two neuroblastoma cell lines. J. Biochem. 94, 303-306. 

Tsuji, S., Yamashita, T., Matsuda, Y., and Nagai, Y. (1992). A novel glycosignaling system GQlb-dependent neuritogenesis of human neuroblastoma cell line, GOTO, is closely associated with GQlb-dependent ecto-type protein phosphorylation. Neurochem. Int. 21, 549-554. 

The delta opioid receptor also could stimulate the PLC activity and increase intracellular Ca2+ level in mechanism other than the activation of Gi/Go proteins. In a human neuroblastoma cell line, SK-N-BE, delta opioid receptors mobilize Ca2+ from intracellular ryanodine-sensitive stores which is independent of the PTX-sensitive G /G0 proteins [94]. Coexpressing the delta opioid receptor with G16, a promiscuous G protein, allows for a PTX-insensitive stimulation of the PLC activity by the opioid agonist [95]. Thus, the ability of the opioid receptors to stimulate PLC(3 is determined in part by the availability of complementary G proteins in any particular cell type. 

Oh JE, Karlmark KR, Shin JH, Poliak A, Freilinger A, Hengstschlager M, et al. Differentiation of neuroblastoma cell line N1E-115 involves several signaling cascades. Neuro-chem Res 2005 30(3) 333-348. 

Urbani A, Poland J, Bernardini S, Bellincampi L, Biroccio A, Schnolzer M, et al. A proteomic investigation into etoposide chemo-resistance of neuroblastoma cell lines. Proteomics 2005 5(3) 796-804. 

Flupirtine was the first compound identified to affect KCNQ channels and has been used in man to treat pain. However, only recently has this drug been shown to be an activator of Kv7 channels (see Munro and Dalby-Brown 2007). The clinical efficacy of flupirtine was originally postulated to occur through receptor-dependent mechanisms, but as noted by Munro and Dalby-Brown (2007) it is likely that Kv7 channel activation occurs at clinically relevant exposure levels. Retigabine, a flupirtine analog, is currently in clinical trials for the treatment of epilepsy and pain and has been the most widely used tool to explore both the in vitro and in vivo effects of Kv7 activation. This compound was initially shown to increase K+ channel activity in a neuroblastoma cell line (Rundfeldt 1997 see also Rundfeldt 1999) and subsequently was... 

Jamaicamides A, B, and G exhibited cytotoxidty to both the H-460 human lung carcinoma and neuro-2a mouse neuroblastoma cell lines. The LG50S were approximately 15 p,M for all three compounds to both cell lines. All three compounds also exhibited sodium charmel-blocking activity at 5 p,M concentration. In the goldfish toxicity assay, a system that has been useful for the detedion of neurotoxic activity in cmde extrads as well as purified compounds, jamaicamide B was the most active (100 % lethality at 5 ppm after 90 min), followed by jamaicamide G (100 % lethality at... 

Seitz G, Gebhardt S, Beck JF, Bohm W, Lode HN, Niethammer D, and Bruchelt G (1998) Ascorbic acid stimulates DOPA synthesis and tyrosine hydroxylase gene expression in the human neuroblastoma cell line SK-N-SH. Neuroscience Letters 244,33-6. 

Leira, E, Alvarez, C., Vieites, IM., Vieytes, M.R., and Botana, L M. 2002. Characterization of distinct apoptotic changes induced by okadaic acid and yessotoxin in the BE(2)-M 17 neuroblastoma cell line. Toxicol in Vitro 16, 23-31. 

Yokosawa, N., Kurokawa, Y., Tsuzuki, K., S mto, B., Fujii, N., Kimura, K., Oguma, K. (1989). Binding of Clostridium botulinum type C neurotoxin to different neuroblastoma cell lines. Infect. Immun. 57 272-7. 

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