Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0027819 (neuroblastoma)
 27,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Lentiviral-mediated gene delivery holds significant promise for sustained gene expression within living systems. Vesicular stomatitis virus glycoprotein-pseudotyped human immunodeficiency virus type 1-based lentiviral vectors can be used to introduce transgenes in a broad spectrum of dividing as well as nondividing cells. In the current study, we construct a lentiviral vector carrying two reporter genes separated by an internal ribosomal entry site and utilize that virus in delivering both genes into neuroblastoma cells in cell culture and into cells implanted in living mice. We utilize two reporter genes, a mutant herpes simplex virus type 1 (HSV1) sr39tk as a reporter gene compatible with positron emission tomography (PET) and a bioluminescent optical reporter gene, firefly luciferase (Fluc), to image expression in living mice by an optical charge-coupled device (CCD) camera. By using this lentivirus, neuroblastoma (N2a) cells are stably transfected and a high correlation (R(2) = 0.91) between expressions of the two reporter genes in cell culture is established. Imaging of both reporter genes using microPET and optical CCD camera in living mice is feasible, with the optical approach being more sensitive, and a high correlation (R(2) = 0.86) between gene expressions is again observed in lentiviral-infected N2a tumor xenografts. Indirect imaging of HSV1-sr39tk suicide gene therapy utilizing Fluc is also feasible and can be detected with increased sensitivity by using the optical CCD. These preliminary results validate the use of lentiviral vectors carrying reporter genes for multimodality imaging of gene expression and should have many applications, including imaging of xenografts, metastasis, and cell trafficking as well as noninvasive monitoring of lentiviral-mediated gene delivery and expression.
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PMID:Noninvasive imaging of lentiviral-mediated reporter gene expression in living mice. 1271 11

Vesicular stomatitis virus (VSV) is a rhabdovirus which causes acute encephalitis in mice after intranasal infection. Because type I interferon (IFN) has been shown to be a potent inhibitor of VSV, we investigated the role of type I IFN in viral replication in neurons in culture. Pre-treatment of NB41A3 neuroblastoma cells or primary neuron cultures with IFN-beta or IFN-alpha strongly inhibits virus replication, with 1000-fold inhibition of infectious virus release occurring at 7 h post-infection, and maximum inhibition of 14,000-fold occurring at 14 h. Type I IFN inhibited both viral protein and RNA synthesis, but not enough to account for the inhibition of infectious virus yield. The influenza virus protein NS1 binds dsRNA and antagonizes induction of PKR activity, an IFN-inducible antiviral protein which phosphorylates and inactivates the elongation factor eIF-2alpha, resulting in cessation of translation. In NS1-expressing neuroblastoma cells, VSV replication was inhibited by IFN-beta as well as in control NB41A3 cells, and eIF-2alpha phosphorylation was blocked, suggesting that PKR activity was not involved in inhibition of viral protein synthesis. Similarly, inhibition of VSV by IFN-beta was not affected by addition of inhibitors of nitric oxide synthase, indicating that IFN-beta activity is not mediated by nitric oxide or superoxide. This contrasts with the essential role of NOS-1 in inhibition of VSV replication when neurons are treated with IFN-gamma. Analysis of cell culture supernatants revealed suppression of release of VSV particles from both NB41A3 cells and primary neurons treated with IFN. The inhibition of virion release closely matched the overall suppression of infectious VSV particle release, suggesting that type I IFN plays a role in inhibition of VSV assembly.
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PMID:VSV replication in neurons is inhibited by type I IFN at multiple stages of infection. 1572 56

Vesicular stomatitis virus (VSV) replication is highly sensitive to interferon (IFN)-induced antiviral responses. Pretreatment of sensitive cultured cells with IFNbeta results in a 10(4)-fold reduction in the release of infectious VSV particles. However, differences exist between the mechanisms of reduced infectious particle titers in cell lines of neuroblastoma and nonneuronal lineage. In L929-fibroblast-derived cells, using immunofluorescence confocal microscopy, infection under control conditions reveals the accumulation of VSV matrix, phosphoprotein (P), and nucleocapsid (N) proteins over time, with induced cellular morphological changes indicative of cytopathic effects (CPEs). Upon observing L929 cells that had been pretreated with IFNbeta, neither detectable VSV proteins nor CPEs were seen, consistent with type I IFN antiviral protection. When using the same techniques to observe VSV infections of NB41A3 cells, a neuroblastoma cell line, aside from similar viral progression in the untreated control cells, IFNbeta-treated cells illustrated a severely attenuated VSV infection. Attenuated VSV progression was observed through detection of VSV matrix, P, and N proteins in isolated cells during the first 8 h of infection. However, by 18-24 h postinfection all neuroblastomas had succumbed to the viral infection. Finally, upon closer inspection of IFNbeta-treated NB41A3 cells, no detectable changes in VSV protein localization were identified compared with untreated, virally infected neuroblastomas. Next, to extend our study to test our hypothesis that virion assembly is compromised within type I IFN-treated neuroblastoma cells, we employed electron microscopy to examine our experimental conditions at the ultrastructural level. Using VSV-specific antibodies in conjunction with immuno-gold reagents, we observed several similarities between the two cell lines, such as identification of viroplasmic regions containing VSV N and P proteins and signs of stress-induced CPEs of VSV-infected cells, which had either been mock-treated or pretreated with interferon-beta (IFNbeta). One difference we observed between nonneuronal and neuroblastoma cells was more numerous actively budding VSV virions across untreated L929 plasma membranes compared with untreated NB41A3 cells. Additionally, IFNbeta-treated, VSV-infected L929 cells exhibited neither cytoplasmic viroplasm nor viral protein expression. In contrast, IFNbeta-treated, VSV-infected NB41A3 cells showed evidence of VSV infection at a very low frequency as well as small-scale viroplasmic regions that colocalized with viral N and P proteins. Finally, we observed that VSV viral particles harvested from untreated VSV-infected L929 and NB41A3 cells were statistically similar in size and shape. A portion of VSV virions from IFNbeta-treated, virally infected NB41A3 cells were similar in size and shape to virus from both untreated cell types. However, among the sampling of virions, pleomorphic viral particles that were identified from IFNbeta-treated, VSV-infected NB41A3 cells were different enough to suggest a misassembly mechanism as part of the IFNbeta antiviral state in neuroblastoma cells.
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PMID:A confocal and electron microscopic comparison of interferon beta-induced changes in vesicular stomatitis virus infection of neuroblastoma and nonneuronal cells. 2011 3