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Sindbis Virus (Sindbi + virus)
Selected AbstractsAltered immune response to CNS viral infection in mice with a conditional knock-down of macrophage-lineage cellsGLIA, Issue 2 2006Jessica Carmen Abstract Neuroadapted Sindbis Virus (NSV) is a neuronotropic virus that causes hindlimb paralysis in susceptible mice and rats. The authors and others have demonstrated that though death of infected motor neurons occurs, bystander death of uninfected neurons also occurs and both contribute to the paralysis that ensues following infection. The authors have previously shown that the treatment of NSV-infected mice with minocycline, an inhibitor that has many functions within the central nervous system (CNS), including inhibiting microglial activation, protects mice from paralysis and death. The authors, therefore, proposed that microglial activation may contribute to bystander death of motor neurons following NSV infection. Here, the authors tested the hypothesis using a conditional knock-out of activated macrophage-lineage cells, including endogenous CNS macrophage cells. Surprisingly, ablation of these cells resulted in more rapid death and similar weakness in the hind limbs of NSV-infected animals compared with that of control animals. Several key chemokines including IL-12 and monocyte chemoattractant protein-1 (MCP-1) did not become elevated in these animals, resulting in decreased infiltration of T lymphocytes into the CNS of the knock-down animals. Either because of the decreased macrophage activation directly or because of the reduced immune cell influx, viral replication persisted longer within the nervous system in knock-down mice than in wild type mice. The authors, therefore, conclude that although macrophage-lineage cells in the CNS may contribute to neurodegeneration in certain situations, they also serve a protective role, such as control of viral replication. © 2006 Wiley-Liss, Inc. [source] The use of gene silencing to study the role of dopa decarboxylase in mosquito melanization reactionsINSECT MOLECULAR BIOLOGY, Issue 3 2005C.-Y. Huang Abstract Mosquito melanization involves hydroxylation of tyrosine to dopa, which then is oxidized to dopaquinone by phenoloxidase, or decarboxylated to dopamine by dopa decarboxlase (DDC). An Armigeres subalbatus cDNA encoding DDC was cloned and real-time PCR analysis revealed increased transcripts in blood-fed and microfilariae (mf)-inoculated mosquitoes. A double subgenomic Sindbis virus was used to silence DDC and assess its role in melanization of mf. DDC transcription and activity were significantly decreased in silenced mosquitoes, as was the degree of mf melanization 48 h postinoculation; however, melanization increased after 72 and 96 h, demonstrating that DDC influences the rate of melanization. DDC-silenced mosquitoes also exhibit high mortality, over-feeding and abnormal movement, consistent with an involvement of DDC in neurotransmission. [source] Development of an orally infectious Sindbis virus transducing system that efficiently disseminates and expresses green fluorescent protein in Aedes aegyptiINSECT MOLECULAR BIOLOGY, Issue 2 2003D. J. Pierro Abstract We have constructed an orally infectious Sindbis virus, ME2/5,2J/GFP, that expresses green fluorescent protein (GFP) in the midgut of Aedes aegypti and in other tissues as the virus disseminates. This virus has two unique features that are improvements over the SIN-based expression systems currently used in mosquitoes. First, a subgenomic RNA promoter and GFP coding sequence is located 5,- to the second subgenomic promoter and structural genes of the virus. Second, the E2 glycoprotein gene of TE/5,2J/GFP is replaced with the E2 gene of MRE16 SIN virus. The first feature enhances virus genome stability during virus dissemination from the midgut to other tissues and the second allows efficient virus entry into the midgut epithelial cells and then spread of the virus throughout the mosquito. [source] The inflammatory cytokine, interleukin-1 beta, mediates loss of astroglial glutamate transport and drives excitotoxic motor neuron injury in the spinal cord during acute viral encephalomyelitisJOURNAL OF NEUROCHEMISTRY, Issue 4 2008Natalie A. Prow Abstract Astrocytes remove glutamate from the synaptic cleft via specific transporters, and impaired glutamate reuptake may promote excitotoxic neuronal injury. In a model of viral encephalomyelitis caused by neuroadapted Sindbis virus (NSV), mice develop acute paralysis and spinal motor neuron degeneration inhibited by the AMPA receptor antagonist, NBQX. To investigate disrupted glutamate homeostasis in the spinal cord, expression of the main astroglial glutamate transporter, GLT-1, was examined. GLT-1 levels declined in the spinal cord during acute infection while GFAP expression was preserved. There was simultaneous production of inflammatory cytokines at this site, and susceptible animals treated with drugs that blocked IL-1, release also limited paralysis and prevented the loss of GLT-1 expression. Conversely, infection of resistant mice that develop mild paralysis following NSV challenge showed higher baseline GLT-1 levels as well as lower production of IL-1, and relatively preserved GLT-1 expression in the spinal cord compared to susceptible hosts. Finally, spinal cord GLT-1 expression was largely maintained following infection of IL-1,-deficient animals. Together, these data show that IL-1, inhibits astrocyte glutamate transport in the spinal cord during viral encephalomyelitis. They provide one of the strongest in vivo links between innate immune responses and the development of excitotoxicity demonstrated to date. [source] Photoinactivation of Sindbis Virus Infectivity Without Inhibition of Membrane FusionPHOTOCHEMISTRY & PHOTOBIOLOGY, Issue 3 2009Wor Thongthai Photoinactivation of enveloped viruses is commonly associated with damage to fusion proteins and inhibition of membrane fusion capacity. Here we show that photobleaching of Sindbis virus labeled with the membrane localized dye, R18 (octadecyl rhodamine B) causes a dramatic loss of infectivity without observable changes in low-pH triggered membrane fusion to liposomes. Sindbis labeled with DiI (1,1,-dioctadecyl-3,3,3,,3,-tetramethylindocarbocyanine perchlorate) also maintains low-pH triggered membrane fusion capacity, but in contrast to R18, extensive photobleaching of DiI-labeled virus has little effect on infectivity. Electrophoretic gel analysis suggests no cross-linking of viral fusion proteins following photobleaching of dye-labeled Sindbis. These observations have implications for live-cell, single particle tracking studies of dye-labeled Sindbis virus. Our observations suggest that R18 and DiI have different propensities for spontaneous flip-flop in lipid bilayers. [source] Amelioration of alphavirus-induced arthritis and myositis in a mouse model by treatment with bindarit, an inhibitor of monocyte chemotactic proteinsARTHRITIS & RHEUMATISM, Issue 8 2009Nestor E. Rulli Objective Alphaviruses such as chikungunya virus, Sindbis virus, o'nyong-nyong virus, Mayaro virus, and Ross River virus (RRV), are commonly associated with arthralgias and overt arthritides worldwide. Understanding the processes by which arthritogenic viruses cause disease is a prerequisite in the quest for better treatments. In this regard, we have recently established that monocyte/macrophages are mediators of alphavirus-induced arthritis in mice. We hypothesized that chemokines associated with monocyte/macrophage recruitment may play an important role in disease. The aim of the present investigations was to determine whether bindarit, an inhibitor of monocyte chemotactic protein (MCP) synthesis, could ameliorate alphavirus-induced rheumatic disease in mice. Methods Using our recently developed mouse model of RRV-induced arthritis, which has many characteristics of RRV disease (RRVD) in humans, the effects of bindarit treatment on RRVD in mice were determined via histologic analyses, immunohistochemistry, flow cytometry, real-time polymerase chain reaction analysis, enzyme-linked immunosorbent assay, and electrophoretic mobility shift assay. Results Bindarit-treated RRV-infected mice developed mild disease and had substantially reduced tissue destruction and inflammatory cell recruitment as compared with untreated RRV-infected mice. The virus load in the tissues was not affected by bindarit treatment. Bindarit exhibited its activity by down-regulating MCPs, which in turn led to inhibition of cell infiltration and lower production of NF-,B and tumor necrosis factor ,, which are involved in mediating tissue damage. Conclusion Our data support the use of inhibitors of MCP production in the treatment of arthritogenic alphavirus syndromes and suggest that bindarit may be useful in treating RRVD and other alphavirus-induced arthritides in humans. [source] Cell permeabilization by poliovirus 2B viroporin triggers bystander permeabilization in neighbouring cells through a mechanism involving gap junctionsCELLULAR MICROBIOLOGY, Issue 8 2010Vanesa Madan Summary Poliovirus 2B protein is a well-known viroporin implicated in plasma membrane permeabilization to ions and low-molecular-weight compounds during infection. Translation in mammalian cells expressing 2B protein is inhibited by hygromycin B (HB) but remains unaffected in mock cells, which are not permeable to the inhibitor. Here we describe a previously unreported bystander effect in which healthy baby hamster kidney (BHK) cells become sensitive to HB when co-cultured with a low proportion of cells expressing poliovirus 2B. Viroporins E from mouse hepatitis virus, 6K from Sindbis virus and NS4A protein from hepatitis C virus were also able to permeabilize neighbouring cells to different extents. Expression of 2B induced permeabilization of neighbouring cell lines other than BHK. We found that gap junctions are responsible mediating the observed bystander permeabilization. Gap junctional communication was confirmed in 2B-expressing co-cultures by fluorescent dye transfer. Moreover, the presence of connexin 43 was confirmed in both mock and 2B-transfected cells. Finally, inhibition of HB entry to neighbouring cells was observed with 18,-glycyrrhethinic acid, an inhibitor of gap junctions. Taken together, these findings support a mechanism involving gap junctional intercellular communication in the bystander permeabilization effect observed in healthy cells co-cultured with poliovirus 2B-expressing cells. [source] Proceedings of the Australian Physiological and Pharmacological Society Symposium: New Frontiers in Muscle Research Gene transfer: manipulating and monitoring function in cells and tissuesCLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 8 2001Rekha G Panchal SUMMARY 1. The ectopic expression of genes has proven to be an extremely valuable tool for biologists. The most widely used systems involve electrically or chemically mediated transfer of genes to immortalized cell lines and, at the other end of the spectrum, transgenic animal models. As would be expected, there are compromises to be made when using either of these broad approaches. Immortalized cell lines have limited ,physiological relevance' and transgenic approaches are costly and out of the reach of many laboratories. There is also significant time required for the de novo generation of a transgenic animal. 2. As a viable alternative to these approaches, we describe the use of recombinant adenovirus and Sindbis virus to deliver genes to cells and tissues. 3. We exemplify this approach with studies from our laboratories: (i) an investigation of Ca2+ handling deficits in cardiac myocytes of hypertrophied hearts using infection with recombinant adenovirus encoding either green fluorescent protein (GFP) or the sarcoplasmic/endoplasmic reticulum calcium-ATPase (Serca2a); (ii) a study of the mechanism of macrophage/microglial migration by infection of embryonic phagocytes with a GFP-encoding virus and coculture with brain slices to then track the movement of labelled cells; and (iii) we are also exploiting the natural tropism of the Sindbis virus to label neurons in hippocampal brain slices in culture to resolve high-resolution structure and to map neuronal connectivity. 4. Further development of these approaches should open new avenues of investigation for the study of physiology in a range of cells and tissues. [source] | |||||||||||||