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Results for “Virus Replication”

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At least 109 records · Page 6Linked to original sources

PubMed1986

Targeting of drug loaded immunoliposomes to herpes simplex virus infected corneal cells: an effective means of inhibiting virus replication in vitro.

The goal of our studies was to develop liposomes containing antiviral drugs and targeted with antiviral antibody (immunoliposomes) that would be effective at inhibiting replication of herpes simplex virus (HSV) in vitro. To achieve this, a monoclonal antibody to glycoprotein D of HSV was derivatized with palmitic acid and was incorporated into the lamellae of dehydration-rehydration vesicles. The gD containing immunoliposomes were shown to bind specifically to HSV-infected rabbit corneal cells in vitro, whereas control immunoliposomes prepared with a monoclonal antibody of the same class as the anti-gD failed to preferentially bind to virus-infected cells. The gD immunoliposome binding was inhibitable by pretreatment with rabbit anti-HSV serum but not by aggregated normal serum. Thus liposome binding was judged to represent an antigen-antibody reaction not binding to Fc receptors expressed by cells infected with HSV. Immunoliposomes loaded with iododeoxyuridine (IUDR) leaked drug rapidly at 37 degrees C, whereas acyclovir (ACV)-loaded liposomes still contained 48% of drug after 24 hr at 37 degrees C. The ACV-liposomes retained 44% of drug after 14 days at 4 degrees C. The ability of immunoliposomes to inhibit virus replication was compared with that of untargeted and empty liposomes by means of virus yield assays in vitro, Immunoliposomes loaded with either IUDR or ACV inhibited virus replication, although ACV-containing immunoliposomes were the most efficacious. The implications of our in vitro results for the development of immunoliposomes suitable for the treatment of ocular herpes infection are briefly discussed.

Acyclovir↗

PubMed1976

Inhibition of influenza virus replication in tissue culture by 2-deoxy-2,3-dehydro-N-trifluoroacetylneuraminic acid (FANA): mechanism of action.

The neuraminidase inhibitor 2-deoxy-2,3-dehydro-N-trifluoroacetylneuraminic acid (FANA) inhibits the mutlicycle replication of influenza viruses in tissue culture. Influenza virus grown in the presence of FANA contains neuraminic acid on its envelope which then serves as receptor for other virus particles causing extensive aggregation. Thus, FANA inhibits influenza virus replication by preventing the enzymatic removal of neuraminic acid from the virus envelope.

Cell Line↗

PubMed2006

Modification of host lipid raft proteome upon hepatitis C virus replication.

Hepatitis C virus (HCV) replication complex resides in detergent-insoluble subcellular domains or lipid rafts. We used two proteomics approaches to characterize the protein content of lipid rafts isolated from Huh7 cells and its modification upon HCV replication. Using two-dimensional electrophoresis and mass spectrometry, we identified approximately 100 protein spots in the isolated lipid rafts; among them, 39 were reproducibly modified in HCV replicon cell lines as compared with control cell lines. We also used stable isotope labeling by amino acids in cell culture (SILAC) combined with one-dimensional electrophoresis separation and mass spectrometry. Using this approach, we identified 1036 individual proteins based on peptides selected with at least 95% confidence; among them, 413 proteins were identified with at least two peptides. Quantification analysis identified 150 proteins modified by at least 2.5-fold (110 up-regulated and 40 down-regulated) in HCV-replicating cells compared with controls. Protein identifications and quantifications obtained by both proteomics approaches were largely concordant. Modulated proteins included a majority of proteins involved in vesicular and protein trafficking and in cell signaling. Remarkably for a large number of proteins, their up-regulation in lipid rafts of HCV replicon cells was due to their relocalization. By using small interfering RNAs directed to the modulated small GTPases Cdc42 and RhoA, we observed an increase in HCV replication, whereas reduction of syntaxin 7 expression resulted in decreased replication of HCV. Our findings indicate that protein subcellular relocalization occurs in HCV-containing cells that can directly affect HCV replication.

Cell Extracts↗

PubMed1993

Hepatitis B virus replication.

Hepatitis B virus (HBV) is the causative agent of B-type hepatitis in humans and the prototypic member of the hepadnaviruses. It is a small enveloped DNA virus that replicates via reverse transcription. Although hepadnaviruses are similar to retroviruses in basic life cycle and genome organization, recent studies have revealed several unique hepadnaviral strategies that optimize exploitation of their extremely small circular DNA genomes.

Animals↗

PubMed1989

Interactions of Pseudorabies virus with swine alveolar macrophages I: virus replication.

The replication of Pseudorabies virus (PRV) in cultured swine alveolar macrophages (AM) was studied using 6 different virus strains. AM were highly permissive to PRV infection, with progeny virus titres of 10(7) TCID50/ml from some strains. Virus progeny titres were higher in cultures infected with the field strains S-62 and 4892 than in cultures infected with the strains Bartha or PRV-C. Virus replication, viral DNA synthesis and the concomitant cell damage were dependent upon virus input m.o.i.s. and virus strain. Furthermore, cells from 7 day old pigs yielded higher virus progeny titres than cells of 6 week old pigs. The results from this study provide support to the premise that PRV infection may predispose anials to respiratory disease.

Aging↗

PubMed1991

Hepatitis B virus replication modulates pathogenesis of hepatitis D virus in chronic hepatitis D.

Hepatitis D virus and hepatitis B virus nucleic acids were detected by Northern and Southern blot hybridization in the sera and livers of 85 chronic carriers of HBsAg and anti-hepatitis D followed up for a mean of 10 yr. We identified three subsets of patients: 13 with hepatitis D virus and hepatitis B virus viremia, 53 with serum hepatitis D virus RNA, but without hepatitis B virus DNA and 19 negative for both nucleic acids. Genomic and subgenomic forms of hepatitis D virus RNA were detected only in patients with hepatitis D virus and hepatitis B virus viremia. Histological findings and disease activity at admission were comparable in the three groups of patients, but the outcome was significantly worse in patients with active replication of both viruses; two of them died of terminal liver failure and hepatocellular carcinoma developed in two; the remaining patients had an uneventful course. These results suggest that active hepatitis B virus replication represents an important previously unrecognized determinant of severe liver damage in patients with chronic hepatitis D virus infection. Since hepatitis B virus provides the means for hepatitis D virus secretion and release from infected cells, active hepatitis B virus multiplication favoring the spread of hepatitis D virus from cell to cell may increase the pathogenetic potential of the defective agent.

Adolescent↗

PubMed1984

Experimental infection of inbred mice with herpes simplex virus. VI. Effect of interferon on in vitro virus replication in macrophages.

Peritoneal exudate cells (PEC) of DBA/2 mice, after 7 days of in vitro preculture and consisting of virtually 100 per cent macrophages, were able to support the replication of Herpes Simplex Virus type 1 strain WAL (HSV). Using a standard medium based on Dulbecco's Modified Eagle Medium (D-MEM), no virus replication was observed in freshly isolated PEC. However a medium based on RPMI 1640 consistently yielded higher virus titres in precultured PEC than the D-MEM medium, and also allowed virus replication in freshly isolated PEC. Macrophages derived from the spleens or the bone marrow, and precultured in the same way as PEC represented a highly pure population and were permissive for infection with HSV. Titres of about 10(6) PFU HSV were observed in PEC 48 hours after infection with 10(3) or 10(6) PFU. However, whereas a complete destruction of the cell monolayer was observed 24 hours after infection with 10(6) PFU, complete cytopathogenicity in PEC infected with 10(3) PFU required at least twice this time. In the latter situation, plaque formation was observed 24 hours after infection. PEC of different strains of mice were compared. Of these, PEC of all mice that are susceptible to HSV infection in vivo replicated HSV to the same degree as PEC of DBA/2 mice, whereas PEC of resistant C57BL/6 and C3H/HeJ mice produced 1000 fold lower titres of viral progeny. Whereas the number of infectious centres were equal in PEC of DBA/2 and C57BL/6 mice, the plaques observed after infection of confluent PEC with a low MOI were considerable smaller in cells from C57BL/6 mice. Furthermore, significantly higher titres of interferon were measured in the supernatants of HSV-infected C57BL/6 macrophages than in those of DBA/2 macrophages, and the former were made fully susceptible by the in vitro addition of an anti-interferon serum.

Animals↗

PubMed1988

The role of cytoskeleton and nuclear matrix in virus replication.

In the light of the cytoarchitecture concept, the presented review attempts to cover what is known of the involvement of cytoskeletal and nucleoskeletal elements in the replication cycle of various viruses and in cell transformation. Our knowledge on the relationship of virus replication with cell architecture has rapidly progressed during the recent years in association with studies on composition of various filaments within cells, their spatial organization and possible functions which focussed much interest on the cytoskeleton and nuclear matrix. The new results indicated that highly specialized elements for normal cell function may be coopted for virus growth. Recently, using various methodical approaches it has been possible to obtain information about the association of virus-structural proteins with cytoskeletal and nucleoskeletal elements, about the reorganization of cytoskeleton during virus replication, about the role of tubulin in transcription and RNA synthesis of negative-strand viruses, about the involvement of cytoskeletal filaments in the transport of viral proteins, virus penetration, virus assembly and release from the infected cell. The role of cytoskeleton in cell transformation and in initiation of DNA synthesis and intracellular signaling to cell proliferation has been also investigated.

Adenoviridae↗

PubMed1994

Hepatitis C virus replication and antibody responses toward specific hepatitis C virus proteins.

We assessed the correlation between hepatitis C virus replication and antibody responses toward hepatitis C virus core (C22-3), NS3 (C33C), NS4 (5-1-1 and C100-3) and NS5 proteins in 59 virus carriers. The concentration of serum hepatitis C virus RNA was determined by a competitive reverse transcription-polymerase chain reaction assay. All 50 patients with high viremic levels of > or = 10(6) copies/mL had antibodies to C22-3 and C33C. Antibodies to 5-1-1, C100-3 and NS5 proteins were detected less frequently (p < 0.01) in 72% (36 of 50), 78% (39 of 50) and 84% (32 of 38) of such patients, respectively. As for the nine patients with low viremic levels of < 10(6) copies/mL, antibodies to C22-3, C33C, 5-1-1 and NS5 proteins were detected in only one patient (11%), which was significantly less than the frequency for highly viremic patients (p < 0.01). Antibody to C100-3 was also found less frequently in only four patients (44%) (p < 0.05). Thus, only four (44%) of the nine low viremic patients tested positive for any antibody compared with all 50 highly viremic patients (p < 0.01). These results indicate that highly viremic carriers can be detected by the presence of hepatitis C virus antibodies, but a considerable proportion of low viremic carriers may not show any serological evidence of hepatitis C virus infection.

Adult↗

PubMed1997

Inhibition of human immunodeficiency virus replication by the herpes simplex virus virion host shutoff protein.

The herpes simplex virus (HSV) virion host shutoff gene (vhs) encodes a protein which nonspecifically accelerates the degradation of mRNA molecules, leading to inhibition of protein synthesis. This ability to inhibit a critical cellular function suggested that vhs could be used as a suicide gene in certain gene therapy applications. To investigate whether vhs might be useful for treatment of AIDS, we tested the ability of both HSV type 1 (HSV-1) and HSV-2 vhs to inhibit replication of human immunodeficiency virus (HIV). Replication of HIV was substantially inhibited when an infectious HIV proviral clone was cotransfected into HeLa cells together with vhs under the control of the cytomegalovirus (CMV) immediate-early promoter. HSV-2 vhs was more active than HSV-1 vhs in these experiments, consistent with previously published studies on these genes. Since expression of vhs from the CMV promoter is essentially unregulated, we also tested the ability of vhs expressed from the HIV long terminal repeat (LTR) promoter to inhibit HIV replication. Wild-type HSV-1 vhs inhibited HIV replication more than 44,000-fold in comparison to a mutant vhs gene encoding a nonfunctional form of the Vhs protein. Production of Vhs in transfected cells was verified by Western blot assays. A larger amount of Vhs was observed in cells transfected with plasmids expressing vhs from the HIV LTR than from the CMV promoter, consistent with the greater inhibition of HIV replication observed with these constructs. Mutant forms of Vhs were expressed at higher levels than wild-type Vhs, most likely due to the ability of wild-type Vhs to degrade its own mRNA. The strong inhibitory activity of the vhs gene and its unique biological properties make vhs an interesting candidate for use as a suicide gene for HIV gene therapy.

Anti-HIV Agents↗

PubMed2003

Recombinant Newcastle disease virus as a viral vector: effect of genomic location of foreign gene on gene expression and virus replication.

Newcastle disease virus (NDV) was examined for its suitability as a vector for the expression and delivery of foreign genes for vaccination and gene therapy. A reporter gene encoding human secreted alkaline phosphatase (SEAP) was inserted as an additional transcription unit at four different positions in the NDV genome, between the NP and P, M and F, and HN and L genes and behind the L gene. Eight infectious recombinant NDV (rNDV) viruses, four in the non-virulent strain NDFL and four in the virulent derivative NDFLtag, were generated by reverse genetics. SEAP expression levels, replication kinetics and virus yield were examined. Replication kinetics of the rNDV viruses in primary chicken embryo fibroblasts showed that the insertion of an additional gene resulted in a delay in the onset of replication. This effect was most prominent when the gene was inserted between the NP and P genes. With the exception of the strain that carried the SEAP gene behind the L gene, all recombinant strains expressed high levels of SEAP, both in cell culture and in embryonated chicken eggs. In embryonated eggs, the rNDV viruses showed a 2.6- to 5.6-fold (NDFL) or 2.1- to 8.1-fold (NDFLtag) reduction in yield compared with the parent strains. These results show that foreign genes can be inserted at different positions in the NDV genome without severely affecting replication efficiency or virus yield.

Alkaline Phosphatase↗

PubMed1974

Primaquine diphosphate: inhibition of Newcastle disease virus replication.

The response of Newcastle disease virus replication to primaquine, an antimalarial drug, was examined in chicken embryo cells (CEC). Virus-induced hemadsorption was completely inhibited by 250 mug of primaquine per ml. At lower concentrations, hemadsorption inhibition was dose dependent. Primaquine retarded virus-induced redistribution of receptor sites on the host cell plasma membrane as shown by the failure of infected, drug-treated CEC to be agglutinated with concanavalin A. The production of infectious progeny virus was substantially inhibited by the addition of primaquine at various times postinfection. When the drug was added early in the virus replication cycle, viral ribonucleic acid (RNA) synthesis was inhibited; however, when the drug was added late in the cycle, stimulation of RNA synthesis was observed. Primaquine was also shown to retard the incorporation of [(14)C]amino acids into proteins of virus-infected CEC. We suggest that the major role of primaquine is inhibition of protein synthesis; this results in changes in: hemadsorption, redistribution of lectin receptors, release of progeny, and virus-induced RNA synthesis.

Agglutination↗

PubMed2006

Role of the amphipathic peptide of Semliki forest virus replicase protein nsP1 in membrane association and virus replication.

Semliki Forest virus RNA replication takes place in association with specific cytoplasmic vacuoles, derived from the endosomal apparatus. Of the four virus-encoded replicase proteins, nsP1 serves as the membrane anchor of the replication complex. An amphipathic peptide segment, G245STLYTESRKLLRSWHLPSV264, has been implicated in the membrane binding of nsP1. nsP1 variants with changes within the peptide were studied after protein expression and in the context of virus infection. Proteins with mutations R253E and W259A accumulated in the cytoplasm and were very poorly palmitoylated. The same mutations also drastically affected the localization of the precursor polyprotein P123, and they were lethal when introduced into the virus genome. Mutations R253A and L255A+L256A partially changed the localization of nsP1, and the respective viruses acquired compensatory changes. L255A+L256A only yielded virus encoding L255A+L256V, indicating the importance of a hydrophobic residue in the central 256 position. When fused to green fluorescent protein, the peptide was required in at least two tandem copies to effect a change in localization, but even then the fusion protein was associated with membranes in a nonspecific manner. Thus, the amphipathic peptide is a crucial element for the membrane association of nsP1 and the replication complex. It provides essential affinity for membranes, and other regions of nsP1 also appear to contribute to the localization of the protein.

Amino Acid Sequence↗

PubMed1981

Inhibiting effects of enflurane and isoflurane anesthesia on measles virus replication: comparison with halothane.

Replication of measles virus in BSC cells was studied in the presence of enflurane (2-chloro-1,1,2-trifluoroethyl difluoromethyl ether), a commonly used volatile anesthetic agent, and its isomer, isoflurane (1-chloro-2,2,2-trifluoroethyl difluoromethyl ether). At clinical concentrations of the anesthetics (up to 4%), cell division was retarded, whereas only minimal toxic cellular effects were observed. The appearance of progeny virus from the cell cultures exposed to these anesthetics was decreased in a dose-related manner. Incorporation of [(3)H]uridine into measles virus nucleocapsids also decreased progressively with increasing anesthetic concentrations. In comparing the inhibition of measles virus replication in the presence of halothane (2-bromo-2-chloro,1,1,1-trifluoroethane), enflurane, or isoflurane, it was found that both inhibition of the appearance of infectious virus at 48 h postinfection and incorporation of [(3)H]uridine into measles virus nucleocapsids were proportional to the anesthetic concentrations. An equivalent degree of effect was produced by anesthetically equivalent concentrations of the three anesthetics (minimal alveolar concentration) but not by absolute concentrations. In addition, recovery of infectious virus synthesis from the inhibition encountered during exposure of infected BSC cells to halothane or isoflurane was also investigated. In cultures exposed to halothane or enflurane, recovery of infectious virus synthesis was rapid and complete. Recovery of virus synthesis was slower after isoflurane removal and did not reach the peak control titers of infected cultures not exposed to the anesthetic. Treatment with halothane resulted in the formation of a preponderance of slowly sedimenting virus nucleocapsid particles which contained less than full-length ribonucleic acids after anesthetic removal. Neither enflurane nor isoflurane treatment of BSC cultures resulted in the formation of significant levels of these slowly sedimenting particles with short genomes after anesthetic removal.

Capsid↗

PubMed1999

Mutations in the retinoblastoma protein-binding LXCXE motif of rubella virus putative replicase affect virus replication.

The rubella virus (RV)-encoded protein NSP90, which contains the retinoblastoma protein (Rb)-binding motif LXCXE, interacts with Rb and RV replication is reduced in cells lacking Rb. Whether the LXCXE motif of RV NSP90 itself is essential for Rb binding and virus replication is not known. Therefore, in the present study, the functional role of this motif was investigated by site-directed mutagenesis in a plasmid from which infectious RV RNA can be produced. Three critical mutations in the motif, two substitutions at the conserved cysteine residue (C --> G and C --> R) and a deletion of the entire motif, were created. A cell-free translated NSP90 C terminus polypeptide containing the deletion did not bind to Rb and a polypeptide carrying the C --> R substitution had barely detectable binding affinity for Rb. Rb binding by the C --> G mutant was reduced significantly compared to that of wild-type protein. Correlating with the binding results, mutant viruses containing the LXRXE and LXGXE motifs had a reduction in replication to < 0.5% and 47% of the wild-type, respectively, while deletion of the motif was found to be lethal. By the first serial passage, replication of the LXRXE-carrying virus had increased from < 0.5% to 2% of the wild-type. Sequencing of the genome of this virus revealed a nucleotide change that altered the motif from LXRXE to LXSXE, which is a known Rb-binding motif in two protein phosphatase subunits. Thus, our results clearly demonstrate that the LXCXE motif is required for efficient RV replication.

Amino Acid Sequence↗

PubMed1972

Inhibition of influenza virus replication by -amanitin: mode of action.

The replication of influenza virus in chick embryo fibroblast cells is inhibited by alpha-amanitin added during the first 2 hr of infection at concentrations similar to those required to inhibit cellular DNA-dependent RNA polymerase form II in vivo. Of two periods of increased RNA synthesis observed in cells infected with influenza virus, only the first, occurring from 0 to 2 hr after infection, is sensitive to alpha-amanitin. During this early period, there is a stimulation of the activity of DNA-dependent RNA polymerase II of nuclei isolated from infected cells. The data suggest that DNA transcription mediated by polymerase II is essential for influenza virus replication.

Animals↗

PubMed2005

Host cell nuclear proteins are recruited to cytoplasmic vaccinia virus replication complexes.

The initiation and termination of vaccinia virus postreplicative transcription have been reported to require cellular proteins, some of which are believed to be nuclear proteins. Vaccinia virus replicates in the cytoplasmic compartment of the cell, raising questions as to whether vaccinia virus has access to nuclear proteins. This was addressed here by following the fate of several nuclear proteins after infection of cells with vaccinia virus. The nuclear transcription factors YY1, SP1, and TATA binding protein were found to colocalize with virus replication complexes in the cytoplasm of infected cells. In addition, the nuclear proteins RNA polymerase II, TAFIIp32, and histone deacetylase 8, but not the structural protein lamin B, also were found in the cytoplasm of the cell. The association of YY1 with replication complexes was dependent on DNA replication and required only the DNA binding domain of the protein, indicating that DNA binding alone may be responsible for the association of nuclear transcription factors with viral replication complexes in the cytoplasm. The cytoplasmic localization of YY1 was resistant to the nuclear export inhibitor leptomycin B. Evidence is presented indicating that nuclear import and export pathways were not adversely affected by vaccinia virus infection. These observations indicate that vaccinia virus replication complexes have ready access to nuclear proteins by allowing leakage from the nucleus.

Animals↗

PubMed1998

Inhibition of Mayaro virus replication by prostaglandin A1 and B2 in Vero cells.

The effect of prostaglandins (PGA1 and PGB2) on the replication of Mayaro virus was studied in Vero cells. PGA1 and PGB2 antiviral activity was found to be dose-dependent. However, while 10 micrograms/ml PGB2 inhibited virus yield by 60%, at the same dose PGA1 suppressed virus replication by more than 90%. SDS-PAGE analysis of [35S]-methionine-labelled proteins showed that PGA1 did not alter cellular protein synthesis. In infected cells, PGA1 slightly inhibited the synthesis of protein C, while drastically inhibiting the synthesis of glycoproteins E1 and E2.

Alphavirus↗