Inhibition Of Virus Multiplication Research Articles

Rescue of SV40 from interferon-treated heterokaryons.

Simian virus 40 (SV40)-transformed nonpermissive cells express only the early products of SV40. Heterokaryons formed by fusion of these transformed cells with uninfected permissive cells support the activation of the resident viral genome leading to subsequent viral DNA replication, late protein synthesis and release of progeny virus. Pretreatment of heterokaryon cultures with either mouse or monkey interferon (IFN) before fusion with polyethylene glycol (PEG) produced a dose-dependent inhibition in the appearance of free viral DNA as well as production of infectious virus. The decreased yield of SV40 in these cultures was similar to the inhibition which was observed in mouse or monkey cells incubated with homologous IFN prior to exogenous infection with SV40. When IFN was added to the cultures at progressively later times after fusion with PEG, there was less inhibition of virus production. Although there was a comparable decrease in the production of virus by pretreatment with either mouse or monkey IFN, monkey IFN exerted the inhibition for a longer period of time when added after heterokaryon formation. These results demonstrate that IFN treatment applied even after initiation of SV40 replication can still inhibit virus multiplication.

Specific high-affinity binding of 125I-labeled mouse interferon to interfevon resistant embryonal carcinoma cells in vitro.

Abstract Multipotential mouse embryonal carcinoma cells are resistant to several biologic effects of mouse interferon: inhibition of viral multiplication and inhibition of cell division. Nevertheless using 125 I-labeled highly purified mouse interferon we have shown that these embryonal carcinoma cells express specific interferon receptors in similar number and of the same affinity as interferon sensitive differentiated embryonal carcinoma cells. Thus, mechanisms of interferon resistance are probably multiple: some cells are resistant because they lack specific binding sites for interferon (interferon resistant mouse L1210 cells) and other cells, as shown herein, are resistant to some of the effects of interferon despite binding of interferon to specific receptor sites. Furthermore, these binding sites may be considered functional, since interferon does induce an increase in 2-5A synthetase in these cells.

Further studies on anti-tumor activity of adenovirus fibre protein on murine sarcoma tumours.

The present study demonstrates that subcutaneous injection of purified adenovirus 12 fibre protein (FP) to 4-week-old Balb/c mice, 4, 2 and 1 day(s) before intramuscular injection of murine sarcoma virus (MSV-M) or 2 and 1 day before and 1 day after MSV-M injection causes significant inhibition of tumour growth (P less than 0.001). The evidence suggests that treatment with FP leading to tumour inhibition is due to neither an interferon mechanism nor to inhibition of virus multiplication of FP in the reticuloendothelial system. It is postulated that immune functions, probably against MSV-induced cell antigens, might be the critical mechanisms underlying inhibition of tumour growth.

14] Isolation and translation of mouse interferon mRNAs

It has been observed that the wheat germ cell-free protein-synthesizing system translates mouse interferon mRNA to protein that was active in inhibiting viral multiplication in mouse cells. Easy availability of the wheat embryo and the simplicity of the procedure for the preparation of the extract make this material a good choice for studies of protein synthesis. Mouse interferon mRNAs isolated by the two methods as discussed in this chapter are translated in the wheat germ extract with high efficiency. The fidelity of the translation is reflected in the production of the biologically active product. A high efficiency of translation of mouse interferon mRNA to biologically active protein in the wheat germ ceil-free system needs the presence of polyamines. Translation of many plant viral RNAs in the wheat germ extract is stimulated by addition of polyamines such as spermine or spermidine. It is important to maximize the translation system for both the ionic requirements and the polyamine concentration. The product of translation of mouse mRNA in the wheat germ extract is characterized as mouse interferon by (a) its inhibition of virus multiplication in homologous cells, (b) its absence of activity in heterologous cells, (c) its neutralization by anti-mouse interferon antiserum, and (d) the absence of inhibitory effects on viral multiplication by the translation products obtained from mRNA isolated from non-induced cells.

Reversal by hypotonic medium of the antiviral state induced by lymphoblastoid interferon in human HeLa cells.

The induction kinetics of the antiviral state in HeLa cells treated with human lymphoblastoid interferon (IFN) was studied. In cells treated with 4-200 U/ml IFN, the antiviral state was fully established in 7-9 h. Inhibition of virus multiplication was more rapid if the concentration of IFN was increased to 1,000 U/ml. This antiviral state gradually disappeared during the 48 h after IFN removal. Several compounds known to act on the cell membrane or on the cytoskeleton were tested for their influence on the establishment and reversal of the antiviral state. None of them were found to influence these two parameters to a significant extent. In contrast, placing HeLa cells in medium lacking NaCl partially reversed the blockade to virus multiplication induced by IFN treatment. Cells treated with IFN and later placed in hypotonic medium synthesized virus proteins after encephalomyocarditis virus infection, although at a reduced level compared to cells that had not been treated with IFN.

(E)-5-(2-Bromovinyl)-2'-deoxyuridine: a potent and selective anti-herpes agent.

Of a series of five newly synthesized 2'-deoxyuridine derivatives, including 5-vinyl-dUrd, 5-ethynyl-dUrd, 5-(1-chlorovinyl)-dUrd, (E)-5-(2-bromovinyl)-dUrd, and (E)-5-(2-iodovinyl)-dUrd, the last two compounds were found to exert a marked inhibitory effect on the replication of herpes simplex virus type 1 [ID50 (mean inhibitory dose), 0.004-0.02 microgram/ml]. Both (E)-5-(2-bromovinyl)-dUrd and (E)-5-(2-iodovinyl)-dUrd were highly selective in their anti-herpes activity in that they did not affect the growth or metabolism of the host (primary rabbit kidney) cells unless drug concentrations were used that were 5,000- to 10,000-fold greater than those required to inhibit virus multiplication. In this sense (E)-5-(2-bromovinyl)-dUrd and (E)-5-(2-iodovinyl)-dUrd proved more selective in their activity against herpes simplex virus type 1 than all other anti-herpes compounds that have been described so far. In animal model systems (namely, cutaneous herpes infections of athymic nude mice), (E)-5-(2-bromovinyl)-dUrd suppressed the development of herpetic skin lesions and mortality therewith associated, whether the compound was administered topically or systemically. Under the same conditions, the standard anti-herpes drug 5-iodo-dUrd (Idoxuridine) offered little, if any, protection. Although the precise mechanism of action of (E)-5-(2-bromovinyl)-dUrd and (E)-5-(2-iodovinyl)-dUrd remains to be established, preliminary findings indicate that they do not specifically act at the thymidylate synthetase step.

Systemic consequences of the local lesion reaction to tobacco mosaic virus in a tobacco variety lacking the N gene for hypersensitivity

White Burley tobacco is a differential host for tobacco mosaic virus. The virus strain vulgare gives a systemic infection. In contrast, the strain flavum is restricted to necrotic local lesions which form round the point of infection. When plants formed local lesions on the lower leaves after inoculation with flavum, the lesions formed on the upper leaves in response to a subsequent challenge inoculation with flavum were smaller and fewer than those formed on comparable lelves of plants which had not received the first inoculation. The differential host therefore showed systemic consequences of the initial local lesion reaction similar to the “acquired systemic resistance” described by other authors for tobacco varieties containing the N gene, which causes a local lesion reaction against all strains of tobacco mosaic virus.Multiplication of tobacco mosaic virus in the challenge-inoculated leaves was followed by measuring the accumulation of viral RNA. It is shown that formation of flavum lesions on the lower leaves did not reduce the accumulation of flavum or the systemic strain vulgare when the upper leaves were challenge-inoculated with these strains. These results imply that “systemic acquired resistance” as measured by reduction in lesion size and number is not resistance as measured by inhibition of viral multiplication. The mechanisms involved in restricting lesion size and number in the “resistant” leaves are discussed.

Selectivity of action of an antiherpetic agent, 9-(2-hydroxyethoxymethyl)guanine

A guanine derivative with an acyclic side chain, 2-hydroxyethoxymethyl, at position 9 has potent antiviral activity [dose for 50% inhibition (ED(50)) = 0.1 muM] against herpes simplex virus type 1. This acyclic nucleoside analog, termed acycloguanosine, is converted to a monophosphate by a virus-specified pyrimidine deoxynucleoside (thymidine) kinase and is subsequently converted to acycloguanosine di- and triphosphates. In the uninfected host cell (Vero) these phosphorylations of acycloguanosine occur to a very limited extent. Acycloguanosine triphosphate inhibits herpes simplex virus DNA polymerase (DNA nucleotidyltransferase) 10-30 times more effectively than cellular (HeLa S3) DNA polymerase. These factors contribute to the drug's selectivity; inhibition of growth of the host cell requires a 3000-fold greater concentration of drug than does the inhibition of viral multiplication. There is, moreover, the strong possibility of chain termination of the viral DNA by incorporation of acycloguanosine. The identity of the kinase that phosphorylates acycloguanosine was determined after separation of the cellular and virus-specified thymidine kinase activities by affinity chromatography, by reversal studies with thymidine, and by the lack of monophosphate formation in a temperature-sensitive, thymidine kinase-deficient mutant of the KOS strain of herpes simplex virus type 1 (tsA1).

ANTIVIRAL ACTION AND SELECTIVITY OF 6‐AZAURIDINE

6-Azauridine (AzUrd) is a broad-spectrum antimetabolite that inhibits both DNA and RNA virus multiplication. Prior work indicated that several AzUrd-sensitive viruses induced an increase in the level of uridine kinase, and this might explain the selective activity of AzUrd on such viruses. Present studies compared AzUrd sensitive and resistant viruses with respect to their orotic acid pathways by labeling cells with [14C]-orotic acid during the latent period of viral infection. No differences were detected by this method with either vaccinia, Newcastle disease, or vesicular stomatitis viruses. AzUrd inhibits transport of orotic acid into the cell by 30%, while incorporation of orotic acid into cellular RNA is inhibited by 50% (taking into consideration the 30% already noted) when the highest concentration of antimetabolite is used. This suggests that, in addition to blocking orotidylic acid decarboxylase, AzUrd may act on some other site (sites) of action in the inhibition of virus multiplication.

Inhibition of Echovirus-12 multiplication by N-carbobenzoxy-D-glucosamine.

The glucosamine derivative, N-carbobenzoxy-D-glucosamine (NCBZG) inhibits the multiplication of Echovirus-12 and the synthesis of both virus RNA and protein at a stage in the virus growth cycle after attachment and penetration. However, the compound does not inhibit virus multiplication after the appearance of progeny virus nor after virus RNA has accumulated. Incorporation of radioactive glucosamine and choline into infected and uninfected cultures is inhibited by NCBZG as is the virus-induced increase in choline incorporation. The compound also prevents the appearance of radioactive choline in isolated membranous structures. The compound did not alter significantly the cellular RNA or protein synthesis, plating efficiency of the cells, their growth over a period of several days, nor the virus-directed inhibition of cellular RNA and protein. These findings suggest that the compound inhibits virus multiplication by its effect on the initiation of biosynthesis which appears to require membrane synthesis.

Inhibition of viral multiplication by homologous methylated ribonucleic acids v. Inhibition of myxoviruses in mouse

Mice respond to intravenous injection of homologous methylated RNA by the production of a circulating interferon-like substance. The treatment with modified RNA induces a significant protection against viral infection. This effect is optimum when the treatment occurs a few hours before viral infection. The protective effect is a peculiar property of homologous methylated RNA as heterologous RNAs do not induce any resistance to the infection with influenza virus.

Effect of tetraethyl thiuram disulfide (disulfiram) on the multiplication of enveloped viruses

Disulfiram at concentrations between 0.1 and 0.3 mM inhibits the multiplication of Semliki Forest virus (SFV), fowl plague virus (FPV), Newcastle disease virus (NDV), vesicular stomatitis virus (VSV), and pseudorabies virus (PRV), when administered 1 hour before and during adsorption. There is, however, no inhibition of virus multiplication, when the drug is added after adsorption onto chick embryo cells. Disulfiram interferes neither with the receptors of the virus nor of erythrocytes, and it does not prevent virus adsorption. Possibly an early step in virus multiplication is affected by disculfiram. Infected cells once treated with the drug recover after some time of incubation in an ingibitor-free medium. The inhibitory state can be maintained, however, if relatively low doses of disulfiram are present in the culture medium also after adsorption. Disulfiram has no effect on macromolecular synthesis of the host cells. It has, however, a marked affect on membrane function. While virus multiplication is readily inhibited by disulfiram when chick embryo or BHK cells were investigated, virus multiplication in HeLa cells is almost resestant against the action of disulfiram.

Inhibition of multiplication of tobacco mosaic virus in protoplasts by antibiotics and its prevention by divalent metals.

At concentrations that inhibit bacterial growth, some antibiotics including gentamicin completely inhibited virus multiplication in protoplasts, and other antibiotics partially inhibited virus multiplication. The inhibition caused by each antibiotic was largely prevented by adding a divalent metal; MnC1(2) was more effective than CaC1(2) and other salts of divalent metals when added at 10 mM to the incubation medium. When added immediately after infection, 1 mug/ml of gentamicin halved the final virus concentration and 3 mug/ml completely inhibited virus multiplication, although 10 mug/ml was required to stop bacterial growth. Gentamicin inhibited virus multiplication even when added 24 h after virus inoculation. Also, when protoplasts were exposed to gentamicin for only 1 or 2 h, either immediately after inoculation or 2 h later, the virus concentration was considerably decreased. Gentamicin seemed not to affect virus multiplication in whole plants. Sap from Dianthus barbatus also strongly inhibited virus multiplication in protoplasts but, unlike gentamicin, it acted in the presence of MnC1(2). By contrast, chelating agents such as 1 mM-EDTA or 5 mM-potassium citrate was strong inhibitors of virus multiplication that were inactive in the presence of MnC1(2). It is suggested that gentamicin and other antibiotics may chelate metals from the protoplast membranes, thus disorganizing their function and affecting virus multiplication.

Inhibition of viral multiplication by homologous methylated ribonucleic acids IV. Subcellular localisation after uptake into fibroblasts and relation between antiviral activity and chain length

Summary The addition of chemically modified homologous ribonucleic acids to the culture induces an inhibition of viral infection. All RNAs, modified or not, enter the cells and are preferentially located at the microsomal level. The differences observed in the antiviral properties cannot be explained by the differences in the way of the uptake or by the differences in the specific resistance of the RNAs to nucleases. On the other hand, the antiviral activity is not a peculiar property of a special class of intact methylated RNA but a general property of chemically modified polynucleotidic chains of a minimal chain length of 40 nucleotides.

Inhibitory Effect of Tea Catechin on the Infectivity of Plant Virus

Inhibitory effects of four kinds of tea catechins on infection with plant virus, TMV and CMV, were investigated.Nicotiana tabacunz (KY-57) was used as the infection plant with CMV, and tomato and N. glutinosa as the infection plant TMV. Each of four tea catechins was added in a concentration of 5 mg/ml. to the sap squeeced from the leaves of tobaco or cucumber infected systemically with the virus. The leaves of the plants were inoculated by rubbing with the mixed inoculum.Inhibitory effects of the concentration of tea catechins. (-) ECg and (-) EGCg, on TMV multiplication, using N. glutinosa as the infection plant, were also investigated.When each of four tea catechins was added to a concentration of 0.5% to 1 % gelatin solution, (-) EC and (-) EGC did not give the emulsion, but (-) ECg and (-) EGCg became immediately muddy to form some precipitates. The same results were obtained when virus inoculant of TMV and CMV were used instead of the gelation solution.At 8-25 days after inoculation, symptoms of virus disease by CMV plus (-) EC or (-) EGC were observed in some cases and were not in the others. However, any symptom could not be observed even at 30 days after by CMV plus (-) ECg or (-) EGCg. These results indicated that such tea catechins as(-) ECg and (-) EGCg which seemed to combine readily with protein, had high inhibitory effects on plant virus.In experiments of systemic infection and local lesion formation on the leaves of N. glutinosa by TMV plus tea catechins, inhibitory effects of (-) EC and (-) EGC could not be observed. and also any symptom was not observed in both systemic and local infection by TMV plus (-) ECg or (-) EGCg. Subsequently it was concluded that these two catechins were almost completely inhibitory against TMV multiplication. The number of local lesions on inoculated leaves were very much decreased in proportion to the concentration of (-) ECg and (-) EGCg in the inoculum and the formation of local lesion could not be observed at 0, 5% of them.Thus, it was supposed that (-) ECg and (-) EGCg combined rapidly with plant virus protein, resulting in inhibition of virus multiplication or decreasing of local lesion formation.

Inhibition of Respiratory Virus Infections of Mice with Aerosols of Synthetic Double-Stranded Ribonucleic Acid

Aerosols of double-stranded complexes of polyinosinic and polycytidylic acids (poly I:C) were useful in protecting mice infected with aerosols of influenza (A(2)/Taiwan/64) and parainfluenza type 1 (Sendai) viruses. Administration of poly I:C as an aerosol offers an advantage, particularly in therapy, by eliminating the risk of pulmonary dissemination of viral infections due to intranasally instilled fluids. Treatment of mice with aerosols of poly I:C reduced the infection rate with influenza virus but did not inhibit virus multiplication in the lungs of most of those animals where infection became established. Sendai virus infection rates were undiminished in mice treated with poly I:C, but lung-virus titers were significantly suppressed as compared with those of untreated animals. The maximum poly I:C doses (40 mug) administered by aerosol produced no evidence of toxicity in the mice.

TUNICAMYCIN, A NEW ANTIBIOTIC. II

Tunicamycin showed antiviral activity in the agar diffusion, cytopathic effect suppression and plaque reduction tests against the multiplication of Newcastle disease virus (NDV) in cultured chick embryo fibroblasts. It also inhibited virus multiplication in embryonated eggs. The antiviral activity was observed whenever the antibiotic was added during a one-step growth cycle of NDV. Reversal of the antiviral activity of tunicamycin was observed after removal of the antibiotic, but treatment for more than 2 hours permitted only slight recovery of the virus growth. Static cells were insensitive to tunicamycin, but the antibiotic effected on growing cells. These effects will be correlated with the mode of action of the antibiotic.

Inhibition of viral multiplication by chemically methylated DNA from the host cells.

THE addition to cell cultures of heterologous ribonucleic acids which have secondary structure induces the biosynthesis of interferon1–3. An analogous response is produced by double stranded synthetic polyribonucleotides4–8. We have shown that the addition to the cell culture medium of chemically modified heterologous or homologous RNA produces resistance to viral infection8–11.

Inhibition de la multiplication virale á l'aide d'acides ribonuclêiques chimiquement modifiès: III Influence de la nature et du taux des modifications

Inhibition of viral multiplication by chemically modified ribonucleic acids III. Influence of the nature and the extent of the chemical modifications The multiplication of an Arbovirus (virus Sindbis) on fibroblasts of chicken embryos in the presence of RNA extracted from chicken embryos and modified by aarious chemical reagents was studied. All these chemically modified RNA's induce on inhibition of the viral multiplication. This effect is dependent both on the quantity vf RNA introduced into the cell culture medium and on the level of the chemical modification. In the case of several modifications such as methylation, allylation, deamination by nitrous acid, chloruration or iodination, an increase in the percentage of modified bases gives rise to an increase in the inhibition of the viral multiplication (90–100%). On the contrary, when the RNA is modified by oxidation with mono-perphthalic acid, the increase in the percentage of modified bases produces a decrease in the level of inhibition. This difference is discussed in relation with the integrity of the primary and secondary structure of the RNA's.

Structural requirements of selective inhibition of enteroviruses by 2-(alpha-hydroxybenzyl)-benzimidazole and related compounds.

A survey of the antiviral activity of forty-one derivatives of 2-(α-hydroxybenzyl)-benzimidazole (HBB) pinpoints the structural features of the HBB molecule essential for selective inhibition of viral multiplication.