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Biomedical subjects

H Tsiang

Publications and source records attributed to H Tsiang.

At least 37 records · Page 2Linked to original sources
PubMed1992

Inhibition of rabies virus transcription in rat cortical neurons with the dissociative anesthetic ketamine.

In a previous study (B. P. Lockhart, H. Tsiang, P. E. Ceccaldi, and S. Guillemer, Antiviral Chem. Chemother. 2:9-15, 1991), we demonstrated an antiviral effect of the general anesthetic ketamine for rabies virus in neuronal cultures and in rat brain. This report describes an attempt to determine at what level ketamine acts on the rabies virus cycle in rat cortical neuron cultures. Immunofluorescence and [35S]methionine labelling of infected neurons showed that ketamine (1 to 1.5 mM) inhibited viral nucleoprotein and glycoprotein syntheses. Northern (RNA) blots of total RNA from drug-treated neurons, hybridized with 32P-labelled oligonucleotide probes for rabies virus nucleoprotein, matrix protein, and glycoprotein genes, showed a marked reduction (5- to 11-fold) in the levels of rabies virus mRNAs, relative to those in untreated neurons. No significant change in the levels of cellular beta-actin mRNA were detected in ketamine-treated cells. A similar antiviral effect was observed with MK-801; however, no inhibition of rabies virus synthesis was observed with the general anesthetic chloral hydrate. The antiviral effect was not complete; a time-dependent recovery of viral transcription and rabies virus protein synthesis was observed, but no infectious virus was released into the culture supernatant. The lack of any modification of cellular protein or mRNA synthesis by ketamine suggests an antiviral mechanism acting at the level of rabies virus genome transcription.

Animals↗
PubMed1991

Sleep alterations in experimental street rabies virus infection occur in the absence of major EEG abnormalities.

Brain electrical activity and sleep organization were investigated in chronically implanted mice during street rabies virus infection. Continuous EEG recordings showed no gross electrical abnormalities until a few hours before the fatal issue. In contrast, alterations of sleep stages were observed at an early stage during the course of rabies virus infection, at a time when clinical signs were absent. Quantification by spectral analysis showed that the main feature was the early decrease of REM-sleep stages and the increase of the duration of waking stages. Neuromuscular disorders which could occur early were also observed during the disease. Comparison of these data with those obtained from fixed rabies virus infection shows that in the latter the EEG recordings demonstrated early alterations and a progressive deterioration with disappearance of both sleep and waking stages, which were replaced by a pathological sleep stage. In order to evaluate the potential role of the host-specific immune response in promoting brain electrophysiological alterations, EEG recordings and spectral analysis were also performed in cyclophosphamide-treated mice. Street rabies virus-infected and immunosuppressed mice showed identical physiopathological changes as those observed in immunocompetent mice. The implication of these viral-induced electrophysiological alterations in the context of the pathogenic mechanisms of rabies virus is discussed.

Animals↗
PubMed1991

Rabies virus infection and transport in human sensory dorsal root ganglia neurons.

Cultured human sensory neurons are directly susceptible to CVS rabies virus infection and produce virus yields of 10(5) p.f.u./ml; infection can persist for more than 20 days without any sign of c.p.e. The use of a compartmentalized two-chamber culture system, with access to either the cell soma or neuritic extensions, permitted the study of viral retrograde transport, which occurs at between 50 and 100 mm/day. Neurons of human origin were more susceptible to virus infection than rat neurons and the axonal transport of rabies virus was more efficient. Electron microscopy allowed virus transport and infection of human dorsal root ganglia neurons to be observed.

Axons↗
PubMed1991

Actin-independent maturation of rabies virus in neuronal cultures.

This study outlines the effects of a modification of the actin-based cytoskeleton on the maturation of rabies virus in human neuroblastoma cell and primary rat cortical neuron cultures. In a Ca(2+)-depleted or an EGTA-containing medium, disruption of microfilaments did not affect intracellular viral nucleoprotein synthesis, as demonstrated by dual-immunofluorescence microscopy, and caused no change in the extracellular titre of rabies virus. Furthermore, the continuous presence of the anti-calmodulin drugs trifluoperazine (1 to 20 microM) and chlorpromazine (1 to 30 microM), or the L-type Ca2+ channel antagonist nifedepine (1 to 10 microM) or the Ca(2+)-specific ionophore A23187 (0.05 to 1.0 microM), did not modify the extracellular titre of rabies virus significantly over a 48 h period. The inference from these studies is that the maturation of rabies virus is independent of the integrity of the microfilament structures and calmodulin-dependent processes of neuronal cells.

Actins↗
PubMed1991

Inhibition of rabies virus infection in cultured rat cortical neurons by an N-methyl-D-aspartate noncompetitive antagonist, MK-801.

A noncompetitive N-methyl-D-aspartate (NMDA) antagonist, MK-801 (0.5 to 2.0 mM), inhibits rabies virus infection in rat primary cortical neurons, whereas the competitive NMDA antagonist AP5 has no effect. The results suggest that MK-801-mediated inhibition of rabies virus replication, although selective, is not operating through the high-affinity binding site mechanism.

2-Amino-5-phosphonovalerate↗
PubMed1990

Continuous delivery of colchicine in the rat brain with osmotic pumps for inhibition of rabies virus transport.

Rabies virus is a neurotropic agent which spreads in the CNS via axonal transport. Previous studies had shown that this axonal transport through the brain could be inhibited by stereotaxic administration of colchicine; however, this inhibition was reversible. We describe here a method to enhance the duration of this colchicine-mediated inhibition by delivering the drug continuously in the rat brain with osmotic pumps.

Animals↗
PubMed1989

Inhibition of the transport of rabies virus in the central nervous system.

The effect of colchicine, an inhibitor of axonal transport, on the spread of rabies virus in the central nervous system was investigated using Wistar rats. Colchicine was inoculated into the striatum at various times before and after inoculation of rabies virus into the same site. Rats were killed at various times after viral inoculation and the spread of rabies virus was monitored by rabies immunofluorescence of selected areas of brain. The most effective inhibitory effect was obtained by colchicine treatment applied two days before virus inoculation. Under these conditions, no fluorescent foci could be detected until day 3 post-infection whereas control rats exhibited infected cells as soon as two days post-infection. This inhibitory effect is reversible and the general consequence seems to be a delay in the rate of viral spread. However, five days after the virus challenge, some major brain areas were still partially preserved from infection (striatum, frontal cortex, pyriform cortex). Ten days after colchicine treatment, the microtubules have recovered their capacity to transport the virus. At the onset of paralysis, the general pattern of infection in brain sections from colchicine-treated rats was not significantly different from that of control rats. This inhibitory effect on the transport of rabies virus can be prolonged by administration of additional colchicine.

Animals↗
PubMed1989

The anterograde transport of rabies virus in rat sensory dorsal root ganglia neurons.

We have previously described the capacity of neurites extending from cultured rat sensory dorsal root ganglia (DRG) neurons to transport rabies virus through axoplasm in the retrograde direction. Here we report the infection of cultured neurons derived from the DRG and the subsequent anterograde transport of rabies virus from the infected cell somas through the extending neurites to its release into the culture supernatant. Viral transport was monitored by titration of the virus yield in the external compartment. Both early and late transport mechanisms of rabies virions were identified. The first one occurred a few hours post-infection and was undetectable 6 h later, before the initiation of viral replication. The velocity of this first wave of infective virions was in the range of 100 to 400 mm/day. The early viral transport was probably the result of a direct translocation of infective virions from the somatic site of entry to the neuritic extensions and subsequent release into the culture medium without replication in the cellular perikaryon. The second virus transport peak was detected 48 h post-infection. In this case, the virions detected in the neuritic compartment were presumably the progeny of the inoculated virus which had replicated in the perikaryon before the viral transport occurs. Using a four-compartment culture device we were able to demonstrate, simultaneously, retrograde and anterograde transport of the virus. The presence of antirabies serum in contact with the exposed neurites did not inhibit either the retrograde or the anterograde transport mechanisms. The viral release from the neuritic extensions after the fast anterograde transport was evaluated to be in the range of 150 to 300 infectious virions per bundle of neurites per day.

Animals↗
PubMed1988

Inhibition of rabies virus infection by a soluble membrane fraction from the rat central nervous system.

This paper describes the inhibitory effect of a normal rat brain solubilized membrane preparation (RBSM-liposomes) on rabies virus infection. Rabies virus was incubated with RBSM-liposomes or their separated components (proteins, phospholipids, gangliosides) before infection of CER or neuroblastoma cells. In addition, both RBSM-liposomes and target cells were treated with enzymes prior to the infection step. All these experimental procedures showed that the active components were mainly lipids.

Animals↗
PubMed1988

Rapid diagnosis of rabies infection by means of a dot hybridization assay.

Dot hybridization was used to detect specific rabies RNA in brains, either from experimental infection in mouse or from brain material to be processed for routine diagnosis. 32P cDNA probes were employed to identify minute amounts of specific viral RNA. Purified RNA was obtained after phenol extraction. The RNA was fixed on nylon membranes and hybridized with a pool of M13 inserts complementary to 200-400 nucleotides of each rabies gene and mRNA. Hybridized, labelled probes were detected by autoradiography. There was strong cross-hybridization between fixed rabies and street rabies virus RNA, which enable the detection of field strains for diagnosis purpose using a fixed rabies (PV strain) cDNA. A positive response was obtained with as little as 80 ng of brain RNA material from a fixed rabies-infected mouse. Detection of viral RNA was still specific 1 week after death, the brain material being left at room temperature. A total correlation was found when the samples were examined in parallel using a fluorescent rabies-specific antibody and by virus isolation on murine neuroblastoma cells. These data show that the use of rabies-specific cDNA probes in a dot-blot hybridization assay has great potential for the diagnosis of rabies.

Animals↗
PubMed1987

The effect of interferon treatment in rabies prophylaxis in immunocompetent, immunosuppressed, and immunodeficient mice.

The development of rabies is modulated by many interacting factors, most of which are dependent on the host immune response. For this reason, we studied the action of interferon (IFN) treatment on street rabies virus infection in mice, immunocompetent or immunosuppressed with cyclophosphamide. In immunocompetent mice, paralysis of hind limbs is the first symptom characteristic of rabies disease before weight loss and general prostration leading to death. Paralysis does not occur in immunosuppressed mice, which develop a shaggy hair and eventually lose weight and die. Administration of interferon (10(5) units, intraperitoneally) 1 h after virus inoculation and every 24 h led to a delay in the onset of first disease signs, but in general did not rescue immunocompetent or immunosuppressed mice from death. In both types of mice, rabies virus production in the brain was reduced by 1 log in response to IFN treatment. In immunocompetent mice treated with IFN, there was a significant increase of antibody synthesis against rabies virus. As expected, antibody synthesis in immunosuppressed mice was almost negligible. However, in mice treated with IFN and cyclophosphamide there was still significant antibody synthesis specific for rabies virus. IFN administered intravenously, subcutaneously, or intraperitoneally crosses the blood-brain barrier to cause enhanced levels of the two double-stranded RNA-dependent enzymes, the protein kinase and 2',5'-oligoadenylate (2-5A) synthetase in the brain. However in spite of this effect, IFN treatment seems to be unable to prevent the evolution of rabies disease in immunocompetent and immunosuppressed mice. Since the suppressing effect of cyclophosphamide is nonselective on both the cellular and humoral immune responses of mice, we investigated the action of IFN in rabies virus-infected athymic nude mice, which lack T cells. Athymic nude mice infected with street rabies virus become cachectic and die without any apparent symptom of paralysis of hind limbs. IFN treatment (administered as above) protected nude mice against rabies infection. Three months after virus inoculation and 2 months after the end of IFN treatment, 7 of 8 IFN-treated mice remained in perfect health. These results illustrate that the efficacy of IFN treatment against the evolution of rabies disease in mice is dependent on the suppression of the T-cell-mediated immune response of the host.

2',5'-Oligoadenylate Synthetase↗
PubMed1987

Rabies virus infection of cultured rat sensory neurons.

The axonal transport of rabies virus (challenge virus strain of fixed virus) was studied in differentiated rat embryonic dorsal root ganglion cells. In addition, we observed the attachment of rabies virus to neuronal extensions and virus production by infected neurons. A compartmentalized cell culture system was used, allowing infection and manipulation of neuronal extensions without exposing the neural soma to the virus. The cultures consisted of 60% large neuronal cells whose extensions exhibited neurofilament structures. Rabies virus demonstrated high binding affinity to unmyelinated neurites, as suggested by assays of virus adsorption and immunofluorescence studies. The rate of axoplasmic transport of virus was 12 to 24 mm/day, including the time required for internalization of the virus into neurites. The virus transport could be blocked by cytochalasin B, vinblastine, and colchicine, none of which negatively affected the production of virus in cells once the infection was established. It was concluded that, for the retrograde transfer of rabies virus by neurites from the periphery to the neuronal soma, the integrity of tubulin- and actin-containing structures is essential. The rat sensory neurons were characterized as permissive, moderately susceptible, but low producers of rabies virus. These neurons were capable of harboring rabies virus for long periods of time and able to release virus into the culture medium without showing any morphological alterations. The involvement of sensory neurons in rabies virus pathogenesis, both in viral transport and as a site for persistent viral infection, is discussed.

Adhesiveness↗
PubMed1986

Electrophysiological and sleep alterations in experimental mouse rabies.

Changes in the spontaneous brain electrical activity and sleep organization were investigated in 5 mice strains during the evolution of experimental fixed rabies infection. Cortical electrodes were chronically implanted for continuous EEG recording and spectral analysis until death. Three evolutionary phases were individualized. The initial phase exhibited alterations of sleep stages, REM sleep disappearance, pseudoperiodic facial myoclonus and first clinical signs. The mature phase was characterized by a generalized EEG slowing (2-4 cycles/s). The terminal phase occurring with extinction of hippocampal rhythmic slow activity showed a flattening of cortical activity. The brain electrical activity ceased about 30 min before the cardiac arrest. Paroxysmal activities appeared during the course of the disease as bursts of rhythmic slow waves, pseudoperiodic spikes or occasionally ictal epileptic discharges. Spectral analysis revealed a progressive and characteristic clustering of the EEG frequency band power values. The spread of infection in the CNS was monitored by specific immunofluorescence studies which revealed the presence of rabies virus antigen in the pons, the cerebellum, the thalamus and the cortex during the initial phase. The pyramidal field of the hippocampus was infected during the mature phase but the gyrus dentatus was never infected even at the terminal phase. These studies show that particular neuronal functions are impaired during rabies virus infection suggesting that neuronal alterations may be involved in the pathogenic mechanisms leading to lethality.

Animals↗
PubMed1986

Infection of cultured rat myotubes and neurons from the spinal cord by rabies virus.

Rabies virus multiplication was investigated in cultured primary rat myotubes and neurons. The susceptibility of these two cell types to fixed rabies challenge virus strain (CVS) was monitored by fluorescence and virus titration. Differentiated rat myotubes were susceptible to rabies virus infection, and showed an increasing accumulation of viral material from day one to day four. However, these cells did not release infective viral particles, nor did they accumulate infectious virions in the cytoplasm. In contrast, infected neurons released large amounts of infectious particles. Electron microscopy observation of infected myotubes showed minor alterations and the presence of typical viral inclusions in the cytoplasm without mature virions assembling viral membranes. Competition binding experiments show that alpha-bungarotoxin inhibits rabies virus infection from 10(-5) to 10(-7) M, whereas lower toxin concentrations failed to have any effect. These data do not confirm the hypothesis of a fixed rabies virus amplification step at the site of the viral entry. On the other hand, the high susceptibility of peripheral neurons to rabies virus infection is an argument for the direct uptake of virions by these cells. The restrictive viral multiplication in the myotubes is an alternative explanation for the local persistence of rabies virus at the site of inoculation.

Animals↗
PubMed1986

Axonal transport of rabies virus in the central nervous system of the rat.

Stereotaxic inoculation of rabies virus into specific nuclei in the central nervous system has been used for the investigation of the central neural transport mechanisms of viral information. The infection was monitored by specific fluorescence and peroxidase studies and the titration of viral infectivity in dissected brain areas. Twenty-four hours after inoculation into the striatum, cortex, or substantia nigra, infected neurons were detected only in cells from areas and nuclei which were related to the site of inoculation. The distribution of infected neurons showed that retrograde axoplasmic flow plays a determining role in the transport of rabies virus 24 hours after delivery of virus to specific target nuclei. Local destruction of neurons by kainic acid at the site of viral inoculation did not prevent the uptake and subsequent retrograde axonal transport of virus. There was an overall correlation between the major neural connections of the inoculated areas (e.g. the striatum) and the infected areas 24 hours later (e.g. the substantia nigra).

Animals↗
PubMed1986

Involvement of gangliosides in rabies virus infection.

The role of gangliosides in rabies virus infection of chick embryo-related (CER) cells was investigated. Cultured cells were pretreated with neuraminidase to render the cells transiently non-susceptible to viral infection. Incubation of these desialylated cells with gangliosides allowed them to incorporate exogenous gangliosides and they recovered their susceptibility to rabies virus infection. Infection of CER cells was monitored by specific fluorescence 24 h after virus inoculation. The use of individual purified gangliosides or mixtures of two gangliosides to restore cellular susceptibility to viral infection showed that GT1b and GQ1b were the most effective. The disialogangliosides were also active, principally GD1b, whereas GM1, GM3 were poorly active and GD3 inactive. Incubation of rabies virus with gangliosides prior to virus infection reduced the percentage of infected cells. The results indicate that highly sialylated gangliosides are part of the cellular membrane receptor structure for the attachment of infective rabies virus. However, it is possible that other glycoconjugates such as glycoproteins or glycolipids also participate as components of a receptor structure for rabies virus.

Animals↗
PubMed1986

Neutralization of interferon produced early during rabies virus infection in mice.

Inoculation of mice (strain C3H/He) with a purified preparation of fixed rabies virus led to the production of interferon with two different peaks of activity detectable in the plasma: an early peak 24 h after inoculation followed by another peak on the 7th day after infection. The level of splenic 2-5A synthetase was enhanced in parallel with the pattern of interferon activity. Neutralization of the first peak of interferon activity by anti-mouse alpha/beta interferon globulin blocked the induction of splenic 2-5A synthetase and modified the development of disease. Infected mice given anti-interferon globulin had a significantly shorter morbidity period than normally infected mice. These results suggest that interferon produced early after virus inoculation plays a role in the onset of rabies disease.

Animals↗