[Antiviral activities of interferon and PML pathway]. / Les mécanismes de l'action antivirale des interférons : la voie PML.

Discovered in 1957 for their antiviral properties, interferons (IFNs) are a growing cytokine family with diverse biological activities including antitumor and immunoregulatory activities. IFN are classified in three types I, II and III. They bind to different specific cell receptors and induce via the Jak/Stat pathway the expression of more than 300 genes, the products of which are believed to mediate their biological effects. Several proteins have been implicated in resistance to viral infection in IFN-treated cells, i.e. the dsRNAdependent protein kinase PKR, the 2'5' oligoadenylate synthetase/RNaseL and Mx proteins. However, it was demonstrated that cells from triple knockout mice lacking PKR, RNase L and Mx are still sensitive to the IFN-induced antiviral state, indicating that other pathways exist. One of these pathways implicates promyelocytic leukemia (PML) protein. This article reviews the potential antiviral activities of the different IFN-induced mediators focusing onPMLpathway and how viruses from different families overcome this defence.

Evidence against a direct cytotoxic effect of alpha interferon and zidovudine in HTLV-I associated adult T cell leukemia/lymphoma.

The combination of the anti-viral agents, zidovudine (AZT) and interferon-alpha (IFN), is a potent treatment of HTLV-I-associated adult T cell leukemia/lymphoma (ATL). In this study we investigate the possible mechanism of action of this combination by examining several cellular parameters including cell proliferation, cell cycle distribution and apoptosis. The ATL-derived T cell lines HuT-102 and MT-2 served as models. HTLV-I negative T cell lines (CEM and Jurkat) were used as controls. No significant modification of cell growth was observed except at suprapharmacological doses of AZT and IFN. Moreover, these effects were less pronounced in HTLV-I-infected cell lines compared to control cell lines. AZT and IFN treatment did not induce any significant modification of the expression of bcl-2 and p53. Interestingly no in vitro cytotoxic effect of AZT/IFN combination was observed on fresh leukemic cells derived from an acute ATL patient at diagnosis despite achievement of in vivo complete remission using the same therapy. These results suggest that the therapeutic effect of AZT and IFN is not through a direct cytotoxic effect of these drugs on the leukemic cells.

Arsenic trioxide and interferon-alpha synergize to induce cell cycle arrest and apoptosis in human T-cell lymphotropic virus type I-transformed cells.

Human T-cell lymphotropic virus type I (HTLV-I) is the causative agent of adult T-cell leukemia/lymphoma (ATL). ATL is an aggressive proliferation of mature activated T cells associated with a poor prognosis. The combination of the antiviral agents, zidovudine (AZT) and interferon (IFN), is a potent treatment of ATL. Recently, arsenic trioxide (As) was shown to be an effective treatment of acute promyelocytic leukemia (APL). We have tested the effects of the combination of As and IFN on cell proliferation, cell cycle phases distribution, and apoptosis in ATL-derived or control T-cell lines. A high synergistic effect between IFN and As was observed in ATL-derived cell lines in comparison to the control cell lines, with a dramatic inhibition of cell proliferation, G1 arrest, and induction of apoptosis. Similar results were obtained with fresh leukemia cells derived from an ATL patient. Although the mechanisms involved are unclear, these results could provide a rational basis for combined As and IFN treatments in ATL.

Effects of serotonin and melanin on in vitro HIV-1 infection.

In this article, we describe the effect of indoleamines: serotonin (5-HT) and synthetic soluble melanin, on the multiplication of HIV-1 in T4 lymphocytic cell lines. The results show that viral production is increased when infected CEM-11 cells are incubated with 5-HT (10(-7) M and 10(-8) M) for 72 hours, whereas at higher doses (10(-3) M and 10(-4) M), there is an inhibition of viral multiplication. As well, when infected CEM cells were cultured in the presence of 5-HT at 10(-4) M, during 15 days, virus production, syncytia formation and cytolytic effect were drastically inhibited. Melanin also inhibits HIV-1 cytopathic effect on MT-2 cells, without cell toxicity, at concentrations of 0.2-10 micrograms/ml. Syncytium formation and cell lysis were also blocked by melanin at concentrations of 0.1 to 10 micrograms/ml, when uninfected MT-2 cells were mixed with HIV-1 chronically infected CEM-11 cells.

Cell abnormalities associated with human papillomavirus-induced squamous intraepithelial cervical lesions. Multivariate data analysis.

One hundred ninety-six cervical scrapings were obtained for simultaneous research of cell abnormalities in Papanicolaou smears and detection of genital human papillomavirus (HPV) genotypes by polymerase chain reaction in extracted DNA from each clinical sample. The samples described by six discriminant cytologic parameters, and a synthetic HPV-presence/absence parameter provided an efficient matrix for multiple correspondence analysis. This statistical analysis displayed a plurality of HPV-related cell abnormalities in squamous intraepithelial lesions, and a high correspondence between HPV infection and the presence of multinucleated squamous cells, morphologically transformed keratinocytes (dyskaryotic cells), koilocytes, and cellular changes related to epithelial maturation. Because of the low proportion of detectable koilocytes in the HPV-infected scrapings, the authors also inferred that cellular changes related to epithelial maturation and morphologically transformed keratinocytes are the most accurate and efficient cell abnormalities, in terms of frequency and correlation, for cytodiagnosis of HPV-induced or associated squamous intraepithelial cervical lesions.

Detection of IAP related transcripts in normal and transformed rat cells.

Intracisternal A-particles (IAP) genes in variable copy number exist in all rodent species studied. Expression is highly repressed in murine normal cells except in embryonic and transformed cells. We searched for IAP related sequences expression in another rodent species cells. Using the more conserved sequence of IAP gene between mouse, Syrian hamster, and rat species (0.4 kb HindIII-PstI fragment from PMIA14), we have been able to detect IAP related transcripts in rat cells. We found that, i) IAP related transcripts are poorly expressed in normal cells, since among 10 tissues tested, only the placenta shows IAP RNA. ii) IAPs are highly expressed in all the transformed cells tested. iii) the detected transcripts appear to have similar sizes in rat cells as in mouse cells (7.2 kb; 5.4 kb). None of the probes corresponding to other regions of the IAP gene nor the entire sequence of PMIA14 allowed us to detect such transcripts.

In vitro induction of early mouse embryo intracisternal particles (epsilon particles) in cultured cell lines.

The experimental induction of epsilon particles, retrovirus-like structures corresponding to the small IA particles of the mouse, was studied by electron microscopy in rodent-cultured cell lines. Among the chemicals tested, only IdUr was shown to be an effective inducer, but not cycloheximide, puromycin , deoxy-fluorouracil or 5-azacytidine. However, only two mouse-derived cell lines: Ki-BALB and FG 10, among 27 cell lines of mouse, rat and mink origins tested, expressed epsilon particles upon IdUr treatment. Epsilon particles thus respond to chemical inducers very differently in comparison with large IAP. Moreover, the addition of interferon previously shown to attenuate IAP production, had no effect on that of epsilon particles.

Electron microscopic characterisation of retrovirus and retrovirus-like particles induced by demethylating agents (5-azacytidine and 5-azadeoxycytidine) in Syrian hamster (Mesocricetus auratus) cells.

In a framework of a study on retrovirus expression in Syrian hamster (mesocricetus auratus), three cell lines were examined by electron microscopy. As we had previously demonstrated the efficacy of demethylating drug in activating the formation of retroviral particles, we used 5-azacytidine to activate endogenous retrovirus expression. Proceeding to a more detailed survey we were able to observe various types of retrovirions. All of them were present in the different activated cell lines. They were: intracisternal R-type particles (IRP), intracisternal A-type particles (IAP), intracytoplasmic A-type particle (ICAP), intramitochondrial A-type particle (IMAP) and M-432 virus-like particles. Some of these particles have never been described before in Syrian hamster cells. The significance of their presence is discussed here.

Effects of in vitro infection of mouse glial and neuroblastoma cells with the scrapie agent.

Mouse glial and neuroblastoma cells were infected with the mouse adapted strains C506 and 139A of scrapie agent. Lysates of the in vitro infected cells (from the 3rd to the 16th passage) intracerebrally inoculated into CD-1 mice, caused the development of a neurological disease, with characteristic signs of scrapie. Morphological changes in scrapie-infected neural cells were observed after about fifteen in vitro passages. In liquid medium, the cloning efficiency of these cells increased. They acquired the capacity to form large tridimensional colonies in agar. Heating the infectious brain extracts at 60 and 80 degrees C for 30 minutes did not inhibit these changes thus showing the involvement of the thermoresistant scrapie agent. Supernatants of scrapie-infected glial cells promoted colony formation in liquid medium with different types of normal cells. Analysis of supernatants of scrapie-infected mouse neuroblastoma cells showed a profound modification of neurotransmitter metabolism.

Regulation of intracisternal A particles in mouse teratocarcinoma cells: involvement of DNA methylation in transcriptional control.

The methylation state of Intracisternal A Particle (IAP) genes in mouse teratocarcinoma cell lines has been investigated as an approach to study the regulation of the expression of these particles. Treatment of the cells with the methylation inhibitor 5-azacytidine induces the production of the particles in all the cells; the induction is particularly striking in an embryonal carcinoma cell line which is normally devoid of IAPs. The induction is accompanied by decrease in DNA methylation as demonstrated by using the methylation sensitive isoschizomer enzymes MspI and HpaII. Hypomethylation of the IAP genes correlates with accumulation of IAP, specific polyadenylated RNA reinforcing the hypothesis that methylation plays an important role in the control of IAP expression.

Intracisternal A particles: RNA expression and DNA methylation in murine teratocarcinoma cell lines.

Two main intracisternal-A-particle-specific RNA species (29 to 30S and 35 to 36S) are variably expressed in particle-producing and particle-nonproducing murine teratocarcinoma cell lines. An analysis of DNA methylation patterns after hybridization with an intracisternal-A-particle-specific probe showed an apparent direct correlation between DNA methylation and RNA expression. However, when the methylation assay was performed on the excised intracisternal A particle genes in one of the particle-rich cell lines (PCC6), some undermethylated sequences were detectable. These results are consistent with the concept that only few of the genes are transcriptionally active.

In vitro propagation of the scrapie agent. I. Transformation of mouse glia and neuroblastoma cells after infection with the mouse-adapted scrapie strain c-506.

Seven cell lines including glia cells from mouse brains and mouse neuroblastoma cells were infected with the mouse-adapted scrapie strain c-506. During the early in vitro passages, a stimulation of growth was already observed but cellular morphology and differentiation did not alter. Later on, after 12-16 passages, six of the seven infected lines displayed cell proliferation and morphological alterations, suggesting an in vitro morphological transformation. At this stage, differentiation was no longer observed in the scrapie-infected neuroblastoma cells and all the scrapie-infected cells formed two to four times more colonies in liquid medium than the controls, and developed large tridimensional colonies in agar. The part played by the scrapie agent in these changes is discussed.

[Effect of 5-azacytidine on the expression of intracisternal A and epsilon particles in various mouse cell lines]. / Action de la 5-azacytidine sur l'expression des particules intracisternales A et epsilon dans différentes lignées cellulaires de Souris.

5-Azacytidine treatment of several Mouse cell lines cultured in vitro induces the synthesis of a high number of intracisternal A particles. This induction could be related to the demethylation effect of the 5-Azacytidine. On the contrary, the epsilon intracisternal particles are not activated by 5-Azacytidine.

[In vitro modification of the morphology and the growth of cells infected with scrapie (author's transl)]. / Modifications in vitro de la morphologie et de la croissance de cellules infectées par l'agent scrapie.

Seven cell lines originated either in brains or in neuroblastomas of Mice, were infected with Scrapie. After 12 to 16 in vitro passages, 6 lines out of 7 showed changes of their morphology, and of their growth, resembling those occurring in the course of a malignant transformation. The Scrapie infected cells acquired the capacity to form 2 to 4 times more colonies in liquid medium than the controls, and to develop large tridimensional colonies in semisolid medium. The role of Scrapie in these changes is discussed.

[Replication of an isolate of human rotavirus on human cells cultured in vitro: observation by electron microscopy]. / Replication d'un isolat de rotavirus humain sur des cellules humaines cultivées in vitro: observation par microscopie électronique.

Replication of human rotaviruses on cultured human cells obtained from an intestinal carcinoid tumor was observed by electron microscopy. No enzymatic pretreatment of either the cell monolayers or the inoculum was needed. The characteristics of the vital progeny are discussed.

[Presence of enteric viruses in non-diarrheic canine stools]. / Présence des virus entériques dans des déjections canines non diarrhéiques.

Rota-, corona- and parvovirus particles have been visualized by direct electron microscopy in canine stools collected at random in Paris streets. A possible involvement of these viruses in gastroenteric diseases is discussed in the light of these findings.

Electron microscopy detection and characterization of viral particles in dog stools.

A number of apparently normal dog stool samples, randomly collected on the sidewalks of Paris were examined by Electron Microscopy. The study revealed the presence of viral particles in 27 cases. Morphological criteria lead to the characterization of rotavirus in 2 specimen, coronavirus in 7 and parvovirus in 5. Rotavirus particles appeared always alone while coronavirus and parvovirus particles were present together in 5 cases. Similar particles have been implicated in animal and human gastroenteritis. The importance of their presence in canine dejections is discussed in view of pollution by dog stools of urban and suburban areas.