Biomedical subjects

M Aguet

Publications and source records attributed to M Aguet.

At least 37 records · Page 2Linked to original sources

PubMed1995

IFN-gamma receptor-deficient mice are hypersensitive to the anti-CD3-induced cytokine release syndrome and thymocyte apoptosis. Protective role of endogenous nitric oxide.

Mice with a disruption of the IFN-gamma receptor alpha-chain gene (IFN-gamma R alpha o/o mice) were found to be significantly more sensitive than their wild-type counterparts to induction of the anti-CD3-induced disease syndrome. Specifically, when given a selected dose of anti-CD3 Ab, IFN-gamma R alpha o/o mice developed severe hypothermia and hypoglycemia, leading to 100% mortality within 72 h. In contrast, wild-type mice failed to develop overt pathologic manifestations and survived. Histologic examination revealed apoptosis in thymuses and spleens, which were significantly more pronounced in the mutant than in the wild-type mice, as confirmed by flow cytometric and DNA electrophoretic analysis. Apoptosis affected mainly CD4+CD8+ and CD4+CD8- thymocytes. Other histologic alterations were steatosis in livers, and erythrocyte extravasation and infiltration of apoptotic cells in lungs, all of which were exclusively observed in IFN-gamma R alpha o/o mice. Blood levels of TNF, IL-2, IL-6, and IL-10 were slightly more elevated in IFN-gamma R alpha o/o mice, but insufficiently so to explain increased disease severity. Thus, even more elevated cytokine levels in wild-type mice receiving high doses of anti-CD3 Ab were not associated with morbidity or apoptosis. Blood levels of IFN-gamma were barely detectable in anti-CD3-challenged wild-type mice, but were relatively high in the mutant mice. Increased susceptibility of IFN-gamma R alpha o/o mice was associated with impaired nitric oxide (NO) production, as indicated by significantly lower plasma nitrite levels and by more transient expression of spleen inducible NO synthase mRNA. Moreover, treatment of wild-type mice with the NO synthase inhibitor N-nitro-L-arginine methylester resulted in increased anti-CD3-induced morbidity and mortality. The data indicate that IFN-gamma R alpha o/o mice produce less NO and are therefore more sensitive than wild-type mice to the deleterious effect of anti-CD3 Ab.

Animals↗

PubMed1995

Role of interferon-gamma in contact hypersensitivity assessed in interferon-gamma receptor-deficient mice.

The role of interferon-gamma (IFN gamma) in contact hypersensitivity induced by the haptens, oxazolone and 2,4,6-trinitrochlorobenzene (TNCB), was investigated in mice with a targeted disruption of the IFN gamma receptor (IFN gamma-R-/-). The 24-h ear-swelling response to oxazolone or TNCB in sensitized animals was not significantly reduced by the disruption of IFN gamma signalling. Dermal mononuclear infiltrates (MN) and epidermal microabscesses, however, were clearly diminished in the mutant mice. The hapten-induced upregulation of intercellular-adhesion molecule-1 (ICAM-1) and major histocompatibility complex (MHC) class I in IFN gamma-R-/- mice was smaller when compared to wild-type mice. It is concluded that oxazolone- and TNCB-induced contact hypersensitivity is partially dependent on a functional IFN gamma system. While the cutaneous oedema is IFN gamma-independent, the mononuclear cell infiltration and epidermal microabscess formation are at least partly IFN gamma-dependent. Therefore, reduced cellular infiltrates are likely due to a reduced upregulation of ICAM-1 and class I antigen expression in IFN gamma-R-/- mice.

Adjuvants, Immunologic↗

PubMed1995

Inducible gene targeting in mice.

A method of gene targeting that allows the inducible inactivation of a target gene in mice is presented. The method uses an interferon-responsive promoter to control the expression of Cre recombinase. Here, Cre was used to delete a segment of the DNA polymerase beta gene flanked by IoxP recombinase recognition sites. Deletion was complete in liver and nearly complete in lymphocytes within a few days, whereas partial deletion was obtained in other tissues. This method can be used for the inducible inactivation of any other gene in vivo.

Animals↗

PubMed1995

Aggressive behavior and altered amounts of brain serotonin and norepinephrine in mice lacking MAOA.

Deficiency in monoamine oxidase A (MAOA), an enzyme that degrades serotonin and norepinephrine, has recently been shown to be associated with aggressive behavior in men of a Dutch family. A line of transgenic mice was isolated in which transgene integration caused a deletion in the gene encoding MAOA, providing an animal model of MAOA deficiency. In pup brains, serotonin concentrations were increased up to ninefold, and serotonin-like immunoreactivity was present in catecholaminergic neurons. In pup and adult brains, norepinephrine concentrations were increased up to twofold, and cytoarchitectural changes were observed in the somatosensory cortex. Pup behavioral alterations, including trembling, difficulty in righting, and fearfulness were reversed by the serotonin synthesis inhibitor parachlorophenylalanine. Adults manifested a distinct behavioral syndrome, including enhanced aggression in males.

Aggression↗

PubMed1995

Theiler's virus infection of 129Sv mice that lack the interferon alpha/beta or interferon gamma receptors.

The Daniels strain of Theiler's virus causes a persistent infection of the white matter of spinal cord of susceptible mice, with chronic inflammation and primary demyelination. Inbred 129Sv mice are resistant to this infection; they present with mild encephalomyelitis and clear the infection within a matter of days. A very different outcome was observed with inbred 129Sv mice whose receptors for interferon alpha/beta or interferon gamma had been inactivated by homologous recombination. The former presented severe encephalomyelitis with acute infection of neurons, particularly in brain and hippocampus, and extensive infection with necrosis of the choroid plexus. Most animals died of this acute disease. The latter, presented the same early encephalomyelitis as the control 129Sv mice. However, they remained persistently infected and developed a very severe late infection of the white matter with extensive primary demyelination. This late disease looked like an exacerbated form of the chronic demyelinating disease observed in susceptible inbred mice such as the SJL/J or FVB strains. Our results show that the two interferon systems play nonredundant roles in the resistance of the 129Sv mouse to the infection by Theiler's virus. They also lend support to the notion that the Ifg gene is involved in the resistance/susceptibility of inbred strains of mice to persistent infection by this picornavirus.

Animals↗

PubMed1995

Development of antimalaria immunity in mice lacking IFN-gamma receptor.

IFN-gamma receptor deficient (IFN-gamma R-/-) mice, immunized with different developmental stages of malaria parasites, were used to define the mechanisms of protection against the various stages of this infection. IFN-gamma R-/- mice failed to develop protective immunity against Plasmodium yoelii sporozoites or liver stages, upon immunization with a single dose of irradiated sporozoites, whereas in immunized wild-type mice, parasite development was strongly inhibited. Immunized wild-type mice expressed high levels of inducible nitric oxide synthase (iNOS) mRNA in their liver, upon challenge with viable sporozoites, whereas only background levels of iNOS were detected in immunized IFN-gamma R-/- mice. In contrast, after immunization with multiple doses of irradiated sporozoites, both IFN-gamma R-/- and wild-type mice mounted an immune response, which strongly inhibited the development of liver stage parasites. In both types of mice, protection occurred in the absence of appreciable expression of liver iNOS mRNA. As for the course of the erythrocytic phase of infection by nonlethal malaria species, P. yoelii yoelii and P. chabaudi adami, we observed only a moderately prolonged parasitemia in IFN-gamma R-/- mice compared with wild-type mice, indicating that IFN-gamma may only play a modest role in immunity against erythrocytic stages. These results indicate that IFN-gamma is the main mediator of the protective mechanism that develops first upon immunization with sporozoites. However, the nature of the anti-parasite mechanism(s) changes in the course of immunization, so that multiple immunizing doses elicit additional protective mechanisms, which are independent of IFN-gamma and its receptor.

Amino Acid Oxidoreductases↗

PubMed1995

Early interleukin 12 production by macrophages in response to mycobacterial infection depends on interferon gamma and tumor necrosis factor alpha.

Interleukin 12 (IL-12) produced by macrophages immediately after infection is considered essential for activation of a protective immune response against intracellular pathogens. In the murine Mycobacterium bovis Bacillus Calmette-Guérin (BCG) model we assessed whether early IL-12 production by macrophages depends on other cytokines. In vitro, murine bone marrow-derived macrophages produced IL-12 after infection with viable M. bovis BCG or stimulation with LPS, however, priming with recombinant interferon gamma (rIFN-gamma) was necessary. In addition, IL-12 production by these macrophages was blocked by specific anti-tumor necrosis factor alpha (TNF-alpha) antiserum. Macrophages from gene deletion mutant mice lacking either the IFN-gamma receptor or the TNF receptor 1 (p55) failed to produce IL-12 in vitro after stimulation with rIFN-gamma and mycobacterial infection. In vivo, IL-12 production was induced in spleens of immunocompetent mice early during M. bovis BCG infection but not in those of mutant mice lacking the receptors for IFN-gamma or TNF. Our results show that IL-12 production by macrophages in response to mycobacterial infection depends on IFN-gamma and TNF. Hence, IL-12 is not the first cytokine produced in mycobacterial infections.

Animals↗

PubMed1995

The stimulatory effects of interleukin (IL)-12 on hematopoiesis are antagonized by IL-12-induced interferon gamma in vivo.

Interleukin (IL)-12 synergizes with other cytokines to stimulate the proliferation and differentiation of early hematopoietic progenitors in vitro. However, in vivo administration of IL-12 decreases peripheral blood counts and bone marrow hematopoiesis. Here, we used interferon (IFN) gamma receptor-deficient (IFN gamma R-/-) mice to investigate whether the in vivo inhibition of hematopoiesis by IL-12 is indirectly mediated by IL-12-induced IFN-gamma. IL-12 administered for 4 d (1 microgram/mouse per day) resulted in lower peripheral blood counts and a 2-fold decrease in bone marrow cellularity in wild-type mice, but not in IFN gamma R-/- mice. Bone marrow hematopoietic progenitors were decreased after IL-12 treatment in wild-type mice, but rather increased in IFN gamma R-/- mice. Splenic cellularity was 2.3-fold higher after IL-12 administration in wild-type mice, largely due to natural killer (NK) cell and macrophage infiltration together with some extramedullary hematopoiesis. In IFN gamma R-/- mice, spleen cellularity was less increased, there were fewer infiltrating NK cells, but a strong extramedullary hematopoiesis. Thus, alterations mediated by IL-12-induced IFN-gamma include reduction in bone marrow cellularity and hematopoietic progenitors, as well as pronounced splenomegaly, largely caused by NK cell infiltration. In the absence of IFN-gamma signaling, IL-12 promotes hematopoiesis, consistent with its in vitro activities.

Animals↗

PubMed1995

Involvement of IFN-gamma in Bacillus Calmette-Guérin-induced but not in tumor-induced sensitization to TNF-induced lethality.

In healthy mice, murine (m) TNF is fairly lethal, whereas human (h) TNF (a selective murine TNF-R55 agonist) is rather harmless. However, we and others observed that mice suffering from a bacterial infection, such as Bacillus Calmette-Guérin (BCG), or bearing i.m. some types of tumor, develop a hypersensitivity to the IL-6-inducing and lethal properties of hTNF. This is a cardinal problem as it severely limits the potential use of hTNF-R55-specific agonists for systemic treatment of human cancer. Using mice carrying a targeted disruption in the gene encoding the IFN-gamma receptor (IFN-gamma Ro/o), we here report that endogenous IFN-gamma plays a crucial role in the development of TNF hypersensitivity during BCG infection. Indeed, both the lethality and the IL-6 induced by hTNF were drastically reduced in IFN-gamma Ro/o mice as compared with control mice. These results demonstrate that the enhancement of TNF effects is at least an equally important mechanism by which IFN-gamma contributes to BCG-induced hypersensitivity as the previously described augmentation of TNF production. Experiments in athymic nude mice, either depleted of NK cells or not, revealed that the latter cell population is an important source of the sensitizing IFN-gamma during BCG infection. In contrast, IFN-gamma Ro/o mice were as susceptible as control mice to the sensitizing effects of tumors. mTNF, which interacts with both mTNF-R55 and mTNF-R75 and causes lethality on its own, is as toxic in IFN-gamma Ro/o mice as in wt control mice; this means that TNF-induced IFN-gamma does not play a role in mTNF-induced lethality.

Animals↗

PubMed1995

Mice from a genetically resistant background lacking the interferon gamma receptor are susceptible to infection with Leishmania major but mount a polarized T helper cell 1-type CD4+ T cell response.

Mice with homologous disruption of the gene coding for the ligand-binding chain of the interferon (IFN) gamma receptor and derived from a strain genetically resistant to infection with Leishmania major have been used to study further the role of this cytokine in the differentiation of functional CD4+ T cell subsets in vivo and resistance to infection. Wild-type 129/Sv/Ev mice are resistant to infection with this parasite, developing only small lesions, which resolve spontaneously within 6 wk. In contrast, mice lacking the IFN-gamma receptor develop large, progressing lesions. After infection, lymph nodes (LN) and spleens from both wild-type and knockout mice showed an expansion of CD4+ cells producing IFN-gamma as revealed by measuring IFN-gamma in supernatants of specifically stimulated CD4+ T cells, by enumerating IFN-gamma-producing T cells, and by Northern blot analysis of IFN-gamma transcripts. No biologically active interleukin (IL) 4 was detected in supernatants of in vitro-stimulated LN or spleen cells from infected wild-type or deficient mice. Reverse transcription polymerase chain reaction analysis with primers specific for IL-4 showed similar IL-4 message levels in LN from both types of mice. The IL-4 message levels observed were comparable to those found in similarly infected C57BL/6 mice and significantly lower than the levels found in BALB/c mice. Anti-IFN-gamma treatment of both types of mice failed to alter the pattern of cytokines produced after infection. These data show that even in the absence of IFN-gamma receptors, T helper cell (Th) 1-type responses still develop in genetically resistant mice with no evidence for the expansion of Th2 cells.

Animals↗

PubMed1995

Dissection of the extracellular human interferon gamma receptor alpha-chain into two immunoglobulin-like domains. Production in an Escherichia coli thioredoxin gene fusion expression system and recognition by neutralizing antibodies.

The extracellular interferon gamma receptor alpha-chain (IFN gamma R) is believed to comprise two discrete approximately 110 amino acid immunoglobulin-like domains, perhaps similar to those seen in the crystal structure of the extracellular human growth hormone receptor [De Vos, A. M., Ultsch, M., & Kossiakoff, A. (1992) Science 255, 306-312], a distant relative in the cytokine receptor superfamily. In accord with this idea, we show that these IFN gamma R immunoglobulin-like domains can be produced separately in a soluble form with a native-like fold. The N-terminal domain (residues 1-108), with a Cys105 to Ser105 mutation, was produced at a high level, in a soluble form, as a thioredoxin-interferon gamma receptor fragment fusion protein in the cytoplasm of Escherichia coli. Upon extraction, the receptor Cys60-Cys68 disulfide bond formed spontaneously, to generate a native-like structure directly without the need for refolding. Cleavage of the fusion protein by enterokinase released the receptor fragment (approximately 12 kDa), which was recognized by several neutralizing antibodies with affinities, measured using surface plasmon resonance technology, that were essentially indistinguishable from those seen with the full length extracellular IFN gamma R produced in eukaryotic cells. Circular dichroism and 1D 1H nuclear magnetic resonance spectra indicated that the receptor fragment adopts a folded state, with mainly beta-sheet and reverse turn secondary structure. The second membrane-proximal Ig-like domain of the IFN gamma R (residues 90-229) was produced, albeit less efficiently, and characterized in a similar way. The production of these two independently folded proteins provides experimental support for the two domain organization of the IFN gamma R and opens new avenues for structural studies on these Ig-like molecules by NMR and crystallographic methods.

Amino Acid Sequence↗

PubMed1995

Variable region cDNA sequences and characterization of murine anti-human interferon gamma receptor monoclonal antibodies that inhibit receptor binding by interferon gamma.

Murine monoclonal antibodies (mAbs) are described that recognize the extracellular human interferon gamma receptor alpha-chain (IFN gamma R) and inhibit the binding to it of interferon gamma. The inhibitory activities (IC50s) of these mAbs, quantified by radioimmunoassay using native receptor on human Raji cells, lie in the range 0.5-24 nM, whereas their relative affinities for the immobilised recombinant extracellular receptor, determined using surface plasmon resonance technology, are in the range 0.6-40.9 nM. Nine mAbs derived from one immunization, were shown by variable region cDNA sequencing to be clonally related, with mAb A6 from this group showing the highest affinity for the receptor. Another two mAbs, gamma R38 and gamma R99, derived from a separate immunization, are clonally unrelated to each other and to those in the A6 family. From the V-region sequences, the L-chains of mAbs A6, gamma R38 and gamma R99 were shown to belong to the V kappa 34C, V kappa 34C and V kappa 1 families, whereas the H-chains belong to the 3069, J606 and J558 families, respectively. The mAbs A6 and gamma R38 recognize overlapping epitopes on the N-terminal Ig-like domain of the IFN gamma R, whereas the gamma R99 epitope is located largely in the membrane proximal Ig-like domain. Sequence comparisons with Ig structures solved by X-ray diffraction allowed deductions concerning likely CDR canonical conformations. These studies provide essential information for crystallographic and mutagenesis experiments aimed at understanding the molecular basis of the interactions of these mAbs with the extracellular IFN gamma R.

Amino Acid Sequence↗

PubMed1995

MHC antigens in interferon gamma (IFN gamma) receptor deficient mice: IFN gamma-dependent up-regulation of MHC class II in renal tubules.

MHC class II gene products in parenchymal cells, such as tubular epithelial cells in kidney, may play a role in the regulation of autoimmune reactions. Expression of MHC class II in renal tubular cells is normally very low, but it increases considerably under various pathologic conditions. The predominant role of IFN gamma in up-regulation of MHC class II expression has been demonstrated repeatedly. We tested the existence of alternative pathways of MHC class II regulation using IFN gamma receptor-deficient (IFN gamma R-/-) mice. Mutant and wild type mice received 50 micrograms bacterial endotoxin (LPS) i.p. Four days later the kidneys were removed for immunofluorescence examination. In agreement with published results LPS provoked an increase of immunoreactivity for MHC class I and MHC class II in proximal tubules of wild type mice. While MHC class I up-regulation was strictly IFN gamma receptor-dependent, up-regulation of MHC II was still evident in mutant mice, although less than in wild type mice. Since injection of IFN gamma induced proximal tubular MHC class II expression in wild type mice but not in IFN gamma R-/- mice, an alternative signaling pathway for IFN gamma does not seem to exist. Thus, up-regulation of MHC class II expression in renal tubules does not necessarily require IFN gamma. The markedly patchy pattern of immunofluorescence in IFN gamma R-/- mice suggests that induction of MHC class II after LPS injection may represent renal injury due to shock.

Animals↗

PubMed1995

CD40 ligand has potent antiviral activity.

For B cells to make antibodies against most antigens, they require help from T cells. T cell help is delivered as two signals to the B cell, one of which is via CD40 and the other can be through receptors for any of a variety of soluble cytokines. We have constructed recombinant vaccinia viruses that express the ligand for CD40 and have shown that the growth of these viruses is dramatically controlled in vivo, even in mice that lack T or B cells. In this paper, we also describe our attempts to analyse the CD40 ligand-mediated antiviral activity by studying the clearance of these viruses in mice that are deficient in important antiviral mechanisms. Thus, the antiviral activity of CD40L may represent a surprising and potent effector mechanism of T cells activated during a virus infection.

Animals↗

PubMed1995

Antiviral activity of autocrine interferon-beta requires the presence of a functional interferon type I receptor.

We and others have previously observed that the antiviral effects of autocrine interferon (IFN)-alpha/beta activity cannot be abolished by neutralizing antibodies, even when present to a large excess. This raises the possibility that the major part of autocrine activity is triggered intracellularly, possibly bypassing the transmembrane IFN-alpha/beta receptor. To examine this possibility, cells derived from IFN-alpha/beta Ro/o knockout mice lacking a functional IFN-alpha/beta receptor were stably transformed with pHMB-KbMuIFN beta or pMFG-MuIFN beta plasmids encoding a constitutively expressed murine IFN-beta gene. Four different clones were isolated and examined for resistance to a retrovirus, MFG-LacZ, and to Semliki Forest virus. Despite the production of autocrine IFN-beta at levels inducing high antiviral resistance in control cells, none of the clones displayed antiviral resistance. Thus, despite its failure to be neutralized by potent antiserum, the antiviral activity of autocrine IFN-beta takes place via the transmembrane IFN-alpha/beta receptor, and no additional pathway is involved.

3T3 Cells↗

PubMed1995

Immune defence in mice lacking type I and/or type II interferon receptors.

Mice lacking the receptor for type I interferon (IFN-alpha beta, A129 mice), for type II interferon (IFN-gamma, G129 mice) or for both receptors (AG129 mice) have been generated by embryonic stem cell mediated gene targeting and inter-crossing A129 x G129, respectively. The role of the two IFN systems in controlling a range of infections has been studied using these mice. Type I IFN is shown to be responsible for the immune defence against most viral infections tested (Lymphocytic Choriomeningitis Virus, Semliki Forest Virus, Theiler's Virus, Vesicular Stomatitis Virus), type II IFN seems to be of little importance. In Vaccinia Virus and Theiler's Virus infection, however, both IFN systems were found to play a nonredundant role. IFN-gamma was critical for the defence against intracellular bacteria (Mycobacterium, Listeria) and parasites (Leishmania), whereas IFN-alpha beta was not. IFN-alpha beta is produced by virus-infected cells within hours and plays an important role in preventing virus spread early. Production of IFN-gamma on the other hand needs activation of the immune system and plays a major role later, i.e. mostly during the immune response. Data obtained with the mice described here show that both IFN systems seem to have evolved to complement each other in the host defence against a wide variety of infectious agents.

Animals↗

PubMed1995

Expansion of gamma delta+ T cells in BALB/c mice infected with Leishmania major is dependent upon Th2-type CD4+ T cells.

T cells belong to either the alpha beta+ or gamma delta+ lineage as defined by their antigen receptor. Although both T-cell subsets have been shown to be involved in the immune response to the parasite Leishmania major, very little is known about possible interactions between these two populations. In this study, using a mouse model of infection with L. major, we showed that expansion of a subset of gamma delta+ T cells in vivo is dependent upon the presence of alpha beta+ CD4+ T cells. Moreover, this effect appears to be mediated via the secretion of lymphokines by CD4+ cells with a T-helper 2 (Th2) functional phenotype. Results showing that activation of Th2-type cells in mice treated with anti-immunoglobulin D antibodies or infected with Nippostrongylus brasiliensis also results in gamma delta+ T-cell expansion suggest that this effect of the Th2-type CD4+ cells is a general phenomenon not restricted to infection with L. major.

Animals↗

PubMed1995

The in vivo antiviral activity of interleukin-12 is mediated by gamma interferon.

The injection of 20 ng of mouse interleukin-12 (IL-12) protects mice from a lethal infection with encephalomyocarditis virus. In vitro, an anti-gamma interferon (anti-IFN-gamma) monoclonal antibody but not an anti-IL-12 monoclonal antibody neutralizes the antiviral activity present in the supernatants of splenocytes stimulated with IL-12. Finally, IL-12 fails to protect 129 Sv/Ev IFN-gamma R0/0 mice against encephalomyocarditis virus infection. These results demonstrate that IL-12 exerts its antiviral activity through the induction of endogenous IFN-gamma.

Animals↗