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Biomedical subjects

M B Oldstone

Publications and source records attributed to M B Oldstone.

At least 109 records · Page 6Linked to original sources
PubMed1995

Virus-induced immunosuppression: immune system-mediated destruction of virus-infected dendritic cells results in generalized immune suppression.

Despite the clinical importance of virus-induced immunosuppression, how virus infection may lead to a generalized suppression of the host immune response is poorly understood. To elucidate the principles involved, we analyzed the mechanism by which a lymphocytic choriomeningitis virus (LCMV) variant produces a generalized immune suppression in its natural host, the mouse. Whereas adult mice inoculated intravenously with LCMV Armstrong rapidly clear the infection and remain immunocompetent, inoculation with the Armstrong-derived LCMV variant clone 13, which differs from its parent virus at only two amino acid positions, by contrast results in persistent infection and a generalized deficit in responsiveness to subsequent immune challenge. Here we show that the immune suppression induced by LCMV clone 13 is associated with a CD8-dependent loss of interdigitating dendritic cells from periarteriolar lymphoid sheaths in the spleen and, functionally, with a deficit in the ability of splenocytes from infected mice to stimulate the proliferation of naive T cells in a primary mixed lymphocyte reaction. Dendritic cells are not depleted in immunocompetent Armstrong-infected mice. LCMV Armstrong and clone 13 exhibit differences in their tropism within the spleen, with clone 13 causing a higher level of infection of antigen-presenting cells in the white pulp, including periarterial interdigitating dendritic cells, than Armstrong, thereby rendering these cells targets for destruction by the antiviral CD8+ cytotoxic T-lymphocyte response which is induced at early times following infection with either virus. Our findings illustrate the key role that virus tropism may play in determining pathogenicity and, further, document a mechanism for virus-induced immunosuppression which may contribute to the clinically important immune suppression associated with many virus infections, including human immunodeficiency virus type 1.

Animals↗
PubMed1995

Optimal lymphocytic choriomeningitis virus sequences restricted by H-2Db major histocompatibility complex class I molecules and presented to cytotoxic T lymphocytes.

Infection with lymphocytic choriomeningitis virus induces the generation of CD8+ cytotoxic T lymphocytes (CTL). In the H-2b mouse, this cellular immune response is directed against three viral structural epitopes (GP1, GP2, and NP) presented by the major histocompatibility complex (MHC) class I H-2Db molecules. This study was undertaken to delineate which sequence of each of these three epitopes is optimal for MHC binding and CTL recognition. The first step was to synthesize the relevant peptides truncated at the N or C terminus and flanking the crucial H-2Db-anchoring Asn residue in position 5. These peptides were then tested (i) for their binding properties in two H-2Db-specific assays with viable cells (upregulation of H-2Db expression on the surface of RMA-S cells and competition against the Db-restricted peptide 125I-gp276-286 on T2-Db cells) and (ii) for their abilities to sensitize H-2b target cells for CTL lysis in vitro. For optimal antigenic presentation, all three epitopes required the MHC-anchoring Asn residue at position 5 of their sequences. The results clearly and unambiguously delineated optimal lengths for two of the epitopes and two options for the third. NP appeared as a conventional 9-amino-acid (aa)-long peptide, np396-404 (FQPQNGQFI). GP2 was defined as a longer peptide (11 aa), gp276-286 (SGVENPGGYCL). Characterization of the GP1 epitope was more complex: the 9-aa-long peptide gp33-41 (KAVYNFATC) and the carboxyl-extended 11-aa-long peptide gp33-43 (KAVYN FATCGI) were both established as possible optimal sequences depending on the cell line used to test binding and lysis.

Amino Acid Sequence↗
PubMed1995

Focal expression of interleukin-2 does not break unresponsiveness to "self" (viral) antigen expressed in beta cells but enhances development of autoimmune disease (diabetes) after initiation of an anti-self immune response.

The participation of IL-2 in insulin-dependent (type 1) diabetes (IDDM) was analyzed in transgenic (tg) mice expressing the nucleoprotein (NP) of lymphocytic choriomeningitis virus and IL-2 under control of the rat insulin promoter focally in beta cells of the islets of Langerhans. Insertion and expression of the viral (self) gene or of the IL-2 gene alone did not lead to IDDM. Infiltration primarily of CD4 and B lymphocytes and increased expression of MHC class I and II molecules occurred in islets where IL-2 was expressed. By contrast, neither cellular infiltrates nor expression of MHC class I or II glycoproteins above base levels was noted in tgs expressing the viral protein alone. Double tg mice expressing both the viral protein and IL-2 in their islets displayed a modest increase in incidence of spontaneous diabetes compared with that of single transgenic mice expressing IL-2 alone. Breaking of immunological unresponsiveness or sensitization to self antigens did not occur. Neither cytotoxic T lymphocytes (CTL) nor antibodies directed against the viral tg (NP) were generated. However, after challenge with lymphocytic choriomeningitis virus, double tg mice developed anti-self (viral) CTL and IDDM (incidence > 95%) within 2 mo. The generation of virus ("self")-specific MHC-restricted CTL was dependent on CD4+ help. In contrast, viral inoculum to single tg mice expressing either the viral protein or IL-2 failed to enhance the incidence of IDDM over 30% for viral protein or 10% for IL-2 after an 8-mo observation period. Hence, in this autoimmune model in situ expression of IL-2 did not break unresponsiveness but markedly enhanced ongoing disease.

Analysis of Variance↗
PubMed1995

Sensitization to self (virus) antigen by in situ expression of murine interferon-gamma.

Autoimmune disease results from inflammatory destruction of tissues by aberrant self-reactive lymphocytes. We studied the autoimmune potential of T lymphocytes immunologically ignorant of viral antigens acting as self antigens and whether the host defense molecule IFN-gamma could stimulate these cells to cytotoxic competency. For this purpose, we produced double transgenic mice expressing pancreatic IFN-gamma as well as lymphocytic choriomeningitis virus (LCMV) nucleoprotein (NP) or glycoprotein (GP) antigen. 100% of the NP+/IFN-gamma+ mice became diabetic before 2 mo of age, while none of the NP single transgenic littermates and only 10% of IFN-gamma single transgenic littermates did. Strikingly, NP+/IFN-gamma+ mice spontaneously developed cytotoxic T lymphocyte activity on LCMV-infected targets and vaccinia virus-NP-infected ones without prior LCMV infection but NP+/IFN-gamma- mice did not, which indicates specific sensitization to the viral antigen by IFN-gamma. These results suggest that lymphocytes ignorant of self antigens can be activated by IFN-gamma released after immunologic stimulation such as viral infection. This mechanism may account for the loss of apparent tolerance to self antigens in autoimmune diseases such as insulin-dependent diabetes mellitus.

Animals↗
PubMed1994

Thymic selection and adaptability of cytotoxic T lymphocyte responses in transgenic mice expressing a viral protein in the thymus.

Upon primary challenge with lymphocytic choriomeningitis virus (LCMV), H-2d (BALB/cByJ) mice mount a cytotoxic T lymphocyte (CTL) response to a single immunodominant domain of the viral nucleoprotein (NP) but no detectable response to the viral glycoprotein (GP). To manipulate this CTL response, the viral NP gene was expressed in the thymus and peripheral T lymphocytes using the murine Thy1.2 promoter. As a result, such Thy1.2-NP (H-2d) transgenic (tg) mice deleted their high-affinity anti-LCMV-NP CTL, but generated equal numbers of lower-affinity NP CTL. Further, they made an alternative anti-LCMV-GP CTL response that is not normally found in non-tg mice indicating a hierarchial control of the CTL response. Unlike the H-2d mice, H-2b (C57Bl/6J) mice normally mount a CTL response to both LCMV-GP and -NP. When the LCMV-NP was expressed using the Thy1.2 promoter in these H-2b mice, the LCMV-NP-specific CTL response was completely aborted and no CTL to new, alternative viral epitopes were generated. Dilutions of H-2b or H-2d NP peptides indicated that 3-4 logs less H-2b NP peptide was required to sensitize syngeneic target cells for CTL-specific lysis, suggesting that the differing affinities of H-2b and H-2d major histocompatibility complex molecules for their peptides likely account for the total removal of NP CTL in the H-2b mice but only partial removal in H-2d mice made to express thymic NP. Thymic grafting experiments done with thymi from newborn Thy1.2-NP tg mice show that selection processes studied in this model are of central (thymic) origin and are not caused by Thy1.2-positive LCMV-NP-expressing T lymphocytes in the periphery.

Adaptation, Physiological↗
PubMed1994

Multiple isoforms of CD46 (membrane cofactor protein) serve as receptors for measles virus.

Measles virus (MV) causes a productive infection in humans and certain simian hosts. Rodent cells such as Chinese hamster ovary (CHO) and murine cell lines normally resist MV infection. Human CD46, or membrane cofactor protein, a complement regulatory protein, recently has been reported as the cellular receptor for MV. Multiple isoforms of the CD46 protein exist; four of these isoforms are commonly expressed on human cells. Expression of each of the four isoforms in CHO cells followed by exposure to MV led to the appearance of viral proteins within the cells and on the cell surface as detected by immunofluorescence. Syncytium formation also was observed in the cultures. CHO cells expressing any of the four isoforms and exposed to MV formed infectious centers when plated on Vero cell monolayers, indicating that the cells can transmit virus to uninfected cells. The murine cell line MC57 expressing the BC1 isoform of CD46 also stained positively for MV antigens and was positive in the infectious center assay after exposure to MV. Treatment of CD46-expressing cells with antibody to human CD46 inhibited MV binding in a dose-dependent manner. These observations indicate that any of the four primary isoforms of CD46 are able to serve as a receptor for MV.

Amino Acid Sequence↗
PubMed1994

Cerebral expression of multiple cytokine genes in mice with lymphocytic choriomeningitis.

The simultaneous expression of multiple cytokine genes was examined within the brain and peripheral organs of euthymic and congenic athymic mice during the course of lymphocytic choriomeningitis (LCM) induced by the intracranial inoculation of LCM virus. In euthymic mice, levels of TNF-alpha, IL-1 alpha, IL-6, and IL-1 beta mRNA in the brain increased early in infection and further increased with the progression of disease; the level of IFN-gamma mRNA was expressed at high levels late in infection and correlated with the onset of clinical disease. In the peripheral organs, TNF-alpha, IL-1 alpha, and IL-1 beta gene expression predominated early and then declined late in disease. In athymic mice (which do not develop LCM), expression of TNF-alpha, IL-1 alpha, IL-6, and IL-1 beta mRNA increased progressively in brain and peripheral organs from days 3 to 6 postinfection. The levels of these transcripts were significantly lower than those in infected euthymic animals. In euthymic mice, in situ hybridization revealed IFN-gamma-, TNF-alpha-, and IL-1 alpha-expressing cells throughout the inflammatory cell infiltrates associated with areas of LCMV nucleic acid. Therefore, in mice with LCM there is simultaneous expression of multiple cytokine genes, the pattern of which is distinct for brain vs peripheral organs and for euthymic vs athymic mice. The close spatiotemporal relationship between expression of cytokine genes and evolution of disease in brain implicate cytokines are not only key coordinators of the host response but also as possible mediators of neuropathology in LCM.

Animals↗
PubMed1994

Failure to detect genomic viral sequences in pancreatic tissues from two children with acute-onset diabetes mellitus.

Two cases of fatal, acute-onset, insulin-dependent diabetes mellitus (IDDM) in children were diagnosed. Epidemiologic and serologic studies, as well as histologic analysis of pancreatic tissue in fatal viral infections, support the contention that a viral infection could cause beta cell destruction, leading to IDDM. The presence of nucleic acid sequences from viral agents considered to be potentially diabetogenic, specifically, cytomegalovirus and mumps, rubella, and coxsackie viruses, were investigated in the pancreatic tissues by reverse transcription followed by polymerase chain reaction (RT-PCR) and Southern blot hybridization. Total pancreatic RNAs extracted from five children who died from nondiabetic causes were included as controls. Viral genetic information from any of these four viral agents was not found. This result indicates that the acute IDDM in these cases was not due to a direct infection of pancreatic beta cells by any of the viral agents studied.

Acute Disease↗
PubMed1994

Mechanism of lymphocytic choriomeningitis virus entry into cells.

The path that the arenavirus lymphocytic choriomeningitis virus (LCMV) uses to enter rodent fibroblastic cell lines was dissected by infectivity and inhibition studies and immunoelectron microscopy. Lysosomotropic weak bases (chloroquine and ammonium chloride) and carboxylic ionophores (monensin and nigericin) inhibited virus entry, assessed as virus nucleoprotein expression at early times post-infection, indicating that the entry process involved a pH-dependent fusion step in intracellular vesicles. That entry occurred in vesicles rather than by direct fusion of virions with the plasma membrane was confirmed by immunoelectron microscopy. The vesicles involved were large (150-300 nm diameter), smooth-walled, and not associated with clathrin. Unlike classical phagocytosis, virus uptake in these vesicles was a microfilament-independent process, as it was not blocked by cytochalasins. LCMV entry into rodent fibroblast cell lines thus involves viropexis in large smooth-walled vesicles, followed by a pH-dependent fusion event inside the cell.

Alkalies↗
PubMed1994

Limiting the available T cell receptor repertoire modifies acute lymphocytic choriomeningitis virus-induced immunopathology.

The invariably fatal immunopathological disease that follows intracerebral injection of CBA/Ca (H-2k) mice with 1000 PFU of lymphocytic choriomeningitis virus (LCMV) generally fails to develop in congenic mice transgenic for a V beta 8.1D beta 2J beta 2.3C beta 2 T cell receptor (TCR) gene. The majority of these LCMV-infected TCR-transgenic mice show a substantial meningitis of delayed onset, that resolves without causing any obvious clinical impairment. This inflammatory process depends on the involvement of V beta 8+ T cells, but does not require the participation of the CD4+ subset. The cytotoxic effectors that develop in both the transgenic mice and the CBA/Ca controls are lytic for target cells infected with a vaccinia construct expressing genes encoding the putative polymerase protein of LCMV. Limiting the available TCR repertoire to lymphocytes with a single V beta phenotype (not required for the generation of potent effectors in wild-type mice) thus modifies the development of the lethal neuropathology characteristic of LCMV infection, although the CD8+ cytotoxic T lymphocyte response is not greatly compromised.

Acute Disease↗
PubMed1994

A transgenic mouse model to assess the interaction of cytotoxic T lymphocytes with virally infected, class I MHC-expressing astrocytes.

Astrocytes provide crucial support for neurons and their impairment by viruses or their interactions with anti-viral or autoimmune responses could contribute to neurological disease. We have developed a transgenic mouse model to assess lymphocyte-astrocyte interactions. The major histocompatibility complex (MHC) class I molecule, Db, was expressed in astrocytes under the transcriptional control of regulatory sequences from the glial fibrillary acidic protein (GFAP) gene. Baseline cerebral MHC class I mRNA levels from transgenic mice were elevated over those of non-transgenic controls, and a prominent increase in cerebral MHC class I expression occurred following focal, injury-induced astroglial activation within transgenic brains but not in non-transgenic controls. FACS analysis of explant astrocyte cultures from established transgenic lines demonstrated astroglial expression of the GFAP-Db fusion gene at the protein level. Functional antigen-presenting capacity was conferred by the Db transgene, as virus-infected primary astrocytes obtained from transgenic BALB/c mice (KdIdDdLd) expressing the Db molecule were lysed by Db-restricted anti-viral CTL.

Animals↗
PubMed1994

How virus induces a rapid or slow onset insulin-dependent diabetes mellitus in a transgenic model.

We developed two distinct transgenic mouse models in which virus induced insulin-dependent (type 1) diabetes mellitus (IDDM). In one of these lines, the unique viral transgene was expressed in the islets of Langerhans and also in the thymus, but in the other line, expression was only in the islets. Insertion and expression of the viral (self) gene, per se, did not lead to IDDM, (incidence < 5%). By contrast, induction of an anti-self (anti-viral) CD8+ CTL response to the same virus later in life caused IDDM (incidence < 90%) in both transgenic lines, although the kinetics and requirements for CD4 help, the affinity and avidity of CD8+ CTL differed in each line. Mice not expressing the viral (self) gene in the thymus developed IDDM 10-14 days after infection. CD4+ T cells played no detectable role, since their depletion failed to alter either the kinetics or incidence of IDDM. By contrast, mice that expressed the viral gene in the thymus required significantly more time to develop IDDM. Their anti-self (viral) CD8+ CTL were of lower affinity and avidity than CD8+ CTL generated by nontransgenic controls. Disease was dependent on T cell help, since deletion of CD4+ cells completely circumvented the IDDM.

Animals↗
PubMed1994

Behavioral effects of persistent lymphocytic choriomeningitis virus infection in mice.

Lymphocytic choriomeningitis virus (LCMV) is a nonlytic murine virus that provides a valuable model system for studying the behavioral correlates of CNS viral infection. Newborn or immunosuppressed mice infected with LCMV develop a persistent tolerant infection characterized by continuous viral production. Virus can be found in various body organs including lung, liver, kidney, and brain. In brain, neurons are the predominant CNS cells infected and the greatest number of persistently infected neurons are found in the cerebral cortex, hippocampus, other limbic structures and parts of the hypothalamus. Despite continuous infection throughout the animal's life, neurons show no structural injury or dropout. Mice from the DBA/2J strain were infected with LCMV (1000 plaque-forming units) within 18 h of birth and tested for behavioral function as adults. Plaque assays indicated persistent infection in virus-injected mice. Mice were tested for their ability to learn a Y-maze spatial discrimination to avoid the onset of a mild footshock (0.43 mA). The number of correct avoidance responses made during training was taken as a measure of acquisition performance. The virus-infected mice showed a deficit in acquisition of the Y-maze discrimination compared to that seen in vehicle-injected and noninjected controls. Following additional training to reach control levels of performance, the infected mice and the controls were injected with the cholinergic antagonist scopolamine. Scopolamine (2.0 mg/kg) disrupted the performance of the infected mice significantly more than control performance, suggesting that a cholinergic dysfunction accounted for some of the learning deficit. A separate group of virus-infected mice exhibited hypoactivity during the first exposure to a locomotor testing apparatus.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

T-cell receptors from virus-specific cytotoxic T lymphocytes recognizing a single immunodominant nine-amino-acid viral epitope show marked diversity.

Following infection of the H-2d mouse by lymphocytic choriomeningitis virus, the newly generated cytotoxic T lymphocyte (CTL) response is focused to a single 9-amino-acid peptide sequence (epitope) of the virus. More than 96% of the primary, secondary, and clonal CTL respond to this lymphocytic choriomeningitis virus nucleoprotein epitope. This unique system affords the opportunity to evaluate the T-cell response to a single viral CTL epitope in a case in which the outcome of infection, either viral clearance or host death, is mediated by the CTLs. Specifically, the molecular structure of the T-cell receptors (TCRs) of CTLs responding to this epitope was analyzed. By using an anchored polymerase chain reaction, the TCR chains of three CTL clones cDNAs were amplified, sequenced, and found to have unique V alpha of V beta chains relative to each other as well as to lack restriction to any particular variable chain. These data indicate that the highly diverse antiviral CTL response is pleomorphic and probably provides an advantage to the host as it limits the emergence of viral variants that could more easily arise if the TCR response were homogeneous.

Amino Acid Sequence↗
PubMed1994

Virus-induced immunosuppression: kinetic analysis of the selection of a mutation associated with viral persistence.

Infection of neonatal mice with lymphocytic choriomeningitis virus (LCMV) strain Armstrong (ARM) results in a lifelong persistent infection. Viral variants (cytotoxic T lymphocyte [CTL] negative, persistence positive [CTL- P+]) can be isolated from the lymphoid tissues of such mice. Adult mice inoculated with these CTL- P+ viruses fail to generate sufficient cytotoxic T lymphocytes to clear the acute infection and become persistently infected. By contrast, inoculation of a similar dose of the parental ARM virus (CTL+ P-) into adult mice leads to the generation of a vigorous virus-specific CTL response that clears the infection. Sequence analysis revealed a phenylalanine (Phe)-to-Leucine (Leu) change at amino acid 260 of the viral glycoprotein (GP) as a marker for variant viruses with the CTL- P+ phenotype. An RNA PCR assay that detects the variant GP sequence and thus allows kinetic studies of the selection of the Leu at position 260 was developed. We found that although CTL- P+ viruses are known to be lymphotropic, mature T and B cells were not required for the generation and selection of the Leu at GP amino acid 260. Kinetically, in mice infected at birth with LCMV ARM, as early as 3 weeks postinfection the Phe-to-Leu change was found in virus in the serum. By 5 weeks, viral nucleic acid obtained from peritoneal macrophages, spleen, lymph nodes, and liver showed the Phe-to-Leu change. At 2 months postinfection, the Leu change was detected in virus from the thymus, heart, lung, and kidney. By contrast, virus replicating in the central nervous system showed only minimal levels of the Leu change by 4 months and as long as 1 year postinfection. In vitro studies showed that the parental LCMV ARM CTL+ P- virus replicates more efficiently and outcompetes CTL- P+ virus in a cultured neuronal cell line, indicating that differential growth properties in neurons are likely the basis for the selection of the parental virus over the CTL- P+ variant in the brain.

Amino Acid Sequence↗
PubMed1994

Activation of cerebral cytokine gene expression and its correlation with onset of reactive astrocyte and acute-phase response gene expression in scrapie.

The pathogenesis of scrapie, a transmissible subacute spongiform encephalopathy, is unclear. However, certain aspects of the known cellular and molecular neuropathology in scrapie led us to hypothesize that cytokines could mediate cerebral pathological changes in this neurodegenerative disease. Therefore, expression of multiple cytokine genes in the brain and peripheral organs of scrapie-infected mice was examined. Late in the course of scrapie, expression of tumor necrosis factor alpha (TNF-alpha), interleukin-1 alpha (IL-1 alpha), and IL-1 beta mRNA was markedly increased in the brain but not the spleen, kidneys, or liver. In time course studies, scrapie-infected mice exhibited increased cerebral expression of the TNF-alpha, IL-1 alpha, and IL-1 beta mRNAs by week 15 postinoculation--a time point that coincided with the onset of clinical symptoms. Thereafter, the levels of these cytokine transcripts increased progressively to the terminal stages of of the disease (week 25). To determine the relationship of the increased cerebral expression of the cytokine mRNAs to the development of pathological changes in scrapie, we examined the expression of the glial fibrillary acidic protein gene (a marker for astrocytosis) and the murine acute-phase response gene homologous to the alpha 1-antichymotrypsin gene (designated EB22/5.3). Markedly increased expression of both the glial fibrillary acidic protein and EB22/5.3 mRNAs was observed in the brain but not the peripheral organs of scrapie-infected mice. The increased expression of both these gene products also occurred at week 15 of infection and, thereafter, increased progressively to the terminal stages of the disease. Therefore, infection of mice with scrapie resulted in significant increases in the expression of the TNF-alpha, IL-1 alpha, and IL-1 beta gene products, whose pattern correlated with the onset and development of molecular and clinical pathological changes. Since scrapie is known not to evoke an immune response, the present findings strongly suggest the existence of a localized cerebral host response to the agent during which proinflammatory cytokines could be key pathogenic mediators.

Acute-Phase Reaction↗
PubMed1994

Virus-specific CD8+ cytotoxic T-lymphocyte activity associated with control of viremia in primary human immunodeficiency virus type 1 infection.

Human immunodeficiency virus type 1 (HIV-1) Env-, Gag-, Pol-, Nef-, and Tat-specific cytotoxic T-lymphocyte (CTL) activities were quantitated temporally in five patients with symptomatic primary HIV-1 infection. A dominant CD8(+)-mediated, major histocompatibility complex class I-restricted CTL response to the HIV-1 envelope glycoprotein, gp160, was noted in four of the five patients studied. The level of HIV-1-specific CTL activity in the five patients paralleled the efficiency of control of primary viremia. Patients who mounted strong gp160-specific CTL responses showed rapid reduction of acute plasma viremia and antigenemia, while in contrast, primary viremia and antigenemia were poorly controlled in patients in whom virus-specific CTL activity was low or undetectable. These results suggest that HIV-1-specific CTL activity is a major component of the host immune response associated with the control of virus replication following primary HIV-1 infection and have important implications for the design of antiviral vaccines.

Amino Acid Sequence↗