CD8 T cell defect of TNF-α and IL-2 in DNAM-1 deficient mice delays clearance in vivo of a persistent virus infection.

DNAM-1 gene-deficient (-/-) mice take significantly longer to clear an acute and persistent LCMV infection in vivo than DNAM-1 +/+ mice. During acute LCMV priming, at the single cell level, DNAM-1 -/- mice made significantly less cytoplasmic CD8 TNF-α and IL-2 but not IFN-γ than their DNAM-1 +/+ counterparts. Restimulated immune memory CD8 T cells from DNAM-1 -/- and DNAM-1 +/+ mice were equivalent in cytolytic activity against LCMV-infected target cells but DNAM-1 -/- CD8 T cells had significant reductions in TNF-α and IL-2 that were associated on adoptive transfer with the inability to terminate the persistent viral infection.

Expanded potential for recombinant trisegmented lymphocytic choriomeningitis viruses: protein production, antibody production, and in vivo assessment of biological function of genes of interest.

The recombinant engineering of trisegmented lymphocytic choriomeningitis virus (LCMV) to express two genes of interest was recently reported. We used this technology to efficiently express green fluorescent protein (GFP) and the immunoregulatory gene product interleukin-10 (IL-10) in vitro, assess IL-10 function in vivo during viral meningitis, and generate specific, robust monoclonal antibody responses to IL-10. Tripartite viruses were attenuated in wild-type and TLR7(-/-) mice.

RNA editing enzyme adenosine deaminase is a restriction factor for controlling measles virus replication that also is required for embryogenesis.

Measles virus (MV), a member of the family Paramyxoviridae and an exclusively human pathogen, is among the most infectious viruses. A progressive fatal neurodegenerative complication, subacute sclerosing panencephalitis (SSPE), occurs during persistent MV infection of the CNS and is associated with biased hypermutations of the viral genome. The observed hypermutations of A-to-G are consistent with conversions catalyzed by the adenosine deaminase acting on RNA (ADAR1).

Mapping and restriction of a dominant viral CD4+ T cell core epitope by both MHC class I and MHC class II.

Virus-specific CD4(+) T cells contribute to effective virus control through a multiplicity of mechanisms including direct effector functions as well as "help" for B cell and CD8(+) T cell responses. Here, we have used the lymphocytic choriomeningitis virus (LCMV) system to assess the minimal constraints of a dominant antiviral CD4(+) T cell response. We report that the core epitope derived from the LCMV glycoprotein (GP) is 11 amino acids in length and provides optimal recognition by epitope-specific CD4(+) T cells.

Mass spectrometry reveals specific and global molecular transformations during viral infection.

Mass spectrometry analysis was used to target three different aspects of the viral infection process: the expression kinetics of viral proteins, changes in the expression levels of cellular proteins, and the changes in cellular metabolites in response to viral infection. The combination of these methods represents a new, more comprehensive approach to the study of viral infection revealing the complexity of these events within the infected cell. The proteins associated with measles virus (MV) infection of human HeLa cells were measured using a label-free approach.

CD4 T cell control primary measles virus infection of the CNS: regulation is dependent on combined activity with either CD8 T cells or with B cells: CD4, CD8 or B cells alone are ineffective.

Measles virus (MV), one of the most infectious of human pathogens, still infects over 30 million humans and causes over 500,000 deaths each year [Griffin, D., 2001. Measles virus.

T cells infiltrate the brain in murine and human transmissible spongiform encephalopathies.

CD4 and CD8 T lymphocytes infiltrate the parenchyma of mouse brains several weeks after intracerebral, intraperitoneal, or oral inoculation with the Chandler strain of mouse scrapie, a pattern not seen with inoculation of prion protein knockout (PrP(-/-)) mice. Associated with this cellular infiltration are expression of MHC class I and II molecules and elevation in levels of the T-cell chemokines, especially macrophage inflammatory protein 1beta, IFN-gamma-inducible protein 10, and RANTES. T cells were also found in the central nervous system (CNS) in five of six patients with Creutzfeldt-Jakob disease.

Lymphotoxin-alpha- and lymphotoxin-beta-deficient mice differ in susceptibility to scrapie: evidence against dendritic cell involvement in neuroinvasion.

Transmissible spongiform encephalopathy or prion diseases are fatal neurodegenerative disorders of humans and animals often initiated by oral intake of an infectious agent. Current evidence suggests that infection occurs initially in the lymphoid tissues and subsequently in the central nervous system (CNS). The identity of infected lymphoid cells remains controversial, but recent studies point to the involvement of both follicular dendritic cells (FDC) and CD11c(+) lymphoid dendritic cells.

Evidence that the hypermutated M protein of a subacute sclerosing panencephalitis measles virus actively contributes to the chronic progressive CNS disease.

Subacute sclerosing panencephalitis (SSPE) is a progressive degenerative disease of the brain uniformly leading to death. Although caused by measles virus (MV), the virus recovered from patients with SSPE differs from wild-type MV; biologically SSPE virus is defective and its genome displays a variety of mutations among which biased replacements of many uridine by cytidine resides primarily in the matrix (M) gene. To address the question of whether the SSPE MVs with M mutations are passive in that they are not infectious, cannot spread within the CNS, and basically represent an end-stage result of a progressive infection or alternatively SSPE viruses are infectious, and their mutations enable them to persist and thereby cause a prolonged neurodegenerative disease, we utilized reverse genetics to generate an infectious virus in which the M gene of MV was replaced with the M gene of Biken strain SSPE MV and inoculated the recombinant virus into transgenic mice bearing the MV receptor.

Common antiviral cytotoxic t-lymphocyte epitope for diverse arenaviruses.

Members of the Arenaviridae family have been isolated from mammalian hosts in disparate geographic locations, leading to their grouping as Old World types (i.e., lymphocytic choriomeningitis virus [LCMV], Lassa fever virus [LFV], Mopeia virus, and Mobala virus) and New World types (i.

Structural and functional identification of major histocompatibility complex class I-restricted self-peptides as naturally occurring molecular mimics of viral antigens. Possible role in CD8+ T cell-mediated, virus-induced autoimmune disease.

Structural similarity (molecular mimicry) between viral epitopes and self-peptides can lead to the induction of autoaggressive CD4(+) as well as CD8(+) T cell responses. Based on the flexibility of T cell receptor/antigen/major histocompatibility complex recognition, it has been proposed that a self-peptide could replace a viral epitope for T cell recognition and therefore participate in pathophysiological processes in which T cells are involved. To address this issue, we used, as a molecular model of viral antigen, the H-2D(b)-restricted immunodominant epitope nucleoprotein (NP)-(396-404) (FQPQNGQFI) of lymphocytic choriomeningitis virus (LCMV).

Differences in affinity of binding of lymphocytic choriomeningitis virus strains to the cellular receptor alpha-dystroglycan correlate with viral tropism and disease kinetics.

alpha-Dystroglycan (alpha-DG) was recently identified as a receptor for lymphocytic choriomeningitis virus (LCMV) and several other arenaviruses, including Lassa fever virus (W. Cao, M. D.

Immunosuppression and resultant viral persistence by specific viral targeting of dendritic cells.

Among cells of the immune system, CD11c(+) and DEC-205(+) splenic dendritic cells primarily express the cellular receptor (alpha-dystroglycan [alpha-DG]) for lymphocytic choriomeningitis virus (LCMV). By selection, strains and variants of LCMV that bind alpha-DG with high affinity are associated with virus replication in the white pulp, show preferential replication in a majority of CD11c(+) and DEC-205(+) cells, cause immunosuppression, and establish a persistent infection. In contrast, viral strains and variants that bind with low affinity to alpha-DG are associated with viral replication in the red pulp, display minimal replication in CD11c(+) and DEC-205(+) cells, and generate a robust anti-LCMV cytotoxic T lymphocyte response that clears the virus infection.

Transgenic mice expressing human HLA and CD8 molecules generate HLA-restricted measles virus cytotoxic T lymphocytes of the same specificity as humans with natural measles virus infection.

Control of primary measles virus (MV) infection in humans and continued maintenance of immune memory that protects against reinfection are mediated primarily through the anti-MV T cell response, as judged by observations of children with defects in antibody formation but competency in making T cells. Further, the failure of T cell responses in those infected with MV most often leads to overwhelming infection. To better define and manipulate the elements involved in human T cell responses to MV, we analyzed the generation of HLA-restricted cytotoxic T lymphocytes (CTL) in a small animal model.

V and C proteins of measles virus function as virulence factors in vivo.

The measles virus (MV) P gene encodes three proteins: the P protein and two nonstructural proteins, C and V. Because the functions of both the C and V protein are unknown, we used MV C (C-) and V (V-) deletion recombinants generated by the MV reverse genetics system (F. Radecke, P.

Measles virus infection in a transgenic model: virus-induced immunosuppression and central nervous system disease.

Measles virus (MV) infects 40 million persons and kills one million per year primarily by suppressing the immune system and afflicting the central nervous system (CNS). The lack of a suitable small animal model has impeded progress of understanding how MV causes disease and the development of novel therapies and improved vaccines. We tested a transgenic mouse line in which expression of the MV receptor CD46 closely mimicked the location and amount of CD46 found in humans.

Use of a high-affinity peptide that aborts MHC-restricted cytotoxic T lymphocyte activity against multiple viruses in vitro and virus-induced immunopathologic disease in vivo.

Binding of a specific peptide(s) from a viral protein to major histocompatibility complex (MHC) class I molecules is a critical step in the activation of CD8(+) cytotoxic T lymphocytes (CTLs). Once activated, CTLs can cause lethal disease in an infected host, for example, by killing virus-containing ependymal and ventricular cells in the central nervous system or viral protein-expressing beta cells in the pancreatic islets of Langerhans. Here we describe the usage of a designed (not natural) high-affinity peptide to compete with viral peptide(s)-MHC binding.

In vivo treatment with a MHC class I-restricted blocking peptide can prevent virus-induced autoimmune diabetes.

We tested the in vivo potential of a MHC class I-restricted blocking peptide to sufficiently lower an anti-viral CTL response for preventing virus-induced CTL-mediated autoimmune diabetes (insulin-dependent diabetes mellitus (IDDM)) in vivo without affecting systemic viral clearance. By designing and screening several peptides with high binding affinities to MHC class I H-2Db for best efficiency in blocking killing of target cells by lymphocytic choriomeningitis virus (LCMV) and other viral CTL, we identified the peptide for this study. In vitro, it selectively lowered CTL killing restricted to the Db allele, which correlated directly with the affinity of the respective epitopes.

Antiviral pressure exerted by HIV-1-specific cytotoxic T lymphocytes (CTLs) during primary infection demonstrated by rapid selection of CTL escape virus.

The HIV-1-specific cytotoxic T lymphocyte (CTL) response is temporally associated with the decline in viremia during primary HIV-1 infection, but definitive evidence that it is of importance in virus containment has been lacking. Here we show that in a patient whose early CTL response was focused on a highly immunodominant epitope in gp 160, there was rapid elimination of the transmitted virus strain and selection for a virus population bearing amino acid changes at a single residue within this epitope, which conferred escape from recognition by epitope-specific CTL. The magnitude (> 100-fold), kinetics (30-72 days from onset of symptoms) and genetic pathways of virus escape from CTL pressure were comparable to virus escape from antiretroviral therapy, indicating the biological significance of the CTL response in vivo.

Coexpression of B7-1 and viral ("self") transgenes in pancreatic beta cells can break peripheral ignorance and lead to spontaneous autoimmune diabetes.

We evaluated the role of the costimulatory molecule B7-1 in overcoming peripheral ignorance in transgenic mice, which expressed the glycoprotein (GP) or nucleoprotein (NP) of lymphocytic choriomeningitis virus (LCMV) as the self-antigen in pancreatic beta cells. The viral transgenes or B7-1 alone did not induce autoimmune diabetes (IDDM). However, in bigenic mice expressing B7-1 and LCMV-GP, anti-self (viral) cytotoxic T lymphocytes (CTL) were activated without viral infection and spontaneous IDDM occurred.

Discriminated selection among viral peptides with the appropriate anchor residues: implications for the size of the cytotoxic T-lymphocyte repertoire and control of viral infection.

Structural characterization of peptides restricted by major histocompatibility complex (MHC) class I molecules has identified residues critical for MHC class I binding and for T-cell receptor recognition. For example, optimal peptides fitting into the murine MHC class I Db groove are 9 to 11 amino acids long and require as MHC anchor residues an Asn (N) at position 5 and also either a hydrophobic residue, a Met (M) or a Cys (C), at the carboxy terminus. The three known Db-restricted peptides of lymphocytic choriomeningitis virus (LCMV) are glycoproteins GP1 (amino acids [aa] 33 KAVYNFATC), GP2 (aa 276 SGVENPGGYCL), and nucleoprotein NP (aa 396 FQPQNGQFI).

An essential role for type 1 interferon-gamma in terminating persistent viral infection.

The mechanism(s) by which infectious material is cleared by the host is an area of intensive study. This is especially so with the realization that persistent viral infection is a cause of chronic disease in humans and presents a major health problem. We have used the murine model of infection with lymphocytic choriomeningitis virus to evaluate immune clearance.

CTL escape viral variants. II. Biologic activity in vivo.

The proteins of lymphocytic choriomeningitis virus (LCMV) contain only three known peptide regions that are processed and then held in place by the MHC class I H-2b (Db) glycoprotein on the cell's surface for recognition by LCMV-specific Db-restricted cytotoxic T lymphocytes (CTL). These peptides are from the glycoprotein (GP), amino acids 33-41 KAVYNFATC (GP1) and 276-286 SGVENPGGYCL (GP2), and the nucleoprotein (NP), 396-404. We have used CTL clones that recognized only GP1, GP2, and NP to select viral variants that upon infecting cells bearing H-2b molecules escaped recognition by virus-specific CTL directed against the viral GP (GP1 + GP2) mutant, termed GPV, or the viral GP and NP (GP1 + GP2 + NP) mutant, termed GPV+NPV.