Hybridoma cell lines producing monoclonal antibodies to the JHM strain of mouse hepatitis virus-4 (MHV-4) were established. By indirect immunofluorescence and immune precipitation, monoclonal antibodies of three viral polypeptide specificities were characterized. Monoclonal antibodies to nucleocapsid reacted in the cytoplasm of infected cells and precipitated the 60,OOOd nucleocapsid polypeptide (VP-4) of MHV-4. Other monoclonal antibodies reacted both in the cytoplasm and on the surface of infected cells and were found to precipitate the 170,OOOd viral glycoprotein (GP-1). A third set of monoclonal antibodies reacted both in the cytoplasm and on the surface of infected cells and precipitated the 25,OOOd viral glycoprotein (GP-5) and its precursor VP-6 (23,OOOd). Anti-GP-1 alone had direct neutralizing activity for MHV-4 virus, while in the presence of complement both anti-GP-1 and anti-GP-5 neutralized virus. Only anti-GP-1 had the ability to inhibit the spread of infection due to fusion in L241 cells. Thus, the viral glycoprotein GP-1 likely contains both the attachment and fusion activities of MHV-4.

Experimental allergic encephalomyelitis (EAE) serves as a model for autoimmune diseases mediated by T lymphocytes. Following sensitization to rat, mouse or guinea pig myelin basic protein (MBP) in complete Freund's adjuvant, inbred mouse strains PL/J (H-2u), SJL/J (H-2s) and (PL/J X SJL/J)F1((PLSJ)F1) develop EAE. Whereas sensitization to the N-terminal 37 amino-acid peptide of rat or guinea pig MBP [MBP(1-37)] induces EAE in PL/J mice, immunization to the C-terminal peptide (89-169) leads to EAE in SJL/J mice. The immune response to MBP in (PLSJ)F1 mice is not co-dominant; sensitization to the N-terminal peptide induces EAE, while sensitization to the C-terminal peptide does not. We have generated MBP-specific T-cell clones restricted to class II (Ia) antigens of the major histocompatibility complex (MHC) from PL/J and (PLSJ)F1 mice following sensitization to rat MBP. Two such I-Au-restricted T-cell clones that proliferate in response to the encephalitogenic N-terminal MBP peptide and recognize a shared determinant with mouse (self) MBP cause paralysis in 100% of (PLSJ)F1 mice tested. Paralysis is induced even when recipients are injected with as few as 1 X 10(5) cloned T cells. Relapsing paralysis followed in two-thirds of the recipients after recovery from acute paralysis, whereas one-third developed chronic persistent paralysis, a form of EAE not usually seen. Histopathology revealed intense perivascular inflammation, demyelination and remyelination within the central nervous system of paralysed mice. The experimental disease induced with these clones shares important features with human demyelinating diseases such as multiple sclerosis. This is the first demonstration that T-cell clones that respond to a defined self-antigen can induce clinical and histological autoimmune disease.

Monoclonal hybridoma antibodies directed against the polypeptides of murine hepatitis virus-4 (JHM strain) were tested for their ability to alter the course of a normally lethal intracerebral virus challenge. Three monoclonal antibodies directed against two distinct epitopes on the E2 glycoprotein of MHV-4 protected mice against lethal virus challenge and converted the infection from fatal encephalomyelitis to demyelination. A single neutralizing antibody directed against a third epitope on E2 as well as seven nonneutralizing antibodies to E2, E1, and N polypeptides did not protect against challenge. In mice which received protective antibody, MHV-4 infection was not blocked, however, virus grew to lower titers in liver and brain, and virus replication in the CNS was more restricted than in unprotected mice. Decreased involvement of neurons in the brains of protected mice was observed, and no evidence of neuronal infection in the spinal cords was found. In contrast, oligodendrocytes were infected in the presence of protective antibody, and evidence of demylination associated with mononuclear cell infiltration was found. These studies demonstrate that antibody to a single epitope on a viral glycoprotein can substantially alter the course and phenotype of disease.

Spinal cord MS plaques are characteristically peripherally located, are less than two vertebral segments in length, and occupy less than half the cross-sectional area of the cord.

Mouse hepatitis virus (JHM strain) type 4 induces acute encephalitis followed by death in many strains of laboratory mice. Immunohistochemical study in vivo and analysis of mouse neuronal cells in vitro both indicate that the target cells in this infection is the neuron. Further, examination of several inbred mouse strains and neuronal cells from them shows that disease expression is controlled by a single autosomal gene action at the level of the neuronal cell. Susceptibility is dominant but not H-2 linked. However, cultured neuronal cells and macrophages from SJL/J mice, which are resistant to this infection, fail to make significant amounts of infectious virus after an appropriate viral inoculation. Apparently the defect is not at the level of the virus-cell receptor, because these cells, in part, express viral antigens.

Quantified lesion scores derived from MRI correlate significantly with neuropsychological testing in patients with multiple sclerosis (MS). Variables used to reflect disease severity include total lesion area (TLA), ventricular-brain ratio, and size of the corpus callosum. We used these general measures of cerebral lesion involvement as well as specific ratings of lesion involvement by frontal, temporal, and parieto-occipital regions to quantify the topographic distribution of lesions and consequent effects upon cognitive function. Lesions were heavily distributed in the parieto-occipital regions bilaterally. Neuropsychological tests were highly related to all generalized measures of cerebral involvement, with TLA being the best predictor of neuropsychological deficit. Mean TLA for the cognitively impaired group was 28.30 cm2 versus 7.41 cm2 for the cognitively intact group (p less than 0.0001). Multiple regression analyses revealed that left frontal lobe involvement best predicted impaired abstract problem solving, memory, and word fluency. Left parieto-occipital lesion involvement best predicted deficits in verbal learning and complex visual-integrative skills. Analysis of regional cerebral lesion load may assist in understanding the particular pattern and course of cognitive deficits in MS.

Monoclonal hybridoma antibodies (MAb) of defined polypeptide specificity and biological activity were used in a competition binding assay to identify antibody binding sites (epitopes) on the glycoproteins of murine hepatitis virus-4 strain JHM (MHV-4). Individual MAb were labeled with horseradish peroxidase (HRP) and used as probes in a competition enzyme immunoassay (EIA). Four topographically distinct antigenic sites were detected on the E2 glycoprotein of MHV-4. Antibodies reacting with these four determinants provisionally designated A(E2), B(E2), C(E2), and D(E2) had corresponding biological activities (M. J. Buchmeier, H. A. Lewicki, P. J. Talbot, and R. L. Knobler (1984) Virology 132, 261-270). Antibodies to sites A(E2) and B(E2) mediated virus neutralization in vitro and passively protected mice against lethal virus challenge in vivo. Antibody to site C(E2) neutralized virus efficiently in vitro but did not alter disease in vivo, while antibody to site D(E2) neither neutralized nor protected. Two major nonoverlapping antigenic sites were defined on the E1 glycoprotein. Overlapping epitopes A(E1) and B(E1) constituted one site and epitope C(E1) the other.

Experimental allergic encephalomyelitis (EAE) 1 is a clear example of an autoimmune disease mediated by class II-restricted T lymphocytes (1-5). Certain forms of EAE are characterized by relapsing paralysis, with histopathology revealing both perivascular lymphocytic cuffs and demyelination. Because of these features, chronic relapsing EAE is often cited as a model for the human disease multiple sclerosis (MS) (6). Further similarities exist between EAE and MS, including the presence of T helper cells in the inflammatory lesions (7,8), and the linkage of susceptibility for both EAE and MS to immune response genes (9, 10). With the capability for cloning antigen-specific T cells (1 1), it is now possible to analyze the precise cellular and immunogenetic mechanisms involved in EAE.Recently, it has been shown (12, 13) that the N-terminal 1-37 amino acid (aa) peptide of rat or guinea pig myelin basic protein (MBP) can induce EAE in PL/J (H-2 ") mice, while the C-terminal 89-169 aa peptide is encephalitogenic in SJL/ J (H-2 s) mice. The (PL/J × SJL/J)F1 [(PLSJ)FI] mice do not respond to MBP in a codominant manner. Instead, only the N-terminal peptide of MBP induces EAE in the (PLSJ)F1 mouse (12, 13).To characterize the T cells involved in the encephalitogenic response, MBPspecific T cell lines and clones have been isolated from (PLSJ)Ft mice after immunization with rat or bovine MBP. The separate determinants recognized by these clones have been investigated using peptides derived from MBP. In addition, the pattern of restriction to I-A or I-E class II molecules has been analyzed for each clone. With both the initial MBP-primed T cell lines and the L.