The CD4-mediated immune response is critical in determining the outcome of infection using Theiler's viruses with VP1 capsid protein point mutations.

Daniel's strain of Theiler's virus (DA) induces a chronic demyelinating disease in the central nervous system (CNS) of susceptible SJL mice, which serves as an excellent model of multiple sclerosis. We previously demonstrated that point mutations near a putative virus receptor-binding site [VP1 99 (Gly to Ser) or 100 (Gly to Asp)] totally attenuate the ability of DA to persist and induce demyelination in SJL mice. The current studies demonstrate that class II-restricted CD4(+) T cells play a major role in clearing VP1 mutant DA viruses from the CNS to prevent demyelination. Infection of SJL CD4((-/-)) mice with DA-VP1-99(Ser) or DA-VP1-100(Asp) resulted in virus persistence and prominent demyelination in the spinal cord. In contrast, infection of SJL CD8((-/-)) mice with DA-VP1-99(Ser) or DA-VP1-100 did not result in virus persistence or demyelination. In addition, no virus-specific cytotoxicity was observed in CNS-infiltrating lymphocytes following infection of SJL mice with VP1 mutant viruses. The mutant DA-VP1-99(Ser) and DA-VP1(100) viruses were in fact neurovirulent when compared to the wild-type DA virus, as they induced an overwhelming encephalitis and early lethality (2 to 4 days postinfection) in mice deficient in the IFN-alpha/beta receptor. Therefore, the nondemyelinating phenotype observed with DA-VP1-99(Ser) and DA-VP1-100(Asp) viruses is dependent in part on the CD4-mediated host immune response.

Assessment of hindlimb gait as a powerful indicator of axonal loss in a murine model of progressive CNS demyelination.

Identifying the role of axonal injury in the development of permanent, irreversible neurologic disability is important to the study of central nervous system (CNS) demyelinating diseases. Our understanding of neurologic dysfunction in demyelinating diseases and the ability to assess therapeutic interventions depends on the development of objective functional assays that can non-invasively measure axonal loss. In this study, we demonstrate in a murine model of progressive CNS demyelination that assessment of the hindlimb width of stride provides a powerful indicator of axonal loss and can dissociate between deficits induced by demyelination versus axonal loss.

CD40L is critical for protection from demyelinating disease and development of spontaneous remyelination in a mouse model of multiple sclerosis.

Theiler's murine encephalomyelitis virus (TMEV) induces acute neuronal disease followed by chronic demyelination in susceptible strains of mice. In this study we examined the role of a limited immune defect (deletion or blocking of CD40 ligand [CD40L]) on the extent of brain disease, susceptibility to demyelination, and the ability of demyelinated mice to spontaneously remyelinate following TMEV infection. We demonstrated that CD40L-dependent immune responses participate in pathogenesis in the cerebellum and the spinal cord white matter but protect the striatum of susceptible SJL/J mice. In mice on a background resistant to TMEV-induced demyelination (C57BL/6), the lack of CD40L resulted in increased striatal disease and meningeal inflammation. In addition, CD40L was required to maintain resistance to demyelination and clinical deficits in H-2b mice. CD40L-mediated interactions were also necessary for development of protective H-2b-restricted cytotoxic T cell responses directed against the VP2 region of TMEV as well as for spontaneous remyelination of the spinal cord white matter. The data presented here demonstrated the critical role of this molecule in both antibody- and cell-mediated protective immune responses in distinct phases of TMEV-mediated pathology.

Quantitative assessment of neurologic deficits in a chronic progressive murine model of CNS demyelination.

The precise factors involved in the development of a progressive motor dysfunction, a hallmark of immune-mediated demyelinating diseases such as multiple sclerosis, are not well defined. The ability to identify neurologic deficits that result in impaired motor performance early in disease may allow for the identification of therapeutic interventions that slow or eliminate the progression toward a permanent dysfunction. Here we describe the use of three objective, quantitative functional assays (spontaneous activity box, rotarod, and footprint analysis) to detect early neurologic deficits following the initiation of a demyelinating disease with Theiler's murine encephalomyelitis virus (TMEV). The results show that the assays are capable of revealing neurologic deficits at the early stages of the demyelinating disease process. These findings are the first to objectively characterize neurologic function in an animal model of progressive CNS demyelination.

Chromosome 14 contains determinants that regulate susceptibility to Theiler's virus-induced demyelination in the mouse.

Theiler's murine encephalomyelitis virus causes a chronic demyelinating disease in susceptible strains of mice that is similar to human multiple sclerosis. Several nonmajor histocompatibility complex-linked genes have been implicated as determinants of susceptibility or resistance to either demyelination or virus persistence. In this study, we used linkage analysis of major histocompatibility complex identical H-2d (DBA/2J x B10.D2) F2 intercross mice to identify loci associated with susceptibility to virus-induced demyelinating disease. In a 20-cM region on chromosome 14, we identified four markers, D14Mit54, D14Mit60, D14Mit61, and D14Mit90 that are significantly associated with demyelination. Because two peaks were identified, one near D14Mit54 and one near D14Mit90, it is possible that two loci in this region are involved in controlling demyelination.

Transplantation of normal bone marrow into a pig with severe von Willebrand's disease.

Bone marrow from a normal male pig was transplanted into a related female pig with severe homozygous von Willebrand's disease (vWd). After engraftment the circulating leukocytes were of the male karyotype, and the platelets were strongly positive for von Willebrand factor (vWF) by indirect immunofluorescence. The average level of vWF was 1.96 U/dl and of ristocetin cofactor was 2.8 U/dl. The ear immersion bleeding time before transplantation was consistently more than 15 min and afterwards varied between 5 min and more than 15 min. Transfused vWF corrected the bleeding time at a level of 10 U/dl, which is lower than that required for a von Willebrand pig. We concluded that: the plasmatic compartment is only minimally replenished by the vWF from platelets and megakaryocytes; and the platelet vWF alone only partially corrects the abnormal tests of the hemostatic mechanism in severe vWd.

Inheritance of a new bleeding disease in a herd of swine with Willebrand's disease.

A herd of swine affected by Willebrand's disease was begun in 1967 at the Mayo Clinic in order to study the inherited hemostatic abnormality in swine as a model for the human disease. Affected individuals have bleeding times in excess of 15 minutes, extremely low levels of Willebrand factor (less than or equal to 0.25 percent of normal), and decreased levels of VIII coagulant activity. Individuals with long bleeding times, higher levels of Willebrand factor and normal levels of VIII coagulant activity began to appear in the colony. It is hypothesized that this new (N) condition is inherited as a simple autosomal recessive (N/n) at a locus separate and independent of the similarly autosomal recessive (A/a) von Willebrand locus. In addition, the Willebrand locus is epistatic to the N locus, i.e., individuals will only express the new condition provided there is at least one normal allele at the von Willebrand locus. Therefore, individuals with genotype aa--are all von Willebrand phenotypically, and A-nn individuals have the new disease.