Takeshi Hosokawa

Monoclonal antibodies specific to the integral membrane protein P0 of bovine peripheral nerve myelin

Two monoclonal antibodies (mAbs), 58A and 46E, were generated against the major protein P0 of bovine peripheral nervous system myelin (PNSM). The reactivities of the mAbs were assessed by enzyme-linked immunosorbent assay (ELISA), Western blot, and immunohistochemistry. Both mAbs, 58A and 46E, reacted to PNSM of bovine, human, rat and rabbit, but not to chicken PNSM or the brains of rat and rabbit. In the Western blot, these mAbs showed specific binding to bovine P0 as well as deglycosylated P0, but not to myelin-associated glycoprotein (MAG) of bovine spinal cord. The analyses of the lysylendopeptidase-digested peptides of bovine P0 revealed that the epitopes for the mAbs 58A and 46E were located on the amino acid residues 68-79 and 210-216, respectively. Since the mAbs 58A and 46E recognize the extracellular domain and the cytoplasmic domain of P0, respectively, they could be useful for studies on P0s role in myelin formation, its adhesive properties, and functions of the N-terminal extracellular and C-terminal cytoplasmic domains of the protein.

Cell-mediated immunity to bovine P2 protein and neuritogenic synthetic peptide in experimental allergic neuritis

Cellular reactivity to bovine P2 protein (P2) and its two synthetic peptides, SP66-78 and SP70-78, was serially examined by the lymphocyte proliferation test in animals with experimental allergic neuritis (EAN). SP66-78 and SP70-78 correspond to residues 66-78 and 70-78 of bovine P2. Proliferative response to SP66-78 as well as P2 appeared at day 7 before the onset of EAN and was clearly manifested at day 14 in the active stage, thereafter disappearing in the stable stage, whereas no response to SP70-78 was detected during the course of the disease. These results suggest that cell-mediated immune response to P2 and the specific part residues 66-78 of P2 play an important role in the pathogenesis of EAN.

Activated T lymphocyte subsets in experimental allergic neuritis

Changes in activated T cell subsets in peripheral blood were examined during the course of experimental allergic neuritis (EAN), using two-color immunofluorescence flow cytometry. Both CD4+ and CD8+ activated T cells decreased transiently before the onset of clinical signs, and increased just around the time of onset of the disease. In contrast, during the recovery phase, the numbers of CD4+ activated T cells returned to the normal range, whereas CD8+ activated T cells continued to increase. These findings imply that activation of CD4+ helper/inducer cells contributes mainly to the evolution of EAN, and that of CD8+ suppressor cells are necessary for recovery.

Serum Antibodies to Peripheral Nerve Antigens in Guillain‐Barré Syndrome

We previously reported that cellular hypersensitivity to P2 protein (P2) might play an important role in the evolution of experimental allergic neuritis’ and the GuillainBarri syndrome (GBS).’ In the present study, we examined serum antibodies to three different nervous antigens, P2, a synthetic tridecapeptide (SP66-78) corresponding to residues 66-78 of bovine P2, and galactocerebroside (GC), in GBS and other neurologic diseases. Myelin fraction was prepared from bovine peripheral nerve roots, as previously described,’ and P2 was purified by acid extraction of myelin using the method of Kitamura et aL4 SP66-78 was synthesized in a liquid phase by stepwise elongation according to the method of Suzuki et aL5 Antibody titers were measured by enzyme-linked immunosorbent assay (ELISA) in the sera of 11 patients with GBS,

Changes in T‐Cell Subsets in Experimental Allergic Neuritis

FIGURE 1. Clinical course of experimental allergic neuritis and immune responses to P2 protein. Upper column shows changes in mean values of clinical scores on the scale of Hughes et ~ l . , ~ with slight modification. The mean time of onset was 10.5 days, and the clinical scores rapidly reached their maximum on day 14, continued to be high until day 16, and thereafter gradually declined. Lower column shows the results of the lymphocyte proliferation test and enzyme-linked immunosorbent assay in peripheral blood from another 15 animals sacrificed on days 7, 12, and 21 after immunization. Vertical bars indicate the mean ? 1 standard deviation. CS = clinical score; SI = stimulated index; OD = optical density at 405 nm.

1126 Characterization of autoantibody specifically recognizing a neuron

Tenascin gene is expressed in the immature astroglia and radial glia in the developing mouse brain prior to the appearance of glial fibrillary acidic protein (GFAP). Thus, astrogliogenesis in the olfactory bulb was investigated by employing in situ hybridization histochemistry for tenascin q RNA as an early marker. Tenascin gene was expressed in the ventricular germinative zone as early as El5 and they began to migrate radially. A subset of the labeled cells were aligned along the forming olfactory glomeruli. Tenascin-immunoreactive radial glial processes extended from the ventricular zone towards the forming olfactory glomeruli. Another set of thinner and elaborate immunopositive processes were found in the forming olfactory glomeruli. On the other hand, tenascin gene expressing cells began to migrate from the rostra1 end of the lateral ventricle towards the olfactory bulb as early as E17. These cells were q itotic and the stream reached the olfactory bulb at PO. At P2, these cells showed GFAP immunoreactivity and extended processes to various direction. This astroglial strand coincides with the migratory stream of granule cells towards the olfactory bulb. Above findings suggest that astroglia in the olfactory bulb have multiple origins, i.e., the ventricular zone within the olfactory bulb and the rostra1 end of the lateral ventricle. Tenascin gene expressing cells aligned early in the olfactory glomeruli might represent the positional information for the formation of glomeruli. On the other hand, tenascin gene expressing cells originating from the rostra1 end of the lateral ventricle might be involved in the guidance of granule cell migration.

Antibody activities to sulfatide, PO protein and oligosaccharide chains of PO protein in polyneuropathies

We h a v e e v a l u a t e d a n t i S u l f a t i d e and PO p r o t e i n IgM a n t i b o d y a c t i v i t i e s i n s e r a f r o m one c a s e o f c h r o n i c p o l y n a u r o p a t h y w i t h IgM p a r a p r o t e i n e m i a ( IgM-CP) , 5 c a s e s o f c h r o n i c i d i o p a t h i c p o l y n e u r o p a t h y ( C I P ) , 6 c a s e s o f ac u t e i d i o p a t h i c p o l y n e u r o p a t h y (AIP) and 6 c a s e s o f c o n t r o l s . ELISA a n a l y s i s f o r a n t i S u l f a t i d e and P0 p r o t e i n IgM a n t i b o d y r e v e a l e d h i g h t i t e r i n IgM-CP and s i g n i f i c a n t l y h i g h e r i n CIP t h a n i n AIP and c o n t r o l s . F u r t h e r m o r e , we c o u l d d e t e r m i n e t h e s t r u c t u r e o f o l i g o a a c c h a r i d a c h a i n s o f P0 p r o t e i n , GP 4 and GP 5. These d i f f e r e n c e s were o n l y n o n r e d u c i n g t e r m i n a l s u g a r o f t h e s e c h a i n s . GP 5 h a s a s u l f a t e d g l u c n r o n i o a c i d and GP 4 h a s a s i a l i c a c i d a t t h e t e r m i n a l . We examined t h e r e a c t i v i t i e a o f e l u t e d IgH f r o m one c a s e o f IgH-CP and 3 c a s e s o f CIP w i t h t h e s e c h a i n s . The IgM o f IgM-CP r e a c t e d w i t h GP 5 o n l y , and t h e IgM o f CIP r e a c t e d w i t h b o t h . These d a t a s u g g e s t e d t h e p r e s e n c e o f a n t i S u l f a t i d e and P0 p r o t e i n IgH a n t i b o d i e s i n t h e s e r a o f IgN-CP and CIP , and t h e e p i t o p e o f t h e se rum o f IgH-CP m i g h t be t h e s u l f a t e d g l u c u r o n i c a c i d i n GP 5.

Immunoglobulin G in multiple sclerosis brain

We extracted free and bound IgG from plaques and normal-appearing white matter of multiple sclerosis (MS) brains. By isoelectric focusing (IEF), three patterns of IgG distribution were seen: (A) a restricted high-pI distribution with a specific band at low pH, (B) a restricted high-pI pattern, and (C) a broad pI pattern similar to that of the unbound IgG extracted at neutral pH. In one MS brain, we compared the IEF pattern of plaque material with that of normal-appearing white matter (NAWM); the low-pH extract of plaque material (PM) had a restricted pattern at high pI. In another MS brain, a specific band of bound IgG was found. These data suggest that MS lesions expose an antigen(s) unique to MS. B cells consequently might be stimulated by a disease-related antigen(s) in the MS lesion.

Changes in T cell subsets during the course of experimental allergic neuritis

Seven ~ormal controls and 40 patients with various neurological disorders including multiple sclerosis, cystercosis, and Guillain13arr~ syndro,ne etc. were quantitatively an4 qualitative-ly studied. identical results from both quantitative and qualitative studies were obtained in 50 patients for positive, and 3 patients and all seven norm:~l controls for negative. ~egative result was obtained in 4 and 5 patients by quantitative an,l q1~alitative study respectively. Our results suggest that a co,ncrehensive study of CSF igG is better than either intrathecal IgG de nero synthesis o~: i~G fraction bands alone.