Biochemical study of glial cytoplasmic inclusions in multiple system atrophy brains

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Glial cytoplamic inclusions (GCI) which appear in oligodendroglial cells are the main pathological feature of multiple system atrophy (MSA). In this study, to determine the main constituent of GCI we tried several assay methods of GCI including immunohistochemical methods and found that Gallyas staining is the best assay method of GCI.Using Gallyas staining we developed the partial purification method of GCI.Furthermore electron micrograph showed the fibrous GCI in the Triton insoluble GCI-enriched fraction which was also observed with Gallyas silver staining. Then we performed 2D gel analysis of the GCI-rich fraction. This analysis revealed the increased amount of the protein which showed MW22kD and p17.0-7.6 corresponding to alpha B-Crystallin. Immunoblots of 2D gel of that fraction were stained with anti-alpha B-Crystallin antibodies and with Gallyas silver staining. Anti-alpha B-Crystallin antibodies reacted with the spots which increased in GCI-enriched fraction corresponding to alpha B-Crystallin. Gallyas silver staining also stained these spots, suggesting that alpha B-Crystallin is one of the Gallyas reactive proteins. These results suggests that one of the main constituent of GCI is alpha B-Crystallin.Furthermore anti-carboxyl methtltransferase antibodies stained GCI,suggesting that the racemization of those abnormal proteins occured in MSA brain.We got several antibodies against the gene products of genetic neurological disorders to study the relationship between these protein and GCI.These antibodies detect abnormal gene products in trinucleotide diseases.