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Progressive accumulation of ubiquitin and disappearance of α-synuclein epitope in multiple system atrophy-associated glial cytoplasmic inclusions: triple fluorescence study combined with Gallyas-Braak method

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Abstract

α-Synuclein (αS) and ubiquitin (Ub) are shared constituents of glial cytoplasmic inclusions (GCIs) and Lewy bodies (LBs), both composed of fibrillary structures. Staining profiles of GCIs were investigated with triple immunofluorescence involving immunostaining for αS and Ub, both amplified with catalyzed reporter deposition, and a fluorochrome, thiazin red (TR) that has an affinity to fibrillary structures. After observation for the triple-fluorescent images, the sections were subsequently stained with the Gallyas-Braak method. Sections of putamen, cerebellar white matter and motor cortex from patients suffering from multiple system atrophy (MSA) with varying duration of the disease (4–15 years) were quantified for these staining profiles of Gallyas-positive GCIs. Although most of GCIs were positive for Ub and variably positive for αS, they were consistently negative for TR. The result was opposite in LBs in Lewy body disease with variable affinity to TR, suggesting that the construction of GCIs is different from that of LBs. These four staining features (αS, Ub, TR and Gallyas) alone failed to exhibit apparent correlation with disease duration, lesion site or severity of degeneration as reported previously. The fraction of αS-negative and Ub-positive GCIs, however, linearly increased along the disease progression, while that of αS-positive and Ub-negative GCIs decreased in contrast. This reciprocal change suggests that αS immunoreactivity in GCIs is being replaced by Ub immunoreactivity during the disease progression, which resulted in the ultimate predominance of αS-negative and Ub-positive GCIs in the most advanced case. Interestingly, this predominance of αS-negative and Ub-positive GCIs was a feature of motor cortex, where degeneration usually remains mild in spite of robust appearance of Gallyas-positive GCIs. Another fraction, αS-positive and Ub-positive GCIs were frequent in cerebellar white matter, suggesting that GCI evolution is heterogeneous and dependent also on area examined. Progressive accumulation of Ub with concomitant disappearance of αS epitope and their colocalization, partly shared with LBs, may represent a process of GCI formation, possibly linked to an aspect of degeneration in MSA.

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Acknowledgements

We are grateful to Prof. Hiroshi Tanaka and Dr. Michihiko Koeda (Department of Bioinformatics, Medical Research Institute, Tokyo Medical and Dental University) for their help in the statistical analyses. This work was supported in part by the grants from the Ministry of Health and Welfare, Japan (Longevity Science H-14–005 to T.U. and H.M.) and the Ministry of Education, Culture, Sports, Science and Technology (grant in aid for scientific research B15300118 to T.U.)

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Authors and Affiliations

  1. Department of Neurology, Tokyo Metropolitan Neurological Hospital, Tokyo, Japan

    Masaki Sakamoto

  2. Department of Neuropathology, Tokyo Metropolitan Institute for Neuroscience, 2-6 Musashi-dai, Fuchu, 183-8526, Tokyo, Japan

    Masaki Sakamoto, Toshiki Uchihara & Ayako Nakamura

  3. Department of Neurology and Neurological Science, Tokyo Medical and Dental University, Tokyo, Japan

    Masaki Sakamoto & Hidehiro Mizusawa

  4. Department of Pathology, Tokyo Metropolitan Neurological Hospital, Tokyo, Japan

    Toshio Mizutani

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  1. Masaki Sakamoto
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Correspondence to Toshiki Uchihara.

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Sakamoto, M., Uchihara, T., Nakamura, A. et al. Progressive accumulation of ubiquitin and disappearance of α-synuclein epitope in multiple system atrophy-associated glial cytoplasmic inclusions: triple fluorescence study combined with Gallyas-Braak method. Acta Neuropathol 110, 417–425 (2005). https://doi.org/10.1007/s00401-005-1066-9

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  • DOI: https://doi.org/10.1007/s00401-005-1066-9

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