Original Article Localization of CHMP2B-immunoreactivity in the brainstem of Lewy body disease Takashi Kurashige, 1 Tetsuya Takahashi, 1 Yuu Yamazaki, 1 Masanori Hiji, 1,2 Yuishin Izumi, 2 Takemori Yamawaki 1 and Masayasu Matsumoto 1 1 Department of Clinical Neuroscience and Therapeutics, Hiroshima University Graduate School of Biomedical Sciences, Hiroshima and 2 Mifukai Viha-ra Hananosato Hospital, Miyoshi, Japan Alpha-synuclein (aS) is one of the major constituents of Lewy bodies (LBs). Several lines of evidence suggest that the autophagy-lysosome pathway (ALP) is involved in the removal of aS. We have previously reported that granulo- vacuolar degeneration (GVD) in neurons involved a subunit of the endosomal sorting complexes required for transport (ESCRT). In this study, we examined the asso- ciation between alpha-synucleinopathy and autophagy through immunohistochemical analysis of charged multi- vesicular body protein 2B (CHMP2B), a component of the ESCRT-pathway. We examined the brainstems of 17 patients with Parkinson’s disease (PD), incidental Lewy body disease (ILBD), multiple system atrophy (MSA), and Alzheimer’s disease (AD) immunohistochemically using antibodies against phosphorylated aS (paS), phosphor- ylated tau and CHMP2B. LBs and a proportion of glial cytoplasmic inclusions (GCIs) were immunopositive for paS and CHMP2B. Neurons containing CHMP2B- immunoreactive granules were detected in PD and ILBD, but not in MSA and AD brains. CHMP2B immunoreac- tivity was increased in the dorsal motor nucleus of the vagus nerve (DMNX) in PD and ILBD brains, relative to that in MSA and AD. These findings indicate that the ESCRT-pathway is implicated in the formation of aS inclusions, especially in PD and ILBD. Key words: alpha-synuclein, CHMP2B (charged multive- sicular body protein 2B), ESCRT (endosomal sorting com- plexes required for transport), glial cytoplasmic inclusion, Lewy body. INTRODUCTION Parkinson’s disease (PD) and dementia with Lewy bodies (DLB) are neurodegenerative diseases characterized by the presence of LBs, Lewy neurites (LNs), and pale bodies in the central, peripheral and autonomic nervous system as pathological features. The major components of LBs and LNs are aggregated alpha-synuclein (aS), ubiquitin (Ub), phosphorylated neurofilaments, components of the ubiquitin-proteasome system, molecular chaperones and lipids. 1–5 MSA is a similar neurodegenerative disease with glial cytoplasmic inclusions (GCIs) and neuronal cytoplas- mic inclusions (NCIs), which can be stained with Gallyas Braak (GB) silver. 6–9 Because LBs, LNs, GCIs, and NCIs contain pathologic aS, the term alpha-synucleinopathy is used to collectively refer to PD, DLB and multiple system atrophy (MSA). 10 Since aS is thought to be processed by the ubiquitin proteasome system (UPS), 11 UPS dysfunction has been implicated in the pathogenesis of PD, and more recently, there has been growing interest in characterizing the role of the autophagy-lysosome pathway (ALP) in PD. 2,12 There are three distinct autophagic pathways: (i) macroautophagy; (ii) microautophagy; and (iii) chaperone- mediated autophagy. Among these pathways, macroau- tophagy, a bulk protein degradation pathway, is thought to be constitutively active and highly efficient in healthy neurons. In response to various stimuli, an isolated mem- brane appears in the cytosol, where it gradually elongates to sequester cytoplasmic constituents. Macroautophagy targets cytoplasmic constituents and organelles to the lyso- some for bulk degradation; 13,14 thus, macroautophagy has been linked to neuronal cell death 15,16 and is activated in mouse models of neurodegeneration. 17,18 In PD patients, Anglade et al. described the ultrastructural characteristics of apoptosis and autophagic degeneration in melanized neurons of the substantia nigra. 19 Similarly, granulovacu- Correspondence: Takashi Kurashige, MD, Department of Clinical Neuroscience and Therapeutics, Hiroshima University Graduate School of Biomedical Sciences, Kasumi 1-2-3, Minami-ku, Hiroshima 734-8551, Japan. Email: takashi-kurashige@hiroshima-u.ac.jp Received 31 March 2012; revised 11 August 2012 and accepted 12 August 2012. Neuropathology 2012; ••, ••–•• doi:10.1111/j.1440-1789.2012.01346.x © 2012 Japanese Society of Neuropathology