A morphological and biochemical study of argyrophilic inclusions in multiple system atrophy

对6例多系统萎缩（MSA）脑的临床病理学研究显示，在初级运动皮质中存在胶质细胞胞浆内含物（GCI）的精确分布和特征性的层状星形胶质细胞增生，GCI不仅对泛素进行了免疫染色，而且对泛素C-末端水解酶和20 S-/26 S-蛋白酶体。由于少突胶质细胞参与铁代谢，而铁与自由基的形成有关，因此我们对铁、铁蛋白和超氧化物歧化酶（SOD）进行了研究。在MSA运动皮质，铁蛋白和Cu/Zu SOD免疫细胞化学定位GCI内的小胶质细胞和巨噬细胞中检测到三价铁。在生化分析中，GCI通过筛网筛分和不连续蔗糖密度梯度离心从MAS脑中部分纯化。GCI主要集中在45%-50%蔗糖界面，含有32 KDa和40 KDa蛋白质。这些蛋白质被抗泛素和抗α B-胱抑素抗体识别，并且在来自对照脑的相同部分中未发现。这是第一个生化证明α B-cystallin可能是泛素化的，作为GCI的一个组成部分。

Clinicopathological study of 6 brains of multiple system atrophy (MSA) revealed the precise distribution of glial cytoplasmic inclusions (GCIs) and a characteristic, laminar astrogliosis in the primary motor cortex, where a decrease in number of small to medium-sized neurons was found for the first time by a morphometric analysis.GCIs were clearly immunostained for not only ubiquitin but also ubiquitin C-terminal hydrolase and 20S-/ 26S-proteasomes. The finding indictes that the main process of formation and degradation of GCIs is via ATP-dependent, non-lysosomal pathway.Since oligodendroglia are involved in iron metabolism and iron is related to free radical formation, we investigated iron, ferritin and superoxide dismutase (SOD). In the MSA motor cortex, ferric iron was detected in microglia and macrophages with immunocytochemical localization of ferritin and Cu/Zu SOD within GCIs. These facts suggest first that GCIs might be related to neural death via oxidative stress.In biochemical analysis, GCIs were partially purified from the MAS brain by sieving with mesh and discontinuous sucrose density gradient centrifugation. GCIs were mainly collected in the 45% to 50% sucrose interface which contained 32KDa and 40KDa proteins. These proteins were recognized with both anti-ubiquitin and anti-alpha B-cystallin antibodies, and not found in the identical fraction from the control brain. This is the first biochemical demonstration of alpha B-cystallin which might be ubiquitinated, as a component of GCI.