AIMS Chaperone-mediated autophagy (CMA) is a pathway involved in the autophagy lysosome protein degradation system. CMA has attracted attention as a contributing factor to neurodegenerative diseases since it participates in the degradation of disease-causing proteins. We previously showed that CMA is generally impaired in cells expressing the proteins causing spinocerebellar ataxias (SCAs). Therefore, we investigated the effect of CMA impairment on motor function and the neural survival of cerebellar neurons using the micro RNA (miRNA)-mediated knockdown of lysosome-associated protein 2A (LAMP2A), a CMA-related protein. METHODS We injected adeno-associated virus serotype 9 vectors, which express green fluorescent protein (GFP) and miRNA (negative control miRNA or LAMP2A miRNA) under neuron-specific synapsin I promoter, into cerebellar parenchyma of 4-week old ICR mice. Motor function of mice was evaluated by beam walking and footprint tests. Immunofluorescence experiments of cerebellar slices were conducted to evaluate histological changes of cerebella. RESULTS GFP and miRNA were expressed in interneurons (satellite cells and basket cells) in molecular layers and granule cells in the cerebellar cortices, but not in cerebellar Purkinje cells. LAMP2A knockdown in cerebellar neurons triggered progressive motor impairment, prominent loss of cerebellar Purkinje cells, interneurons, granule cells at the late stage, and astrogliosis and microgliosis from the early stage. CONCLUSIONS CMA impairment in cerebellar interneurons and granule cells triggers the progressive ataxic phenotype, gliosis, and the subsequent degeneration of cerebellar neurons, including Purkinje cells. Our present findings strongly suggest that CMA impairment is related to the pathogenesis of various SCAs.

中文翻译：

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共济失调表型和神经变性是由小脑神经元中伴侣介导的自噬受损引发的

AIMS 分子伴侣介导的自噬 (CMA) 是参与自噬溶酶体蛋白质降解系统的途径。CMA 作为神经退行性疾病的一个促成因素引起了人们的关注，因为它参与了致病蛋白质的降解。我们之前表明，在表达导致脊髓小脑性共济失调 (SCA) 的蛋白质的细胞中，CMA 通常受损。因此，我们使用微 RNA (miRNA) 介导的溶酶体相关蛋白 2A (LAMP2A)（一种 CMA 相关蛋白）的敲低研究了 CMA 损伤对运动功能和小脑神经元神经存活的影响。方法 我们注射腺相关病毒血清型 9 载体，其在神经元特异性突触蛋白 I 启动子下表达绿色荧光蛋白 (GFP) 和 miRNA（阴性对照 miRNA 或 LAMP2A miRNA），进入 4 周龄 ICR 小鼠的小脑实质。通过横梁行走和足迹测试评估小鼠的运动功能。进行小脑切片的免疫荧光实验以评估小脑的组织学变化。结果 GFP 和 miRNA 在小脑皮质分子层的中间神经元（卫星细胞和篮状细胞）和颗粒细胞中表达，但不在小脑浦肯野细胞中表达。小脑神经元中的 LAMP2A 敲低引发了进行性运动障碍，晚期小脑浦肯野细胞、中间神经元、颗粒细胞的显着丧失，以及早期的星形胶质细胞增生和小胶质细胞增生。结论 小脑中间神经元和颗粒细胞中的 CMA 损伤会引发进行性共济失调表型、神经胶质增生以及随后的小脑神经元变性，包括浦肯野细胞。我们目前的研究结果强烈表明 CMA 损伤与各种 SCA 的发病机制有关。