1 Centre for Dementia Research, Nathan Kline Institute, 140 Old Orangeburg Road, Orangeburg, NY 10962, USA 2 Department of Psychiatry, New York University Langone Medical Centre, 550 First Avenue, New York, NY 10016, USA
1 Centre for Dementia Research, Nathan Kline Institute, 140 Old Orangeburg Road, Orangeburg, NY 10962, USA.
Brain. 2014 Dec;137(Pt 12):3300-18. doi: 10.1093/brain/awu278. Epub 2014 Sep 29.
Autophagy, the major lysosomal pathway for the turnover of intracellular organelles is markedly impaired in neurons in Alzheimer's disease and Alzheimer mouse models. We have previously reported that severe lysosomal and amyloid neuropathology and associated cognitive deficits in the TgCRND8 Alzheimer mouse model can be ameliorated by restoring lysosomal proteolytic capacity and autophagy flux via genetic deletion of the lysosomal protease inhibitor, cystatin B. Here we present evidence that macroautophagy is a significant pathway for lipid turnover, which is defective in TgCRND8 brain where lipids accumulate as membranous structures and lipid droplets within giant neuronal autolysosomes. Levels of multiple lipid species including several sphingolipids (ceramide, ganglioside GM3, GM2, GM1, GD3 and GD1a), cardiolipin, cholesterol and cholesteryl esters are elevated in autophagic vacuole fractions and lysosomes isolated from TgCRND8 brain. Lipids are localized in autophagosomes and autolysosomes by double immunofluorescence analyses in wild-type mice and colocalization is increased in TgCRND8 mice where abnormally abundant GM2 ganglioside-positive granules are detected in neuronal lysosomes. Cystatin B deletion in TgCRND8 significantly reduces the number of GM2-positive granules and lowers the levels of GM2 and GM3 in lysosomes, decreases lipofuscin-related autofluorescence, and eliminates giant lipid-containing autolysosomes while increasing numbers of normal-sized autolysosomes/lysosomes with reduced content of undigested components. These findings have identified macroautophagy as a previously unappreciated route for delivering membrane lipids to lysosomes for turnover, a function that has so far been considered to be mediated exclusively through the endocytic pathway, and revealed that autophagic-lysosomal dysfunction in TgCRND8 brain impedes lysosomal turnover of lipids as well as proteins. The amelioration of lipid accumulation in TgCRND8 by removing cystatin B inhibition on lysosomal proteases suggests that enhancing lysosomal proteolysis improves the overall environment of the lysosome and its clearance functions, which may be possibly relevant to a broader range of lysosomal disorders beyond Alzheimer's disease.
自噬是细胞内细胞器降解的主要溶酶体途径，在阿尔茨海默病和阿尔茨海默病小鼠模型的神经元中明显受损。我们之前曾报道过，在 TgCRND8 阿尔茨海默病小鼠模型中，严重的溶酶体和淀粉样神经病理学以及相关的认知缺陷可以通过遗传缺失溶酶体蛋白酶抑制剂胱抑素 B 来改善溶酶体蛋白水解能力和自噬通量。在这里，我们提供的证据表明，巨自噬是脂质周转的重要途径，在 TgCRND8 大脑中，脂质作为膜结构和巨自噬溶酶体中的脂滴积累，该途径存在缺陷。在 TgCRND8 大脑中，多种脂质物种的水平升高，包括几种鞘脂（神经酰胺、神经节苷脂 GM3、GM2、GM1、GD3 和 GD1a）、心磷脂、胆固醇和胆固醇酯，这些脂质存在于自噬小体和溶酶体分离物的自噬小体部分中。在野生型小鼠的双免疫荧光分析中，脂质定位于自噬体和自噬溶酶体中，在 TgCRND8 小鼠中，神经溶酶体中检测到异常丰富的 GM2 神经节苷脂阳性颗粒，其共定位增加。在 TgCRND8 中缺失胱抑素 B 可显著减少 GM2 阳性颗粒的数量，降低溶酶体中的 GM2 和 GM3 水平，减少脂褐素相关的自发荧光，并消除含有大量脂质的自噬溶酶体，同时增加正常大小的自噬溶酶体/溶酶体，其未消化成分的含量减少。这些发现确定了巨自噬作为一种以前未被认识到的途径，将膜脂质递送到溶酶体进行周转，这一功能迄今为止被认为仅通过内吞作用途径介导，并表明 TgCRND8 大脑中的自噬溶酶体功能障碍会阻碍溶酶体对脂质和蛋白质的周转。通过去除胱抑素 B 对溶酶体蛋白酶的抑制作用，改善 TgCRND8 中的脂质积累表明，增强溶酶体蛋白水解可改善溶酶体的整体环境及其清除功能，这可能与阿尔茨海默病以外的更广泛的溶酶体疾病有关。
