Monitoring the itinerary of lysosomal cholesterol in Niemann-Pick Type C1-deficient cells after cyclodextrin treatment
Monitoring the itinerary of lysosomal cholesterol in Niemann-Pick Type C1-deficient cells after cyclodextrin treatment
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DOI：
10.1194/jlr.ra119000571
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发表时间：
2020-03-01
影响因子：
6.5
通讯作者：
Schaffer, Jean E.
中科院分区：
文献类型：
--
作者：
Feltes, McKenna;Gale, Sarah E.;Schaffer, Jean E.
Niemann-Pick病C型(NPC)是一种脂质储存障碍，由NPC蛋白编码基因突变引起，并导致溶酶体胆固醇积聚。2-羟丙基-β-环糊精(CD)已被证明能降低溶酶体胆固醇水平并增强固醇稳态反应，但CD的作用机制尚不清楚。最近的研究提供了Cd刺激溶酶体胞吐的证据，增加了溶酶体胆固醇在外切体中释放的可能性。然而，治疗浓度的镉不会改变细胞总胆固醇，内质网的胆固醇稳态反应与内质网膜胆固醇的增加最为一致。为了解决这些不同的发现，我们在这里使用稳定同位素标记来跟踪NPC1缺陷的U2OS细胞中脂蛋白胆固醇货物对Cd的响应。尽管可以检测到释放的胆固醇，但它与细胞外小泡无关。相反，我们证明了溶酶体胆固醇运输到质膜(PM)，在那里它以CD依赖的方式与脂蛋白结合的胆固醇交换。我们发现，在没有合适的细胞外胆固醇受体的情况下，胆固醇交换被取消，胆固醇在PM中积累，内质网的再酯化增加。这些结果支持一种模型，在该模型中，在恢复胆固醇稳态反应期间，Cd促进溶酶体胆固醇在细胞内的重新分布，但不促进胆固醇的胞吐或外流。
Niemann-Pick disease type C (NPC) disease is a lipid-storage disorder that is caused by mutations in the genes encoding NPC proteins and results in lysosomal cholesterol accumulation. 2-Hydroxypropyl-beta-cyclodextrin (CD) has been shown to reduce lysosomal cholesterol levels and enhance sterol homeostatic responses, but CD's mechanism of action remains unknown. Recent work provides evidence that CD stimulates lysosomal exocytosis, raising the possibility that lysosomal cholesterol is released in exosomes. However, therapeutic concentrations of CD do not alter total cellular cholesterol, and cholesterol homeostatic responses at the ER are most consistent with increased ER membrane cholesterol. To address these disparate findings, here we used stable isotope labeling to track the movement of lipoprotein cholesterol cargo in response to CD in NPC1-deficient U2OS cells. Although released cholesterol was detectable, it was not associated with extracellular vesicles. Rather, we demonstrate that lysosomal cholesterol trafficks to the plasma membrane (PM), where it exchanges with lipoprotein-bound cholesterol in a CD-dependent manner. We found that in the absence of suitable extracellular cholesterol acceptors, cholesterol exchange is abrogated, cholesterol accumulates in the PM, and reesterification at the ER is increased. These results support a model in which CD promotes intracellular redistribution of lysosomal cholesterol, but not cholesterol exocytosis or efflux, during the restoration of cholesterol homeostatic responses.