{"doi":"10.1194/jlr.ra119000571","title":"Monitoring the itinerary of lysosomal cholesterol in Niemann-Pick Type C1-deficient cells after cyclodextrin treatment","abstract":"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-β-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. 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-β-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. Cholesterol is an essential component of mammalian cell membranes that plays a major role in tuning membrane fluidity, thickness, and permeability to regulate membrane function and support the needs of specific organelles. Different cellular membranes vary widely in cholesterol content, ranging from the cholesterol-rich plasma membrane (PM) and endosomes to the cholesterol-poor ER and mitochondria (1Wüstner D. Solanko K. How cholesterol interacts with proteins and lipids during its intracellular transport.Biochim. Biophys. Acta. 2015; 1848: 1908-1926Crossref PubMed Scopus (47) Google Scholar, 2Chen F.W. Li C. Ioannou Y.A. Cyclodextrin induces calcium-dependent lysosomal exocytosis.PLoS One. 2010; 5: e15054Crossref PubMed Scopus (78) Google Scholar, 3Das A. Brown M.S. Anderson D.D. Goldstein J.L. Radhakrishnan A. Three pools of plasma membrane cholesterol and their relation to cholesterol homeostas","journal":"Journal of Lipid Research","year":2020,"id":94073,"datarank":1.3448707785929064,"base_score":3.6635616461296463,"endowment":3.6635616461296463,"self_citation_contribution":0.5495342469194471,"citation_network_contribution":0.7953365316734592,"self_endowment_contribution":0.5495342469194471,"citer_contribution":0.7953365316734592,"corpus_percentile":null,"corpus_rank":null,"citation_count":38,"citer_count":38,"citers_with_citation_signal":33,"citers_with_endowment":33,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.954,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2020-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":469142,"name":"Sarah E. Gale","orcid":"0000-0001-8762-4497","position":1,"is_corresponding":false},{"id":470443,"name":"Samantha Moores","orcid":null,"position":2,"is_corresponding":false},{"id":310078,"name":"Daniel S. Ory","orcid":null,"position":3,"is_corresponding":false},{"id":340224,"name":"Jean E. Schaffer","orcid":"0000-0002-4954-6659","position":4,"is_corresponding":false},{"id":469141,"name":"McKenna Feltes","orcid":"0000-0002-4178-8055","position":0,"is_corresponding":true}],"reference_count":28,"raw_metadata":{"citation_network_status":"fetched"},"created_at":"2026-07-18T22:32:04.778261Z","pmid":"31988149","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}