NPC1L1-dependent transport of 27-alkyne cholesterol in intestinal epithelial cells
| Autor: | Daesung Lee, Seema Saksena, Waddah A. Alrefai, Pradeep K. Dudeja, Pooja Malhotra, Ravinder K. Gill, Arivarasu Natarajan Anbazhagan, Nathan Calzadilla, Hyunjin Lee, Alexander L. Ticho |
|---|---|
| Rok vydání: | 2021 |
| Předmět: |
0301 basic medicine
Drug Physiology media_common.quotation_subject Alkyne 030204 cardiovascular system & hematology 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine Ezetimibe medicine Animals Humans Intestinal Mucosa media_common chemistry.chemical_classification Cholesterol Anticholesteremic Agents Membrane Transport Proteins Biological Transport Epithelial Cells Cell Biology Endocytosis Mice Inbred C57BL 030104 developmental biology chemistry Biochemistry Intestinal Absorption Intestinal cholesterol absorption Click chemistry Molecular targets lipids (amino acids peptides and proteins) Caco-2 Cells medicine.drug Research Article |
| Zdroj: | Am J Physiol Cell Physiol |
| ISSN: | 1522-1563 |
| Popis: | Niemann-Pick C1 Like-1 (NPC1L1) mediates the uptake of micellar cholesterol by intestinal epithelial cells and is the molecular target of the cholesterol-lowering drug ezetimibe (EZE). The detailed mechanisms responsible for intracellular shuttling of micellar cholesterol are not fully understood due to the lack of a suitable NPC1L1 substrate that can be traced by fluorescence imaging and biochemical methods. 27-Alkyne cholesterol has been previously shown to serve as a substrate for different cellular processes similar to native cholesterol. However, it is not known whether alkyne cholesterol is absorbed via an NPC1L1-dependent pathway. We aimed to determine whether alkyne cholesterol is a substrate for NPC1L1 in intestinal cells. Human intestinal epithelial Caco2 cells were incubated with micelles containing alkyne cholesterol in the presence or absence of EZE. Small intestinal closed loops in C57BL/6J mice were injected with micelles containing alkyne cholesterol with or without EZE. Alkyne cholesterol esterification in Caco2 cells was significantly inhibited by EZE and by inhibitor of clathrin-mediated endocytosis Pitstop 2. The esterification was similarly reduced by inhibitors of the acyl-CoA cholesterol acyltransferase (ACAT). Alkyne cholesterol efficiently labeled the apical membrane of Caco2 cells and the amount retained on the membrane was significantly increased by EZE as judged by accessibility to exogenous cholesterol oxidase. In mouse small intestine, the presence of EZE reduced total alkyne cholesterol uptake by ∼75%. These data show that alkyne cholesterol acts as a substrate for NPC1L1 and may serve as a nonradioactive tracer to measure cholesterol absorption in both in vitro and in vivo models. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|