Role of 6-O-α-maltosyl-β-cyclodextrin in lysosomal cholesterol deprivation in Npc1-deficient Chinese hamster ovary cells. (2nd January 2018)

• Record Type: Journal Article
• Authors: Okada, Yasuyo; Ueda, Erika; Kondo, Yuki; Ishitsuka, Yoichi; Irie, Tetsumi; Higashi, Taishi; Motoyama, Keiichi; Arima, Hidetoshi; Matuso, Muneaki; Higaki, Katsumi; Ohno, Kousaku; Nishikawa, Junichi; Ichikawa, Atsushi
• Is Part Of: Carbohydrate research. Volume 455(2017)
• Page Start: 54
• Page End: 61
• Publication Date: 2018-01-02
• Subjects: 6-O-α-maltosyl-β-cyclodextrin (Mal-βCD) -- Npc1-deficient cells -- Lysosomal unesterified cholesterol -- Cholesterol/efflux -- Endocytosis -- Liquid chromatography-mass spectrometry; βCD β-cyclodextrin -- CHO Chinese hamster ovary -- DAOS (hydroxy-3-sulfopropyl)-3, 5-dimethoxyaniline sodium salt -- Glc-βCD 6-O-α-D-glucosyl-βCD -- HP-βCD 2-hydroxypropyl-β-cyclodextrin -- LDH lactate dehydrogenase -- Mal-βCD 6-O-α-maltosyl-βCD -- NPC Niemann-Pick Type C; Carbohydrates -- Periodicals; Chemistry, Organic -- Periodicals; Biochemistry -- Periodicals; Carbohydrates -- Periodicals; Chimie organique -- Périodiques; Glucides -- Périodiques; Biochemistry; Carbohydrates; Chemistry, Organic; Periodicals; Electronic journals; 507.78
• DOI: 10.1016/j.carres.2017.11.003
• Languages: English
• ISSNs: 0008-6215

Abstract

Abstract: We aimed to investigate whether 6- O -α-maltosyl-β-cyclodextrin (Mal-βCD) is incorporated into cells and lysosomes during the release of unesterified cholesterol in Npc1- deficient Chinese hamster ovary (CHO) cells ( Npc1 KO cells) and CHO-JP17 cells (JP17 cells). Internalization of Mal-βCD in cells and lysosomes and extracellular release of lysosomal unesterified cholesterol were demonstrated by LC/MS/MS and LC/MS, respectively. Internalization of Mal-βCD was greater in Npc1 KO cells than in JP17 cells. The majority of internalized Mal-βCD in both cell types was metabolized by lysosomal α-glucosidase to 6- O -α-D-glucosyl-β-cyclodextrin (Glc-βCD). However, Mal-βCD did not directly enter the lysosomes prepared from cell homogenates. Mal-βCD-treated Npc1 KO cells and JP17 cells both released Mal-βCD and Glc-βCD, together with unesterified cholesterol, out of cells. The release of unesterified cholesterol by Mal-βCD in Npc1 KO cells was much greater than that in JP17 cells. This study is the first to report the influx of Mal-βCD into the lysosomes of Npc1 KO cells and JP17 cells, followed by generation of Glc-βCD, and the efflux of Mal-βCD/Glc-βCD and unesterified cholesterol to the extracellular fluid, based on the quantitative LC/MS analysis. Graphical abstract: Highlights: 6- O -α-Maltosyl(Mal)-βCD reduces lysosomal cholesterol storage of Npc1 -mutant cells. Mal-βCD is endocytosed, and accumulated in the lysosome of Npc1- mutant cells. Mal-βCD accumulated in the lysosome is partly metabolized to Glc-βCD. Mal/Glc-βCD and cholesterol are released from the lysosome and Npc1- mutant cells. Intracellular behavior of Mal-βCD and cholesterol is quantitated by LC/MS.