Confocal Microscopy Confirmed that in Phosphatidylcholine Giant Unilamellar Vesicles with very High Cholesterol Content Pure Cholesterol Bilayer Domains Form

被引:11
作者
Raguz, Marija [1 ,2 ]
Kumar, Suresh N. [3 ]
Zareba, Mariusz [2 ,4 ]
Ilic, Nada [5 ]
Mainali, Laxman [2 ,6 ]
Subczynski, Witold K. [2 ]
机构
[1] Univ Split, Sch Med, Dept Med Phys & Biophys, Split, Croatia
[2] Med Coll Wisconsin, Dept Biophys, Milwaukee, WI 53226 USA
[3] Med Coll Wisconsin, Dept Pathol, CRI Imaging Core, Translat & Biomed Res Ctr, Wauwatosa, WI USA
[4] Med Coll Wisconsin, Inst Eye, Dept Ophthalmol, 8700 W Wisconsin Ave, Milwaukee, WI 53226 USA
[5] Univ Split, Dept Phys, Fac Sci, Split, Croatia
[6] Boise State Univ, Dept Phys, 1910 Univ Dr, Boise, ID 83725 USA
基金
美国国家卫生研究院;
关键词
Confocal microscopy; Cholesterol; Cholesterol bilayer domain; Membrane; Giant unilamellar vesicles; CELL PLASMA-MEMBRANES; OXYGEN PERMEABILITY; LIPID DOMAINS; LENS LIPIDS; EPR; PHASE; TRANSPORT; CATARACT; AGE; MANIFESTATIONS;
D O I
10.1007/s12013-019-00889-y
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The cholesterol (Chol) content in the fiber cell plasma membranes of the eye lens is extremely high, exceeding the solubility threshold in the lenses of old humans. This high Chol content forms pure Chol bilayer domains (CBDs) and Chol crystals in model membranes and membranes formed from the total lipid extracts from human lenses. CBDs have been detected using electron paramagnetic resonance (EPR) spin-labeling approaches. Here, we confirm the presence of CBDs in giant unilamellar vesicles prepared using the electroformation method from Chol/1-palmitoyl-2-oleoylphosphocholine and Chol/distearoylphosphatidylcholine mixtures. Confocal microscopy experiments using phospholipid (PL) analog (1,1 '-dioctadecyl-3,3,3 ',3 '-tetramethylindocarbocyanine-5,5 '-disulfonic acid) and cholesterol analog fluorescent probes (23-(dipyrrometheneboron difluoride)-24-norcholesterol) were performed, allowing us to make three major conclusions: (1) In all membranes with a Chol/PL mixing ratio (expressed as a molar ratio) >2, pure CBDs were formed within the bulk PL bilayer saturated with Chol. (2) CBDs were present as the pure Chol bilayer and not as separate patches of Chol monolayers in each leaflet of the PL bilayer. (3) CBDs, presented as single large domains, were always located at the top of giant unilamellar vesicles, independent of the change in sample orientation (right-side-up/upside-down). Results obtained with confocal microscopy and fluorescent Chol and PL analogs, combined with those obtained using EPR and spin-labeled Chol and PL analogs, contribute to the understanding of the organization of lipids in the fiber cell plasma membranes of the human eye lens.
引用
收藏
页码:309 / 317
页数:9
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