A new model system for lipid interactions in stratum corneum vesicles: effects of lipid composition, calcium, and pH.
We prepared large unilamellar vesicles (LUVs) with three different stratum corneum lipid compositions: constant amounts of ceramides (55 wt %) and fatty acids (15%) with varying amounts of cholesterol sulfate (0-15%) and cholesterol (15-30%). One of the compositions served as a model for normal stratum corneum, while the second one served as a model for recessive X-linked ichthyosis stratum corneum. The third composition consisted of no cholesterol sulfate. Intervesicle lipid interactions in these LUVs were monitored by fluorescence methods for content leakage, and contents mixing at pH 9, in the absence and presence of Ca2+, and at pH 6. Since the content leakage and contents mixing assays were originally developed for phospholipid vesicles, we characterized the probe binding and the probe quenching properties for stratum corneum LUV systems, and modified the assays slightly accordingly. The time-dependent fluorescence intensity changes in the probe-containing LUVs at pH 9 and 6 and in response to the addition of calcium were monitored. Our results demonstrated that all three types of LUVs were relatively stable at pH 9. Addition of Ca2+ or decreasing the pH to 6 activated intervesicle lipid mixing followed by vesicle fusion and lysis. We found that the LUVs with no cholesterol sulfate and 30% cholesterol exhibited a more extensive Ca2+- or low-pH-activated intervesicle lipid interaction than LUVs with either 5% cholesterol sulfate and 25% cholesterol or 15% cholesterol sulfate and 15% cholesterol. These results suggest that fusogenic agents such as Ca2+ and H+ act to neutralize the fatty acids in the lipid bilayer of stratum corneum vesicles. The inclusion of 5-15% cholesterol sulfate helps to prevent the collapse of fused vesicles into other structures.
Hatfield
R M
RM
Department of Chemistry, Loyola University of Chicago, Illinois 60626, USA.
Fung
L W
LW
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry* / Q000378:metabolism; D002417:Cattle; D002855:Chromatography, Thin Layer; D005454:Fluorescence Polarization; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007057:Ichthyosis / Q000378:metabolism; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D050356:Lipid Metabolism; D008055:Lipids / Q000737:chemistry; D051379:Mice; D008956:Models, Chemical; D009281:Naphthalenes / Q000737:chemistry; D011726:Pyridinium Compounds / Q000737:chemistry; D012867:Skin / Q000737:chemistry* / Q000378:metabolism; D013050:Spectrometry, Fluorescence
D005456:Fluorescent Dyes; D008051:Lipid Bilayers; D008055:Lipids; D009281:Naphthalenes; D011726:Pyridinium Compounds; C046466:N,N'-4-xylylenebis(pyridinium); C046465:8-amino-1,3,6-naphthalenetrisulfonic acid; D002118:Calcium