The transbilayer cholesterol distribution of human erythrocytes was examined by two independent techniques, quenching of dehydroergosterol fluorescence and fluorescence photobleaching of NBD-cholesterol. Dehydroergosterol in conjunction with leaflet selective quenching showed that, at equilibrium, 75% of the sterol was localized to the inner leaflet of resealed erythrocyte ghosts and intact erythrocytes. The fractional contribution of the fast and slow diffusion values in both resealed ghosts and intact erythrocytes. The fractional contribution of the fast and slow diffusion constants of NBD-labelled cholesterol represent its inner and outer leaflet distribution. At room temperature the plasma membrane inner leaflet of erythrocyte ghosts as well as intact erythrocytes cells contained 78% of the plasma membrane sterol. The erythrocyte membrane transbilayer distribution of sterol was independent of temperature. In conclusion, dehydroergosterol and NBD-cholesterol data are consistent with an enrichment of cholesterol in the inner leaflet of the human erythrocyte.
Bibliographical noteFunding Information:
This work was supportedin part by grantsf romthe USPHS (DK41402a nd ,~07292)a nd by the Medical ResearchS ervicea nd theG eriatricR esearchE, duca-
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- Fluorescence photobleaching
- Fluorescence quenching
- Membrane domain