Fluorescence studies on the age related changes in bovine and human lens membrane structure

Bovine and human lens fiber cell plasma membranes were isolated as the urea-insoluble fraction for a study of age related changes. The changes in fluorescence intensity, both intrinsic (tryptophan) and extrinsic probes ANS (1-anilinonaphthalene-8-sulfonic acid) and DPH (1,6-diphenyl 1,3,5-hexatriene), as well as DPH anisotropy and lifetime were measured. The results of tryptophan fluorescence indicate that tryptophan residues in membrane proteins are in a very hydrophobic environment and do not show a change with aging. ANS reacts with surface protein in the polar-apolar interface, while DPH penetrates into the interior of membranes. Both probes show a decrease in fluorescence intensity in the old membranes. The decrease in ANS fluorescence may result from the conversion of MP26 to MP22, while the decrease of DPH fluorescence intensity may indicate a decrease in accessibility of lipid to DPH. To further delineate the change, fluorescence anisotropy and lifetime data of the lipid probe DPH were obtained. While lifetimes do not change with age, anisotropy shows a definite age-dependent increase. Anisotropy is related to the degree of lipid structural order. Greater anisotropy values were found for older membrane samples, indicating an increased rigidity with age, which may be partially caused by the increased cholesterol/phospholipid (C/P) ratio as reported in the literature.