Abstract
Purpose: :
To examine membrane fluidity in human lenses of different ages and the factors that influence it.
Methods: :
Equatorial sections from 23 pairs of lenses, aged 22-83 years, were assessed using Laurdan with two-photon confocal microscropy. Laurdan intensity images were converted to Generalised Polarization (GP) images and GP values at specific distances from the central point were obtained. Cholesterol was measured in the contralateral lenses of those used for Laurdan staining. The effect of mild thermal stress on lens membrane fluidity and soluble protein content were also studied. Three lens pairs (30y, 41y, and 63y) were used with one lens of each pair incubated at 50°C and the other not heated. All lenses were sectioned and treated with Laurdan. Gel filtration HPLC was used to analyse soluble crystallins after sectioning. Dihydrosphingomyelin vesicles were pre-loaded with Laurdan and alpha, beta, or gamma crystallins and GP values determined using fluorimetry.
Results: :
Membrane fluidity in the centre of human lenses increased substantially as the lens ages in a manner that correlated with the loss of cytosolic proteins. Furthermore, membrane fluidity could be increased simply by exposing intact human lenses to mild thermal stress; conditions which decrease the concentration of these cytosolic proteins. Vesicles binding experiment showed that alpha and beta, but not gamma, crystallins markedly affected fluidity.
Conclusions: :
Cell membranes from the lens centre become significantly more fluid with age. Alpha and beta crystallins play an important role in regulating the fluidity of cell membranes. Age-dependent loss of these crystallins may affect membrane integrity and contribute to the recognised dysfunction of the lens in older people.
Keywords: aging • crystallins • lipids