CHOLESTEROL, PHOSPHOLIPID, AND PROTEIN-CHANGES IN FOCAL OPACITIES IN THE HUMAN EYE LENS

PURPOSE. Focal opacities are signs of early cataractogenesis in the hu man lens. They progress slowly over a lifetime and may be precursors o f mature cataracts. The authors analyzed changes in proteins, phosphol ipids, and cholesterol in these opacities using in situ techniques: Ra man microspectroscopy, filipin cytochemistry for cholesterol, and tran smission electron microscopy (TEM). METHODS. Human lenses with verifie d focal opacities were fixed in 1% paraformaldehyde. Slabs with opacit ies were analyzed using confocal Raman spectroscopy, then filipin Rama n analysis of cholesterol, and finally TEM. RESULTS. Compared with nor mal fibers, opacities consistently showed elevated levels of cholester ol and aliphatic chains, increased phospholipid acyl chain disorder, a nd changes in phospholipid lateral packing. Disulfide bridges of speci fic geometry (trans-gauche-trans) were found. Although protein content was unchanged, compared with normal fibers, aromatic amino acid conte nt was significantly lower. The hydrophobicity of tyrosine residues sh owed a significant decrease, and a change in the tryptophan indole rin g angle was found. The changes were abrupt and sharply delineated foca l opacities. TEM confirmed this sharp boundary and showed that the opa cities were densely packed with vesicles of varying size and electron density embedded in a homogenous matrix. CONCLUSIONS. The Raman and TE M analyses of opacities showed that early cataractogenic events led to disruption of fiber membranes, formation of vesicles from the membran e constituents, and protein changes. The aberrant morphology of the me mbranes enveloping the focal opacities may have segregated the affecte d fibers from the surrounding normal tissue, thus explaining the stati onary or slowly progressing character of these opacities.