MICRODISSECTED CATARACTOUS LENSES

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DESCRIPTION (Applicant's abstract): The long term objective of this proposal is to better understand the biochemical and biophysical changes that occur in the human lens with aging, and which ultimately lead to cataract. The proposed studies will be useful for achieving, in the future, the ultimate goal of significantly retarding the formation of aging-dependent cataract as well as eliminating the formation of secondary cataract which is commonly encountered following cataract surgery. Specific Aim I: To characterize human lens eptithelial proteins: A key player in the maintenance of lens homeostasis is the epithelial layer. Freshly excised human epithelium which will be obtained following cataract surgery will be used. Measurements will be performed on individual epithelium. A portion of the epithelium will be use for enzyme activity determination, whereas the remaining piece will be used for determination of mRNA levels of the specific enzyme using PCR technology. The construction of human epithelial cDNA library will allow us to characterize various human epithelial proteins, and to find if there are lens- specific isoforms for any of the proteins. Specific Aim II: To elucidate the newly discovered molecular chaperone function of alpha-crystallins: Molecular chaperones are very efficient in suppressing aggregation of denatured protein, a property the aging lens needs. Alpha-crystallin was found by us to be a molecular chaperone. This suggests that it has a dual role in the lens. One as a refractive element and a second one as a chaperone protecting other proteins from aggregation. The physical properties of alpha-crystallins will be studied by various techniques. The interaction of alpha-crystallin with other lens proteins such as beta, gamma, and MP-26 will be evaluated. The effects of aging and cataract on the chaperone function of alpha-crystallin will be examined in normal and cataractous human lenses.