The goal of this project is to identify the events involved in lens fiber cell differentiation and to use this information to understand the basis of lens cataract formation. We will purify to homogeneity and prepare antibodies against lentropin, a protein factor found in vitreous humor which causes embryonic lens epithelial cells differentiate into lens fiber cells. Lentropin has already been purified greater than 1,000X and its basis properties described. Antibodies to lentropin will be used to identify the cells responsible for secreting lentropin and to prepare antibody affinity columns for bulk purification of lentropin. Using labelled lentropin preparations, the lentropin receptor on lens epithelial cells will be identified and quantitated. Preliminary steps will be taken to characteriza and purify the receptor. When lentropin binds to lens epithelial cells there is a rapid stimultion of phospholipid methylation which appears essential for further fiber cell differentiation. Several agents which alter the metabolism of phospholipids or their fatty acid components can block lentropin-stimulated cell differentiation or cause differentiation in the absence of lentropin. The phospholipid metabolites resulting from lentropin stimulation will be identified, isolated and tested for their ability to affect fiber cell formation. Lens cell elongation during fiber differentiation appears to be caused by an increase in cell volume. The molecular mechanisms responsible for this volume regulation will be examined with respect to changes in transmembrane transport of small molecules and the role of protein synthesis in volume regulation. Since cell volume regulation plays an important role in lens fiber formation, changes in cell volume during x-ray cataract formation have been measured. Lens epithelial cells increase in volume prior to fiber cell swelling and cataract formation. The importance of this increase in volume will be determined by studies that will diminish the possible role of inflammation in this phenomonon. In addition, tests will be performed to establish whether damage to dividing epithelial cells for disruption of fiber cell differentiation is the primary cause of radiation cataracts.
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