Development of appropriate animal models of glaucoma is a critical step for understanding the molecular mechanisms of this blinding disease. We are developing a novel mouse model of glaucoma using a transgenic approach. It is now well established that mutations in the myocilin gene may lead to juvenile open-angle glaucoma and in some cases to adult onset glaucoma. The most severe mutations in this gene interfere with secretion of the encoded protein, and may compromise a secretory pathway in the tissues of the eye angle. Previous experiments demonstrated that eye morphology and intraocular pressure were normal in myocilin-null mice and in mice overexpressing myocilin in the eye tissues. We demonstrated that a BAC clone, containing the mouse myocilin gene 20 kb of 5?- flanking and 85 kb of 3?-flanking sequence, reproduced well the expression pattern of the endogenous myocilin gene. Since the Tyr437His mutation in human myocilin correlates with the most severe cases of glaucoma, a similar mutation has been introduced into the mouse protein. While mouse and human wild-type myocilin were secreted from transfected cells, mutants were not secreted. The Tyr437His point-mutation was introduced into the above mentioned BAC clone (in collaboration with Dr. S. Sharan, NCI), and several transgenic lines containing the mutated BAC clone were created. Two transgenic lines containing the full-size BAC clone insert are currently under investigation. One of them contains 1-2 copies of the mutated BAC clone, while the second line contains 20-30 copies. Changes in the expression pattern of myocilin in the eyes of transgenic animals may lead to morphological changes and changes in intraocular pressure. These effects are now under investigation. We continue to study properties of a novel olfactomedin-containing protein, optimedin, which we previously identified. We have demonstrated that optimedin is a secreted protein, and that mutations in this gene may prevent secretion of the encoded protein. We have shown that optimedin may interact with several olfactomedin-containing proteins including myocilin, noelin-1, and a latrotoxin receptor, Lec3. It may also interact with thioredoxin 2, as judged by yeast two-hybrid screen (in collaboration with Dr. K. Mitton, Oakland University). Changes in the gene expression levels in the human glaucomatous retina were studied using Affimetrix arrays. These changes were compared with those observed in the retina of rats with experimentally elevated IOP (in collaboration with Dr. J. Morrison, Casey Eye Institute). Analysis of these data may lead to the identification of common features, as well as differences between glaucoma in humans and in a rat model of glaucoma. For example, similar changes in the expression levels of gamma-synuclein were observed in a rat model of glaucoma and in some glaucoma patients. Characterization of gene expression in the human trabecular meshwork allowed the identification of several candidate genes for glaucoma.
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