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Abstract #12050 Published in IGR 7-1
Overexpression and properties of wild-type and Tyr437His mutated myocilin in the eyes of transgenic mice
Zillig M; Wurm A; Grehn FJ; Russell P; Tamm ERInvestigative Ophthalmology and Visual Science 2005; 46: 223-234
See also comment(s) by Douglas Johnson • Beatrice Yue •
PURPOSE: To obtain experimental in vivo information on the functional properties of myocilin for aqueous humor (AH) outflow and to study in vivo the processing of mutated Tyr437His myocilin. Myocilin is a secreted glycoprotein that is mutated in some forms of primary open-angle glaucoma (POAG), and patients with the Tyr437His mutation have severe phenotypes. METHODS: The chicken betaB1-crystallin promoter was used to overexpress wild-type human myocilin and mutated Tyr437His myocilin in the lenses of transgenic mice. Expression of transgenic mRNA was monitored by Northern blot analysis and in situ hybridization. The localization and secretion of transgenic myocilin was investigated by Western blot analysis and light and electron microscopy. Intraocular pressure (IOP) was measured by anterior chamber cannulation. RESULTS: Two independent lines were established with each of the constructs that showed a strong expression of transgenic mRNA in their lenses. Transgenic expression resulted in a 4.7 ± 1.8-fold increase of secreted normal myocilin in mouse AH, compared with its concentration in human AH. Immunoreactivity for transgenic myocilin was observed along the surfaces of lens and corneal endothelium, and in the chamber angle. At 12 weeks of age, the ultrastructure of the trabecular meshwork in mice expressing normal myocilin was not different from that of control eyes, and IOP of transgenic animals did not significantly differ from that of control littermates. In contrast, mutated Tyr437His myocilin was not secreted from lens fibers, but accumulated in dilated cisterns of rough endoplasmic reticulum. Although no structural changes were observed in lenses of animals expressing normal myocilin, lenses with Tyr437His expression developed nuclear cataracts, completely lost transparency, and eventually ruptured. Structural changes in lenses of Tyr437His expressing mice were correlated with a significant increase in IOP. CONCLUSIONS: The results do not support the concept that increasing amounts of myocilin in the outflow tissues obstruct the system and directly cause an increase in outflow resistance. Mutated Tyr437His myocilin is not secreted in vivo and causes severe alterations of cellular structure and function. A comparable mechanism may cause POAG in patients with myocilin mutations.
Dr. M. Zillig, Department of Anatomy, Molecular Anatomy and Embryology, University of Erlangen-Nurnberg, Erlangen, Germany
Classification:
5 Experimental glaucoma; animal models
3.5 Molecular biology incl. SiRNA (Part of: 3 Laboratory methods)
2.6.2 Outflow (Part of: 2 Anatomical structures in glaucoma > 2.6 Aqueous humor dynamics)
