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S.I. Tomarev, V.V. Senatorov, I.V. Malyukova, O.V. Grinchuk, E.F. Wawrousek, R.N. Fariss, S. Swaminathan, S. Sharan; Development of a Novel Mouse Model of Glaucoma by the Expression of Mutated Myocilin in the Trabecular Meshwork of Transgenic Mice . Invest. Ophthalmol. Vis. Sci. 2005;46(13):2371.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Previous experiments indicated that myocilin acts by a gain of function mechanism, since a complete absence (Mol. Cell. Biol. 21, 7707) or 10–15 fold overexpression of myocilin (Mol.Cell. Biol. 24, 9019) in the mouse eye led neither to elevation of intraocular pressure (IOP) nor to detectable changes in the retina and optic nerve. We produced transgenic (TG) mice expressing mutated mouse and human myocilin genes to develop a genetic mouse model of glaucoma. Methods: The Tyr423His and Tyr437His point mutations were introduced into BAC DNAs containing the full length mouse and human myocilin genes, respectively, using oligonucleotide–based recombining in E. coli. These BAC DNAs were used to generate TG mice. Expression of the myocilin gene was analyzed by in situ hybridization and by real–time PCR. The distribution of myocilin was investigated in western blotting experiments and by immunohistochemistry. Changes in the mRNA levels from different genes were analyzed by array hybridization. Retinal ganglion cells (RGC) were counted in whole mount retinas and paraffin sections of retina after immunostaining with cell–specific antibodies. Apoptosis was assessed using a TUNEL technique. IOP was measured using a non–invasive fiber–optic pressure transducer. Results:One line of TG animals expressing mutated mouse myocilin was analyzed in more detail. In the eyes of these mice, the ratio of mutated to wild type (WT) myocilin mRNAs was about 1:3. mRNA levels of about 60 genes were changed in the eyes of TG mice, compared to WT littermates. Although no significant changes were detected in the central retina of 6 to 8 month old TG mice, consistent signs of degeneration were found in the peripheral RGC layer of TG animals, as judged by appearance of TUNEL–positive cells. TUNEL–positive cells were not detected in the retinas of WT mice. Immunostaining for RGC markers, Brn3b and NF68, showed the loss of about 20% of RGC in 6 to 12 month old TG animals, compared to WT littermates. 16 month old TG mice had higher IOP than their non–transgenic littermates (13.3 ± 0.4 vs 11.2 ± 0.3 mmHg, P<0.05). Conclusions:Changes in the eyes of TG mice resemble those observed in human glaucoma. These TG mice represent a novel animal model of glaucoma.
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