advertisement
Abstract #6297 Published in IGR 3-2
Glucocorticoid induction of the glaucoma gene MYOC in human and monkey trabecular meshwork cells and tissues
Clark AF; Steely HT; Dickerson JE Jr; English-Wright S; Stropki K; McCartney MD; Jacobson N; Shepard AR; Clark JI; Matsushima HInvestigative Ophthalmology and Visual Science 2001; 42: 1769-1780
PURPOSE: To examine the intracellular and extracellular expression of myocilin in the human and primate trabecular meshwork (TM) in the presence and absence of glucocorticoids. METHODS: Myocilin expression was examined in cultured human TM cells by Northern blot analysis and myocilin antibody-mediated immunoprecipitation. Myocilin expression was quantified using high-resolution two-dimensional polyacrylamide gel electrophoresis of rabiolabelled proteins from human TM cells, TM tissue explants, and perfused human anterior segments cultured with and without dexamethasone (DEX) for 14-21 days, as well as TM tissue from pigtailed monkeys treated orally for one year with cortisone acetate. Immunofluorescence with anti-myocilin antibodies was used to localize cellular and extracellular expression of myocilin in cultured human TM cells. RESULTS: Glucocorticoid treatment caused a significant induction of myocilin mRNA, a tetrad of cell-associated proteins, and eight to 20 secreted proteins (molecular mass (Mr 56 and 59 kDa and isoelectric point (pI) 5.2 and 5.3) in some of, but not all, the cultured human TM cells and explanted tissues. Western immunoblot analysis using anti-myocilin peptide antibodies identified these proteins as encoded by the MYOC gene. There was significant induction of the myocilin proteins in three perfusion-cultured human eyes, in which DEX-induced elevated intraocular pressure developed. Monkeys treated for one year with cortisol acetate showed steroid glaucoma-like morphological changes in the TM that correlated with the induction of myocilin in the TM. Immunofluorescence analysis of cultured TM cells localized myocilin intracellularly in discrete perinuclear and cytoplasmic vesicular deposits, as well as extracellularly on the cell surface associated with the extracellular matrix. In several DEX-treated TM cell lines, there were significant levels of myocilin secreted into the media. Enzymatic deglycosylation of proteins in the TM media converted the higher molecular weight isoforms of myocilin (~57 kDa) to the lower molecular weight isoforms (~55 kDa). CONCLUSIONS: Although the function of myocilin is unknown, induction of these TM proteins was found in eyes in which glucocorticoid-induced ocular hypertension developed. Therefore, myocilin may play an important pathogenic role in ocular hypertension, in addition to its role in certain forms of primary open-angle glaucoma.
Dr A.F. Clark, Glaucoma Research R2-41, Alcon Research Ltd., 6201 South Freeway, Forth Worth, TX 76135, USA. abe.clark@alconlabs.com
Classification:
1.2 Population genetics (Part of: 1 General aspects)
