Glucocorticoid Induction of the Glaucoma Gene MYOC in Human and Monkey Trabecular Meshwork Cells and Tissues

• Published in: Investigative ophthalmology & visual science Vol. 42; no. 8; pp. 1769 - 1780
• Main Authors: Clark, Abbot F, Steely, H. Thomas, Dickerson, Jaime E., Jr, English-Wright, Sherry, Stropki, Karen, McCartney, Mitchell D, Jacobson, Nasreen, Shepard, Allan R, Clark, John I, Matsushima, Hiroyuki, Peskind, Elaine R, Leverenz, James B, Wilkinson, Charles W, Swiderski, Ruth E, Fingert, John H, Sheffield, Val C, Stone, Edwin M
• Format: Journal Article
• Language: English
• Published: Rockville, MD ARVO 01.07.2001; Association for Research in Vision and Ophtalmology
• Subjects: Aged; Aged, 80 and over; Animals; Biological and medical sciences; Blotting, Northern; Blotting, Western; Cells, Cultured; Cortisone - analogs & derivatives; Cortisone - pharmacology; Cytoskeletal Proteins; Dexamethasone - pharmacology; Electrophoresis, Gel, Two-Dimensional; Eye Proteins - biosynthesis; Eye Proteins - genetics; Fluorescent Antibody Technique, Indirect; Gene Expression - drug effects; Glaucoma and intraocular pressure; Glucocorticoids - pharmacology; Glycoproteins - biosynthesis; Glycoproteins - genetics; Humans; Intraocular Pressure - drug effects; Macaca nemestrina; Medical sciences; Middle Aged; Ocular Hypertension - chemically induced; Ocular Hypertension - metabolism; Ocular Hypertension - pathology; Ophthalmology; RNA, Messenger - biosynthesis; Trabecular Meshwork - drug effects; Trabecular Meshwork - metabolism; Trabecular Meshwork - ultrastructure; Human; Cell culture; Open angle glaucoma; Steroid hormone; Pathogenesis; Ocular hypertension; Monkey; Gene expression; Glucocorticoid; Protein; Eye; Eye disease; Vertebrata; Myocilin; Mammalia; Gene; Animal; Adrenal hormone; Primates; Trabecular Meshwork
• ISSN: 0146-0404; 1552-5783

To examine the intracellular and extracellular expression of myocilin in the human and primate trabecular meshwork (TM) in the presence and absence of glucocorticoids. 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 radiolabeled proteins from human TM cells, TM tissue explants, and perfused human anterior segments cultured with and without dexamethasone (DEX) for 14 to 21 days, as well as TM tissue from pigtailed monkeys treated orally for 1 year with cortisone acetate. Immunofluorescence with anti-myocilin antibodies was used to localize cellular and extracellular expression of myocilin in cultured human TM cells. Glucocorticoid treatment caused a significant induction of myocilin mRNA, a tetrad of cell-associated proteins, and 8 to 20 secreted proteins (molecular mass [M(r)] 56 and 59 kDa and isoelectric point [pI] 5.2 and 5.3) in some, 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 1 year with cortisol acetate showed steroid glaucoma-like morphologic 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 (approximately 57 kDa) to the lower molecular weight isoforms ( approximately 55 kDa). 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 POAG.