EFFECT OF TRANSFORMING GROWTH FACTOR BETA-2 SIGNALING AND GREMLIN INDUCTION ON FIBRONECTIN, OCULAR HYPERTENSION, AND OPTIC NERVE DAMAGE

Purpose: Transforming growth factor β2 (TGFβ2) induces extracellular matrix (ECM) remodeling and alters the cytoskeleton, which likely contribute to the inefficient function of the trabecular meshwork (TM) tissue leading to glaucomatous phenotypes. Bone morphogenetic proteins (BMPs) inhibit these profibrotic effects of TGFβ2. The BMP antagonist gremlin is elevated in glaucomatous TM cells and increases intraocular pressure (IOP) in an ex vivo perfusion culture model. The purpose of this study was to determine whether TGFβ2 and gremlin regulate ECM proteins in the TM, induce ocular hypertension, and cause optic nerve damage in mice. Methods: Ad5.hTGFβ226/228 or Ad5.Gremlin (2 uL, 2 X 10^7 pfu) was injected intravitreally into one eye of A/J mice (n=7-13 mice per group), with the uninjected contralateral eye serving as the control eye. Conscious IOP measurements were taken using a TonoLab rebound tonometer. Optic nerve damage was assessed using the optic nerve damage score of PPD stained optic nerve cross sections. TGFβ2, fibronectin, and gremlin protein expression in the TM was determined by immunofluorescence and immunohistochemistry. Transduction of the TM with viral vector Ad5.hTGFβ2226/228 caused a prolonged, reproducible, and statistically significant IOP elevation. IOPs increased to approximately 25 mm Hg for 8 weeks (p<0.001). IOPs were stable (12-15 mm Hg) in the uninjected control eyes. The TGFP2 induced ocular hypertension also caused significant optic nerve damage with optic nerve damage scores (ONDS) > 3 (p<0.001) in the injected eye. Intraocular administration of viral vector Ad5.Gremlin also caused significant IOP elevation in A/J mice for 3 weeks (n=9, injected eye 23.2 +/- 5.6 mmmHg, uninjected eye 15.5 +/- 2.4; p<0.01). In addition, immunofluorescence and immunohistochemistry demonstrated that intraocular injection of Ad5.hTGFβ226/228 and Ad5.Gremlin increased TGFβ2 and fibronectin expression in the TM. Conclusions: These results demonstrate that intravitreal injections of Ad5.hTGFβ226/228 and Ad5.Gremlin in A/J mice elevate IOP and upregulate the ECM protein fibronectin. In addition, Ad5.hTGFβ226/228 expression induced significant optic nerve damage. These data demonstrate for the first time gremlin's role in inducing ocular hypertension in an in vivo model system and emphasize the importance of the TGFβ2 signaling pathway in ocular hypertension.