Eye / Vision
Permanent URI for this collectionhttps://hdl.handle.net/20.500.12503/31256
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Browsing Eye / Vision by Subject "glaucoma"
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Item EFFECT OF TRANSFORMING GROWTH FACTOR BETA-2 SIGNALING AND GREMLIN INDUCTION ON FIBRONECTIN, OCULAR HYPERTENSION, AND OPTIC NERVE DAMAGE(2013-04-12) McDowell, ColleenPurpose: 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.Item EFFECTS OF TGF-BETA2 ON THE ELASTIC MODULUS OF TRABECULAR MESHWORK TISSUE(2013-04-12) Baradia, HusseinPurpose: The primary risk factor for developing glaucoma is elevated intraocular pressure (IOP.) The key outflow passage for aqueous humor (AH) in the human eye involves the trabecular meshwork (TM) and the Schlemm's canal (SC) with the main regulator of IOP being at the junction of the two. Elevated IOP has been attributed to increased resistance to flow at the TM/SC junction. Aqueous humor levels of transforming growth factor beta-2 (TGF-b2) are elevated in glaucoma patients, suggesting its contribution to the progression of the disease. Studies using cultured TM cells as well as ex vivo tissue have shown that TGF-b2 induces extracellular matrix (ECM) remodeling. A separate study used atomic force microscopy (AFM) to measure the elastic modulus (e.g. stiffness) of TM tissue obtained from glaucoma patients compared to age-matched controls. A marked increase in stiffness in the glaucoma tissue was observed compared to non-glaucomatous controls. Taken together, these findings imply that an elevated level of TGF-b2 may lead to ECM remodeling and increased stiffness thus reducing AH outflow and elevating IOP. The hypothesis of this study was that increased expression of TGF-b2 will cause increased stiffness (e.g. increased elastic modulus) of the TM. The goal of our study was to compare the stiffness of the TM cultured overnight with or without TGF-b2. Methods: Bovine eyes were obtained from an abattoir and the TM dissected from the anterior segment. The dissected TM was then cut into halves. One half was cultured in control media and the other half cultured in media containing TGF-b2 (5 ng/ml). Both TM halves were incubated at 37oC for a period of 72 hours. The TM was then loaded onto a force transducer and contraction induced with carbachol to measure stiffness. Results: The raw data obtained indicated that the TM exposed to TGF-b2 contracted with greater force than control indicating that TGF-b2 reduced the stiffness of the TM. However, using a Students T test at a p of 0.05 the results showed no statistical significance. Conclusions: The results obtained are from one experiment with an N =8 and since no statistical significance was observed, the data has to be considered inconclusive. The force transducer used was actually that designed for skeletal muscles which may not be the best system to test trabecular meshwork which has some smooth muscle components. The future goal of the study is to actually use force microscopy, the current standard for such experiments.