TGFβ2-TLR4 Crosstalk Signaling in the Glaucomatous Trabecular Meshwork

dc.contributor.authorCurry, Stacy
dc.contributor.authorClark, Abbot F.
dc.contributor.authorMcDowell, Colleen
dc.creatorRoberts, Amanda
dc.description.abstractPurpose: Glaucoma is a group of optic neuropathies and the leading cause of irreversible blindness worldwide. Primary open angle glaucoma (POAG) is the most prevalent type of glaucoma. Elevated intraocular pressure (IOP) is a major risk factor for the development of POAG. Elevated IOP is caused by aqueous humor fluid not draining properly through the drainage structures in the eye and leads to vision loss. Discovering potential new targets to lower IOP is necessary to develop novel and effective drug therapies. Here we explore a novel molecular mechanism involved in the development of glaucomatous trabecular meshwork (TM) damage. The TM regulates aqueous humor outflow and IOP. The effects of transforming growth factor beta (TGFβ)signaling pathways on the TM’s extracellular matrix (EÇM) have been extensively studied. Recently, we identified TGFβ2 and toll-like receptor 4 (TLR4) signaling crosstalk regulates changes in the TM ECM and mutation in Tlr4 rescues TGFβ2-induced ocular hypertension in mice. Here, we investigated the role of an endogenous TLR4 ligand, FN-EDA, and a downstream signaling molecule of TLR4, NFκB, in TGFβ2-induced ocular hypertension in mice. Methods: B6.FN-EDA+/+, B6.TLR4-/-, B6.FN-EDA-/-, B6.FN-EDA+/+/TLR4-/-, B6.FN-EDA-/-/TLR4-/-, and C57BL/6J mice were intravitreally injected with 2.0μL Ad5.TGFβ2 (2.5x107pfu) in one eye and the contralateral uninjected eye was used as a negative control. Likewise, we tested mice lacking the p50 subunit of NFκB (B6.Cg-NFκB1tm1Bal/J) and C57BL/6J mice. IOP was measured once per week using a TonoLab rebound tonometer on isoflurane-anesthetized mice 42 or 49 days post-injection. Significance determined by one-way ANOVA at each time point. Eyes were harvested, fixed in 4% paraformaldehyde, and sectioned for immunohistochemistry to access total fibronectin and FN-EDA isoform expression. Results: Ad5.TGFβ2significantly induced ocular hypertension in C57BL/6J mice and enhanced ocular hypertension in B6.EDA+/+ mice. Mutations in Tlr4,FN-EDA, and NFκB blocked Ad5.TGFβ2induced ocular hypertension with no significant IOP elevation at any time point. Total FN and FN-EDA isoform expression increased in Ad5.TGFβ2 injected C57BL/6J mice. Data suggest the onset of ocular hypertension developed in uninjected B6.EDA+/+ mice at 14 weeks of age and Ad5.TGFβ2 enhanced elevated IOP levels. Conclusions: TLR4, FN-EDA, and NFkB are necessary for TGFβ2 induced ocular hypertension in mice. In the absence of Ad5.TGFb2 constitutively active EDA (FN-EDA+/+) mice develop ocular hypertension and in the presence of Ad5.TGFβ2 ocular hypertension is enhanced. These data demonstrate that the crosstalk between TGFβ2 and TLR4 is involved in the glaucomatous development within the trabecular meshwork. In addition, it provides potential new targets to lower IOP and to further explore mechanisms involved in the development of glaucomatous TM damage.
dc.titleTGFβ2-TLR4 Crosstalk Signaling in the Glaucomatous Trabecular Meshwork