Abstract
Purpose :
Primary open angle glaucoma (POAG) is the most common form of glaucoma. Pathology changes of trabecular meshwork (TM) in POAG is largely responsible for the increased outflow resistance of aqueous humor, which will result in IOP elevation. Morphological and biochemical analyses of the TM of POAG patients revealed loss of cells, increased accumulation of extracellular matrix (ECM), changes in the cytoskeleton but the molecular mechanisms that drive remodeling in POAG TM remain poorly understood. Here, we investigated the plasticity of known mechanosensitive pathways in cells subjected to mechanical stress and glaucoma.
Methods :
TM cells were obtained from healthy and POAG patients. Primary cells were stimulated with biaxial strains (6%) for 1-3 hours and/or exposed to TRPV4 agonists & antagonists or modulators of the Rho signaling pathway. Genes/proteins encoding mechanosensitive channels, cell specific markers, cytoskeletal/focal adhesion, ECM and cell-cell adhesion components were tracked and measured in TM with qRT-PCR, Western blots and immunohistochemistry. [Ca2+]TM levels using standard optical imaging.
Results :
Glaucomatous TM cells exhibited altered transcript and protein expression of putative mechanosensitive channels, myocilin and focal adhesion (FA) proteins (e.g., Focal Adhesion Kinase (FAK), vinculin) compared to healthy cells. Radial stretch time-dependently upregulated transcript and protein levels of mechanosensors and FA proteins, with differential effects observed in POAG samples. Immunolabeling and Ca2+-imaging experiments revealed that some of the glaucomatous TM exhibited elevated protein levels of fibronectin and Zonula occludens-1 (ZO1) and decreased phalloidin signal, yet others showed similar amplitudes and responder percentage with increased fibronectin and phalloidin, with lower level of ZO1.
Conclusions :
We confirmed strong expression of mechanosensitive proteins and pathways in human TM cells and documented a range of expression differences between healthy and POAG samples. Overall, stretch-activated channels such as TRPV4, ECM proteins (fibronectin, ZO1) and the cytoskeleton show complex, stage-dependent responses to TM/SC remodeling. The dysregulation of these mechanisms in POAG could contribute to dynamic and chronic reorganization of conventional outflow resistance.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.