Targeting YAP/TAZ mechanoregulation with statins to reverse glaucomatous trabecular meshwork stiffening

Abstract

Purpose : Trabecular meshwork (TM) stiffening in primary open-angle glaucoma (POAG) arises from elevated ROCK-mediated TM cell contraction and extracellular matrix (ECM) deposition/crosslinking. YAP/TAZ play central roles in TM mechanoregulation and increased YAP/TAZ activity has been linked to glaucomatous TM stiffening. Common cholesterol-lowering statins competitively inhibit HMG-CoA reductase (HMGCR), which potently decreases YAP/TAZ activity. Statins also inhibit ROCK activity upstream of YAP/TAZ. Here, we investigate whether statins can reverse glaucomatous TM cell dysfunction using a tissue-engineered TM hydrogel model.

Methods : Normal TM/glaucomatous GTM cells were isolated from surgical discard corneal rims/POAG globes. Cells were plated on glass coverslips or encapsulated in photocrosslinked hydrogels (collagen type I, elastin-like polypeptide and hyaluronic acid). Glaucomatous conditions were induced with DEX/TGF-β2, then treated with cerivastatin or simvastatin; GTM cells were used for validation and proven ROCK inhibitor Y27632 served as treatment control. Cell morphology and cytoskeletal organization were assessed by phalloidin staining for f-actin. Immunoblot and immunostaining were used to evaluate YAP/TAZ expression/activity and changes in cell contractile force regulation. TM hydrogel contraction and stiffness were determined by longitudinal imaging and oscillatory rheology.

Results : Induced TM and GTM cells exhibited decreased cytoplasmic (inactive) pYAP and increased nuclear (active) YAP/TAZ with increased downstream TGM2 (an ECM crosslinking enzyme) vs. controls (p<0.01); this was significantly reduced with either statin in a dose- and time-dependent manner. These effects were reversed by supplementation of mevalonate, thus bypassing statin-mediated HMGCR inhibition. Statin treatment restored glaucomatous actin stress fibers, and pMLC/α-SMA levels to baseline. Immunostaining revealed significant statin-mediated reduction of glaucomatous fibronectin deposition. Pathologic TM hydrogel contraction and stiffening were potently rescued with statin treatment; constructs were significantly relaxed and softened (p<0.01), comparable to direct ROCK inhibition.

Conclusions : Our data suggest that indirectly targeting the YAP/TAZ mechanoregulatory axis with statins presents an intriguing avenue for treating glaucomatous TM dysfunction with high translational potential.

This is a 2021 ARVO Annual Meeting abstract.