Activation of canonical Wnt signaling pathway inhibits dexamethasone-induced glucocorticoid receptor signaling in the trabecular meshwork

Abstract

Purpose : Dexamethasone (Dex) is frequently used to treat inflammation. Though Dex has many advantages, it has side effects including elevated intraocular pressure (IOP) and glaucoma. About 40% of general population may develop ocular hypertension (OHT) upon prolonged glucocorticoid treatment which is due to damages in the trabecular meshwork (TM). This type of secondary glaucoma is called glucocorticoid-induced glaucoma (GIG). Identifying the key molecules responsible for glucocorticoid-induced OHT and GIG is crucial since they can be used to develop novel glucocorticoids without causing side effects. Our published studies using perfusion cultured bovine eyes suggested that inhibition of the canonical Wnt signaling pathway promotes glucocorticoid-induced OHT and glucocorticoid responsiveness. Therefore, we hypothesize that activation of canonical Wnt signaling prevents the side effects of glucocorticoids.

Methods : Aqueous humor and TM tissues were collected from patients and donor eyes for analyzing DKK1 expression using ELISA and immunofluorescence, respectively. Luciferase assays were performed in GTM3 and NTM5 cells using the GRE or TCF reporter kit to study the activity of the glucocorticoid receptor (GR) signaling or Wnt signaling pathway. Primary human TM (HTM) cells were treated with or without Dex, Wnt3a and/or GSK3β inhibitors, and whole cell lysates and conditional medium were used for western immunoblotting (WB).

Results : We found elevated canonical Wnt signaling inhibitor Dkk1 in the aqueous humor and TM of glaucoma patients. At signaling level, we found that Dkk1 positively regulated GR signaling. We also observed that Wnt signaling activators decreased GR signaling in HTM cells using luciferase assays. Similarly, activation of GR signaling inhibited Wnt signaling. At protein level, Dex-induced extracellular matrix (ECM) was inhibited by Wnt activation using Wnt activators or Dkk1 knockdown in primary HTM cells. In contrast, inhibition of canonical Wnt signaling by β-catenin knockdown increased Dex-induced ECM proteins. At physiological level, we found that adenovirus-mediated Wnt3a expression decreased Dex-induced OHT in mouse eyes.

Conclusions : Wnt and GR signaling inhibit each other in the TM, and canonical Wnt signaling activators have the potential to prevent the side effect of glucocorticoids.

This is a 2021 ARVO Annual Meeting abstract.