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Yong Yuan, Chia-Yang Liu, Winston Kao; Dexamethasone induces stress fiber re-arrangement through non-canonical Wnt signaling in trabecular meshwork cells. Invest. Ophthalmol. Vis. Sci. 2013;54(15):3532.
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Dexamethasone (DEX) regulates aqueous humor outflow by inducing a re-organization of the cytoskeleton to form cross-linked actin networks (CLAN) in the trabecular meshwork (TM) cells. Myosin Light Chain Kinase (MLCK) has been demonstrated to play an important role in this process, but the upstream components of the pathway leading to MLCK activation are still not clearly defined. Attempts were made to determine if non-canonical Wnt signaling mediates the DEX-induced cytoskeletal changes in TM cells.
Primary TM cells were treated with 100 nM DEX in low serum medium. The medium was changed every 3 days. After 7-10 days of culture, the cells were harvested and subjected to molecular biology analysis for the expression of Wnt ligands by degenerate PCR and real-time qRT-PCR. Morphology and immunofluorescence analysis were also performed for stress fiber (phalloidin stain), MLC-P and Wnt5a. Lentivirus-based shRNA against non-canonical Wnt receptor (ROR2) was used to determine the role of non-canonical Wnt signaling in DEX induced cytoskeleton changes.
DEX induced stress fiber re-arrangement in TM cells. Increased level of myosin light chain phosphorylation was observed in DEX-treated cells. Non-canonical Wnt ligand (Wnt5a) was up-regulated by DEX as demonstrated by real time qRT-PCR and immunostaining. Knockdown ROR2, the receptor of non-canonical Wnt signaling, abolished the effects of DEX on the formation of CLAN and the increased phosphorylation of MLC seen in Dex-treated TM cells.
Our data suggests that DEX induces the up-regulation of non-canonical Wnt ligand Wnt5a. Wnt 5a activates myosin light kinase through non-canonical Wnt receptor ROR2. Given the similarities between DEX-induced glaucoma and primary open angle glaucoma, our results provided mechanism of action for applying ROCK inhibitor to treat primary open-angle glaucoma.
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