Attenuation of EMT in RPE cells and subretinal fibrosis by an RAR-γ agonist

Subretinal fibrosis contributes to the loss of vision associated with age-related macular degeneration (AMD). Retinal pigment epithelial (RPE) cells play a key role in the pathogenesis of AMD including the fibrotic reaction. We examined the role of retinoic acid receptor-gamma (RAR-gamma) in the epithelial-mesenchymal transition (EMT) and other fibrosis-related processes in mouse RPE cells cultured in a type I collagen gel. Transforming growth factor-beta 2 (TGF-beta 2)-induced collagen gel contraction mediated by the RPE cells was inhibited by the RAR-gamma agonist R667 in a concentration- and time-dependent manner. Expression of the mesenchymal markers alpha-smooth muscle actin and fibronectin, the release of interleukin-6, and the phosphorylation of paxillin, mitogen-activated protein kinases (ERK, p38, and JNK), Smad2, and AKT induced by TGF-beta 2 were also suppressed by the RAR-gamma agonist. Furthermore, gelatin zymography and immunoblot analysis revealed that the TGF-beta 2-induced release of matrix metalloproteinase (MMP)-2, MMP-3, MMP-8, and MMP-9 from RPE cells was inhibited by R667, and the MMP inhibitor GM6001 attenuated TGF-beta 2-induced RPE cell contraction. Finally, immunohistofluorescence analysis with antibodies to glial fibrillary acidic protein showed that R667 inhibited the development of subretinal fibrosis in a mouse model in vivo. Our results thus suggest that RAR-gamma agonists may prove effective for the treatment of subretinal fibrosis associated with AMD.