Downregulating the Myocardin-related transcription factor/ Serum response factor (MRTF/SRF) pathway is a novel therapeutic approach to prevent post-surgical fibrosis in glaucoma

Purpose Glaucoma is the first leading cause of irreversible blindness worldwide and fibrosis is the main cause of failure of glaucoma surgery. There are currently no available anti-fibrotic treatments in the eye. We have previously described how the MRTF/SRF transcription pathway is intricately linked to all the key pathways in ocular fibrosis. We thus hypothesised that inhibiting the MRTF/SRF pathway would be a good therapeutic target to prevent fibrosis in the eye. Methods MRTF and SRF expression were knocked down in human Tenon's fibroblasts using small interfering RNAs, and three-dimensional fibroblast-populated collagen gels were used to measure contraction in vitro. Cytotoxicity was assessed using the live-dead assay. Live cell imaging was used to study the cell protrusive behaviour during contraction and quantification was performed using a custom parameter, the dynamic index. Reflection confocal microscopy and DQ collagen were also used to measure matrix degradation. Real-time qPCR was used to compare the gene expression of key matrix metalloproteinases in contracting gels. Results MRTF and SRF silencing significantly decreased collagen matrix contraction by 80% and 87% respectively. MRTF silencing did not impair cell viability compared to controls. Knocking down MRTF markedly reduced the protrusive behaviour of human Tenon's fibroblasts (dynamic index = 0.19 as compared to 0.66 for controls). MRTF silencing also significantly decreased matrix degradation and the expression of MMP-1, MMP-2, MMP-9, and MMP-14 genes. Conclusions Our study is the first to show that inhibiting the MRTF/SRF pathway significantly decreases collagen matrix contraction, cell protrusive behaviour of fibroblasts, and matrix degradation in the conjunctiva, and thus represents a promising new therapeutic approach to prevent post-surgical fibrosis in glaucoma.