SPARC-YAP/TAZ inhibition prevents the fibroblasts-myofibroblasts transformation

Abstract

BackgroundFibrotic scar is a severe side effect of trabeculectomy, resulting in unsatisfactory outcomes for glaucoma surgery. Accumulating evidence showed human Tenon's fibroblasts (HTFs) play an important role in fibrosis formation. We previously reported that the aqueous level of secreted protein acidic and rich in cysteine (SPARC) was higher in the patients with primary angle closure glaucoma, which was associated with the failure of trabeculectomy. In this study, the potential effect and mechanism of SPARC in promoting fibrosis were explored by using HTFs. MethodsHTFs were employed in this study and examined under a phase-contrast microscope. Cell viability was determined by CCK-8. The expressions of SPARC-YAP/TAZ signaling and the fibrosis-related markers were examined with reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), Western blot, and immunofluorescence, subcellular fractionation was conducted to further determined the variation of YAP and phosphorylated YAP. The differential gene expressions were analyzed with RNA sequencing (RNAseq), followed by Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses. ResultsExogenous SPARC induced HTFs-myofibroblast transformation, as evidenced by the increased expression of α-SMA, collagen I and fibronectin in both protein and mRNA levels. SPARC knockdown decreased the expressions of the above genes in TGF-β2-treated HTFs. KEGG analysis showed that the Hippo signaling pathway was mostly enriched. SPARC treatment increased the expressions of YAP, TAZ, CTGF and CYR61 as well as enhanced YAP translocation from cytoplasm to nucleus, and decreased the phosphorylation of YAP and LAST1/2, which was reversed by SPARC knockdown. Knockdown of YAP1 decreased the fibrosis-related markers, such as α-SMA, collagen I and Fibronectin, in SPARC-treated HTFs. ConclusionsSPARC induced HTFs-myofibroblast transformation via activating YAP/TAZ signaling. Targeting SPARC-YAP/TAZ axis in HTFs might provide a novel strategy for inhibiting fibrosis formation after trabeculectomy.