ABSTRACT
Vocal fold (VF) fibrosis is a major cause of intractable voice-related disability and reduced quality of life. Excision of fibrotic regions is suboptimal and associated with scar recurrence and/or further iatrogenic damage. Non-surgical interventions are limited, putatively related to limited insight regarding biochemical events underlying fibrosis, and downstream, the lack of therapeutic targets. YAP/TAZ integrates diverse cell signaling events and interacts with signaling pathways related to fibrosis, including the TGF-ß/SMAD pathway. We investigated the expression of YAP/TAZ following vocal fold injury in vivo as well as the effects of TGF-ß1 on YAP/TAZ activity in human vocal fold fibroblasts, fibroblast-myofibroblast transition, and TGF-ß/SMAD signaling. Iatrogenic injury increased nuclear localization of YAP and TAZ in fibrotic rat vocal folds. In vitro, TGF-ß1 activated YAP and TAZ in human VF fibroblasts, and inhibition of YAP/TAZ reversed TGF-ß1-stimulated fibroplastic gene upregulation. Additionally, TGF-ß1 induced localization of YAP and TAZ in close proximity to SMAD2/3, and nuclear accumulation of SMAD2/3 was inhibited by a YAP/TAZ inhibitor. Collectively, YAP and TAZ were synergistically activated with the TGF-ß/SMAD pathway, and likely essential for the fibroplastic phenotypic shift in VF fibroblasts. Based on these data, YAP/TAZ may evolve as an attractive therapeutic target for VF fibrosis.
Subject(s)
Intracellular Signaling Peptides and Proteins/metabolism , Signal Transduction , Smad Proteins/metabolism , Vocal Cord Dysfunction/metabolism , Vocal Cords/metabolism , Adaptor Proteins, Signal Transducing/metabolism , Animals , Female , Fibrosis , Humans , Rats , Rats, Sprague-Dawley , Transcription Factors/metabolism , Transcriptional Coactivator with PDZ-Binding Motif Proteins , Vocal Cord Dysfunction/pathology , Vocal Cords/pathology , YAP-Signaling ProteinsABSTRACT
Fibrosis of the vocal folds poses a substantive clinical challenge potentially underlying the rapid proliferation of direct steroid injections into the upper airway. The variable clinical response to glucocorticoids (GCs) in the vocal folds is likely related to diversity inherent to GCs and patient-specific, and upstream, cell-specific responses to GCs. Broadly, we hypothesize the disparity in clinical outcomes are due to undesirable effects of GCs on resident fibroblasts. Transcriptome analysis identified significant GC-mediated modulation of Hippo signaling, a known regulator of fibrotic gene expression. Subsequent analysis confirmed GC-mediated YAP activation, a transcriptional co-factor in the Hippo signaling pathway. YAP inhibition attenuated ACTA2 expression in GC-treated human vocal fold fibroblasts. Nuclear localization and phosphorylation at Ser211, however, was not affected by YAP inhibition, suggesting nuclear translocation of YAP is indirectly driven by GR. RNA-seq analysis confirmed the influence of GCs on Wnt signaling, and canonical Wnt signaling target genes were upregulated by GCs. These data implicate YAP and its downstream targets as putative mediators of a pro-fibrotic response to GCs. Therapeutic YAP inhibition may ultimately be clinically relevant and warrants further consideration.