TRPV4 regulates matrix stiffness and TGFβ1-induced epithelial-mesenchymal transition.

Shweta Sharma,Rishov Goswami,Shaik O Rahaman,David X Zhang

doi:10.1111/jcmm.13972

Abstract

Substrate stiffness (or rigidity) of the extracellular matrix has important functions in numerous pathophysiological processes including fibrosis. Emerging data support a role for both a mechanical signal, for example, matrix stiffness, and a biochemical signal, for example, transforming growth factor β1 (TGFβ1), in epithelial‐mesenchymal transition (EMT), a process critically involved in fibrosis. Here, we report evidence showing that transient receptor potential vanilloid 4 (TRPV4), a mechanosensitive channel, is the likely mediator of EMT in response to both TGFβ1 and matrix stiffness. Specifically, we found that: (a) genetic ablation or pharmacological inhibition of TRPV4 blocked matrix stiffness and TGFβ1‐induced EMT in normal mouse primary epidermal keratinocytes (NMEKs) as determined by changes in morphology, adhesion, migration and alterations of expression of EMT markers including E‐cadherin, N‐cadherin (NCAD) and α‐smooth muscle actin (α‐SMA), and (b) TRPV4 deficiency prevented matrix stiffness‐induced EMT in NMEKs over a pathophysiological range. Intriguingly, TRPV4 deletion in mice suppressed expression of mesenchymal markers, NCAD and α‐SMA, in a bleomycin‐induced murine skin fibrosis model. Mechanistically, we found that: (a) TRPV4 was essential for the nuclear translocation of YAP/TAZ (yes‐associated protein/transcriptional coactivator with PDZ‐binding motif) in response to matrix stiffness and TGFβ1, (b) TRPV4 deletion inhibited both matrix stiffness‐ and TGFβ1‐induced expression of YAP/TAZ proteins and (c) TRPV4 deletion abrogated both matrix stiffness‐ and TGFβ1‐induced activation of AKT, but not Smad2/3, suggesting a mechanism by which TRPV4 activity regulates EMT in NMEKs. Altogether, these data identify a novel role for TRPV4 in regulating EMT.

Highlights

The key findings described are: (a) transient receptor potential vanilloid 4 (TRPV4), a mechanosensitive Ca2+‐permeable channel, regulates matrix stiffness and transforming growth factor β1 (TGFβ1)‐induced epithelial‐mesenchymal transition (EMT); (b) TRPV4 deletion suppresses expression of mesenchymal markers in a murine skin fibrosis model and (c) fibrosis induced in skin tissues with bleomycin is associated with increased co‐localization of TRPV4 with mesenchymal markers and decreased co‐localization of TRPV4 with epithelial markers
We found that: (a) TRPV4 is essential for the nuclear translocation of YAP and TAZ in response to matrix stiffness and TGFβ1 in normal mouse primary epidermal keratinocytes (NMEKs); (b) TRPV4 deletion blocks both matrix stiffness‐induced and TGFβ1‐induced expression of YAP and TAZ proteins and (c) TRPV4 deletion abrogates matrix stiffness‐induced and TGFβ1‐induced activation of AKT, but not Smad2/3, suggesting a mechanism by which TRPV4 activity regulates EMT
We studied normal mouse keratinocytes on collagen‐coated polyacrylamide hydrogels, adjusting substrate stiffness in culture, which enabled us to determine the effect of normal or fibrotic matrix on TRPV4‐dependent EMT

Summary

| INTRODUCTION

Substrate stiffness (or rigidity) of the extracellular matrix (ECM) has important functions in numerous physiological and pathological processes including development, wound healing, oncogenesis and tissue fibrosis.[1,2,3,4,5,6,7,8] Cells detect differences in matrix stiffness and adjust expression of genes and proteins in response, a capacity that has increasingly become a subject of research because of its importance in critical cellular processes including cell differentiation, migration and proliferation.[1,2,3,4,5,6,7,8] The epithelial‐mesenchymal transition. | 763 results identify a novel regulatory role for TRPV4 in EMT induced by both TGFβ1 and pathophysiological matrix stiffness

| RESULTS

| DISCUSSION

Findings

| MATERIALS AND METHODS