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Ca 2+ permeation through TRPV4 in fibroblasts is associated with pathological matrix degradation. In human gingival fibroblasts, IL-1β binding to its signaling receptor (IL-1R1) induces activation of extracellular regulated kinase (ERK) and MMP1 expression, processes that require Ca 2+ flux across the plasma membrane. It is not known how IL-1R1, which does not conduct Ca 2+ , generates Ca 2+ signals in response to IL-1. We examined whether TRPV4 mediates the Ca 2+ fluxes required for ERK signaling in IL-1 stimulated gingival fibroblasts. TRPV4 was immunostained in fibroblasts of human gingival connective tissue and in focal adhesions of cultured mouse gingival fibroblasts. Human gingival fibroblasts treated with IL-1β showed no change of TRPV4 expression but there was increased MMP1 expression. In mouse, gingival fibroblasts expressing TRPV4, IL-1 strongly increased [Ca 2+ ] i . Pre-incubation of cells with IL-1 Receptor Antagonist blocked Ca 2+ entry induced by IL-1 or the TRPV4 agonist GSK101. Knockout of TRPV4 or expression of a non-Ca 2+ -conducting TRPV4 pore-mutant or pre-incubation with the TRPV4 inhibitor RN1734, blocked IL-1-induced Ca 2+ transients and expression of the mouse interstitial collagenase, MMP13. Treatment of mouse gingival fibroblasts with GSK101 phenocopied Ca 2+ and ERK responses induced by IL-1; these responses were absent in TRPV4-null cells or cells expressing a non-conducting TRPV4 pore-mutant. Immunostained IL-1R1 localized with TRPV4 in adhesions within cell extensions. While TRPV4 immunoprecipitates analyzed by mass spectrometry showed no association with IL-1R1, TRPV4 associated with Src-related proteins and Src co-immunoprecipitated with TRPV4. Src inhibition reduced IL-1-induced Ca 2+ responses. The functional linkage of TRPV4 with IL-1R1 expands its repertoire of innate immune signaling processes by mediating IL-1-driven Ca 2+ responses that drive matrix remodeling in fibroblasts. Thus, inhibiting TRPV4 activity may provide a new pharmacological approach for blunting matrix degradation in inflammatory diseases., (© 2024 The Author(s). The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

Transient Receptor Potential Vanilloid-type 4 (TRPV4) is a mechanosensitive, Ca 2+ -permeable plasma membrane channel that associates with focal adhesions, influences collagen remodeling, and is associated with fibrotic processes through undefined mechanisms. While TRPV4 is known to be activated by mechanical forces transmitted through collagen adhesion receptors containing the β1 integrin, it is not understood whether TRPV4 affects matrix remodeling by altering β1 integrin expression and function. We tested the hypothesis that TRPV4 regulates collagen remodeling through its impact on the β1 integrin in cell-matrix adhesions. In cultured fibroblasts derived from mouse gingival connective tissues, which exhibit very rapid collagen turnover, we found that higher TRPV4 expression is associated with reduced β1 integrin abundance and adhesion to collagen, reduced focal adhesion size and total adhesion area, and reduced alignment and compaction of extracellular fibrillar collagen. The reduction of β1 integrin expression mediated by TRPV4 is associated with the upregulation of miRNAs that target β1 integrin mRNA. Our data suggest a novel mechanism by which TRPV4 modulates collagen remodeling through post-transcriptional downregulation of β1 integrin expression and function., (© 2023 The Authors. The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)