Mechanically activated Piezo1 channels of cardiac fibroblasts stimulate p38 mitogen-activated protein kinase activity and interleukin-6 secretion

J Biol Chem. 2019 Nov 15;294(46):17395-17408. doi: 10.1074/jbc.RA119.009167. Epub 2019 Oct 4.

Abstract

Piezo1 is a mechanosensitive cation channel with widespread physiological importance; however, its role in the heart is poorly understood. Cardiac fibroblasts help preserve myocardial integrity and play a key role in regulating its repair and remodeling following stress or injury. Here we investigated Piezo1 expression and function in cultured human and mouse cardiac fibroblasts. RT-PCR experiments confirmed that Piezo1 mRNA in cardiac fibroblasts is expressed at levels similar to those in endothelial cells. The results of a Fura-2 intracellular Ca2+ assay validated Piezo1 as a functional ion channel that is activated by its agonist, Yoda1. Yoda1-induced Ca2+ entry was inhibited by Piezo1 blockers (gadolinium and ruthenium red) and was reduced proportionally by siRNA-mediated Piezo1 knockdown or in murine Piezo1 +/- cells. Results from cell-attached patch clamp recordings on human cardiac fibroblasts established that they contain mechanically activated ion channels and that their pressure responses are reduced by Piezo1 knockdown. Investigation of Yoda1 effects on selected remodeling genes indicated that Piezo1 activation increases both mRNA levels and protein secretion of IL-6, a pro-hypertrophic and profibrotic cytokine, in a Piezo1-dependent manner. Moreover, Piezo1 knockdown reduced basal IL-6 expression from cells cultured on softer collagen-coated substrates. Multiplex kinase activity profiling combined with kinase inhibitor experiments and phosphospecific immunoblotting established that Piezo1 activation stimulates IL-6 secretion via the p38 mitogen-activated protein kinase downstream of Ca2+ entry. In summary, cardiac fibroblasts express mechanically activated Piezo1 channels coupled to secretion of the paracrine signaling molecule IL-6. Piezo1 may therefore be important in regulating cardiac remodeling.

Keywords: IL-6; calcium; cardiac fibroblast; fibroblast; heart; ion channel; mechanotransduction; mitogen-activated protein kinase (MAPK); p38 MAPK; patch clamp; signal transduction.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Calcium Signaling / genetics
  • Endopeptidases / genetics
  • Endothelial Cells / chemistry
  • Endothelial Cells / metabolism
  • Fibroblasts / metabolism
  • Gene Expression Regulation / genetics
  • Gene Knockdown Techniques
  • Humans
  • Interleukin-6 / chemistry
  • Interleukin-6 / genetics*
  • Ion Channels / chemistry
  • Ion Channels / genetics*
  • MAP Kinase Signaling System / genetics
  • Mechanotransduction, Cellular / genetics
  • Mice
  • Myocardium / chemistry
  • Myocardium / metabolism*
  • Phosphorylation / genetics
  • RNA, Messenger / genetics
  • RNA, Small Interfering / genetics
  • Signal Transduction / genetics
  • Thiolester Hydrolases / genetics
  • p38 Mitogen-Activated Protein Kinases / chemistry
  • p38 Mitogen-Activated Protein Kinases / genetics*

Substances

  • Interleukin-6
  • Ion Channels
  • PIEZO1 protein, human
  • RNA, Messenger
  • RNA, Small Interfering
  • p38 Mitogen-Activated Protein Kinases
  • YOD1 protein, human
  • Thiolester Hydrolases
  • Endopeptidases