TRPV4 channel opening mediates pressure-induced pancreatitis initiated by Piezo1 activation
Sandip M. Swain, … , Steven R. Vigna, Rodger A. Liddle
Sandip M. Swain, … , Steven R. Vigna, Rodger A. Liddle
Published January 30, 2020
Citation Information: J Clin Invest. 2020;130(5):2527-2541. https://doi.org/10.1172/JCI134111.
View: Text | PDF
Research Article Gastroenterology Article has an altmetric score of 12

TRPV4 channel opening mediates pressure-induced pancreatitis initiated by Piezo1 activation

Abstract

Elevated pressure in the pancreatic gland is the central cause of pancreatitis following abdominal trauma, surgery, endoscopic retrograde cholangiopancreatography, and gallstones. In the pancreas, excessive intracellular calcium causes mitochondrial dysfunction, premature zymogen activation, and necrosis, ultimately leading to pancreatitis. Although stimulation of the mechanically activated, calcium-permeable ion channel Piezo1 in the pancreatic acinar cell is the initial step in pressure-induced pancreatitis, activation of Piezo1 produces only transient elevation in intracellular calcium that is insufficient to cause pancreatitis. Therefore, how pressure produces a prolonged calcium elevation necessary to induce pancreatitis is unknown. We demonstrate that Piezo1 activation in pancreatic acinar cells caused a prolonged elevation in intracellular calcium levels, mitochondrial depolarization, intracellular trypsin activation, and cell death. Notably, these effects were dependent on the degree and duration of force applied to the cell. Low or transient force was insufficient to activate these pathological changes, whereas higher and prolonged application of force triggered sustained elevation in intracellular calcium, leading to enzyme activation and cell death. All of these pathological events were rescued in acinar cells treated with a Piezo1 antagonist and in acinar cells from mice with genetic deletion of Piezo1. We discovered that Piezo1 stimulation triggered transient receptor potential vanilloid subfamily 4 (TRPV4) channel opening, which was responsible for the sustained elevation in intracellular calcium that caused intracellular organelle dysfunction. Moreover, TRPV4 gene–KO mice were protected from Piezo1 agonist– and pressure-induced pancreatitis. These studies unveil a calcium signaling pathway in which a Piezo1-induced TRPV4 channel opening causes pancreatitis.

Authors

Sandip M. Swain, Joelle M.-J. Romac, Rafiq A. Shahid, Stephen J. Pandol, Wolfgang Liedtke, Steven R. Vigna, Rodger A. Liddle

×

Figure 2

Piezo1 activation induces mitochondrial depolarization and trypsinogen activation in pancreatic acini.

Options: View larger image (or click on image) Download as PowerPoint
Piezo1 activation induces mitochondrial depolarization and trypsinogen a...
(A) Brightfield and fluorescence images of TMRE-loaded pancreatic acini 0 and 12 minutes after Yoda1 (50 μM) application. (B) Traces of live-cell TMRE fluorescence intensity of single acinar cells over time following the administration of Yoda1 (arrow). FD is the decrease in TMRE fluorescence, and F0 is the baseline TMRE intensity. Acini from WT or Piezo1aci-KO mice were treated with the mitochondrial uncoupler FCCP (10 μM), Yoda1 (50 μM), Yoda1 + GsMTx4 (5 μM), or CCK (10 nM). A representative tracing from 3 experiments is shown. (C) Mean decrease in TMRE intensity from B is depicted. n = 3–5 independent experiments with 15–20 cells in each experiment. (D) Decrease in TMRE intensity of pancreatic acini is shown in response to CCK (10 nM) from 60 cells over 20 minutes. (E) Decrease in fluorescence intensity of TMRE in pancreatic acinar cells upon Yoda1 application with or without preincubation of BAPTA-AM (20 μM) for 20 minutes is shown from 29–45 cells. (F) Live-cell imaging of intracellular trypsin activation with CCK (10 nM) and Yoda1 (50 μM) treatments. Scale bar: 10 μm. (G) Time course of Yoda1-induced trypsin activation is shown for 3 experiments. (H) Peak BZiPAR fluorescence was measured in acini from WT mice treated with Yoda1 in the absence or presence of GsMTx4 (5 μM) or in acini from Piezo1aci-KO mice. n = 3–5 independent experiments with 16–31 cells. (I) Pancreatic acini from Piezo1aci-KO mice had a trypsin activation response to CCK (10 nM) similar to that of WT mice. n = 3 from 16–20 cells. Statistical analyses were performed using Student’s t test (D) and 1-way ANOVA (C, E, H, and I). Data are shown as mean ± SEM. **P ≤ 0.01; ***P ≤ 0.001; ****P ≤ 0.0001.