Tu170 - TRPV2 Plays a Critical Role in Hepoxilin a3-medited Neutrophil Transepithelial Chemotaxis

Merran Dunford – University of Bath; Rebecca Ferrisi – Universita di Pisa; Francesca Gado – Universita Di Pisa; Clementina Manera – Universita Di Pisa; Keyi Liu – Applied Molecular Transport; Beth McCormick – University of Massachusetts; Randall Mrsny – Life Sciences – University of Bath

Abstract Text:

Purpose: Neutrophil transepithelial migration and accumulation at mucosal surfaces is a hallmark of many inflammatory conditions. Recent studies have demonstrated lipids secreted from the luminal surface of intestinal epithelial cells to regulate this event with N-acyl ethanolamine-type (NAE) endocannabinoids (eCBs) suppressing and the eicosanoid hepoxilin A3 (HxA3) activating the chemotaxis of neutrophils across this mucosal barrier. We hypothesized that directional chemotaxis of neutrophils mediated by HxA3 should involve cell-surface receptor(s). Here, we identify a role for the transient receptor potential cation channel subfamily V member 2 (TRPV2) in HxA3-mediated chemotaxis that drives neutrophil transepithelial migration.

Methods: Human promyelocytic cells (HL-60), differentiated (dHL-60) into a neutrophil-like phenotype, were used in a surface internalization-biased protocol, and proteomic analysis of internalized materials following HxA3 exposure identified receptor candidates. Cell migration assays were performed using dHL-60 cells, followed by the addition of chemoattractant HxA3 or fMLP or 2-aminoethoxydiphenylborane (2-APB). siRNA-knock-down of CB2R and TRPV2 genes were performed for their ability to block HxA3- dHL-60 cell migration.

Results: TRPV2 was identified in the proteomic screen. The CB2R agonists individually blocked HxA3 and 2-APB-induced migration of dHL-60 cells. Our data suggests TRPV2 interacts with CB2R, linking these two elements in a potentially coordinated chemotactic function that could be used to position neutrophils in the lamina propria in a state of readiness for transepithelial migration induced by the apical epithelial secretion of HxA3.

Conclusions: We have identified TRPV2 as a promising candidate receptor for HxA3 to drive the transepithelial migration of neutrophils using the dHL-60 cell system.