AB0067 Allosteric receptor modulation of a free fatty acid receptor turns natural agonists into potent activators of the superoxide generating neutrophil nadph-oxidase
Authors: L. Björkman, J. Mårtensson, A. Holdfeldt, H. Forsman, M. Gabl, M. Sundqvist, C. Dahlgren
Journal: Annals of the Rheumatic Diseases (2018)

<h3>Background</h3> Short chain fatty acids are generated in the colon by bacterial fermentation of dietary fibres and serve as natural agonists for the free fatty acid receptor 2 (FFA2R/GPR43) which belongs to the large family of G-protein coupled receptors. We have earlier shown that acetate triggers an increase in the cytosolic concentration of free Ca<sup>2+</sup> in neutrophils without any assembly/activation of the superoxide generating NADPH-oxidase (Mol Cell Biol. 2016 Sep 26;36(20):2583–95). Allosteric modulators bind to receptors at sites topographically distinct from the agonist/antagonist binding site and can regulate receptor functions positively or negatively. <h3>Objectives</h3> We undertook this study to determine whether an FFA2R selective modulator affects the neutrophil response induced by natural FFA2R agonists. <h3>Methods</h3> Neutrophils were collected from healthy blood donors. Release of superoxide anions generated by the assembled/activated NADPH-oxidase was recorded by sensitive isoluminol/HRP amplified chemiluminescence method. Intracellular Ca<sup>2+</sup>transients were measured with FURA 2-AM labelled neutrophils. <h3>Results</h3> The allosteric modulator lacked a direct activating effect on neutrophils, but turned natural FFA2R agonists into potent activating agonists that triggered not only a transient rise in the cytosolic concentration of free Ca<sup>2+</sup> ions but also an assembly of the NADPH-oxidase. The NADPH-oxidase activity induced by the combined effect of the allosteric modulator and the natural agonist acetate could be further increased in neutrophils treated with the pro-inflammatory cytokine TNF-α. The receptor selectivity was demonstrated through the inhibition of the neutrophil activity by the novel FFA2R antagonist CATPB. In addition, the allosteric modulator lacked effect on neutrophil responses triggered by a novel and selective agonist for the closely related GPR84, a receptor that recognises medium chain fatty acids. <h3>Conclusions</h3> Allosteric modulators that positively co-operate with natural FFA2R agonists and prime neutrophils in their response to such agonists may serve as good tools for further unravelling the physiological functions of the FFA2R and its involvement in various diseases. In this study, allosteric modulation of FFA2R is introduced as a receptor selective mechanism to prime neutrophils to produce increased amounts of reactive oxygen species. <h3>Reference</h3> [1] Björkman L, Mårtensson J, Winther M, Gabl M, Holdfeldt A, Uhrbom M, Bylund J, Højgaard Hansen A, Pandey SK, Ulven T, Forsman H, Dahlgren C. The Neutrophil Response Induced by an Agonist for Free Fatty Acid Receptor 2 (GPR43) Is Primed by Tumor Necrosis Factor Alpha and by Receptor Uncoupling from the Cytoskeleton but Attenuated by Tissue Recruitment. Mol Cell Biol. 2016Sep 26;36(20):2583–95. doi:10.1128/MCB.00161-16 [Print 2016 Oct 15]. <h3>Disclosure of Interest</h3> None declared