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Dissecting the Therapeutic Mechanisms of Sphingosine-1-Phosphate Receptor Agonism during Ischaemia and Reperfusion

Lookup NU author(s): Dr Georgie WilkinsORCiD, Dr Jenny Gilmour, Eirini Giannoudaki, Emeritus Professor John Kirby, Professor Neil SheerinORCiD, Professor Simi Ali



This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).


Abstract: Sphingosine 1-phosphate (S1P) and S1P receptors (S1PR) regulate many cellular processes, including lymphocyte migration and endothelial barrier function. As neutrophils are major mediators of inflammation, their transendothelial migration may be the target of therapeutic approaches to inflammatory conditions such as ischaemia–reperfusion injury (IRI). The aim of this project was to assess whether these therapeutic effects are mediated by S1P acting on neutrophils directly or indirectly through the endothelial cells. First, our murine model of peritoneum cell recruitment demonstrated the ability of S1P to reduce CXCL8-mediated neutrophil recruitment. Mechanistic in vitro studies revealed that S1P signals in neutrophils mainly through the S1PR1 and 4 receptors and induces phosphorylation of ERK1/2; however, this had no effect on neutrophil transmigration and adhesion. S1P treatment of endothelial cells significantly reduced TNF-α-induced neutrophil adhesion under flow (p < 0.01) and transendothelial migration towards CXCL8 during in vitro chemotaxis assays (p < 0.05). S1PR1 agonist CYM5442 treatment of endothelial cells also reduced neutrophil transmigration (p < 0.01) and endothelial permeability (p < 0.005), as shown using in vitro permeability assays. S1PR3 agonist had no effects on chemotaxis or permeability. In an in vivo mouse model of renal IRI, S1PR agonism with CYM5442 reduced endothelial permeability as shown by reduced Evan’s Blue dye extravasation. Western blot was used to assess phosphorylation at different sites on vascular endothelial (VE)–cadherin and showed that CYM5442 reduced VEGFmediated phosphorylation. Taken together, the results of this study suggest that reductions in neutrophil infiltration during IRI in response to S1P are mediated primarily by S1PR1 signalling on endothelial cells, possibly by altering phosphorylation of VE–cadherin. The results also demonstrate the therapeutic potential of S1PR1 agonist during IRI.

Publication metadata

Author(s): Wilkins GC, Gilmour J, Giannoudaki E, Kirby JA, Sheerin NS, Ali S

Publication type: Article

Publication status: Published

Journal: International Journal of Molecular Sciences

Year: 2023

Volume: 24

Issue: 13

Online publication date: 07/07/2023

Acceptance date: 21/06/2023

Date deposited: 10/07/2023

ISSN (print): 1661-6596

ISSN (electronic): 1422-0067

Publisher: MDPI AG


DOI: 10.3390/ijms241311192


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Funder referenceFunder name
Kidney Research UK
PG/18/57/33941British Heart Foundation