GPR15-dependent T cell recruitment in the acute phase of viral myocarditis is associated with improved virus elimination and outcome
Authors: Lindner D , Brauninger H , Stoffers B , Bacmeister L , Kupsch S , Vico T , Marchini T , Escher F , Klingel K , Kirchhof P , Blankenberg S , Zeller T , Wolf D , Hilgendorf I , Westermann D 
Published: May 2024

Abstract

Funding Acknowledgements
Type of funding sources: Public Institution(s). Main funding source(s): DZHK (German Centre for Cardiovascular Research)

Background
Myocarditis is an inflammatory heart disease that is primarily caused by viral pathogens such as Coxsackievirus B3 (CVB3). The clinical course of myocarditis has a wide spectrum of outcomes, ranging from complete recovery to cardiac dysfunction and dilated cardiomyopathy. The G protein-coupled receptor 15 (GPR15) has been identified as a T cell homing receptor associated with inflammatory diseases.

Purpose
The aim of this study was to address the following questions: Does GPR15 deficiency (i) lead to differences in immune cell infiltration or inflammation in the acute phase of myocarditis; (ii) does it affect elimination of CVB3 and thus outcome; (iii) does it affect immune cell chemotaxis or functionality? Therefore, GPR15-deficient (Gpr15gfp/gfp) and WT mice were infected with CVB3 to study the acute and subacute phase of myocarditis.

Results
In the subacute phase of myocarditis 16 days p.i., virus persistence was observed in more than 85% of Gpr15gfp/gfp mice, while more than 70% of WT mice successfully cleared the virus. In addition, Gpr15gfp/gfp mice showed reduced cardiac function accompanied by increased fibrosis compared to WT mice. Based on these findings, we analysed the acute phase of myocarditis in more detail. Comparing the infected genotypes, no significant differences were found at 6 days p.i. in contrast to 5 and 7 days p.i.. After 5 days p.i., Gpr15gfp/gfp mice showed significantly lower gene expression of inflammatory cytokines and immune cell markers, especially regulatory T cells. In contrast, after 7 days p.i., inflammatory cytokines and markers for regulatory and cytotoxic T cells were significantly increased in infected Gpr15gfp/gfp mice compared to infected WT mice. RNASeq confirmed that the response to the virus did not decrease from day 6 to 7 in infected Gpr15gfp/gfp mice, as observed in infected WT mice. Furthermore, gene ontology (GO) analyses revealed altered chemotaxis and increased cytotoxic T cell-related GO terms in Gpr15gfp/gfp mice 7 days p.i.. In addition, we demonstrated in vitro that chemotaxis of both regulatory and cytotoxic T cells depends on the interaction between GPR15 and its ligand GPR15L and that loss of GPR15 abrogates chemotaxis of T cells towards GPR15L.

Conclusion
Here we show that GPR15 deficiency leads to reduced virus elimination in the heart and thus to adverse remodelling and impaired cardiac function. Delayed recruitment of regulatory T cells was associated with prolonged persistence of cytotoxic and regulatory T cells in Gpr15gfp/gfp mice. In addition, RNASeq revealed a prolonged inflammatory response and altered chemotaxis in Gpr15gfp/gfp mice. Through in vitro studies, we identified GPR15L as an important chemokine for the recruitment of regulatory and cytotoxic T cells. In conclusion, insufficient virus elimination could be caused by delayed recruitment of T cells, leading to a prolonged inflammatory response and an adverse outcome in GPR15-deficient mice.