Leptin-triggered Eosinophil-Macrophage crosstalk: paracrine activation of macrophage lipid metabolism mediated by eosinophil-derived PGD2/CCL5

Eosinophils and macrophages (MØs) cross-talking orchestrates adipose tissue homeostasis. Previously, we have observed that eosinophils potentiate leptin-induced MØs activation in in vitro co-cultures, increasing lipid droplet (LD) formation after leptin stimulation (50 nM; 2 h). Here, we studied if this effect depends on an eosinophil-driven paracrine activity. To this end, we evaluated the impact of supernatants of BALB/c in vitro bone marrow-derived eosinophils on resident peritoneal MØs. Our results demonstrated that both sEos and sEosLep (supernatants from non- and leptin-stimulated eosinophils) were able to induce LTC₄ synthesis, IL-10 secretion and LD biogenesis in MØs, with sEosLep being more potent (n=5; p<0.05). As leptin triggers PGD₂ and CCL5 (RANTES) secretion by eosinophils, to study their potential paracrine role, we used the following strategies: (i) pharmacological inhibition of PGD₂ synthesis in eosinophils pretreated with HQL-79 (10 µM); (ii) inhibition of protein Gi-coupled receptors (GPCRs) in MØs using pertussis toxin (100 ng/mL), and inhibition of CCL5 receptors using the metRANTES analog; and (iii) assessment of the direct induction of LD biogenesis in MØs stimulated with exogenous PGD₂ (25 nM) and/or CCL5 (100 ng/mL). Treatment of MØs with pertussis toxin (n=2) inhibited MØ activation (both LD biogenesis and LTC4 synthesis), indicating that eosinophils activate MØs via the secretion of agonists of GPCRs expressed by MØs. Both HQL-79 approach (n=3) and metRANTES antagonism (n=3) inhibited macrophage LD formation induced by the eosinophil supernatants. Only the combination of exogenous CCL5 and PGD₂ (n=3). but neither mediator alone, was able to activate MØs, suggesting a PGD₂/CCL5 synergistic action. Our data indicates that eosinophil/MØs crosstalk in response to leptin are partly mediated by soluble eosinophil-derived factors, with the CCL5/PGD₂ combination representing a key molecular mechanism. CEUA: 036/23.