Nonredundant protective properties of FPR2/ALX in polymicrobial murine sepsis

Sepsis is characterized by overlapping phases of excessive inflammation temporally aligned with an immunosuppressed state, defining a complex clinical scenario that explains the lack of successful therapeutic options. Here we tested whether the formyl-peptide receptor 2/3 (Fpr2/3)—ortholog to human FPR2/ALX (receptor for lipoxin A4)—exerted regulatory and organ-protective functions in experimental sepsis. Coecal ligature and puncture was performed to obtain nonlethal polymicrobial sepsis, with animals receiving antibiotics and analgesics. Clinical symptoms, temperature, and heart function were monitored up to 24 h. Peritoneal lavage and plasma samples were analyzed for proinflammatory and proresolving markers of inflammation and organ dysfunction. Compared with wild-type mice, Fpr2/3^−/− animals exhibited exacerbation of disease severity, including hypothermia and cardiac dysfunction. This scenario was paralleled by higher levels of cytokines [CXCL1 (CXC receptor ligand 1), CCL2 (CC receptor ligand 2), and TNFα] as quantified in cell-free biological fluids. Reduced monocyte recruitment in peritoneal lavages of Fpr2/3^−/− animals was reflected by a higher granulocyte/monocyte ratio. Monitoring Fpr2/3^−/− gene promoter activity with a GFP proxy marker revealed an over threefold increase in granulocyte and monocyte signals at 24 h post-coecal ligature and puncture, a response mediated by TNFα. Treatment with a receptor peptido-agonist conferred protection against myocardial dysfunction in wild-type, but not Fpr2/3^−/−, animals. Therefore, coordinated physio-pharmacological analyses indicate nonredundant modulatory functions for Fpr2/3 in experimental sepsis, opening new opportunities to manipulate the host response for therapeutic development.Sepsis is a clinical syndrome expression of the host reaction to pathogen invasion, as a consequence of either direct dissemination into the bloodstream or postsurgical trauma and gut ischemia/reperfusion-mediated pathogen translocation. The complexity of sepsis is due to multiple local and systemic immune responses that involve release of soluble mediators such as cytokines, bioactive lipid mediators, and cell stress markers, leading to multiple organ failure and ultimately death (1). Originally believed to result exclusively from an overzealous inflammatory response (e.g., cytokine storm), the lack of efficacy of anticytokine therapy in several clinical trials demonstrated that the pathogenesis of sepsis is complex. Notwithstanding the difficulty in clinical cases to establish the beginning of the infection (and the temporal recruitment of failing organs), it is now appreciated that the systemic inflammatory response syndrome (SIRS) can overlap with a compensatory anti-inflammatory response syndrome (CARS) (2). Immunosuppression associated with CARS may explain the failure of classical anti-inflammatory strategies in patients (3, 4).The acute inflammatory reaction against pathogens is in many cases successful, leading to healing and recovery of biological function. To achieve this end point, specific mediators and pathways of endogenous protection must be engaged by the host to promote what is now referred to as “resolution of inflammation” (5). Proresolving mediators share a set of properties that are emerging as paradigmatic (6); these include modulation of immune cell recruitment [inhibition of polymorphonuclear (PMN) migration and promotion of monocyte influx], augmentation of phagocytosis (leading to bacteria containment), promotion of apoptosis and efferocytosis, and eventually tissue/organ repair with restoration of physiological function (6, 7). It is perhaps for these organic and multifactorial biological actions that proresolving mediators like the protein annexin A1 (AnxA1) and the bioactive lipids lipoxin A₄ (LXA₄) and resolvin D₂ exert protection in models of experimental sepsis (8–10). Of relevance, the receptor target for AnxA1 and LXA₄ is a G protein-coupled receptor that belongs to the formyl-peptide receptor (FPR) family, termed FPR2/ALX. To establish the validity of FPR2/ALX for the development of innovative therapeutic approaches, proof-of-concept data within loss-of-function settings should be established.In the mouse, the human FPR2/ALX gene corresponds to two genes, termed Fpr2 and Fpr3, which share the first of the two exons (11). LXA₄ and AnxA1 are largely inactive in a transgenic mouse that lacks both murine genes (12) as shown in models of acute inflammation and ischemia-reperfusion injury (12–15). Herein we establish the patho-pharmacology of Fpr2/3 in experimental polymicrobial sepsis as a way to validate the human ortholog as a genuine receptor target for innovative treatments in sepsis.