Resolvin D4 disengages emergency granulopoiesis

In this issue of Blood, Libreros et al¹ report that increase in resolvin D4 (RvD4) from distant bacterial infection disengages emergency granulopoiesis and limits excessive neutrophil accumulation at the inflamed site, thereby promoting return to homeostasis following infections.

Neutrophils play a central role in host defense against microbial infections and tissue injury. They have a short lifespan in the blood and are constantly replenished in steady-state conditions from hematopoietic stem cells (HSCs) and granulocyte-monocyte progenitor cells (GMPs) in the bone marrow.² During severe infections or tissue injury, neutrophils are consumed in larger quantities. The hematopoietic system senses and rapidly adapts to the increased neutrophil demand by switching from steady-state to emergency granulopoiesis. Emergency granulopoiesis is differently regulated at the transcriptional level from steady-state conditions and ensures de novo generation of large quantities of neutrophils from GMPs in the bone marrow (and extramedullary sites), which is critical for host survival.² Because persisting excessive neutrophil production is a hallmark of diverse inflammatory and autoimmune diseases,^3,4 identifying the molecular signals that switch emergency granulopoiesis back to steady-state conditions is of utmost importance.

Resolution of the acute inflammatory response is an active process governed by production of endogenous specialized proresolving lipid mediators (lipoxins, resolvins, protectins, and maresins), proteins (eg, annexin A1), and gaseous mediators (hydrogen sulfite and carbon monoxide), predominantly acting on phagocytes and other immune cells.^5,6 These mediators of active inflammation resolution regulate neutrophil trafficking into the inflamed tissue, enhance antimicrobial defense and phagocytosis, and promote neutrophil apoptosis and clearance of apoptotic neutrophils and cellular debris by macrophages.^5,6 

Comparing lipid profiles in the bone marrow in mice without active infection and mice during self-resolving bacterial peritonitis, Libreros et al identified and verified different temporal changes in the n-3 docosapentaenoic acid–derived resolvin D1 (RvD1) and RvD4, distinctly separate from other D and E series resolvins. RvD1 levels were elevated during the initiation phase (0-12 hours) and remained elevated during the resolution phase of peritonitis (24-72 hours), whereas RvD4 levels were below baseline levels during the entire observation period. Coinciding with these changes, distal infection evoked increases in bone marrow preneutrophils, immature neutrophils, and mature neutrophils along with an increase in the multipotent myeloid progenitors and GMPs, indicating emergency granulopoiesis.

The authors employed single-cell mass cytometry to map immune cell populations in the bone marrow following systemic administration of RvD1 or RvD4 to mice with peritonitis. They show that RvD4 promoted HSC differentiation toward B and T lymphocytes and controlled granulopoiesis, as evidenced by downregulation in granulocyte lineage trajectory in lineage^–Sca-1⁺c-Kit⁺ cells, GMPs, preneutrophils, and immature neutrophils without affecting the trajectory of terminally differentiated cells, mature neutrophils, and circulating neutrophils. Furthermore, RvD4 also attenuated leukotriene B₄–stimulated neutrophil deployment and promoted apoptosis and removal by bone marrow macrophages via efferocytosis of aged neutrophils. By contrast, RvD1 had no effects on granulopoiesis, but facilitated differentiation of GMPs into Ly6C^low reparative monocytes. More important, like RvE1 and RvD1,^7,8 RvD4 given together with Escherichia coli markedly reduced neutrophil accumulation in the peritoneal cavity, yet increased bacterial clearance and accelerated neutrophil apoptosis and efferocytosis, indicating efficient control of infection. The main implication of these findings is that by controlling HSC differentiation and excessive neutrophil mobilization into the circulation, and enhancing phagocytosis and efferocytosis, RvD4 can ensure rapid resolution and prevent bystander injury to the host. Additional studies will, however, be necessary to assess the therapeutic potential and optimal timing of RvD4 given after infection. In humans, phagocytosing neutrophils and macrophages appear to be the major, but not the sole, sources of RvD4.⁹ Hence, it is tempting to speculate that RvD4 enhancement of neutrophil phagocytosis and efferocytosis by macrophages may beget further increases in the synthesis of RvD4 (and other proresolving mediators),^8,10 thereby forming a positive feed-forward loop to drive efficient resolution and timely return to homeostasis.

Proresolving lipids exhibit stereoselective actions in human leukocytes through specific G-protein–coupled receptors (GPCRs), sometimes in a cell type–specific manner. For instance, resolvin E1 signals through the leukotriene B₄ receptor leukotriene B4 receptor 1 and chemerin receptor 23 in neutrophils and macrophages, respectively, to enhance phagocytosis.^5,7 The authors’ data imply involvement of distinct receptors for RvD1 and RvD4. RvD1 is known to signal through the lipoxin A4 receptor/formyl peptide receptor 2 (ALX/FPR2),⁵ and this was confirmed by Libreros et al using ALX/FPR2-deficient mice. RvD4 did not activate this GPCR for signaling in either neutrophils or monocytes. In human macrophages, RvD4-stimulated phagocytosis was cholera toxin sensitive, suggesting a role for a GPCR that has yet to be identified. Future studies should also address whether RvD4 acts through the same GPCR in neutrophils and macrophages as well as the potential interplays among RvD1 and RvD4 or other proresolving circuits. For example, proresolving agonists of ALX/FPR2 trigger synthesis of lipoxin A4 and RvD1, setting in motion integrated actions that result in resolution.^5,6,8 How RvD4 signals is incompletely understood. The authors did show that regardless of the GPCR involved, RvD4 stimulated phosphorylation of extracellular signal-regulated kinase 1/2 and STAT3 in the bone marrow, which are critical in granulocyte differentiation and mobilization into circulation.² Phosphorylation of STAT3 is also known to facilitate apoptosis in human neutrophils.¹⁰ Additional studies are required to explore RvD4 signaling in HSCs and efferocytic macrophages.

Libreros et al identify a novel regulatory mechanism by specialized proresolving mediators of host defense during bacterial infection (see figure). This study builds on and expands the works of the Serhan laboratory over the years, with identifying the functions of the superfamily of specialized proresolving lipid mediators, including resolvins.⁵ Herein, the group provides compelling evidence on the multipronged actions of RvD4, which partly overlap with those of RvD1 and other proresolving lipids,^5-8 but also include controlling emergency granulopoiesis and preventing excessive mobilization of neutrophils into circulation, while facilitating clearance of bacteria and apoptotic neutrophils within the infected site and bone marrow. These findings suggest that there is therapeutic potential for RvD4 for mitigating pathologic conditions, such as idiopathic arthritis³ and severe sepsis,⁴ linked to excessive emergency granulopoiesis and neutrophil-mediated tissue injury.

Conflict-of-interest disclosure: The author declares no competing financial interests.