Atypical chemokine receptors (ACKRs) are regulators of leukocyte traffic, inflammation, and immunity. ACKR2 is a scavenger for most inflammatory CC chemokines and is a negative regulator of ...inflammation. Here we report that ACKR2 is expressed in hematopoietic precursors and downregulated during myeloid differentiation. Genetic inactivation of ACKR2 results in increased levels of inflammatory chemokine receptors and release from the bone marrow of neutrophils with increased anti-metastatic activity. In a model of NeuT-driven primary mammary carcinogenesis ACKR2 deficiency is associated with increased primary tumor growth and protection against metastasis. ACKR2 deficiency results in neutrophil-mediated protection against metastasis in mice orthotopically transplanted with 4T1 mammary carcinoma and intravenously injected with B16F10 melanoma cell lines. Thus, ACKR2 is a key regulator (checkpoint) of mouse myeloid differentiation and function and its targeting unleashes the anti-metastatic activity of neutrophils in mice.
The plasma membrane tetraspan molecule MS4A4A is selectively expressed by macrophage-lineage cells, but its function is unknown. Here we report that MS4A4A was restricted to murine and human ...mononuclear phagocytes and was induced during monocyte-to-macrophage differentiation in the presence of interleukin 4 or dexamethasone. Human MS4A4A was co-expressed with M2/M2-like molecules in subsets of normal tissue-resident macrophages, infiltrating macrophages from inflamed synovium and tumor-associated macrophages. MS4A4A interacted and colocalized with the β-glucan receptor dectin-1 in lipid rafts. In response to dectin-1 ligands, Ms4a4a-deficient macrophages showed defective signaling and defective production of effector molecules. In experimental models of tumor progression and metastasis, Ms4a4a deficiency in macrophages had no impact on primary tumor growth, but was essential for dectin-1-mediated activation of macrophages and natural killer (NK) cell-mediated metastasis control. Thus, MS4A4A is a tetraspan molecule selectively expressed in macrophages during differentiation and polarization, essential for dectin-1-dependent activation of NK cell-mediated resistance to metastasis.
Inflammatory CC chemokines have long been associated with cancer, but unequivocal evidence of a role in clinically relevant models of carcinogenesis is lacking. D6, a promiscuous decoy receptor that ...scavenges inflammatory CC chemokines, plays a non-redundant role in reducing the inflammatory response in various organs. As inflammation is a key player in the development of inflammatory bowel disease (IBD) and IBD-associated colorectal cancer, we investigated D6 expression in human colitis and colon cancer, and its role in experimental colitis and inflammation-associated colon cancer.
In humans, D6 was mainly expressed by lymphatic vessels and leukocytes in the mucosa of individuals with IBD and colon cancer, as well as the mucosa of control individuals. Mice lacking expression of D6 were significantly more susceptible to experimental colitis than wild-type mice and failed to resolve colitis, with significantly higher levels of several pro-inflammatory chemokines. In bone marrow chimeric mice, the ability of D6 to regulate colitis was tracked to the stromal/lymphatic compartment, with no contribution of haemopoietic cells. Finally, after administration of the carcinogen azoxymethane, D6(-/-) mice showed increased susceptibility to colitis-associated cancer in the distal segment of the colon compared with wild-type mice.
D6 expressed on lymphatic vessels plays a key role in the control of intestinal inflammation and the development of inflammation-associated colon cancer. Our results reveal a new unexpected role for the lymphatic system in the pathogenesis of IBD and intestinal cancer, and candidate chemokines as novel players in tumour promotion and progression.
Adult-type diffuse gliomas are malignant primary brain tumors characterized by very poor prognosis. Dendritic cells (DCs) are key in priming antitumor effector functions in cancer, but their role in ...gliomas remains poorly understood.
In this study, we characterized tumor-infiltrating DCs (TIDCs) in adult patients with newly diagnosed diffuse gliomas by using multi-parametric flow cytometry and single-cell RNA sequencing.
We demonstrated that different subsets of DCs are present in the glioma microenvironment, whereas they are absent in cancer-free brain parenchyma. The largest cluster of TIDCs was characterized by a transcriptomic profile suggestive of severe functional impairment. Patients undergoing perioperative corticosteroid treatment showed a significant reduction of conventional DC1s, the DC subset with key functions in antitumor immunity. They also showed phenotypic and transcriptional evidence of a more severe functional impairment of TIDCs.
Overall, the results of this study indicate that functionally impaired DCs are recruited in the glioma microenvironment. They are severely affected by dexamethasone administration, suggesting that the detrimental effects of corticosteroids on DCs may represent one of the mechanisms contributing to the already reported negative prognostic impact of steroids on glioma patient survival.
The atypical chemokine receptor ACKR2 promotes resolution of acute inflammation by operating as a scavenger receptor for inflammatory CC chemokines in several experimental models of inflammatory ...disorders, however its role in the brain remains unclear. Based on our previous reports of increased expression of inflammatory chemokines and their corresponding receptors following traumatic brain injury (TBI), we hypothesised that ACKR2 modulates neuroinflammation following brain trauma and that its deletion exacerbates cellular inflammation and chemokine production. We demonstrate increased CCL2 and ACKR2 mRNA expression in post-mortem human brain, whereby ACKR2 mRNA levels correlated with later times post-TBI. This data is consistent with the transient upregulation of ACKR2 observed in mouse brain after closed head injury (CHI). As compared to WT animals, ACKR2-/- mice showed a higher mortality rate after CHI, while the neurological outcome in surviving mice was similar. At day 1 post-injury, ACKR2-/- mice displayed aggravated lesion volume and no differences in CCL2 expression and macrophage recruitment relative to WT mice. Reciprocal regulation of ACKR2 and CCL2 expression was explored in cultured astrocytes, which are recognized as the major source of CCL2 and also express ACKR2. ACKR2 mRNA increased as early as 2 hours after an inflammatory challenge in WT astrocytes. As expected, CCL2 expression also dramatically increased at 4 hours in WT astrocytes but was significantly lower in ACKR2-/- astrocytes, possibly indicating a co-regulation of CCL2 and ACKR2 in these cells. Conversely, in vivo, CCL2 mRNA/protein levels were increased similarly in ACKR2-/- and WT brains at 4 and 12 hours after CHI, in line with the lack of differences in cerebral macrophage recruitment and neurological recovery. In conclusion, ACKR2 is induced after TBI and has a significant impact on mortality and lesion development acutely following CHI, while its role in chemokine expression, macrophage activation, brain pathology, and neurological recovery at later time-points is minor. Concordant to evidence in multiple sclerosis experimental models, our data corroborate a distinct role for ACKR2 in cerebral inflammatory processes compared to its reported functions in peripheral tissues.
Background D6 is an atypical chemokine receptor involved in chemokine degradation and resolution of acute inflammatory responses in mice. Emerging evidence suggests that D6 might behave differently ...in human chronic inflammatory conditions. We, therefore, investigated the involvement of D6 in the immune responses in COPD, a chronic inflammatory condition of the lung. Methods D6 expression was quantified by immunohistochemistry in surgical resected lung specimens from 16 patients with COPD (FEV1 , 57% ± 6% predicted) and 18 control subjects with normal lung function (nine smokers and nine nonsmokers). BAL was also obtained and analyzed by flow cytometry, immunofluorescence, and molecular analysis for further assessment of D6 involvement. Results D6 expression in the lung was mainly detected in alveolar macrophages (AMs). The percentage of D6+ AMs was markedly increased in patients with COPD as compared with both smoker and nonsmoker control subjects ( P < .0005 for both). D6 expression was detected at both transcript and protein level in AMs but not in monocyte-derived macrophages. Finally, D6 expression was positively correlated with markers of immune activation (CD8+ T lymphocytes, IL-32, tumor necrosis factor-α, B-cell activating factor of the tumor necrosis factor family, phospho-p38 mitogen-activated protein kinase) and negatively with lung function (FEV1 , FEV1 /FVC). Conclusions D6 is expressed in AMs from patients with COPD, and its expression correlates with the degree of functional impairment and markers of immune activation. Upregulation of D6 in AMs could indicate that, besides its known scavenger activity in acute inflammation, D6 may have additional roles in chronic inflammatory conditions possibly promoting immune activation.
The atypical chemokine receptor ACKR2, formerly named D6, is a scavenger chemokine receptor with a non-redundant role in the control of inflammation and immunity. The scavenging activity of ACKR2 ...depends on its trafficking properties, which require actin cytoskeleton rearrangements downstream of a β-arrestin1-Rac1-PAK1-LIMK1-cofilin-dependent signaling pathway. We here demonstrate that in basal conditions, ACKR2 trafficking properties require intact actin and microtubules networks. The dynamic turnover of actin filaments is required to sustain ACKR2 constitutive endocytosis, while both actin and microtubule networks are involved in processes regulating ACKR2 constitutive sorting to rapid, Rab4-dependent and slow, Rab11-dependent recycling pathways, respectively. After chemokine engagement, ACKR2 requires myosin Vb activity to promote its trafficking from Rab11-positive recycling endosomes to the plasma membrane, which sustains its scavenging activity. Other than cofilin phosphorylation, induction of the β-arrestin1-dependent signaling pathway by ACKR2 agonists also leads to the rearrangement of microtubules, which is required to support the myosin Vb-dependent ACKR2 upregulation and its scavenging properties. Disruption of the actin-based cytoskeleton by the apoptosis-inducing agent staurosporine results in impaired ACKR2 internalization and chemokine degradation that is consistent with the emerging scavenging-independent activity of the receptor in apoptotic neutrophils instrumental for promoting efficient efferocytosis during the resolution of inflammation. In conclusion, we provide evidence that ACKR2 activates a β-arrestin1-dependent signaling pathway, triggering both the actin and the microtubule cytoskeletal networks, which control its trafficking and scavenger properties.
The chemokine decoy receptor D6 controls inflammatory responses by selective recognition and degradation of most CCR1 to CCR5 agonistic ligands. CCL14 is a homeostatic chemokine present at high ...concentrations in the serum with a weak agonist activity on CCR1. Under inflammatory conditions, plasmin and UPA-mediated truncation of 8 amino acids generates the potent CCR1/CCR3/CCR5 isoform CCL14(9–74), which is further processed and inactivated by dipeptidyl peptidase IV/CD26 that generates CCL14(11–74). Here we report that D6 efficiently binds both CCL14 and its truncated isoforms. Like other D6 ligands, the biologically active CCL14(9–74) induces adaptive up-regulation of D6 expression on the cell membrane and is rapidly and efficiently degraded. In contrast, the D6-mediated degradation of the biologically inactive isoforms CCL14(1–74) and CCL14(11–74) is very inefficient. Thus, D6 cooperates with CD26 in the negative regulation of CCL14 by the selective degradation of its biologically active isoform. Analysis of a panel of CC chemokines and their truncated isoforms revealed that D6-mediated chemokine degradation does not correlate with binding affinity. Conversely, degradation efficiency is positively correlated with D6 adaptive up-regulation. Sequence analysis indicated that a proline residue in position 2 of D6 ligands is dispensable for binding but crucial for D6 adaptive up-regulation and efficient degradation.
The decoy receptor D6 plays a nonredundant role in the control of inflammatory processes through scavenging of inflammatory chemokines. However it remains unclear how it is regulated. Here we show ...that D6 scavenging activity relies on unique trafficking properties. Under resting conditions, D6 constitutively recycled through both a rapid wortmannin (WM)–sensitive and a slower brefeldin A (BFA)–sensitive pathway, maintaining low levels of surface expression that required both Rab4 and Rab11 activities. In contrast to “conventional” chemokine receptors that are down-regulated by cognate ligands, chemokine engagement induced a dose-dependent BFA-sensitive Rab11-dependent D6 re-distribution to the cell membrane and a corresponding increase in chemokine degradation rate. Thus, the energy-expensive constitutive D6 cycling through Rab11 vesicles allows a rapid, ligand concentration–dependent increase of chemokine scavenging activity by receptor redistribution to the plasma membrane. D6 is not regulated at a transcriptional level in a variety of cellular contexts, thus ligand-dependent optimization of its scavenger performance represents a rapid and unique mechanism allowing D6 to control inflammation.
► The atypical chemokine receptor D6 scavenges most inflammatory CC chemokines. ► D6 signals through a Gαi-independent and β-arrestin-1 dependent pathway. ► D6 regulates cofilin activity to optimize ...its chemokine scavenging activity. ► D6 contains inflammatory response allowing proper development of adaptive immunity. ► D6 controls accumulation of myeloid-derived suppressor cells to lymphoid organs.
The atypical chemokine receptor D6 was initially called “silent” on the basis of lack of conventional signaling events that lead to directional cell migration. It has emerged that D6 is able to bind and drive to degradative compartments most inflammatory CC chemokines and that is able to convey G-protein independent signaling events to optimize its scavenging activity. We here summarize the knowledge available today on D6 structural and signaling properties and its essential role for the control of inflammatory cells traffic and proper development of the adaptive immune response.
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