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Sialic acid-binding immunoglobulin-like lectin 8 (Siglec-8) is an activating receptor mediating ?2-integrin-dependent function in human eosinophils.

ABSTRACT: BACKGROUND:Siglec-8 is a CD33 subfamily cell-surface receptor selectively expressed on human eosinophils. After cytokine priming, Siglec-8 mAb or glycan ligand binding causes eosinophil apoptosis associated with reactive oxygen species (ROS) production. Most CD33-related Siglecs function as inhibitory receptors, but the ability of Siglec-8 to stimulate eosinophil ROS production and apoptosis suggests that Siglec-8 might instead function as an activating receptor. OBJECTIVE:We sought to determine the role of IL-5 priming and identify the signaling molecules involved in Siglec-8 function for human eosinophils. METHODS:We used an mAb and/or a multimeric synthetic sulfated sialoglycan ligand recognizing Siglec-8 in combination with integrin blocking antibodies, pharmacologic inhibitors, phosphoproteomics, and Western blot analysis to define the necessity of various proteins involved in Siglec-8 function for human eosinophils. RESULTS:Cytokine priming was required to elicit the unanticipated finding that Siglec-8 engagement promotes rapid ?2-integrin-dependent eosinophil adhesion. Also novel was the finding that this adhesion was necessary for subsequent ROS production and apoptosis. Siglec-8-mediated ROS was generated through reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation because pretreatment of eosinophils with catalase (an extracellular superoxide scavenger) or NSC 23766 (a Rac GTPase inhibitor) completely inhibited Siglec-8-mediated eosinophil apoptosis. Finally, engagement of Siglec-8 on IL-5-primed eosinophils resulted in increased phosphorylation of Akt, p38, and c-Jun N-terminal kinase 1 that was also ?2-integrin dependent; pharmacologic inhibition of these kinases completely prevented Siglec-8-mediated eosinophil apoptosis. CONCLUSIONS:These data demonstrate that Siglec-8 functions uniquely as an activating receptor on IL-5-primed eosinophils through a novel pathway involving regulation of ?2-integrin-dependent adhesion, NADPH oxidase, and a subset of protein kinases.

SUBMITTER: Carroll DJ

PROVIDER: S-EPMC5839929 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature

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Sialic acid-binding immunoglobulin-like lectin 8 (Siglec-8) is an activating receptor mediating β<sub>2</sub>-integrin-dependent function in human eosinophils.

Carroll Daniela J DJ O'Sullivan Jeremy A JA Nix David B DB Cao Yun Y Tiemeyer Michael M Bochner Bruce S BS

The Journal of allergy and clinical immunology 20170906 6

<h4>Background</h4>Siglec-8 is a CD33 subfamily cell-surface receptor selectively expressed on human eosinophils. After cytokine priming, Siglec-8 mAb or glycan ligand binding causes eosinophil apoptosis associated with reactive oxygen species (ROS) production. Most CD33-related Siglecs function as inhibitory receptors, but the ability of Siglec-8 to stimulate eosinophil ROS production and apoptosis suggests that Siglec-8 might instead function as an activating receptor.<h4>Objective</h4>We soug ...[more]

PMID: 28888781

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Project description:Recently, ‘don’t eat me’-signals like CD47 have emerged as novel innate immune checkpoints, enabling cancer cells to evade clearance by phagocytes such as microglia (MG) or monocyte-derived cells (MdCs). Here, we aim at defining the role of inhibitory Siglec-9 in human and its mouse homologue Siglec-E in innate-centered immunotherapy against GBM. TCGA RNA-sequencing data revealed a significant correlation between high expression of immunoinhibitory SIGLEC9 and poor survival in GBM patients (log-rank p = 0.02). Using a CT-2A orthotopic GBM mouse model with MG-specific spatio-temporal deletion of Siglece (Sall1CreERT2 x Sigleceflox), we observed high MG-proliferation upon Siglece knockout (Ki-67+ MG 14.8% in Creneg vs. 34.9% in Crepos, p < 0.0001) accompanied by an enhanced microglial GBM-cell uptake (5.6% in Creneg vs. 12.3% in Crepos, p < 0.001). By extending the Siglece knockout to the MdC compartment (Cx3cr1CreERT2 x Sigleceflox) we observed a significantly prolonged survival in the Crepos population (21d in Creneg vs. 27d post-tumor injection in Crepos, p = 0.018), which could be further promoted by combining Siglece knockout with CD47 blockade (11% long-term remission in Crepos + anti-CD47). Proteomics analysis revealed increased antigen processing and presentation capabilities of SigleceKO MdCs which was confirmed by ex-vivo OT-1 cross-presentation assays. This increased T cell priming upon MdC SigleceKO was further boosted by addition of anti-PD1 antibody to the SigleceKO + anti-CD47 combination. Resulting in 23% of animals experiencing long-term remission in the triple treatment arm, even after tumor re-challenge. Genetic targeting of sialic acids, the ligand for Siglec receptors, on CT-2A cells (GNE-KO), resulted in increased GBM-cell phagocytosis by MG and MdCs and less exhausted tumor-infiltrating CD8+ T cells (14.8% in WT vs. 5.9% in GNE-KO, p = 0.003). In a translational approach, we are currently testing anti-Siglec-9 treatment regimens in patient GBM explants, cultured for 5 days in perfusion bioreactors.

Dataset Information

Sialic acid-binding immunoglobulin-like lectin 8 (Siglec-8) is an activating receptor mediating ?2-integrin-dependent function in human eosinophils.

Publications

Sialic acid-binding immunoglobulin-like lectin 8 (Siglec-8) is an activating receptor mediating β<sub>2</sub>-integrin-dependent function in human eosinophils.

Similar Datasets

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