Dataset Information

Neuronal signal-regulatory protein alpha drives microglial phagocytosis by limiting microglial interaction with CD47 in the retina.

ABSTRACT: Microglia utilize their phagocytic activity to prune redundant synapses and refine neural circuits during precise developmental periods. However, the neuronal signals that control this phagocytic clockwork remain largely undefined. Here, we show that neuronal signal-regulatory protein alpha (SIRPα) is a permissive cue for microglial phagocytosis in the developing murine retina. Removal of neuronal, but not microglial, SIRPα reduced microglial phagocytosis, increased synpase numbers, and impaired circuit function. Conversely, prolonging neuronal SIRPα expression extended developmental microglial phagocytosis. These outcomes depended on the interaction of presynaptic SIRPα with postsynaptic CD47. Global CD47 deficiency modestly increased microglial phagocytosis, while CD47 overexpression reduced it. This effect was rescued by coexpression of neuronal SIRPα or codeletion of neuronal SIRPα and CD47. These data indicate that neuronal SIRPα regulates microglial phagocytosis by limiting microglial SIRPα access to neuronal CD47. This discovery may aid our understanding of synapse loss in neurological diseases.

SUBMITTER: Jiang D

PROVIDER: S-EPMC9772037 | biostudies-literature | 2022 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Neuronal signal-regulatory protein alpha drives microglial phagocytosis by limiting microglial interaction with CD47 in the retina.

Jiang Danye D Burger Courtney A CA Akhanov Viktor V Liang Justine H JH Mackin Robert D RD Albrecht Nicholas E NE Andrade Pilar P Schafer Dorothy P DP Samuel Melanie A MA

Immunity 20221114 12

Microglia utilize their phagocytic activity to prune redundant synapses and refine neural circuits during precise developmental periods. However, the neuronal signals that control this phagocytic clockwork remain largely undefined. Here, we show that neuronal signal-regulatory protein alpha (SIRPα) is a permissive cue for microglial phagocytosis in the developing murine retina. Removal of neuronal, but not microglial, SIRPα reduced microglial phagocytosis, increased synpase numbers, and impaired ...[more]

PMID: 36379210

Similar Datasets

Project description:CD47 is a cell surface protein that transmits an anti-phagocytic signal, known as the "don't-eat-me" signal, to macrophages upon engaging its receptor signal regulatory protein α (SIRPα). Molecules that antagonize the CD47-SIRPα interaction by binding to CD47, such as anti-CD47 antibodies and the engineered SIRPα variant CV1, have been shown to facilitate macrophage-mediated anti-tumor responses. However, these strategies targeting CD47 are handicapped by large antigen sinks in vivo and indiscriminate cell binding due to ubiquitous expression of CD47. These factors reduce bioavailability and increase the risk of toxicity. Here, we present an alternative strategy to antagonize the CD47-SIRPα pathway by engineering high affinity CD47 variants that target SIRPα, which has restricted tissue expression. CD47 proved to be refractive to conventional affinity maturation techniques targeting its binding interface with SIRPα. Therefore, we developed a novel engineering approach, whereby we augmented the existing contact interface via N-terminal peptide extension, coined "Velcro" engineering. The high affinity variant (Velcro-CD47) bound to the two most prominent human SIRPα alleles with greatly increased affinity relative to wild-type CD47 and potently antagonized CD47 binding to SIRPα on human macrophages. Velcro-CD47 synergizes with tumor-specific monoclonal antibodies to enhance macrophage phagocytosis of tumor cells in vitro, with similar potency as CV1. Finally, Velcro-CD47 interacts specifically with a subset of myeloid-derived cells in human blood, whereas CV1 binds all myeloid, lymphoid, and erythroid populations interrogated. This is consistent with the restricted expression of SIRPα compared with CD47. Herein, we have demonstrated that "Velcro" engineering is a powerful protein-engineering tool with potential applications to other systems and that Velcro-CD47 could be an alternative adjuvant to CD47-targeting agents for cancer immunotherapy.

Project description:BackgroundSevere combined immunodeficient (SCID) pigs are an emerging animal model being developed for biomedical and regenerative medicine research. SCID pigs can successfully engraft human-induced pluripotent stem cells and cancer cell lines. The development of a humanized SCID pig through xenotransplantation of human hematopoietic stem cells (HSCs) would be a further demonstration of the value of such a large animal SCID model. Xenotransplantation success with HSCs into non-obese diabetic (NOD)-derived SCID mice is dependent on the ability of NOD mouse signal regulatory protein alpha (SIRPA) to bind human CD47, inducing higher phagocytic tolerance than other mouse strains. Therefore, we investigated whether porcine SIRPA binds human CD47 in the context of developing a humanized SCID pig.MethodsPeripheral blood mononuclear cells (PBMCs) were collected from SCID and non-SCID pigs. Flow cytometry was used to assess whether porcine monocytes could bind to human CD47. Porcine monocytes were isolated from PBMCs and were subjected to phagocytosis assays with pig, human, and mouse red blood cell (RBC) targets. Blocking phagocytosis assays were performed by incubating human RBCs with anti-human CD47 blocking antibody B6H12, non-blocking antibody 2D3, and nonspecific IgG1 antibody and exposing to human or porcine monocytes.ResultsWe found that porcine SIRPA binds to human CD47 in vitro by flow cytometric assays. Additionally, phagocytosis assays were performed, and we found that porcine monocytes phagocytose human and porcine RBCs at significantly lower levels than mouse RBCs. When human RBCs were preincubated with CD47 antibodies B6H12 or 2D3, phagocytosis was induced only after B6H12 incubation, indicating the lower phagocytic activity of porcine monocytes with human cells requires interaction between porcine SIRPA and human CD47.ConclusionsWe have shown the first evidence that porcine monocytes can bind to human CD47 and are phagocytically tolerant to human cells, suggesting that porcine SCID models have the potential to support engraftment of human HSCs.

Project description:Phagocytosis is essential to maintain tissue homeostasis in a large number of inflammatory and autoimmune diseases, but its role in the diseased brain is poorly explored. Recent findings suggest that in the adult hippocampal neurogenic niche, where the excess of newborn cells undergo apoptosis in physiological conditions, phagocytosis is efficiently executed by surveillant, ramified microglia. To test whether microglia are efficient phagocytes in the diseased brain as well, we confronted them with a series of apoptotic challenges and discovered a generalized response. When challenged with excitotoxicity in vitro (via the glutamate agonist NMDA) or inflammation in vivo (via systemic administration of bacterial lipopolysaccharides or by omega 3 fatty acid deficient diets), microglia resorted to different strategies to boost their phagocytic efficiency and compensate for the increased number of apoptotic cells, thus maintaining phagocytosis and apoptosis tightly coupled. Unexpectedly, this coupling was chronically lost in a mouse model of mesial temporal lobe epilepsy (MTLE) as well as in hippocampal tissue resected from individuals with MTLE, a major neurological disorder characterized by seizures, excitotoxicity, and inflammation. Importantly, the loss of phagocytosis/apoptosis coupling correlated with the expression of microglial proinflammatory, epileptogenic cytokines, suggesting its contribution to the pathophysiology of epilepsy. The phagocytic blockade resulted from reduced microglial surveillance and apoptotic cell recognition receptor expression and was not directly mediated by signaling through microglial glutamate receptors. Instead, it was related to the disruption of local ATP microgradients caused by the hyperactivity of the hippocampal network, at least in the acute phase of epilepsy. Finally, the uncoupling led to an accumulation of apoptotic newborn cells in the neurogenic niche that was due not to decreased survival but to delayed cell clearance after seizures. These results demonstrate that the efficiency of microglial phagocytosis critically affects the dynamics of apoptosis and urge to routinely assess the microglial phagocytic efficiency in neurodegenerative disorders.

Dataset Information

Neuronal signal-regulatory protein alpha drives microglial phagocytosis by limiting microglial interaction with CD47 in the retina.

Publications

Neuronal signal-regulatory protein alpha drives microglial phagocytosis by limiting microglial interaction with CD47 in the retina.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets