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Primary phagocytosis of viable neurons by microglia activated with LPS or Aβ is dependent on calreticulin/LRP phagocytic signalling

BACKGROUND: Microglia are resident brain macrophages that can phagocytose dead, dying or viable neurons, which may be beneficial or detrimental in inflammatory, ischaemic and neurodegenerative brain pathologies. Cell death caused by phagocytosis of an otherwise viable cell is called ‘primary phagocy...

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Autores principales: Fricker, Michael, Oliva-Martín, María José, Brown, Guy C
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3481398/
https://www.ncbi.nlm.nih.gov/pubmed/22889139
http://dx.doi.org/10.1186/1742-2094-9-196
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author Fricker, Michael
Oliva-Martín, María José
Brown, Guy C
author_facet Fricker, Michael
Oliva-Martín, María José
Brown, Guy C
author_sort Fricker, Michael
collection PubMed
description BACKGROUND: Microglia are resident brain macrophages that can phagocytose dead, dying or viable neurons, which may be beneficial or detrimental in inflammatory, ischaemic and neurodegenerative brain pathologies. Cell death caused by phagocytosis of an otherwise viable cell is called ‘primary phagocytosis’ or ‘phagoptosis’. Calreticulin (CRT) exposure on the surface of cancer cells can promote their phagocytosis via LRP (low-density lipoprotein receptor-related protein) on macrophages, but it is not known whether this occurs with neurons and microglia. METHODS: We used primary cultures of cerebellar neurons, astrocytes and microglia to investigate the potential role of CRT/LRP phagocytic signalling in the phagocytosis of viable neurons by microglia stimulated with lipopolysaccharide (LPS) or nanomolar concentrations of amyloid-β peptide(1-42) (Aβ). Exposure of CRT on the neuronal surface was investigated using surface biotinylation and western blotting. A phagocytosis assay was also developed using BV2 and PC12 cell lines to investigate CRT/LRP signalling in microglial phagocytosis of apoptotic cells. RESULTS: We found that BV2 microglia readily phagocytosed apoptotic PC12 cells, but this was inhibited by a CRT-blocking antibody or LRP-blocking protein (receptor-associated protein: RAP). Activation of primary rat microglia with LPS or Aβ resulted in loss of co-cultured cerebellar granule neurons, and this was blocked by RAP or antibodies against CRT or against LRP, preventing all neuronal loss and death. CRT was present on the surface of viable neurons, and this exposure did not change in inflammatory conditions. CRT antibodies prevented microglia-induced neuronal loss when added to neurons, while LRP antibodies prevented neuronal loss when added to the microglia. Pre-binding of CRT to neurons promoted neuronal loss if activated microglia were added, but pre-binding of CRT to microglia or both cell types prevented microglia-induced neuronal loss. CONCLUSIONS: CRT exposure on the surface of viable or apoptotic neurons appears to be required for their phagocytosis via LRP receptors on activated microglia, but free CRT can block microglial phagocytosis of neurons by acting on microglia. Phagocytosis of CRT-exposing neurons by microglia can be a direct cause of neuronal death during inflammation, and might therefore contribute to neurodegeneration and be prevented by blocking the CRT/LRP pathway.
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spelling pubmed-34813982012-10-27 Primary phagocytosis of viable neurons by microglia activated with LPS or Aβ is dependent on calreticulin/LRP phagocytic signalling Fricker, Michael Oliva-Martín, María José Brown, Guy C J Neuroinflammation Research BACKGROUND: Microglia are resident brain macrophages that can phagocytose dead, dying or viable neurons, which may be beneficial or detrimental in inflammatory, ischaemic and neurodegenerative brain pathologies. Cell death caused by phagocytosis of an otherwise viable cell is called ‘primary phagocytosis’ or ‘phagoptosis’. Calreticulin (CRT) exposure on the surface of cancer cells can promote their phagocytosis via LRP (low-density lipoprotein receptor-related protein) on macrophages, but it is not known whether this occurs with neurons and microglia. METHODS: We used primary cultures of cerebellar neurons, astrocytes and microglia to investigate the potential role of CRT/LRP phagocytic signalling in the phagocytosis of viable neurons by microglia stimulated with lipopolysaccharide (LPS) or nanomolar concentrations of amyloid-β peptide(1-42) (Aβ). Exposure of CRT on the neuronal surface was investigated using surface biotinylation and western blotting. A phagocytosis assay was also developed using BV2 and PC12 cell lines to investigate CRT/LRP signalling in microglial phagocytosis of apoptotic cells. RESULTS: We found that BV2 microglia readily phagocytosed apoptotic PC12 cells, but this was inhibited by a CRT-blocking antibody or LRP-blocking protein (receptor-associated protein: RAP). Activation of primary rat microglia with LPS or Aβ resulted in loss of co-cultured cerebellar granule neurons, and this was blocked by RAP or antibodies against CRT or against LRP, preventing all neuronal loss and death. CRT was present on the surface of viable neurons, and this exposure did not change in inflammatory conditions. CRT antibodies prevented microglia-induced neuronal loss when added to neurons, while LRP antibodies prevented neuronal loss when added to the microglia. Pre-binding of CRT to neurons promoted neuronal loss if activated microglia were added, but pre-binding of CRT to microglia or both cell types prevented microglia-induced neuronal loss. CONCLUSIONS: CRT exposure on the surface of viable or apoptotic neurons appears to be required for their phagocytosis via LRP receptors on activated microglia, but free CRT can block microglial phagocytosis of neurons by acting on microglia. Phagocytosis of CRT-exposing neurons by microglia can be a direct cause of neuronal death during inflammation, and might therefore contribute to neurodegeneration and be prevented by blocking the CRT/LRP pathway. BioMed Central 2012-08-13 /pmc/articles/PMC3481398/ /pubmed/22889139 http://dx.doi.org/10.1186/1742-2094-9-196 Text en Copyright ©2012 Fricker et al.; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Research
Fricker, Michael
Oliva-Martín, María José
Brown, Guy C
Primary phagocytosis of viable neurons by microglia activated with LPS or Aβ is dependent on calreticulin/LRP phagocytic signalling
title Primary phagocytosis of viable neurons by microglia activated with LPS or Aβ is dependent on calreticulin/LRP phagocytic signalling
title_full Primary phagocytosis of viable neurons by microglia activated with LPS or Aβ is dependent on calreticulin/LRP phagocytic signalling
title_fullStr Primary phagocytosis of viable neurons by microglia activated with LPS or Aβ is dependent on calreticulin/LRP phagocytic signalling
title_full_unstemmed Primary phagocytosis of viable neurons by microglia activated with LPS or Aβ is dependent on calreticulin/LRP phagocytic signalling
title_short Primary phagocytosis of viable neurons by microglia activated with LPS or Aβ is dependent on calreticulin/LRP phagocytic signalling
title_sort primary phagocytosis of viable neurons by microglia activated with lps or aβ is dependent on calreticulin/lrp phagocytic signalling
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3481398/
https://www.ncbi.nlm.nih.gov/pubmed/22889139
http://dx.doi.org/10.1186/1742-2094-9-196
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