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Degradation of Photoreceptor Outer Segments by the Retinal Pigment Epithelium Requires Pigment Epithelium-Derived Factor Receptor (PEDF-R)
PURPOSE: To examine the contribution of pigment epithelium-derived factor receptor (PEDF-R) to the phagocytosis process. Previously, we identified PEDF-R, the protein encoded by the PNPLA2 gene, as a phospholipase A2 in the retinal pigment epithelium (RPE). During phagocytosis, RPE cells ingest abun...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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The Association for Research in Vision and Ophthalmology
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7900850/ https://www.ncbi.nlm.nih.gov/pubmed/33605986 http://dx.doi.org/10.1167/iovs.62.2.30 |
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author | Bullock, Jeanee Polato, Federica Abu-Asab, Mones Bernardo-Colón, Alexandra Aflaki, Elma Agbaga, Martin-Paul Becerra, S. Patricia |
author_facet | Bullock, Jeanee Polato, Federica Abu-Asab, Mones Bernardo-Colón, Alexandra Aflaki, Elma Agbaga, Martin-Paul Becerra, S. Patricia |
author_sort | Bullock, Jeanee |
collection | PubMed |
description | PURPOSE: To examine the contribution of pigment epithelium-derived factor receptor (PEDF-R) to the phagocytosis process. Previously, we identified PEDF-R, the protein encoded by the PNPLA2 gene, as a phospholipase A2 in the retinal pigment epithelium (RPE). During phagocytosis, RPE cells ingest abundant phospholipids and protein in the form of photoreceptor outer segment (POS) tips, which are then hydrolyzed. The role of PEDF-R in RPE phagocytosis is not known. METHODS: Mice in which PNPLA2 was conditionally knocked out (cKO) in the RPE were generated. Mouse RPE/choroid explants were cultured. Human ARPE-19 cells were transfected with siPNPLA2 silencing duplexes. POSs were isolated from bovine retinas. The phospholipase A2 inhibitor bromoenol lactone was used. Transmission electron microscopy, immunofluorescence, lipid labeling, pulse–chase experiments, western blots, and free fatty acid and β-hydroxybutyrate assays were performed. RESULTS: The RPE of the cKO mice accumulated lipids, as well as more abundant and larger rhodopsin particles, compared to littermate controls. Upon POS exposure, RPE explants from cKO mice released less β-hydroxybutyrate compared to controls. After POS ingestion during phagocytosis, rhodopsin degradation was stalled both in cells treated with bromoenol lactone and in PNPLA2-knocked-down cells relative to their corresponding controls. Phospholipase A2 inhibition lowered β-hydroxybutyrate release from phagocytic RPE cells. PNPLA2 knockdown also resulted in a decline in fatty acids and β-hydroxybutyrate release from phagocytic RPE cells. CONCLUSIONS: PEDF-R downregulation delayed POS digestion during phagocytosis. The findings imply that the efficiency of RPE phagocytosis depends on PEDF-R, thus identifying a novel contribution of this protein to POS degradation in the RPE. |
format | Online Article Text |
id | pubmed-7900850 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | The Association for Research in Vision and Ophthalmology |
record_format | MEDLINE/PubMed |
spelling | pubmed-79008502021-03-03 Degradation of Photoreceptor Outer Segments by the Retinal Pigment Epithelium Requires Pigment Epithelium-Derived Factor Receptor (PEDF-R) Bullock, Jeanee Polato, Federica Abu-Asab, Mones Bernardo-Colón, Alexandra Aflaki, Elma Agbaga, Martin-Paul Becerra, S. Patricia Invest Ophthalmol Vis Sci Biochemistry and Molecular Biology PURPOSE: To examine the contribution of pigment epithelium-derived factor receptor (PEDF-R) to the phagocytosis process. Previously, we identified PEDF-R, the protein encoded by the PNPLA2 gene, as a phospholipase A2 in the retinal pigment epithelium (RPE). During phagocytosis, RPE cells ingest abundant phospholipids and protein in the form of photoreceptor outer segment (POS) tips, which are then hydrolyzed. The role of PEDF-R in RPE phagocytosis is not known. METHODS: Mice in which PNPLA2 was conditionally knocked out (cKO) in the RPE were generated. Mouse RPE/choroid explants were cultured. Human ARPE-19 cells were transfected with siPNPLA2 silencing duplexes. POSs were isolated from bovine retinas. The phospholipase A2 inhibitor bromoenol lactone was used. Transmission electron microscopy, immunofluorescence, lipid labeling, pulse–chase experiments, western blots, and free fatty acid and β-hydroxybutyrate assays were performed. RESULTS: The RPE of the cKO mice accumulated lipids, as well as more abundant and larger rhodopsin particles, compared to littermate controls. Upon POS exposure, RPE explants from cKO mice released less β-hydroxybutyrate compared to controls. After POS ingestion during phagocytosis, rhodopsin degradation was stalled both in cells treated with bromoenol lactone and in PNPLA2-knocked-down cells relative to their corresponding controls. Phospholipase A2 inhibition lowered β-hydroxybutyrate release from phagocytic RPE cells. PNPLA2 knockdown also resulted in a decline in fatty acids and β-hydroxybutyrate release from phagocytic RPE cells. CONCLUSIONS: PEDF-R downregulation delayed POS digestion during phagocytosis. The findings imply that the efficiency of RPE phagocytosis depends on PEDF-R, thus identifying a novel contribution of this protein to POS degradation in the RPE. The Association for Research in Vision and Ophthalmology 2021-02-19 /pmc/articles/PMC7900850/ /pubmed/33605986 http://dx.doi.org/10.1167/iovs.62.2.30 Text en Copyright 2021 The Authors http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License. |
spellingShingle | Biochemistry and Molecular Biology Bullock, Jeanee Polato, Federica Abu-Asab, Mones Bernardo-Colón, Alexandra Aflaki, Elma Agbaga, Martin-Paul Becerra, S. Patricia Degradation of Photoreceptor Outer Segments by the Retinal Pigment Epithelium Requires Pigment Epithelium-Derived Factor Receptor (PEDF-R) |
title | Degradation of Photoreceptor Outer Segments by the Retinal Pigment Epithelium Requires Pigment Epithelium-Derived Factor Receptor (PEDF-R) |
title_full | Degradation of Photoreceptor Outer Segments by the Retinal Pigment Epithelium Requires Pigment Epithelium-Derived Factor Receptor (PEDF-R) |
title_fullStr | Degradation of Photoreceptor Outer Segments by the Retinal Pigment Epithelium Requires Pigment Epithelium-Derived Factor Receptor (PEDF-R) |
title_full_unstemmed | Degradation of Photoreceptor Outer Segments by the Retinal Pigment Epithelium Requires Pigment Epithelium-Derived Factor Receptor (PEDF-R) |
title_short | Degradation of Photoreceptor Outer Segments by the Retinal Pigment Epithelium Requires Pigment Epithelium-Derived Factor Receptor (PEDF-R) |
title_sort | degradation of photoreceptor outer segments by the retinal pigment epithelium requires pigment epithelium-derived factor receptor (pedf-r) |
topic | Biochemistry and Molecular Biology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7900850/ https://www.ncbi.nlm.nih.gov/pubmed/33605986 http://dx.doi.org/10.1167/iovs.62.2.30 |
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