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Functional Evaluation of AMD-Associated Risk Variants of Complement Factor B

PURPOSE: The 32W and 32Q variants of complement factor B (CFB) are associated with reduced risk of developing neovascular age-related macular degeneration (AMD) compared with the common 32R allele. The objective of this study was to determine if the most protective R32Q variant affects the neovascul...

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Autores principales: Pilotti, Camilla, Greenwood, John, Moss, Stephen E.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The Association for Research in Vision and Ophthalmology 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7405614/
https://www.ncbi.nlm.nih.gov/pubmed/32407521
http://dx.doi.org/10.1167/iovs.61.5.19
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author Pilotti, Camilla
Greenwood, John
Moss, Stephen E.
author_facet Pilotti, Camilla
Greenwood, John
Moss, Stephen E.
author_sort Pilotti, Camilla
collection PubMed
description PURPOSE: The 32W and 32Q variants of complement factor B (CFB) are associated with reduced risk of developing neovascular age-related macular degeneration (AMD) compared with the common 32R allele. The objective of this study was to determine if the most protective R32Q variant affects the neovascular process in a manner consistent with the reported reduced disease association. METHODS: The 32R, 32W, and 32Q human CFB variants were expressed in human embryonic kidney 293T cells and purified from culture supernatant. The ex vivo mouse fetal metatarsal explant model was used to investigate the effect of these three human CFB variants on angiogenesis. Metatarsal bones were isolated from mouse embryos and cultured in the presence of the three CFB variants, and angiogenesis was measured following immunostaining of fixed samples. ELISAs were used to quantify C3 and VEGF protein levels in metatarsal culture and quantitative PCR to measure Cfb, C3, and Vegf expression. RESULTS: We show here that the three CFB variants have different biological activities in the mouse metatarsal assay, with CFB(R32) exhibiting significantly greater angiogenic activity than CFB(Q32) or CFB(W32), which were broadly similar. We also observed differences in macrophage phenotype with these two variants that may contribute to their activities in this experimental model. CONCLUSIONS: We have demonstrated that the biological activities of CFB(R32), CFB(W32), and CFB(Q32) are consistent with their AMD risk association, and we provide functional evidence of roles for these variants in angiogenesis that may be relevant to the pathogenesis of the neovascular form of AMD.
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spelling pubmed-74056142020-08-19 Functional Evaluation of AMD-Associated Risk Variants of Complement Factor B Pilotti, Camilla Greenwood, John Moss, Stephen E. Invest Ophthalmol Vis Sci Retinal Cell Biology PURPOSE: The 32W and 32Q variants of complement factor B (CFB) are associated with reduced risk of developing neovascular age-related macular degeneration (AMD) compared with the common 32R allele. The objective of this study was to determine if the most protective R32Q variant affects the neovascular process in a manner consistent with the reported reduced disease association. METHODS: The 32R, 32W, and 32Q human CFB variants were expressed in human embryonic kidney 293T cells and purified from culture supernatant. The ex vivo mouse fetal metatarsal explant model was used to investigate the effect of these three human CFB variants on angiogenesis. Metatarsal bones were isolated from mouse embryos and cultured in the presence of the three CFB variants, and angiogenesis was measured following immunostaining of fixed samples. ELISAs were used to quantify C3 and VEGF protein levels in metatarsal culture and quantitative PCR to measure Cfb, C3, and Vegf expression. RESULTS: We show here that the three CFB variants have different biological activities in the mouse metatarsal assay, with CFB(R32) exhibiting significantly greater angiogenic activity than CFB(Q32) or CFB(W32), which were broadly similar. We also observed differences in macrophage phenotype with these two variants that may contribute to their activities in this experimental model. CONCLUSIONS: We have demonstrated that the biological activities of CFB(R32), CFB(W32), and CFB(Q32) are consistent with their AMD risk association, and we provide functional evidence of roles for these variants in angiogenesis that may be relevant to the pathogenesis of the neovascular form of AMD. The Association for Research in Vision and Ophthalmology 2020-05-14 /pmc/articles/PMC7405614/ /pubmed/32407521 http://dx.doi.org/10.1167/iovs.61.5.19 Text en Copyright 2020 The Authors http://creativecommons.org/licenses/by/4.0/ This work is licensed under a Creative Commons Attribution 4.0 International License.
spellingShingle Retinal Cell Biology
Pilotti, Camilla
Greenwood, John
Moss, Stephen E.
Functional Evaluation of AMD-Associated Risk Variants of Complement Factor B
title Functional Evaluation of AMD-Associated Risk Variants of Complement Factor B
title_full Functional Evaluation of AMD-Associated Risk Variants of Complement Factor B
title_fullStr Functional Evaluation of AMD-Associated Risk Variants of Complement Factor B
title_full_unstemmed Functional Evaluation of AMD-Associated Risk Variants of Complement Factor B
title_short Functional Evaluation of AMD-Associated Risk Variants of Complement Factor B
title_sort functional evaluation of amd-associated risk variants of complement factor b
topic Retinal Cell Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7405614/
https://www.ncbi.nlm.nih.gov/pubmed/32407521
http://dx.doi.org/10.1167/iovs.61.5.19
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