title: Complement Factor H Expressed by Retinal Pigment Epithelium Cells Can Suppress Neovascularization of Human Umbilical Vein Endothelial Cells: An in vitro Study.
abstract: Complement factor H (CFH) is one of the most important soluble complement regulatory proteins and is closely associated with age-related macular degeneration (AMD), the leading cause of irreversible central vision loss in the elderly population in developed countries. Our study searches to investigate whether CFH expression is changed in oxidative damaged retinal pigment epithelium (RPE) cells and the role of CFH in the in vitro neovascularization. First, it was confirmed by immunofluorescence staining that CFH was expressed by ARPE-19 cells. CFH mRNA and protein in oxidative (H2O2) damaged ARPE-19 cells were both reduced, as determined by Real-time PCR and Western blotting analysis. Enzyme-linked immunosorbent assay (ELISA) also showed that ARPE-19 cells treated with H2O2 caused an increase in C3a content, which indicates complement activation. Then, wound assays were performed to show that CFH expression suppression promoted human umbilical vein endothelial cell (HUVECs) migration. Thereafter, ARPE-19 cells were transfected with CFH-specific siRNA and CFH knockdown was confirmed with the aid of Real-time PCR, immunofluorescence staining and Western blotting. The ELISA results showed that specific CFH knockdown in ARPE-19 cells activated the complement system. Finally, in vitro matrigel tube formation assay was performed to determine whether change of CFH expression in RPE would affect tube formation by HUVECs. More tubes were formed by HUVECs co-cultured with ARPE-19 cells transfected with CFH specific-siRNA when compared with controls. Our results suggested that RPE cells might be the local CFH source, and RPE cell injuries (such as oxidative stress) may cause CFH expression suppression, which in turn may lead to complement activation and promotion of tube formation by HUVECs. This finding is of importance in elucidating the role of complement in the pathogenesis of ocular neovascularization including choroidal neovascularization.
authors.affiliation: Department of Ophthalmology, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Department of Ophthalmology, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China.
authors.forename: Qing
authors.initials: Q
authors.lastname: Huang
authors.affiliation: Department of Ophthalmology, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China.
authors.forename: Min
authors.initials: M
authors.lastname: Tang
authors.affiliation: Department of Ophthalmology, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China.
authors.forename: Junjun
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Department of Ophthalmology, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Fan
mesh_terms: D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects; D002470:Cell Survival; D002478:Cells, Cultured; D003167:Complement Activation / Q000235:genetics / Q000276:immunology; D017242:Complement Factor H / Q000235:genetics* / Q000378:metabolism; D015870:Gene Expression; D055785:Gene Knockdown Techniques; D061307:Human Umbilical Vein Endothelial Cells / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism / Q000494:pharmacology; D066298:In Vitro Techniques; D018919:Neovascularization, Physiologic / Q000235:genetics*; D018384:Oxidative Stress; D055213:Retinal Pigment Epithelium / Q000276:immunology / Q000378:metabolism*
chemical_list: D017242:Complement Factor H; D006861:Hydrogen Peroxide