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W Fan, G Enzmann, PS Boro, NS Bora, HJ Kaplan, BJ McLaughlin; Complement and Complement Regulatory Proteins in RPE-Bruch’s Membrane-Choroidal Layers . Invest. Ophthalmol. Vis. Sci. 2002;43(13):2814.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:To investigate the immunolocalization and expression of complement and complement regulatory proteins in retinal pigment epithelium (RPE)-Bruch’s membrane- choroidal layers. Methods:Sections of the RPE/choroid from human donor eyes were examined by immunohistochemistry using antibodies directed against specific complement components (C3c, C5 and C5b-9) and complement regulatory proteins (membrane co-factor protein (MCP), decay accelerating factor (DAF) and CD59). Expression of MCP, DAF and CD59 was also examined by Western blot using RPE from donor eyes. In addition, expression of C3, C5, MCP, DAF and CD59 was examined using total RNA for RT-PCR and immunofluorescence microscopy in primary cultured human RPE and choroidal endothelium and ARPE-19 cell lines. Results:In situ studies with immunohistochemistry demonstrated C3c and C5 in the RPE, drusen and choroid with a greater prevalence for C3c compared to C5 in the RPE. Western blots showed the expression of MCP, DAF and CD59 in RPE from donor eyes. Immunohistochemistry confirmed the presence of these complement regulatory proteins with a preferential expression of MCP on the basal RPE surface and not the choroid. In contrast, DAF and CD59 were not localized to the basal RPE surface and were present in the choroid. In vitro studies using mRNA and protein assays revealed the presence of C3, C5, MCP, DAF and CD59 in both RPE and choroid. Conclusion:Our results establish the presence of the major complement components C3 and C5 in the RPE and choroid, as well as complement regulatory proteins. Additionally, there is a preferential distribution of MCP on the basal surface of the RPE. The functional significance of this observation is being explored.
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