Complement-Mediated Regulation of Apolipoprotein E in Cultured Human RPE Cells.
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Complement activation is implicated in the pathogenesis of age-related macular degeneration (AMD). Apolipoprotein E (ApoE) and complement activation products such as membrane attack complex (MAC) are present in eyes of individuals with AMD. Herein, we investigated the effect of complement activation on induction of ApoE accumulation in human retinal pigment epithelial (RPE) cells.Cultured human RPE cells were primed with a complement-fixing antibody followed by treatment with C1q-depleted (C1q-Dep) human serum to elicit alternative pathway complement activation. Controls included anti-C5 antibody-treated serum and heat-inactivated C1q-Dep. Total protein was determined on RPE cell extracts, conditioned media, and extracellular matrix (ECM) by Western blot. ApoE and MAC colocalization was assessed on cultured RPE cells and human eyes by immunofluorescent stain. ApoE mRNA expression was evaluated by quantitative PCR (qPCR).Complement challenge upregulated cell-associated ApoE, but not apolipoprotein A1. ApoE accumulation was blocked by anti-C5 antibody and enhanced by repetitive complement challenge. ApoE mRNA levels were not affected by complement challenge. ApoE was frequently colocalized with MAC in complement-treated cells and drusen from human eyes. ApoE was released into complement-treated conditioned media after a single complement challenge and accumulated on ECM after repetitive complement challenge.Complement challenge induces time-dependent ApoE accumulation in RPE cells. An understanding of the mechanisms by which complement affects RPE ApoE accumulation may help to better explain drusen composition, and provide insights into potential therapeutic targets.
Gene Expression Regulation
Retinal Pigment Epithelium
Real-Time Polymerase Chain Reaction
Published Version (Please cite this version)10.1167/iovs.16-20083
Publication InfoYang, Ping; Skiba, Nikolai P; Tewkesbury, Grace M; Treboschi, Victoria M; Baciu, Peter; & Jaffe, Glenn J (2017). Complement-Mediated Regulation of Apolipoprotein E in Cultured Human RPE Cells. Investigative ophthalmology & visual science, 58(7). pp. 3073-3085. 10.1167/iovs.16-20083. Retrieved from https://hdl.handle.net/10161/17218.
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Robert Machemer M.D. Distinguished Professor of Ophthalmology
I have an active basic and clinical research program. I have been principal investigator on several funded clinical studies including investigations of an oral ganciclovir prodrug to treat CMV retinitis and a study to determine the safety of a cyclosporine sustained drug delivery implant in the treatment of uveitis. I am principal investigator of an ongoing multicenter trial of a fluocinolone sustained drug delivery implant to treat patients with severe uveitis and a trial of this same i
Associate Professor in Ophthalmology
My research focuses on applying mass spectrometry based proteomics to study proteins in eye tissues, cells and sub-cellular compartments to understand mechanisms of vision. An important aspect of my research is to identify proteins in different compartments of retinal photoreceptor cells, their amount and modification status at different cell states defined by the light conditions, genotype, disease etc. This information can be valuable in understanding molecular mechanisms of vision and bi
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