title: Oxidative stress renders retinal pigment epithelial cells susceptible to complement-mediated injury.
abstract: Uncontrolled activation of the alternative pathway of complement is thought to be associated with age-related macular degeneration (AMD). The alternative pathway is continuously activated in the fluid phase, and tissue surfaces require continuous complement inhibition to prevent spontaneous autologous tissue injury. Here, we examined the effects of oxidative stress on the ability of immortalized human retinal pigment epithelial cells (ARPE-19) to regulate complement activation on their cell surface. Combined treatment with H(2)O(2) (to induce oxidative stress) and complement-sufficient serum was found to disrupt the barrier function of stable ARPE-19 monolayers as determined by transepithelial resistance (TER) measurements. Neither treatment alone had any effect. TER reduction was correlated with increased cell surface deposition of C3, and could be prevented by using C7-depleted serum, an essential component of the terminal complement pathway. Treatment with H(2)O(2) reduced surface expression of the complement inhibitors DAF, CD55, and CD59, and impaired regulation at the cell surface by factor H present within the serum. Combined treatment of the monolayers with H(2)O(2) and serum elicited polarized secretion of vascular epidermal growth factor (VEGF). Both, secretion of VEGF and TER reduction could be attenuated using either an alternative pathway inhibitor or by blocking VEGF receptor-1/2 signaling. Regarded together, these studies demonstrate that oxidative stress reduces regulation of complement on the surface of ARPE-19 cells, increasing complement activation. This sublytic activation results in VEGF release, which mediates disruption of the cell monolayer. These findings link oxidative stress, complement activation, and apical VEGF release, which have all been associated with the pathogenesis of AMD.
authors.affiliation: From the Department of Medicine, University of Colorado Denver School of Medicine, Denver, Colorado 80045.
authors.forename: Joshua M
authors.initials: JM
authors.lastname: Thurman
authors.affiliation: From the Department of Medicine, University of Colorado Denver School of Medicine, Denver, Colorado 80045.
authors.forename: Brandon
authors.initials: B
authors.lastname: Renner
authors.affiliation: Departments of Ophthalmology and Neurosciences, Division of Research, Charleston, South Carolina 29425.
authors.forename: Kannan
authors.initials: K
authors.lastname: Kunchithapautham
authors.affiliation: Department of Biochemistry, University of Texas Health Sciences Center, Tyler, Texas 75708.
authors.forename: Viviana P
authors.initials: VP
authors.lastname: Ferreira
authors.affiliation: Department of Biochemistry, University of Texas Health Sciences Center, Tyler, Texas 75708.
authors.forename: Michael K
authors.initials: MK
authors.lastname: Pangburn
authors.affiliation: Departments of Ophthalmology and Neurosciences, Division of Research, Charleston, South Carolina 29425.
authors.forename: Zsolt
authors.initials: Z
authors.lastname: Ablonczy
authors.affiliation: Department of Microbiology and Immunology, Medical University of South Carolina, Charleston, South Carolina 29425.
authors.forename: Stephen
authors.initials: S
authors.lastname: Tomlinson
authors.affiliation: From the Department of Medicine, University of Colorado Denver School of Medicine, Denver, Colorado 80045.
authors.forename: V Michael
authors.initials: VM
authors.lastname: Holers
authors.affiliation: Departments of Ophthalmology and Neurosciences, Division of Research, Charleston, South Carolina 29425. Electronic address: rohrer@musc.edu.
authors.forename: Bärbel
authors.initials: B
authors.lastname: Rohrer
mesh_terms: D018958:CD55 Antigens / Q000378:metabolism; D018957:CD59 Antigens / Q000378:metabolism; D002460:Cell Line; D017242:Complement Factor H / Q000378:metabolism; D003170:Complement Pathway, Alternative; D003165:Complement System Proteins / Q000378:metabolism*; D004847:Epithelial Cells / Q000276:immunology / Q000378:metabolism; D006801:Humans; D008268:Macular Degeneration / Q000209:etiology / Q000276:immunology / Q000378:metabolism; D018384:Oxidative Stress; D055213:Retinal Pigment Epithelium / Q000166:cytology / Q000276:immunology / Q000293:injuries* / Q000378:metabolism*; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
chemical_list: D018958:CD55 Antigens; D018957:CD59 Antigens; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; C051590:CD59 protein, human; D017242:Complement Factor H; D003165:Complement System Proteins