Oxidative stress in retinal pigment epithelial cells increased endogenous complement-dependent inflammatory and angiogenic responses - independent from exogenous complement sources

Abstract

15 16 Page 2 of 35 HIGHLIGHTS 17  Oxidative stress accumulates complement proteins and receptors in RPE cells 18  Oxidative stress activates the RPE inflammasome without external complement proteins 19  Oxidative stress increases foxp3 expression and IL-8/VEGF secretion in RPE cells 20  Olaparib enhances pro-inflammatory response of RPE 21 22 ABSTRACT 23Oxidative stress-induced damage of the retinal pigment epithelium (RPE) together with chronic 24 inflammation has been suggested as major contributors to retinal diseases. Here, we examine the 25 effects of oxidative stress and endogenous complement components on the RPE and its pro-26 inflammatory and -angiogenic responses. 27The RPE cell line, ARPE-19, treated with H 2 O 2 reduced cell-cell contacts, increased marker for 28 epithelial-mesenchymal transition but showed less cell death. Stressed ARPE-19 cells increased the 29 expression of complement receptors CR3 and C5aR1. CR3 was co-localized with cell-derived 30 complement protein C3, which was observed in its activated form in ARPE-19 cells. C3 as well as its 31 regulators CFH and properdin accumulated in ARPE-19 cells after oxidative stress independent from 32 external complement sources. This cell-associated complement accumulation promoted nlrp3 and 33 foxp3 expression and subsequent increased secretion of pro-inflammatory and pro-angiogenic factors. 34The complement-associated ARPE-19 reaction to oxidative stress, independent from external 35 complement source, was increased by the PARP-inhibitor olaparib. 36Our results indicated that RPE cell-derived complement proteins and receptors are involved in RPE cell 37 homeostasis following oxidative stress and should be considered as targets for treatment 38 developments for retinal degeneration. 39 40 Page 4 of 35 GRAPHICAL ABSTRACT 41 42We show a functional link between oxidative stress, complement receptors, endogenous complement 43 proteins, pro-angiogenic and -inflammatory responses in ARPE-19 cells. These effects are independent 44 from extracellularly added complement proteins or receptor ligands. We suggest an oxidative stress-45 associated autocrine mechanism of complement receptor regulation in ARPE-19 cells in connection 46 with upregulated intracellular proteases. 47