Complement Factor H-Related 3 Enhanced Inflammation and Complement Activation in Human RPE Cells.

Nicole Schäfer,Anas Rasras,Volker Enzmann,Diana Pauly,Sabine Amslinger,Herbert Jägle,Delia M Ormenisan

doi:10.3389/fimmu.2021.769242

Abstract

Complement Factor H-Related 3 (FHR-3) is a major regulator of the complement system, which is associated with different diseases, such as age-related macular degeneration (AMD). However, the non-canonical local, cellular functions of FHR-3 remained poorly understood. Here, we report that FHR-3 bound to oxidative stress epitopes and competed with FH for interaction. Furthermore, FHR-3 was internalized by viable RPE cells and modulated time-dependently complement component (C3, FB) and receptor (C3aR, CR3) expression of human RPE cells. Independently of any external blood-derived proteins, complement activation products were detected. Anaphylatoxin C3a was visualized in treated cells and showed a translocation from the cytoplasm to the cell membrane after FHR-3 exposure. Subsequently, FHR-3 induced a RPE cell dependent pro-inflammatory microenvironment. Inflammasome NLRP3 activation and pro-inflammatory cytokine secretion of IL-1ß, IL-18, IL-6 and TNF-α were induced after FHR-3-RPE interaction. Our previously published monoclonal anti-FHR-3 antibody, which was chimerized to reduce immunogenicity, RETC-2-ximab, ameliorated the effect of FHR-3 on ARPE-19 cells. Our studies suggest FHR-3 as an exogenous trigger molecule for the RPE cell “complosome” and as a putative target for a therapeutic approach for associated degenerative diseases.

Highlights

Studied for more than one century, but still not fully understood – the evolutionary ancient complement system is one of the first defense line of our body to protect from pathogens, cellular debris or dead cells
Due to our additional results, showing that retinal pigment epithelium (RPE) cells in response to stress increased the expression of complement components and pro-inflammatory factors [30], we proposed that Factor H-Related 3 (FHR-3) could be a stress factor for the RPE in the aged retina promoting retinal degeneration [9]
Homogenous age-related macular degeneration (AMD)-risk Single nucleotide polymorphisms (SNP) could not be detected in the examined RPE cells, instead heterozygous SNPs were present in the complement factor H (CFH) and C3 gene of human adult retinal pigment epithelium cells (ARPE-19) and CFH, C2/CFB, CFI and ARMS gene of Human primary retinal pigment epithelial cells (hpRPE) cells (Supplementary Figure 1A)

Summary

Introduction

Studied for more than one century, but still not fully understood – the evolutionary ancient complement system is one of the first defense line of our body to protect from pathogens, cellular debris or dead cells. The complement system is tightly regulated to prevent dysregulation leading to progression of different degenerative diseases such as age-related macular degeneration (AMD) [1]. One major group of soluble regulators are Abbreviations: ACTA2, a‐smooth muscle actin; AMD, age-related macular degeneration; ARPE-19, human adult retinal pigment epithelium cells; BSA, bovine serum albumin; CCl, cellular capacitance; CD11b, cluster of differentiation molecule 11B (CR3 subunit); CEP, w−carboxyethyl)pyrrole; COL1A1, collagen type I, alpha 1; CR3, complement receptor 3; EMT, epithelial-mesenchymal transition; FH, factor H; FHL-1, factor-H like protein 1; FHR-1 – 5, factor H-related proteins 1 – 5; FOXP3, forkhead box P3; FP, properdin; hpRPE, primary human RPE cells; MAA, malondialdehydeacetaldehyde; MDA, malondialdehyde; NLRP3, NLR family pyrin domain containing 3; OSE, oxidation-specific epitopes; RETC-2, REgensburg Therapy Complement 2 (anti-FHR-3 antibody); RETC-2-ximab, chimerized REgensburg Therapy Complement 2 (anti-FHR-3 antibody); RPE, retinal pigment epithelium; SNP, single nucleotide polymorphisms; TER, Transepithelial Resistance; TNF, tumor necrosis factor; VIM, vimentin; ZO-1, zonula occludens 1. While the most relevant CFH SNP (Y402H) contributed to AMD progression, a joint deletion of the complement factor-H related 1 and 3 (DCFHR3/1) genes was protective for the disease [7, 8]. A local expression could be reported for FHR-3 by retinal macrophages/microglia cells [9] and for FH and FHL-1 by retinal pigment epithelium (RPE) cells [10]

Methods

Results

Conclusion