Maturation-dependent complement production and C3 processing in human retinal pigment epithelium cells

Induced pluripotent stem cell-derived retinal pigment epithelial (iPSC-RPE) cells, closely resembling healthy RPE, offer valuable insights for retinal disease modelling. This study evaluates immature and mature phenotypes of iPSC-RPE and ARPE-19 cells, comparing RPE characteristics, cell-associated complement profiles, and TGF-{beta}1-mediated stress responses across transcript expression, secretion, and protein levels. Statistical analyses were performed using independent means t-test and Wilcoxon rank-sum test. Mature iPSC-RPE cells exhibited characteristic RPE-morphology, with elevated secretion of complement components (C3, FH/FHL-1, FI), unique FB secretion, and apical complement localisation. Intracellular C3 processing revealed cleavage products different from known blood-derived C3 fragments and showed maturation-dependent differences; only mature iPSC-RPE cells secreted active C3 forms (C3(H2O), C3a) and exhibited the intact C3 {beta}-chain. Mature ARPE-19 and iPSC-RPE cells demonstrated resistance to TGF-{beta}1 treatment, which reduced complement secretion without affecting C3a release. In conclusion, ARPE-19 and iPSC-RPE cells demonstrated local production of complement components and maturation-dependent intracellular processing of C3 into active forms. These findings highlight the impact of RPE maturation on local C3 activation, providing a basis for future studies on C3 functionality in the RPE and its potential pathological effects. Impact statementCell state-dependent local processing of complement C3 in retinal pigment epithelium cells generates active C3 forms that may evade anti-C3 therapies, influencing age-related macular degeneration and treatment outcomes. Funding statementThis project has received funding from the PRO RETINA, Amberg, Germany, under grant agreement number "Pro-Re/Projekt/Schikora - Pauly.05-2022", and Deutsche Forschungsgemeinschaft (DFG), under grant agreement number "498244102 (MA 6139/5-1)". Open access funding provided by the Open Access Publishing Fund of University Marburg. Funding sources were not involved in study design, data collection and interpretation, or the decision to submit the work for publication. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=149 SRC="FIGDIR/small/636432v1_ufig1.gif" ALT="Figure 1"> View larger version (67K): org.highwire.dtl.DTLVardef@6c63b3org.highwire.dtl.DTLVardef@58010org.highwire.dtl.DTLVardef@11ab625org.highwire.dtl.DTLVardef@ac4924_HPS_FORMAT_FIGEXP M_FIG C_FIG