Overexpression of angiotensin-converting enzyme 2 ameliorates amyloid β-induced inflammatory response in human primary retinal pigment epithelium

Abstract

PURPOSE. Amyloid-b (Ab) is a major constituent of drusen, which is a hallmark of early AMD. The purpose of this study was to investigate whether enhancement of ACE2, an important component of the protective angiotensin-converting enzyme 2 (ACE2)/Ang-(1-7)/Mas axis of the renin angiotensin system (RAS), ameliorates Ab-induced inflammatory response in human RPE cells. METHODS. Annexin-V FITC/propidium iodide assay for detecting apoptosis rate was used to determine the optimum concentration and incubation time of Ab1-42. ACE2 plasmid was transfected into primary cultured human RPE (hRPE) and ARPE-19 cells for 6 hours followed by stimulation with Ab1-42 at the concentration of 1 lM for 48 hours. Gene expression was detected by real-time PCR and protein levels were determined by Western blotting or ELISA. A779, an antagonist of Ang-(1-7), was used to further confirm the involvement of ACE2/Ang-(1-7)/Mas axis. RESULTS. Flow cytometry showed that the optimal concentration of Ab1-42 was 1 lM and the optimal incubation time was 48 hours. Ab1-42 upregulated the expression of IL-1b and monocyte chemoattractant protein-1. ACE2 plasmid significantly upregulated the expression of ACE2 and Ang-(1-7) in hRPE and ARPE-19 cells. Activation of ACE2 reduced the overproduction of inflammatory cytokines at both mRNA and protein levels in hRPE and ARPE-19 cells stimulated with Ab1-42. Furthermore, an antagonist of Ang-(1-7), A779 reversed the anti-inflammatory effect of ACE2. CONCLUSIONS. Overexpression of ACE2 ameliorates Ab-induced inflammatory response by activating the ACE2/Ang-(1-7)/Mas axis in human RPE cells. Our data suggest that ACE2/Ang-(1-7)/Mas axis may be a promising target for developing novel therapies for inflammation response in AMD.