Interaction between CD177 and platelet endothelial cell adhesion molecule-1 downregulates membrane-bound proteinase-3 (PR3) expression on neutrophils and attenuates neutrophil activation induced by PR3-ANCA

BACKGROUND: A recent study found that CD177 served as a receptor of membrane-bound proteinase-3 (mPR3) in a subset of neutrophils. Furthermore, CD177 has been identified as a high-affinity heterophilic binding partner for the endothelial cell platelet endothelial cell adhesion molecule-1 (PECAM-1). The current study aimed to investigate whether the interaction between PECAM-1 and CD177 could influence mPR3 expression as well as PR3-antineutrophil cytoplasmic antibody (ANCA)-induced neutrophil activation and glomerular endothelial cell (GEnC) injury. METHODS: The effect of interaction between CD177 and PECAM-1 on mPR3 expression was explored by enzyme-linked immunosorbent assay (ELISA) and flow cytometry. The effect of PECAM-1 on neutrophil activation and GEnC injury induced by PR3-ANCA-positive immunoglobulin (Ig)Gs was evaluated by dihydrorhodamine (DHR) assay and ELISA. CD177-negative neutrophils were selected by magnetic cell sorting (MACS), and the inhibitory effect of PECAM-1 on CD177-negative and mixed neutrophils was explored by measuring neutrophil degranulation. RESULTS: The level of specific interaction between CD177 and PECAM-1 was elevated with increasing CD177 concentration. The expression of mPR3 significantly decreased in neutrophils preincubated with PECAM-1 in a dose-dependent manner. Consistently, the levels of respiratory burst and degranulation induced by PR3-ANCA-positive IgGs in recombinant human tumor necrosis factor-alpha (TNF-α)-primed neutrophils was significantly reduced by preincubation with PECAM-1 (440.6 ± 123.0 vs. 511.4 ± 95.5, p < 0.05; and 3155.0 ± 1733.0 ng/ml vs. 5903.0 ± 717.5 ng/ml, p < 0.05, respectively). In CD177-negative neutrophils incubated with PR3-ANCA-positive IgGs, the level of degranulation was not significantly changed by preincubation with PECAM-1. However, in mixed neutrophils, PECAM-1 significantly decreased the level of degranulation induced by PR3-ANCA-positive IgGs (1015.9 ± 229.2% vs. 1725.2 ± 412.4%, p < 0.01). Furthermore, with preincubation of TNF-α-primed neutrophils with PECAM-1, the level of soluble intercellular cell adhesion molecule-1 (sICAM-1), a marker of endothelial cell activation and injury, in the supernatant of GEnCs treated with primed neutrophils plus PR3-ANCA-positive IgGs was significantly attenuated (112.7 ± 24.2 pg/ml vs. 167.5 ± 27.7 pg/ml, p < 0.05). CONCLUSIONS: PECAM-1 can decrease the level of mPR3 expression on neutrophils, resulting in attenuation of neutrophil activation and subsequent GEnC injury induced by PR3-ANCA-positive IgGs.