Apolipoprotein B binds to enolase-1 and aggravates inflammation in rheumatoid arthritis

Abstract

Background: Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by systemic inflammatory process that eventually leads to joint destruction. Monocytes and synovial macrophages are key players in the inflammatory process of RA. Enolase-1 (ENO1) is a multifunctional glycolytic enzyme located in the cytoplasm of cells. It is also present on the cell surface as plasminogen receptor. The majority of cells expressing ENO1 in peripheral blood mononuclear cells (PBMCs) and synovial fluid mononuclear cells (SFMCs) derived from RA patients are known to be CD14-positive monocytes and macrophages.

Objective: The objective of this study was to identify novel ENO1 binding protein present in RA synovial fluid and determine the functional role of interaction between apolipoprotein B (apoB), a novel ligand of cell surface expressed ENO1, and ENO1 in RA.

Methods: ENO1 binding protein present in RA synovial fluid (SF) was identified by affinity-based mass spectrometry analysis. Interaction between ENO1 and apoB was evaluated by ligand blotting assay, ligand binding assay, surface plasmon resonance (SPR), and confocal microscopy. Production levels of pro-inflammatory cytokines in PBMCs from RA or healthy control (HC) after stimulation with apoB were evaluated by enzyme-linked immunosorbent assay (ELISA). Signaling pathways involved in the inductions of cytokines after stimulation with apoB were identified by using specific inhibitors. Pro-inflammatory effect of apoB was evaluated using K/BxN serum transfer arthritis mouse model.

Results: Characterization of physical interaction between ENO1 and apoB showed that apoB was a novel ligand of ENO1. Interaction between surface ENO1 and apoB induced higher levels of pro-inflammatory cytokines in RA PBMCs compared to that in HC PBMCs. Intracellular p38 MAPK and NF-κB pathways were found to be key signaling pathways upon ENO1 activation. In K/BxN serum transfer arthritis mouse model, apoB aggravated arthritis severity. Moreover, apoB derived peptides showed agonistic or antagonistic actions.

Conclusion: ApoB is a novel ligand of ENO1. It might enhance chronic inflammation in RA patients.