Introduction Rheumatoid arthritis (RA) is a chronic inflammatory disease characterised by immune complex (IC) deposition in the synovium, leading to increased bone destruction. In RA, joint destruction has been associated with high levels of low density lipoproteins (LDL) and enhanced LDL oxidation (oxLDL). Apolipoprotein E (Apoe) is an important regulator of LDL transportation and its absence strongly elevates circulating LDL levels, which may lead to increased oxLDL levels during inflammation.
Objectives In this study, we investigated the effects of high LDL levels on bone destruction during antigen-induced arthritis (AIA) and how increased LDL/oxLDL levels affect osteoclast formation.
Methods AIA was induced by injection of methylated BSA (mBSA) into the right knee joint of Apoe-/- and wild type (WT) control mice previously immunised with mBSA and complete Freund’s adjuvant (CFA). WT and Apoe-/- Hoxb8 myeloid precursor cells were differentiated into osteoclasts using 20 ng/mL RANKL and 30 ng/mL M-CSF, then stimulated for 24 hour with 10 µg/mL LDL/oxLDL. Oil Red O staining was performed to assess lipid uptake by osteoclasts. mRNA levels of c-Fos, RANK, NFATc1, DC-STAMP, TRAP, CTR, ClC-7 and Cat K were measured by qPCR. Bone erosion was quantified by histological analysis using an arbitrary scale from 0 to 3 and TRAP+ cells were determined using immunohistochemistry.
Results Apoe-/- mice showed significantly higher LDL serum levels than WT controls. At day 21 after AIA induction, bone erosion was significantly decreased in Apoe-/- mice (25% reduction from 1.5±0.2 to 1.1±0.1). In line with that, the number of osteoclasts within the knee joints was 36% lower in Apoe-/- mice, as determined by image analysis of TRAP staining. To study the role of Apoe and high LDL levels on osteoclastogenesis in more detail, we differentiated WT and Apoe-/- myeloid precursor cells (Hoxb8) into osteoclasts and found similar mRNA levels of osteoclast markers. Whereas LDL stimulation did not affect osteoclast formation, oxLDL strongly impaired cell fusion keeping them in a mononuclear state. mRNA levels of DC-STAMP were significantly down-regulated in both WT and Apoe-/- osteoclasts (1.4 and 2.3 fold decrease, respectively) as well as TRAP activity (49% and 58% reduction in WT and Apoe-/- osteoclasts), underlining a major role of oxLDL in inhibiting osteoclastogenesis.
Conclusions High LDL/oxLDL levels by Apoe deficiency affect bone destruction during AIA by reducing the number of osteoclasts in the synovium, probably by interfering with osteoclastogenesis.
Disclosure of interest None declared
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