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OP0152 Synovial Macrophages Promote TGF-β Signaling After Intra-Articular Injections of Oxidized LDL in Naïve Murine Knee Joints, Preventing Production of Pro-Inflammatory Factors S100A8/9, Chemokines and Aggrecanase-Induced Neo-Epitopes
  1. W. de Munter1,
  2. A.B. Blom1,
  3. P.M. van der Kraan1,
  4. J. Roth2,
  5. T. Vogl2,
  6. W.B. van den Berg1,
  7. P.L. van Lent1
  1. 1Experimental Rheumatology (272), Radboud university medical center, Nijmegen, Netherlands
  2. 2Intitute of Immunology, Muenster, Germany

Abstract

Background In previous studies we found that synovial macrophages regulate joint pathology during experimental inflammatory osteoarthritis (OA) and that high systemic levels of Low density lipoproteins (LDL) aggravate OA joint pathology.[1-4] LDL in inflamed synovium is prone to be oxidized (oxLDL) and taken-up by macrophages, leading to an activated phenotype.[5]

Objectives In this study, we investigate whether direct injection of oxLDL into a murine knee joint induces pathology and elucidate the role of synovial macrophages in that process.

Methods Knee joints of C57BL/6 mice were injected five consecutive days with 1.2 mg/mL oxLDL, LDL, or an equal volume of vehicle (PBS). This same procedure was performed in mice depleted of synovial macrophages by intra-articular injection of clodronate liposomes seven days prior to the consecutive injections. Joint pathology was investigated by immunohistochemistry and RNA expression and protein production by synovium were determined using RT-PCR and luminex, respectively. TGF-β activity was measured using a functional CAGA-luciferase assay. Data are depicted as mean ± standard deviation.

Results LDL and oxLDL injection in naïve knee joints did neither increase synovial thickening, nor production of pro-inflammatory factors (IL-1β, IL-6 and S100A8/9) compared to vehicle injection. TGF-β activity in synovial wash-outs was, however, significantly increased by 33% (from 84.7 ng/mL/g synovium ±14.4 to 113.0 ng/mL/g synovium ±33.3; p<0.05). Immunohistochemistry of total knee joints showed that oxLDL injection decreased formation of aggrecanase-induced neo-epitopes (NITEGE) compared with vehicle injections, especially in areas along the bone margins that are prone to develop osteophytes (from arbitrary score 1.19±0.57 to 0.33±0.30; p<0.05).

Repeated injections of oxLDL in macrophage-depleted knee joints led to a 3.1 fold increase of synovial thickening, compared with injection of vehicle (p<0.01), while LDL injections did not alter synovial thickening. Protein levels of S100A8/A9, markers for inflammation, were significantly increased in synovial wash-outs of oxLDL injected joints, compared with LDL (fold increase 5.6; p<0.05) or vehicle (fold increase 8.3; p<0.01) injection. RNA levels of chemokines MCP-1 and MIP-1α were also significantly upregulated after oxLDL injections (6.7 fold and 4.6 fold, respectively; p<0.01). No raise in TGF-β activity was measured in macrophage-depleted joints. NITEGE expression was markedly increased (fold increase 1.92) in the synovium-cartilage contact areas after oxLDL injection (p<0.05).

Conclusions Synovial macrophages promote anabolic effects after oxLDL injections, supporting earlier studies which show increased ectopic bone formation during LDL-rich conditions in experimental osteoarthritis.[4] In absence of synovial macrophages, however, oxLDL induces cell influx, production of pro-inflammatory mediators and aggrecanase activity.

References

  1. Bondeson J, Arthritis and rheumatism 2010;62:647-657

  2. Blom AB, Arthritis and rheumatism 2007;56:147-157

  3. van Lent PL, Arthritis and rheumatism 2004;50:103-111

  4. de Munter W, Arthritis research & therapy 2013;15:R178

  5. Maiolino G, Mediators of inflammation 2013;2013:714653

Disclosure of Interest None declared

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