Human rheumatoid synovial fibroblasts promote osteoclastogenic activity by activating RANKL via TLR-2 and TLR-4 activation
Introduction
The innate immune system plays a decisive role in host defense and self-tolerance [1]. Cells of the innate immune system express pattern-recognition receptors, such as the toll-like receptors (TLRs) that recognize certain highly conserved molecules shared by a broad range of bacteria and viruses. The recognition of specific microbial molecules, such as lipopolysaccharides (LPSs) result in the upregulation of costimulatory molecules in antigen-presenting cells, providing the second signal necessary for generating efficient T-cell responses against invading pathogens [2]. The innate immune system controls the subsequent adaptive immune response. Because of this important regulatory role of TLRs, it has been speculated that aberrant TLR signaling may be involved in the generation of autoimmunity.
Rheumatoid arthritis (RA) is an inflammatory autoimmune disease and the most frequent autoimmune form of arthritis. The destruction of cartilage and bone in RA is mediated by a variety of proinflammatory cytokines. Although the exact mechanisms responsible for this destruction remain obscure, the complex interactions among lymphocytes, macrophages, and synovial fibroblasts may elaborate proinflammatory cytokines that activate matrix metalloproteinase (MMP) and osteoclast activity [3]. RA is characterized by chronic synovial inflammation and hyperplasia culminating in joint destruction, which is a major cause of disability in this disease [4], [5], [6].
Osteoclasts are of macrophage–monocyte origin and are giant multinucleated cells with the capacity to degrade bone matrix. Under normal conditions, bone homeostasis depends on the balanced action of bone-forming osteoblasts and bone-resorbing osteoclasts. However, in inflammatory bone disorders characterized by local bone loss, such as RA, osteoclastogenesis is accelerated relative to osteoblastic activity at diseased sites outside the marrow cavity, resulting in abnormal bone resorption and causing bony destruction [7], [8]. The receptor activator of nuclear factor-κB ligand (RANKL), a member of the tumor necrosis factor (TNF) family of cytokines, is a key molecule involved in the differentiation of osteoclasts in the presence of macrophage colony-stimulating factor (M-CSF). Because activated synoviocytes and T lymphocytes also express a large amount of RANKL, the immune system and skeletal system may be closely interrelated [9], [10]. Page and Miossec [18] reported that treatment of synoviocytes with TNF-α or IL-1β in combination with IL-17 was particularly potent at inducing RANKL expression.
Activation/proliferation of synoviocytes, infiltration and retention of leucocytes and angiogenesis lead to the formation of an invasive pannus that destroys articular cartilage and bone [46]. The present study was designed to examine whether synovial fibroblasts can cause monocytes to differentiate into osteoclasts without the help of T cells. A number of TLR ligands are present in RA joints and these ligands stimulate the RA fibroblast-like synoviocytes (RA-FLS). TLRs are involved in the regulation of the inflammatory responses in the synovial compartment. TLRs recognize many exogenous and host-derived molecules; the latter are present in RA serum and synovial fluid. Evidence for a role of TLRs in RA pathogenesis originates from findings that inducible Hsp70, which is a TLR-4 ligand, is increased in RA synovial fluid and in dendritic cells (DCs) isolated from RA synovial fluid [47]. Proinflammatory cytokines increase and regulate the expression of TLR-2 and TLR-4. However, it is not clear whether human synovial fibroblasts activate osteoclastogenesis by producing RANKL-stimulated TLR ligation. Furthermore, we tested the hypothesis that stimulation of TLR-2 and TLR-4 has a synergistic effect on RANKL production by RA-FLS.
CpG DNA also interacts with osteoblastic TLR-9 and elicits intracellular events that lead to the increased expression of molecules regulating osteoclastogenesis [16]. A number of lines of evidence demonstrate the importance of TLR signaling in RA pathogenesis; however, the direct involvement of the TLR pathway in osteoclastogenesis induced bone destruction remains obscure. Because of the importance of TLR signaling in the development of arthritis and the expression of RANKL in RA-FLS, we hypothesized that activation of the innate immune system through TLRs may affect the osteoclastogenic activity of RA-FLS by modulating RANKL expression.
To explore the functional consequences of TLR signaling in RA joints, we assessed the effect of TLR-2 and TLR-4 ligation on the expression of RANKL in RA-FLS. We also evaluated the effect of proinflammatory cytokines on RANKL expression induced by TLR ligation. Here, we show that ligation of TLR-2 and TLR-4 induces RANKL expression in RA-FLS, and that this is mediated partly by IL-1β. We also show that TLR-mediated RANKL expression in RA-FLS may facilitate the differentiation of osteoclasts from monocytes in RA.
Section snippets
Patients and reagents
Seven RA patients fulfilling the 1987 revised criteria of the American College of Rheumatology (formerly, the American Rheumatism Association) took part in this study. The seven RA patients were compared with seven age- and sex-matched control patients with osteoarthritis (OA). Informed consent was obtained from all patients and the experimental protocol was approved by the Catholic University of Korea Human Research Ethics Committee. Samples of rheumatoid synovium were isolated from the seven
Expression of TLR-2 and TLR-4 in RA synovium and FLS
To investigate the role of TLRs in arthritic joint destruction, we used real-time PCR and immunohistochemical staining to examine and compare the expression of TLR-2 and TLR-4 in synovial tissues and cultured FLS derived from patients with RA and OA, and normal skin fibroblasts, which served as a control fibroblast cell line. Fig. 1A shows that the expression of TLR-2 and TLR-4 mRNA was significantly greater in FLS from patients with RA than in FLS from patients with OA and normal skin
Discussion
We investigated the effect of TLR signaling on RANKL-induced osteoclastogenesis in RA. Our data suggest that TLR stimulation through TLR-2 and TLR-4 augments the expression of RANKL in synovial fibroblasts, and that this increased expression facilitates osteoclastogenesis from osteoclast precursor cells. This effect may be mediated partly by increased IL-1β through TLR-2,4 signaling and increased TNF-α through TLR 2 signaling.
Previous studies have highlighted the contribution of innate immunity
Acknowledgement
This work was supported by a grant (R11-2002-098-05001-0) from the Korea Science and Engineering Foundation through the Rheumatism Research Center (RhRC) at The Catholic University of Korea.
References (45)
- et al.
Rheumatoid arthritis
Cell
(1996) - et al.
Expression and regulation of Toll-like receptor 2 in rheumatoid arthritis synovium
Am J Pathol
(2003) - et al.
CpG oligodeoxynucleotides modulate the osteoclastogenic activity of osteoblasts via toll-like receptor 9
J Biol Chem
(2003) - et al.
Cytokine mRNA quantification by real-time PCR
J Immunol Methods
(2002) - et al.
Identification of subpopulations of human macrophages through the generation of human macrophage hybridomas
J Immunol Methods
(1990) - et al.
Activated T lymphocytes support osteoclast formation in vitro
Biochem Biophys Res Commun
(1999) - et al.
Interleukin 1 induces multinucleation and bone-resorbing activity of osteoclasts in the absence of osteoblasts/stromal cells
Exp Cell Res
(1999) - et al.
Novel signal transduction pathway utilized by extracellular HSP70: role of toll-like receptor (TLR) 2 and TLR4
J Biol Chem
(2002) - et al.
HSP70 as endogenous stimulus of the toll/interleukin-1 receptor signal pathway
J Biol Chem
(2002) - et al.
A new mechanism of bone destruction in rheumatoid arthritis: synovial fibroblasts induce osteoclastogenesis
Biochem Biophys Res Commun
(1997)
Inferences, questions and possibilities in toll-like receptor signaling
Nature
Toll-like receptors control activation of adaptive immune responses
Nat Immunol
The expression of toll-like receptors 3 and 7 in rheumatoid arthritis synovium is increased and costimulation of toll-like receptors 3, 4, and 7/8 results in synergistic cytokine production by dendritic cells
Arthritis Rheum
Rheumatoid arthritis Pathophysiology and implications for therapy
N Engl J Med
Clinical improvement and radiological deterioration in rheumatoid arthritis: evidence that the pathogenesis of synovial inflammation and articular erosion may differ
Br J Rheumatol
Pathogenesis of bone and cartilage destruction in rheumatoid arthritis
Rheumatology (Oxford)
The molecular mechanism of osteoclastogenesis in rheumatoid arthritis
Arthritis Res
OPGL is a key regulator of osteoclastogenesis, lymphocyte development and lymph-node organogenesis
Nature
The murine mutation osteopetrosis is in the coding region of the macrophage colony stimulating factor gene
Nature
Interleukin 1 receptor dependence of serum transferred arthritis can be circumvented by toll-like receptor 4 signaling
J Exp Med
Toll-like receptor 2 pathway drives streptococcal cell wall-induced joint inflammation: critical role of myeloid differentiation factor 88
J Immunol
Bacterial peptidoglycans but not CpG oligodeoxynucleotides activate synovial fibroblasts by toll-like receptor signaling
Arthritis Rheum
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Contributed equally to this work.