REVIEWNew Perspective in Osteoarthritis:
The OPG and RANKL System as a Potential Therapeutic Target?
Steeve Kwan Tat,1 Jean-Pierre Pelletier,1 Carmen Ruiz Velasco,2Marc Padrines2 and Johanne Martel-Pelletier1

Osteoarthritis Research Unit, University of Montreal Hospital Centre,
Notre-Dame Hospital, Montreal, Quebec, Canada
2
Université de Nantes, Laboratoire de Physiopathologie de la Résorption Osseuse et Thérapie desTumeurs Osseuses Primitives, Nantes Cedex 01, France
1(Received for publication on June 24, 2008)(Revised for publication on September 3, 2008)(Accepted for publication on October 9, 2008)

Abstract
Bone remodelling is tightly regulated by a molecular triad composed of OPG/RANK/RANKL.
The receptor activator of NF-κB ligand (RANKL) (localized on osteoblasts) enhances osteoclastogenesis via interaction with its receptor RANK (localized on osteoclasts), whereas osteoprotegerin (OPG) (produced by osteoblasts) inhibits this osteoclastogenesis by binding to
RANKL. The equilibrium between OPG and RANKL plays a crucial role in the pathophysiology of bone. Although some studies have shown the efficacy of OPG as a therapeutic agent
against bone resorption, its bioavailability and mechanism of action after binding to RANKL
have only recently been studied. A mechanistic investigation based on what becomes of
OPG after binding to cells expressing membranous RANKL demonstrated an internalization
process of OPG through the clathrin pathway prior to proteasomal and/or lysosomal degradation. Interestingly, the OPG internalization process reduced the half-life of RANKL. Recent evidence has shown that subchondral bone alterations in osteoarthritis (OA) are intimately involved in cartilage degradation, and that OPG and RANKL may be implicated.
Data show that human OA subchondral bone osteoblasts have abnormal OPG and RANKL
levels and consequently an altered OPG/RANKL ratio. Further data also reveal the involvement of some osteotropic factors in these altered levels and that some of these factors generally target RANKL with a differential modulation of the RANKL isoforms. Altogether,data suggest that this system could be targeted as a new strategy for the treatment of OA.
(Keio J Med 58 (1) : 29－40, March 2009)

Keywords: OPG, RANKL, osteoarthritis, osteoblasts, subchondral bone
Introduction

activity The equilibrium between the activities of thesetwo cells maintains the mineral homeostasis via the release of calcium and phosphorous molecules into the circulation (Fig. 1).1 The bone remodelling cycle includes
several distinct phases, the main ones being activation,
resorption, reversal and formation. Physiological events
such as mechanical force and stress induce bone remodelling, in which osteoclast precursors and mature osteoclasts are recruited from the circulation to the bone re-Bone is a specialized connective tissue made up of
several components including the specific cells osteoblasts, osteocytes and osteoclasts, inorganic non-collagenic substances such as proteoglycans, and a collagenic
component of which collagen type I is the major constituent. Bone remodelling involving bone matrix formationand degradation occurs through osteoblast and osteoclast

Presented at the 1559th meeting of the keio Medical Society in Tokyo, April 25, 2008
Reprint request to: Johanne Martel-Pelletier, PhD, Osteoarthritis Research Unit, University of Montreal Hospital Centre, Notre-Dame Hospital, 1560 Sherbrooke Street East, Montreal, Quebec, Canada, H2L 4M1, Tel: +1-514 890 8000 (ext. 26658), Fax: +1-514 412 7582, Email: jm@martelpelletier.ca
29

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Kwan Tat S, et al: OPG and RANKL Involvement during Osteoarthritis

Fig. 1 Bone remodelling is associated with osteoblast activity in bone formation and osteoclast activity in bone resorption. Suchequilibrium maintains the phosphocalcic homeostasis. Imbalance favouring excess osteoclast activity is often described during various
osteolytic pathologies.

modelling unit, thereby initiating the resorption of themineralized matrix. Following this process, the osteoclasts undergo apoptosis, leaving an opening for osteoblasts to deposit newly synthesized matrix.2
Bone remodelling takes place through a coordinatedaction between osteoclasts and osteoblasts. Osteoclasts
are multinucleated giant cells deriving from the hematopoietic progenitors of the monocyte-macrophage lineage.3 Their main function is to resorb the mineralized
tissues of the bone microstructure. Osteoclasts can be
distinguished by their specific morphology and features
such as a polarized morphology and an apical differentiation adjoining the calcified matrix in resorption (ruffledborder). These cells are responsible for the acidificationof the extracellular matrix before the release of proteolytic enzymes involved in the degradation of bone tissues.3 Osteoblasts are derived from the bone marrowcells and are responsible for the synthesis of new matrix.
These cells are progressively transformed into osteocytes
when trapped in their own secreted matrix.4 Secondarycell types such as monocytes/macrophages and endothelial cells also contribute to bone remodelling by acting
either directly with the osteogenic cells or indirectly
through the release of soluble factors, including cytokines, growth factors, and others.
The bone remodelling process becomes unbalanced ina variety of pathologic situations, thereby affecting thegeneral skeleton structure. Diseases such as post-menopausal osteoporosis, bone metastasis, and rheumatoid arthritis include local and/or systemic alterations in which
the level of hormones and pro-inflammatory cytokines
known to stimulate or inhibit bone resorption are highly
involved (Fig. 1).5 In this context, bone stimuli (mechan-ical load, hormones, and growth factors) can act directlyon osteogenic cells as well as on their precursors to control the differentiation, formation and functions (matrix
formation, mineralization, resorption, etc.) of these cells.
This review focuses on OPG (osteoprotegerin), RANK
(receptor activator of nuclear factor-κB) and RANKL(RANK ligand), a molecular triad that has been demonstrated to be of particular importance in the control ofbone pathophysiology. The molecular and cellular mechanisms of action of OPG and RANKL will be reviewed.
Moreover, this triad will be addressed in the context ofosteoarthritis (OA). As substantial evidence is now available suggesting that subchondral bone remodelling prevails early in the disease process, the involvement of
OPG and RANKL in the subchondral bone during OA
will be reviewed. Finally, the possible involvement of
OPG and RANKL as specific therapeutics for OA will
also be discussed.
OPG/RANK/RANKL Molecular Triad
It has been shown that some members of the tumor necrosis factor (TNF) superfamily play major roles in regulating bone metabolism. The molecular triad composedof OPG/RANK/RANKL, members of this superfamily,
has been described as a key cytokine system for controlling the differentiation and function of osteoclast biology.5 Moreover, independent of the bone system, studies
have also revealed new and important functions of thistriad in other pathologies and tissues including mammary
gland development and lactation, the immune and vascular systems, and the proliferation of cancer cells.5

Keio J Med 2009; 58 (1):29－40

31Fig. 2 OPG/RANK/RANKL molecular and cellular mechanisms of action. RANKL is produced by the osteoblast cells in membranous(mRANKL) or soluble (sRANKL) form. Its receptor RANK is expressed on the cell surface of pre-osteoclastic cells and the interaction
between RANK-RANKL induces the differentiation and formation of multinucleated mature osteoclasts. The third protagonist, OPG
is also produced by the osteoblasts and exerts an inhibitory effect on the pre-osteoclast differentiation process. OPG, by binding
to RANKL, inhibits the interaction of RANK-RANKL and the subsequent osteoclastogenesis. Hence, RANKL is described as a
pro-resorptive and OPG as an anti-resorptive agent.

OPG/RANK/RANKL Cellular andMolecular Mechanisms
In the bone remodelling process, the members of themolecular triad OPG/RANK/RANKL are closely linked
to each other. RANKL is synthesized either in membranous or soluble form, primarily by the osteoblastic lineage
cells, the immune cells and some cancer cells. This factor
is essential for mediating bone resorption through osteoclastogenesis and the activation of mature osteoclasts.
RANKL stimulates osteoclastogenesis and osteoclast activity by binding to the cell surface receptor RANK, located on osteoclasts (precursor and mature).6, 7 The bindingof RANKL to the extracellular RANK leads to the activation of specific signalling pathways involved in the formation and survival of osteoclasts, hence bone resorption.8, 9
OPG is secreted by the stromal cells and other cell types,
including osteoblasts, and acts as a soluble decoy receptor for RANKL. OPG, by interacting with RANKL, inhibits the binding of RANKL to RANK, thereby preventing RANK activation and subsequent osteoclastogenesis
and, as a result, inhibits bone resorption (Fig. 2).10
In the immune system, the molecular and cellularmechanisms of this triad are important for maintaining
an immune response. In this system, RANKL expressedby activated T cells interacts with RANK, which is expressed by the dendritic cells, and such binding of
RANKL to RANK on dendritic cells regulates the function and survival of these cells.11-1 3 OPG, on the other

hand, is produced by the B lymphocytes as well as dendritic cells and maintains an equilibrium in this system.
The OPG/RANK/RANKL system, however, is not the
only cytokine mechanism that controls the immune response. Indeed, the cross-talk between T cells and dendritic cells also involves CD40 and CD40L interaction.12, 13
There is accumulating evidence of the potential role of
OPG and RANKL in other tissues including the vascularsystem. OPG and RANKL have been found to be associated with pathologies such as coronary artery disease,atherosclerosis and vascular calcification in patients.
Indeed, experiments done on OPG knockout mice haverevealed the anti calcification property of OPG.1 4-1 6 In
this system, the absence of OPG favours the medial calcification of the aorta as well as calcification of the renalarteries. Treatment of these knockout mice with OPG
significantly reduced the calcified lesion areas.
OPG
OPG belongs to the TNF receptor (TNFR) superfamily, but differs from the common membranous TNFR inthat it is a soluble receptor.1 7 OPG functions as an endogenous antagonist receptor that prevents the biologicaleffects of RANKL, both membranous and soluble, andthus acts as an inhibitor of bone remodelling/resorption.
OPG is highly expressed in the adult lung, heart, kidney,
liver, thymus, lymph nodes and bone marrow, and is syn-

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Kwan Tat S, et al: OPG and RANKL Involvement during Osteoarthritis

thesized by several cells including stromal cells, osteoblasts, vascular smooth muscle cells, B lymphocytes, andarticular chondrocytes.1 8, 1 9 OPG is produced as a proprotein of 401 amino acids and undergoes an intracellular cleavage resulting in a final 380 amino acids mature
protein.6
OPG production by specific cells is modulated by several cytokines, vitamins, estrogens and other molecules,
thereby modulating osteoclastogenesis and bone resorption. OPG production is induced by 1α, 25-dihydroxyvitamin D3, estrogens, pro-inflammatory cytokines such as
interleukin-1 (IL-1) and TNF-α as well as transforming
growth factor-β (TGF-β), whereas parathyroid hormone(PTH) and glucocorticoids inhibit OPG production.20-26
The importance of OPG was illustrated on OPG-deficient mice, in which osteoclast formation was greatly enhanced, and also on an overexpressing-OPG mousemodel, in which severe osteopetrosis with reduced numbers of osteoclasts was observed.18, 27
Moreover, OPG can directly inhibit osteoclast activity,
independently of RANKL, through interactions with stilluncharacterized receptors present on osteoclasts.2 8 The
biological effects of OPG on bone cells include the inhibition of the terminal stages of osteoclast differentiation,
suppression of mature osteoclast activation and the induction of apoptosis of these cells.

minor participation in controlling osteoclast formation.31, 32 Similar to TRAF-6 deficiency, mice expressing
neither RANK nor the NF-κB subunits p50 or p52, developed severe osteopetrosis due to a total absence of osteoclasts.33, 34 NFAT, identified as one of the downstream
targets of RANK activation, is increased in the presenceof RANKL, and in cooperation with c-Fos leads to theterminal differentiation of the osteoclasts by promoting
the expression of osteoclast-specific genes such as tartrate-resistant acid phosphatase (TRAP) and calcitoninreceptors, and its lack leads to an inhibition of osteoclast
numbers.35, 36
Hence, the activation of RANK stimulates the differentiation of osteoclastic precursors into mature osteoclasts as well as the activation of mature osteoclasts.
RANK activation by RANKL involves distinct signalling cascades and mice ablated of these components
were found to have an osteopetrotic phenotype.

RANK
RANK, like OPG, belongs to the TNFR superfamily. It
is synthesized as a type I transmembrane protein of 616
amino acids and assembles itself into a functional trimer. 2 9 RANK is considered a natural receptor of
RANKL and is also expressed by several different tissues such as skeletal muscle, thymus, liver, colon, mammary glands, prostate, pancreas, and cells of themonocyte/macrophage lineage including precursors andmature osteoclasts, B and T lymphocytes, dendritic cells,
fibroblasts, and articular chondrocytes.6 Activation of
RANK, like other TNFR-related proteins, induces a cascade of intracellular signalling factors including the recruitment of TNFR-associated factor proteins (TRAF),
the transcription factors NF-κB subunits p50 and p52,
AP-1 and nuclear factor of activated T cells (NFAT), themitogen-activated protein (MAP) kinase cascades including ERK1/2, JNK and p38, and the induction of Srcand phosphatidylinositol 3- kinase-dependent Akt activation.30
The cytoplasmic domain of RANK possesses independent TRAF interaction motifs that bind to TRAF-2, -5
and -6. RANK mutants and selective deletions of RANK
demonstrated that TRAF-6/RANK interaction is themain adaptor molecule for RANK signalling. 9 With
TRAF-2 and TRAF-5 deficiency, only a slight reductionin osteoclastogenesis was observed, thus showing theirRANKLRANKL belongs to the TNF superfamily and is synthesized as a mature protein of 317 amino acids. RANKL
is largely expressed in bone, bone marrow and lymphoidtissues.37-39 The predominant roles of RANKL in bone
physiology are the stimulation of osteoclastdifferentiation/activation and the inhibition of osteoclast
apoptosis.6 In addition, this cytokine in association withmacrophage-colony stimulating factor (M-CSF) is involved in the osteoclastogenesis process. The importanceof RANKL was demonstrated in RANKL knockout mice
which presented severe osteopetrosis and total loss ofosteoclasts.27
Human RANKL has recently been shown to exist as 3
isoforms: RANKL1, 2, and 3. RANKL1 and 2 encode
for transmembrane forms with an absence of intracellular domain for human RANKL2, whereas RANKL3
lacks the intracellular and transmembrane domain.4 0
Murine RANKL has also been described to exist in 3
isoforms; however, murine RANKL2 possesses a shortcytoplasmic domain, which is completely absent in human RANKL2. These different isoforms are able to differentially regulate osteoclastogenesis. 4 0- 4 2 In cells,
RANKL forms homo- or heterotrimer structures between
the isoforms,4 3, 4 4 and the trimeric combination of theisoforms is crucial for orientating the RANKL membranous localization, thereby controlling the osteoclast formation and differentiation process. Hence, the trimeric
structure of RANKL1 alone or in association withRANKL2 is translated into a membranous localization.
When RANKL1 or 2 is co-transfected with RANKL3,
however, a reduced level of membranous localization isobserved, indicating that RANKL3, by preventing themembranous localization of RANKL, acts as an inhibitorof osteoclastogenesis. 4 0- 4 2 Moreover, NIH3T3 cells

Keio J Med 2009; 58 (1):29－40

33

Fig. 3 Exogenous OPG is internalized through the clathrin pathway by membranous RANKL before being degraded via theproteasome and the lysosome.

transfected with human RANKL2 alone were unable toinduce osteoclastogenesis, whereas those transfectedwith murine RANKL2 induced osteoclastogenesis. Such
difference could be explained by the fact that murine
RANKL2 possesses an intra-cytoplasmic domain allowing the membranous localization of RANKL whereas an
absence of such intra-cellular domain for trimerized human RANKL2 may explain its inability to be expressedat the cell surface.40, 42
RANKL exists in both a soluble and a membranousform.41 The soluble form of RANKL, which corresponds
to the C-terminal domain of membranous RANKL, may
be produced either directly by the cell through an alternative splicing followed by a secretion extracellularly or
by a proteolytic cleavage of membranous RANKL. So
far, various proteases have been proposed to be involvedin the shedding process. Among these, the tumor necrosis factor converting enzyme (TACE), the membranous
metalloprotease MT1-MMP, MMP-7, and ADAM-19
have all been shown to proteolytically cleave membranous RANKL.45-49
THE OPG-RANKL Molecular Complex
Bone remodelling is tightly controlled by OPG andRANKL; however, the ratio of OPG/RANKL is considered to better reflect environmental signals. A high ratioof OPG/RANKL is indicative of promoting bone formation while a low ratio favours bone resorption. Such imbalance has been encountered in some osteolytic lesions
where RANKL is up-regulated as well as in osteoblastic
disorders where OPG is up-regulated.50
Although several studies have proven the beneficial effect of administering OPG as a specific therapeutic agent
against bone resorption as well as against arterial calcification in cardiovascular disorders, there is very little data
concerning the intimate modulation of exogenous OPG
bioavailability in the extracellular environment. It has

been shown that OPG, through its heparin binding domain interacts with some proteoglycans such as syndecan-1 and that OPG binding to cells expressing high levels of membranous syndecan-1 is internalized, then degraded.51, 52 Similarly, with RANKL expressing cells, thebinding of OPG to membranous RANKL is also internalized, yet through another process, the clathrin coat formation pathway, which occurs before proteasomal andlysosomal degradation (Fig. 3).53 Thus, these two OPG
internalization mechanisms, through the lipid rafts bymembranous syndecan-1 or the clathrin pathway, bymembranous RANKL are involved in the control of thebioavailability of extracellular OPG. Interestingly, on
RANKL expressing cells, OPG was also demonstrated
to regulate the half-life of membranous RANKL; bybinding to RANKL, OPG induces the degradation of
RANKL.53
Further investigations on the effect of exogenous soluble glycoaminoglycans (GAGs) on the binding of OPG
to RANKL showed that GAGs such as heparin, heparansulfate, chondroitin sulfate and dermatan sulfate bind to
OPG via the heparin binding domain. Such OPG-GAG
binding inhibits OPG-RANKL interaction, thereby preventing OPG internalization mediated by membranous
RANKL. In vivo, GAGs and proteoglycans of the microenvironment could interact with endogenous OPG, thereby modulating the bioavailability of OPG and RANKL.54
In most of the cytokine-cytokine receptor systems,
such as the TNF/TNFR family, the cytokine interacts
with its transmembranous receptor before being internalized. Thus, OPG-RANKL interaction represents an original mechanism in which OPG, being the receptor, is internalized by its ligand RANKL. This internalization
process is of particular importance for future therapeutic
involvement of OPG. In fact, although OPG has beensuccessfully used as a therapeutic anti-resorptive agent
against osteolytic diseases such as postmenopausal osteoporosis, multiple myeloma, tumor associated humoral

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Kwan Tat S, et al: OPG and RANKL Involvement during Osteoarthritis

hypercalcemia and rheumatoid arthritis,5 5- 5 8 the decrease in extracellular OPG concentration observed in
RANKL-expressing cells raises the question of its bioavailability during osteolytic disease therapies, especially when RANKL is abundantly expressed. In this particular situation, denosumab, a fully human monoclonal antibody against RANKL, is a potent anti-resorptive
agent.59 Its inhibition of the RANK-RANKL interaction
as well as the OPG internalization, thereby enhancing
the circulating concentration and bioavailability of endogenous OPG, could contribute to an additive effect ininhibiting bone resorption.
The OPG-RANKL mechanism is much more complex
than previously thought. Indeed, the anti-resorptive effect of OPG can be explained not only by its properties
as a decoy receptor (competitor inhibitor of RANK/RANKL binding) but also as a modulator of RANKL
half-life and the RANKL shedding mechanism process.
As RANKL controls the bioavailability of OPG and viceversa, the equilibrium between membranous RANKL
and soluble OPG will be determinant of a curative application of OPG. Approaches should target the maintainingof an optimal soluble extracellular OPG concentration by
reducing OPG internalization.

a secondary phenomenon related to multiple factors including cartilage matrix degradation. Moreover, studies
have confirmed that the subchondral bone during OA is
the site of several dynamic morphological changes thatappear to be part of the disease process.61 These changes
are associated with a number of local abnormal biochemical pathways related to the altered osteoblast metabolism in this tissue.
Some clinical studies in OA patients have suggested
that the indices of bone resorption are increased early inthe disease process. This concurs with the in vivo findings of a study performed on an induced OA dog model6 2 allowing the chronological analysis of the diseaseprogression. At an early stage of the disease process, increased bone loss and resorption with subchondral bone
exhibiting surface and trabecular thickness reduction andan increased number of osteoclasts, as well as productionof catabolic factors including cathepsin K and MMP-13
were observed.6 2 Moreover, an in vivo experiment performed at an early stage on the same dog model demonstrated an inhibition of the subchondral bone loss upontreatment with bisphosphonates (an inhibitor of osteoclast function).6 3 Also reported on this dog model at alater stage, 54 months after OA induction, was that thehistomorphometric analysis of the subchondral bone revealed a marked increase in volume with active boneformation.64
Human OA subchondral bone osteoblasts have demonstrated abnormal phenotypes, including elevated alkaline
phosphatase activity and increased release of osteocalcin.6 5 Other factors such as TGF-β, insulin-like growth
factor-1 (IGF-1) and the urokinase-type plasminogen activator (uPA)/plasmin system have also been found atabnormal levels in OA subchondral bone.6 6 Moreover,
our group has also reported that OA subchondral boneosteoblasts, which otherwise demonstrate no different
phenotypic features, can be discriminated into two subgroups identified by their level (low [L] or high [H]) of
endogenous prostaglandin E2 (PGE2).6 7-7 1 Hence, OA
patients can be discriminated into groups named L-OA
or H-OA.

Osteoarthritis
The articular joint is made up of several tissues, themain ones involved during osteoarthritis (OA) being the
cartilage, synovial membrane, and the subchondral bone,
all of which are closely linked. The cartilage is the toughelastic material that covers and protects the ends of bone.
In a healthy joint it acts as a shock absorber when weightis exerted on the joint and its slippery surface allows thejoints to move smoothly. Joint degeneration does occur,
however, and OA is among the most common joint disorders, affecting about 65% of individuals over 60 yearsof age. The disease causes pain and functional disability,
resulting in a significant social and economic burden.
Despite the high prevalence of this disease, its precise
etiopathogenesis is not yet completely understood, although significant progress has been made during thelast few decades. The joints commonly affected by OA
are the base of the thumb, the end joints or middle jointsof the fingers, the hips, the knees and the joints at thebase of the big toe, although the neck (cervical spine)
and the lower back (lumbar spine) can also be affected.
OA is considered a complex illness in which all the tissues of the joint play significant roles.60 Even though articular cartilage degeneration is a major characteristic of
OA, we still do not know what initiates it. Synovial
membrane inflammation is believed to play an importantrole in the progression of joint tissue lesions; however,there is a general consensus that synovial inflammation
in OA is not the primary cause of the disease, but rather

Involvement of OPG
and RANKL during Osteoarthritis
In OA, a subchondral bone resorption/formation process has been shown to occur, in which there are phasesof bone degradation and others of bone formation. Interestingly, factors such as OPG and RANKL, which constitute specific components capable of influencing thebone remodelling process, have been found to be expressed and modulated in human OA subchondral bone.
In addition to subchondral bone, the OPG/RANK/RANKL triad has also been observed to be expressed by
the other articular cells. Indeed, articular chondrocytes

Keio J Med 2009; 58 (1):29－40

also express each factor of the triad but with specific localization in the cartilage, which depends on whether the
cartilage is normal or diseased. OPG, RANK andRANKL have been detected in the superficial zone ofnormal cartilage whereas during OA their expression
was found to extend to the middle zone.7 2 At present,
however, the functional consequence of their expression
by the articular chondrocytes remains unknown. Nevertheless, besides the mechanism of action of OPG binding
to RANKL thus decreasing the RANKL activity, a recent
study on an OPG transgenic mouse model of OA suggested that the in vivo beneficial effect of the administration of OPG could also be due to its capacity to bind tumor necrosis factor-related apoptosis-inducing ligand(TRAIL), an inducer of chondrocyte apoptosis. Hence,
OPG, by binding to TRAIL, is able to prevent chondrocyte apoptosis.7 3 Of note, the beneficial anti-apoptotic
effect of OPG on some cells could be somewhat detrimental when applied to other systems. For example, OA
as well as rheumatoid arthritic synovial membrane demonstrate hyperplasia, and one of the envisaged therapeutic strategies involved increasing apoptosis of cells inthis tissue, in which case, the anti-apoptotic effect of
OPG would be unbeneficial as it could further increase
the number of cells in the synovial membrane.
Synovial cells have also been observed to express OPG
and RANKL, and RANKL appears actively involved in
the erosion of bone during rheumatoid arthritis.7 4-7 6 In
the presence of cytokines commonly involved during
rheumatoid arthritis, synoviocytes were able to induce
osteoclastogenesis by up-regulating the production of
RANKL.74, 75, 77 Moreover, synovial fluid of rheumatoid
arthritis patients showed an induced osteoclast formation
due to excessive production of RANKL.7 8 Such regulation of the OPG/RANKL ratio favours destruction ofmineralized tissue including the cartilage and the corticalbone. In vivo studies performed on animal models of arthritis have proven the beneficial effect of OPG injection
at reducing cortical bone erosion.79, 80 In another arthritic
disease, psoriatic arthritis, dysregulation of OPG andRANKL toward an elevation of RANKL was also
shown.81, 82
Although these factors are gaining importance in the
progression of inflammatory joint diseases (rheumatoid
arthritis and psoriatic arthritis), their participation in OA
has until now been unclear. Recently, efforts have been
devoted to a better comprehension of the implication of
OPG and RANKL during the different stages of OA. Improved understanding is without doubt of major importance in better identifying a specific strategic treatment
for this disease. It was recently demonstrated that during
longstanding OA and rheumatoid arthritis, the OPG/RANKL ratio in the synovial fluid is much more elevated in OA compared to rheumatoid arthritis.8 3, 8 4 Hence,
cortical bone destruction is much more prevalent in rheu-

35

matoid arthritis compared to OA, an observation whichis correlated with clinical observation. In this latter study,
however, the remodelling process of the subchondralbone was not evaluated. This data could have been of
great importance in establishing a close relationship between the OPG/RANKL ratio and the subchondral bonechanges.
As described previously, studies performed on OA patients revealed two distinct phenotypes of subchondralbone osteoblasts, L-OA and H-OA.67-71 In situ histological examination of the subchondral bone revealed thatthe thickness of this tissue was reduced in the L-OA andincreased in the H-OA patients compared to normal individuals.70 Further experiments in which the bone remodelling factors OPG and RANKL were determined in OA
subchondral bone osteoblasts compared to normal revealed that in both the L-OA (having low endogenouslevels of PGE2) and H-OA (having high endogenous levels of PGE2) osteoblasts, OPG and RANKL were abnormally expressed and consequently the OPG/RANKL ratio was reduced in the L-OA, suggesting increased subchondral bone resorption, and increased in H-OA, indicating a shift toward bone formation.70 These data were
further strengthened by an osteoclastogenesis experimentin which the L-OA osteoblasts induced in the pre-osteoclastic murine cell line RAW 264.7 a significantly higher
level of mature osteoclasts compared to the H-OA. 7 0
These findings are summarized in Figure 4.
Further studies carried out on the L-OA and H-OA osteoblasts investigated the membranous localization of
RANKL, and the expression level of RANKL isoforms
demonstrated that membranous RANKL levels were significantly increased in the L-OA osteoblasts compared tonormal and H-OA.7 1 The gene expression level of theRANKL1 profile followed the same trend as the membranous RANKL level. RANKL3 gene expression was
lower on the H-OA osteoblasts compared to the normal
and the L-OA osteoblasts; however, the overall outcome
favoured RANKL1, hence the localization of RANKL at
the membrane. Treatment of the L-OA osteoblasts withthe osteotropic factors 1α,25 dihydroxyvitamin D3, IL-1β,
TNF-α, PGE2, and IL-6 showed a significant increase inmembranous RANKL and the RANKL1 gene expression
level followed the same trend with the exception of treatment with TNF-α. Upon treatment with TNF-α, the increased membranous RANKL and the absence of effecton the RANKL1 gene expression strongly suggest themodulation of the other RANKL isoforms. Of great interest is that in the H-OA subpopulation, the above osteotropic factors had no effect on either membranous
RANKL or its isoforms. Such findings demonstrate thateach of the human OA subchondral bone subpopulations(L-OA and H-OA) has attained a different metabolicstate; L-OA being enriched with factors promoting bone
resorption and H-OA having reduced resorptive proper-Kwan Tat S, et al: OPG and RANKL Involvement during Osteoarthritis

36

Fig. 4 Osteoarthritic (OA) subchondral bone osteoblasts can be discriminated into two subpopulations, Low-OA or High-OA, basedon their endogenous production of PGE2. Differences between these two categories of osteoblasts include the OPG/RANKL ratio, thecapacity to induce osteoclast formation, and the correlation to the thickness of OA subchondral bone.

ties, with the metabolism of the latter cells favouringbone formation. Thus, in humans, the presence of L-OA
and H-OA subchondral bone osteoblasts could reflect
different stages of attempts to repair this damaged tissue
in this disease: an increase in bone resorption followedby bone formation.
OPG and RANKL as a Specific Therapeutic
Target during Osteoarthritis
OA is a disease that tends to get worse over time as the
cartilage continues to wear away. Unfortunately, no curative therapeutics (that is, agents that stop the disease progression) are currently available for this disease. The
main properties of the available treatment are to control
the pain and improve the joint’s ability to function. Somerisk factors have been described for the development of
OA and they include age, family history, excessive
weight, joint injury, and as shown recently by magnetic
resonance imaging, meniscal extrusion and bone marrow
edema.85-89 Some non-medication therapies are also avery important component of the treatment of OA. These
include moderate exercise to reduce pain and stiffness
and to maintain or increase muscle tone, the applicationof heat and cold to provide relief from muscle and joint
pain and a well-balanced diet, which is also essential for
maintaining a healthy weight.
Medications commonly prescribed for OA consist of
non-steroidal anti-inflammatory drugs (NSAIDs), as wellas corticosteroids and viscosupplementation. Patients arealso sometimes given complementary therapy including
glucosamine and chondroitin sulfate. Finally, even if apatient is undergoing therapy, when OA becomes severe,

surgery remains the last option. The most common surgical procedures for OA include hip and knee replacement.
Recently, the Glucosamine/chondroitin Arthritis Intervention Trial (GAIT) reported, following exploratory
analyses, that the combination of glucosamine and chondroitin sulfate was effective on symptoms in a subpopulation of OA patients with moderate to severe knee
pain.9 0 Interestingly, a recent in vitro study carried out
on human L-OA subchondral bone osteoblasts demonstrated that the combination of glucosamine and chondroitin sulfate modulated the OPG/RANKL ratio, in
which there was an increase in favour of reduced bone
resorption.69 This was further supported by data showingthat the combination of these compounds decreased theresorptive activity of L-OA osteoblasts.6 9 These data
may help to explain how these two drugs exert a positiveeffect on OA pathophysiology.
Conclusion
In conclusion, this review emphasizes that the internalization and degradation of OPG is dependent on thepresence of membranous RANKL or on cells expressing
high levels of syndecan-1. The inhibitory effect of OPG
on RANKL can be explained by its role not only as a decoy receptor but also as a modulator of RANKL halflife. Hence, as membranous RANKL controls the bioavailability of exogenous OPG, the equilibrium between
RANKL and OPG is crucial for future therapeutic use of
OPG. An ideal therapeutic approach would be to maintain an optimal soluble OPG concentration associatedwith a low internalization process.
Data also point to specific molecular mechanisms that

Keio J Med 2009; 58 (1):29－40

appear to be operating during human OA progression.
Studies demonstrate that the abnormal OPG and RANKL
levels in human OA subchondral bone osteoblasts are
dependent on the metabolic state of these cells and thatthe different abnormal expression of OPG and RANKL
indicates different stages of attempts to repair the damaged subchondral bone.
Future therapeutic approaches could target reducing
bone remodelling changes that take place in human OA
subchondral bone, and data suggest that the modulationof OPG and RANKL could be an interesting target forsuch treatment. Since L-OA patients are in a remodelling/
resorptive phase, the inhibitory effect of OPG on bone
resorption could therefore be proposed for this subpopulation; however, even though the administration of OPG
in vivo in a murine OA model seems effective for thetreatment of OA,73 care should be taken since full-length
OPG is known to be a survival factor for tumor cells.9 1
Moreover, the anti-apoptotic effect of OPG via TRAIL
can promote OA synovial membrane hyperplasia. An alternative approach is to inhibit RANKL by using an antiRANKL antibody or by acting on upstream factors responsible for the increased membranous localization of
RANKL, for example by modulating one of the RANKL
isoforms. In regard to the OA subpopulation, the additionof OPG or the inhibition of RANKL would be beneficial
on the subchondral bone of the L-OA as this tissue is in aresorptive phase. However, in the H-OA patients, antiresorptive agents are expected to be less effective since
the subchondral bone was shown to be in a formation
phase. Nonetheless, further experiments exploring theeffects of adding or increasing OPG or inhibiting
RANKL on the subchondral bone pathophysiologicalpathways of each OA subpopulation are required.
Acknowledgments
The authors are grateful to Virginia Wallis from the
Osteoarthritis Research Unit of the University of Montreal Hospital Centre for her assistance with the manuscript preparation.
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