Bone remodelling at a
Julie C. Crockett*, Michael J.
Rogers, Fraser P. Coxon, Lynne J.
Hocking and Miep H. Helfrich
Musculoskeletal Research Programme, Institute of
Medical Sciences, University of Aberdeen,
Foresterhill, Aberdeen AB25 2ZD, UK
*Author for correspondence
Journal of Cell Science 124, 991-998
2011. Published by The Company of Biologists Ltd
The bone remodelling cycle (see Poster panel
“The bone remodelling cycle”) maintains the
integrity of the skeleton through the balanced
activities of its constituent cell types. These are
the bone-forming osteoblast, a cell that produces
the organic bone matrix and aids its
mineralisation (Karsenty et al., 2009); the bone-
degrading osteoclast, a unique type of exocrine
cell that dissolves bone mineral and
enzymatically degrades extracellular matrix
(ECM) proteins (Teitelbaum, 2007); and the
osteocyte, an osteoblast-derived post-mitotic
cell within bone matrix that acts as a
mechanosensor and an endocrine cell
(Bonewald and Johnson, 2008). A fourth
cell type, the bone lining cell, is thought to have
a specific role in coupling bone resorption to
bone formation (Everts et al., 2002), perhaps
by physically defining bone remodelling
compartments (Andersen et al., 2009).
Molecular dissection of genetic disorders of
highly increased or reduced bone mass has
identified many of the crucial proteins
controlling the activity of these bone cell types.
This information has resulted in both novel
ways to treat or diagnose more common bone
disorders and a better understanding of the
common genetic variants that lead to differences
in bone density in the general population.
In this poster article, we illustrate the crucial
signalling pathways involved in bone cell
differentiation, function and survival, and
describe how the coupled activities of the cells
in bone are maintained through intercellular
interactions. We pay particular attention to the
factors and signalling processes that have been
found to be indispensable for the maintenance of
healthy bones through the study of rare genetic
diseases of bone.
Osteoblasts: differentiation and
Osteoblast differentiation is achieved by the
concerted expression of a number of key
transcription factors (see Poster panel
“Osteoblast lineage”), and bone formation by
osteoblasts is controlled both locally and
systemically during bone modelling in
development (Box 1) and throughout life.
Studies of diseases associated with defects in
bone formation, such as developmental limb
disorders and high bone mass conditions, have
demonstrated the crucial importance of local
bone formation control by bone morphogenetic
protein (BMP) (Cao and Chen, 2005) and
wingless (Wnt) (Day et al., 2005) signalling
pathways for osteoblast differentiation and
function. In the adult, BMP2 can act as a potent
stimulator of ectopic bone formation (Chen et
al., 1997) and it is used clinically to enhance
bone formation, for example, during fracture
repair (Govender et al., 2002). BMP signalling
through the recruitment and activation of
(See poster insert)
© Journal of Cell Science 2011 (24, pp. 991–998)
Bone Remodelling at a Glance
Julie C. Crockett, Michael J. Rogers, Fraser P. Coxon, Lynne J. Hocking and Miep H. Helfrich
, 1,25 dihydroxy vitamin D
; AP, alkaline phosphatase; AP-1, activator protein-1; Arf6, ADP-ribosylation
factor 6; Arp2/3, actin-related proteins 2 and 3; ATF4, activating transcription factor 4; αvβ3, vitronectin receptor; β2 AR, β2 adrenergic
receptor; BMP, bone morphogenic protein; BSP, bone sialoprotein; CAII, carbonic anhydrase II; CATK, cathepsin K; ClC7, chloride
channel 7; CLOCK, circadian locomotor output cycles kaput; CREB, cAMP responsive binding protein; CTR, calcitonin receptor; DAP12,
DNAX-activating protein of 12 kDa; DC-STAMP, dendritic cell-specific transmembrane protein; DKK1, dickkopf-1; DMP1, dentin matrix
protein 1; EE, early endosome; EphB4, Eph receptor B4; ER, oestrogen receptor; ERK, extracellular signal-regulated kinase; FcRγ,
Fc receptor γ chain; FGF, fibroblast growth factor; Fra1, fos-related antigen 1; GIT2, G-protein-coupled receptor kinase-interactor 2;
GM-CSF, granulocyte-macrophage colony stimulating factor; Grb2, growth factor receptor-bound protein 2; HDAC6, histone deacetylase 6;
Htr1b, 5-hydroxytryptamine receptor 1B; IGF1, insulin-like growth factor 1; IGF BP, IGF-binding protein; IL, interleukin; IR, insulin receptor;
LE/L, late endosome or lysosome; LIFR, leukemia inhibitory factor receptor; LRP, lipoprotein-related protein; Mcl-1, myeloid cell leukemia
sequence 1, isoform 1; MCSF, macrophage colony-stimulating factor; mDia2, mammalian diaphanous protein 2; MITF, microphthalmia-
associated transcription factor; MMP-9, matrix metalloproteinase-9; mTOR, mammalian target of rapamycin; MyoD, myoblast determination
protein 1; NFATc1, nuclear factor of activated T-cells cytoplasmic 1; NOS, nitric oxide synthase; OPG, osteoprotegerin; OSCAR,
osteoclast-associated receptor; OSM, oncostatin M; OSMR, oncostatin M receptor; PGE2, prostaglandin E2; PI3K, phosphoinositide
3-kinase; PLC-γ, phospholipase Cγ; Plekhm, pleckstrin homology domain containing family M; PPARγ, peroxisome proliferator activator
γ; PTH, parathyroid hormone; PTHR, parathyroid hormone receptor; Pyk2, proline-rich tyrosine kinase 2; RANK, receptor activator of
NF-κB; RANKL, receptor activator of NF-κB ligand; Runx2, runt-related transcription factor 2; RXR, retinoic acid receptor; SIRPβ1,
signal-regulatory protein β1; SOST, sclerostin; SV, secretory vesicle; Syk, spleen tyrosine kinase; TCF/LEF, T-cell factor and lymphoid
enhancer factor family of transcription factors; TGFβ, transforming growth factor β; TLSP, thymic stromal lymphopoietin; TNF-α, tumour
necrosis factor α; TRAcP, tartrate-resistant acid phosphatase; TRAF6, TNF-receptor associated factor 6; TREM2, triggering receptor
expressed in myeloid cells-2; TV, transcytotic vesicle; Ub, ubiquitin; VDR, vitamin D receptor; WASP, Wiskott–Aldrich syndrome protein.
β1 or β3
Blood vessel within Haversian canal
RANK signalling pathway
N-cadherin or cadherin11
N-cadherin or cadherin11
Type I collagen
Cell polarisation, actin reorganisation,
degraded bone matrix
Undercarboxylated osteocalcin (active)
(stimulates insulin secretion)
Type 1 collagen
Bone lining cells
Modified from Lian et al., 2003. See text for full citation.
IL-4, TNFα, IL-15, TLSP
Polarised resorbing osteoclast
Immature dendritic cell
FusionCommitment Polarisation Activation/survival
The bone remodelling cycle
a Microdamage or mechanical stress
d Osteoblastic bone formation
b Osteoclastic bone resorption
mechanical stress (a)
tiation and activation
of osteoclasts that
resorb the damaged
bone (b). Osteoclasts
die by apoptosis (c),
migrate to the area
of resorbed bone
and replace it with
which then becomes
Mutated in rare human diseases
Cell Science at a Glance
Journal of Cell Science
heterodimeric Smad proteins controls the
expression of Runt-related transcription factor
2 (Runx2), also known as core binding
factor alpha1 (cbfa1), a transcription factor
indispensable for osteoblast differentiation
(Ducy et al., 1997). The canonical Wnt
signalling pathway is indispensable for
osteoblast differentiation during skeletogenesis
and continues to have important roles in mature
osteoblasts (Box 2). Although the major
function of circulating parathyroid hormone
(PTH) is to regulate plasma calcium (see
below), it also has an important role in bone
formation and prevents osteoblast and osteocyte
apoptosis. Intermittent administration of low
levels of PTH increases osteoblast number, bone
formation and bone mass, and is an established
anabolic treatment for osteoporosis.
The exact mechanisms involved in the
anabolic effects of PTH on bone formation are
not fully understood, but might involve Wnt
signalling (Box 2) as well as insulin-like growth
factor 1 (IGF-1). IGF-1, which is released by
the liver in response to growth hormone, has a
role in the commitment of mesenchymal stem
cells to osteoprogenitor cells. IGF-1 also
regulates osteoclastogenesis both directly,
through the IGF receptor (IGFR) present
on osteoclasts, and by upregulating the crucial
osteoclast differentiation factor receptor
activator of nuclear factor B ligand
Another pathway by which osteoblast
function is regulated is the sympathetic nervous
system (Elefteriou et al., 2005). Sympathetic
stimulation through the
located on osteoblasts inhibits bone formation
and increases bone resorption, thereby resulting
in a reduction in bone mass.
Osteoclasts: differentiation and
Osteoclasts are large, multinucleated cells that
form through fusion of mononuclear precursors
of the hematopoietic lineage (see Poster panel
“Osteoclast lineage”). Osteoclast differentiation
initially depends on signalling through c-fms,
the receptor for macrophage colony stimulating
factor (MCSF), in mononuclear precursor cells,
which upregulates expression of RANK
(Crockett et al., 2011). Its ligand, RANKL, is
expressed in osteoblasts and stromal cells in
response to PTH and stimulation by the active
dihydroxy form of vitamin D
(1,25 Vit D
(Leibbrandt and Penninger, 2008). Signalling
through RANK and c-fms in mononuclear
precursors is the key driver of osteoclast
formation. Together with co-stimulation by the
immunoreceptor tyrosine-based activation
motif (ITAM)-containing adaptors DAP12
(DNAX-activating protein of 12 kDa) and FcR
(Fc receptor chain), this leads to activation of
the transcription factors nuclear factor B
(NF-B), activator protein 1 (AP-1) and nuclear
factor of activated T-cells cytoplasmic 1
(NFATc1) (Humphrey et al., 2005). These in
turn regulate expression of essential osteoclast
genes, such as dendritic cell-specific
transmembrane protein (DC-STAMP), tartrate-
resistant acid phosphatase (TRAcP), cathepsin
K, matrix metalloproteinase 9 (MMP-9) and 3
integrin, which allow the final differentiation
and fusion of the precursors and function of the
resulting multinucleated osteoclast.
Loss-of-function mutations of RANKL and
RANK result in the high bone mass disease
osteopetrosis because osteoclast formation is
completely impaired (Table 1) (Guerrini et al.,
2008; Sobacchi et al., 2007). Throughout the
lifespan of the mature osteoclast (several
weeks), continued signalling through c-fms and
RANK is required for osteoclast survival.
RANK signalling is tightly regulated by a decoy
receptor for RANKL, osteoprotegerin (OPG),
which is produced by osteoblasts and stromal
cells, and prevents interaction of RANKL with
RANK. These factors have been explored as
potential therapeutic targets. Although anabolic
treatment with OPG is no longer pursued, an
antibody against RANKL, which is a powerful
anti-catabolic, has recently been launched to
treat diseases in which either osteoclast
formation or osteoclast function is excessive
(Rizzoli et al., 2010).
Osteoclast formation is upregulated in
inflammatory conditions associated with bone
loss, such as rheumatoid arthritis, through the
synergistic action of pro-inflammatory
cytokines, including tumour necrosis factor
(TNF) and RANKL, and by the transdifferenti-
ation of dendritic cells into osteoclasts (Rivollier
et al., 2004). The immune system also regulates
bone loss that is associated with osteoporosis.
Oestrogen deficiency leads to upregulation of
interleukin 7 (IL-7), which induces T-cell
activation and a complex cascade of pathways
all producing cytokines and reactive oxygen
species, thereby resulting in increased RANKL
and TNF production (Weitzmann and Pacifici,
Osteoclast numbers in bone are controlled not
only through formation, but also through the
regulation of their lifespan. Normally,
osteoclasts die by apoptosis, a process that
involves signalling pathways that include
extracellular-signal-regulated kinase (ERK), the
serine/threonine protein kinase Akt and
mammalian target of rapamycin (mTOR), which
regulate the expression of apoptotic factors,
such as B-cell lymphoma-extra large (BclX
the BH3-only family member Bim and myeloid
cell leukaemia sequence 1 (Mcl-1) (Akiyama et
al., 2003; Xing and Boyce, 2005; Bradley et al.,
2008; Sutherland et al., 2009). However,
osteoclast survival is thought to be increased in
pathological conditions that are associated with
increased osteoclast numbers, such as Paget’s
disease of bone (Chamoux et al., 2009).
Pathways leading to osteoclast activation
and initiation of bone resorption
Bone resorption is the process of osteoclast-
mediated destruction of bone matrix. When
Journal of Cell Science 124 (7)
Box 1. Bone formation and function
During embryogenesis, long bones are formed initially as cartilage that becomes gradually
replaced by bone, a process known as endochondral bone formation. By contrast, flat
bones, such as the skull, are formed directly from mesenchymal condensation through a
process called intramembranous ossification. During early childhood, both bone modelling
(formation and shaping) and bone remodelling (replacing or renewing) occurs, whereas in
adulthood bone remodelling is the predominant process to maintain skeletal integrity, with
the exception of massive increases in bone formation that occur after a fracture. Most
bones consist of a mixture of dense outer cortical bone and inner trabecular (spongy) bone,
enabling the optimal compromise between strength and weight. In addition to providing
support, attachment sites for muscles and protection for vulnerable internal organs, bone
also provides a home for bone marrow and acts as a reservoir for minerals.
Osteoblasts produce bone by synthesis and directional secretion of type I collagen,
which makes up over 90% of bone matrix protein. This, together with some minor types of
collagen, proteoglycans, fibronectin and specific bone proteins, such as osteopontin, bone
sialoprotein and osteocalcin, becomes the unmineralised flexible osteoid on which the
osteoblasts reside. Rigidity of bone, which distinguishes it from other collagenous matrices,
is provided by the bone mineral. Mineralisation is achieved by the local release of
phosphate, which is generated by phosphatases present in osteoblast-derived, membrane-
bound matrix vesicles within the osteoid. Together with the abundant calcium in the
extracellular fluid, this results in nucleation and growth of crystals of hydroxyapatite
]. The proportion of organic matrix to mineral (in adult human cortical
bone approximately 60% mineral, 20% organic material, 20% water) is crucial to ensure the
correct balance between stiffness and flexibility of the skeleton.
Journal of Cell Science
osteoclasts are activated to resorb (see
“Polarising osteoclast” in the Poster), they
polarise and form distinct and unique
membrane domains, including the sealing zone
(SZ), the ruffled border (RB) and the functional
secretory domain (FSD) (Mulari et al., 2003).
Importantly, osteoclasts generate these
domains only when they are in contact with
mineralised matrix and do not form an RB
when cultured in vitro on plastic or glass (Saltel
et al., 2004). Osteoclast polarisation involves
rearrangement of the actin cytoskeleton to
form an F-actin ring that comprises a dense
continuous zone of highly dynamic podosomes
(Luxenburg et al., 2007), thereby isolating an
area of membrane that develops into the RB.
The v3-integrin (the vitronectin receptor)
mediates the attachment of podosomes to the
ECM through the formation of a signalling
complex consisting of the tyrosine kinases
c-Src, proline-rich tyrosine kinase 2 (PYK2)
and spleen tyrosine kinase (Syk), as well as a
number of scaffold proteins, including the E3
ligase c-Cbl, paxillin and the Crk-associated
substrate (CAS) family member p130
leads to activation of signalling pathways that
involve phosphoinositide 3-kinase (PI3K) and
phospholipase C (PLC) (which are also
activated by c-fms and complement v3
signalling), and activation of the small
GTPases Rac and Cdc42 by guanine
nucleotide-exchange factors (GEFs) such as
Vav3 (Novack and Faccio, 2009), combined
with deactivation of ADP-ribosylation factor 6
(Arf6) through its GTPase-activating protein
(GAP) GIT2 (G-protein-coupled receptor
kinase-activator 2) (Heckel et al., 2009).
Together with changes in the activity of Rho
and downstream effects on microtubule
acetylation and stabilisation, the combined
action of these pathways promotes actin and
microtubule reorganisation, thereby leading to
the formation of the SZ and subsequently the
RB. Whereas integrin v3 continues to
mediate signalling between the ECM and the
cytoskeleton during resorption, it is likely that
the tight adhesion to bone in the SZ is mediated
through other proteins such as CD44
(Lakkakorpi et al., 1991; Chabadel et al.,
Pathways involved in continued bone
resorption: role of the ruffled border
Maintenance of the RB is essential for
osteoclastic bone resorption (see “Resorbing
osteoclast” in the Poster). The RB is a highly
convoluted membrane that forms as a result of
directed transport of late endosomes and/or
lysosomes, and serves to deliver the proteins
involved in the resorption process. These are the
-ATPase (V-ATPase), whose
role is to acidify the space beneath the RB (the
resorption lacuna), thus enabling dissolution of
bone mineral, and the cysteine protease
cathepsin K, which degrades type I collagen.
The chloride-proton antiporter ClC-7 (Graves et
al., 2008) acts in concert with the V-ATPase at
the RB (as on lysosomes) by transporting
chloride ions into the resorption lacuna (Kornak
et al., 2001). Loss-of-function mutations of all
these proteins and Ostm1, a subunit of ClC-7,
are the underlying basis of most cases of the
high bone mass disease osteoclast-rich
osteopetrosis (Table 1). In this type of
osteopetrosis, osteoclasts form normally, but are
unable to generate an RB and do not resorb
(Villa et al., 2009; Lange et al., 2006). The
trafficking of lysosomal and endosomal
components that results in the formation of the
RB, together with the presence of V-ATPase,
ClC-7 and other lysosomal proteins at the RB
(Palokangas et al., 1997), indicates that this
unusual membrane domain is more akin to an
intracellular lysosomal membrane than to a
plasma membrane (Salo et al., 1996).
Accordingly, formation of the RB is dependent
on the lysosomal small GTPase Rab7 (Zhao et
al., 2001). Another Rab family GTPase, Rab3D,
is also necessary, but localises to a poorly
characterised secretory compartment that is
distinct from the Rab7-regulated lysosomal
compartment (Pavlos et al., 2005).
The bone resorption process creates a high
concentration of degraded collagen fragments,
in addition to calcium and phosphate, within the
resorption lacuna, which are endocytosed by
osteoclasts and then transported through the cell
and released at the FSD (Nesbitt et al., 1997;
Salo et al., 1997) before finally reaching the
bloodstream. During transcytosis, these
collagen fragments are further proteolytically
degraded by cathepsin K (Yamaza et al., 1998)
and TRAcP, an osteoclast-specific enzyme that
is activated by cathepsin K (Ljusberg et al.,
2005). These enzymes are probably
endocytosed from the resorption lacuna together
with the collagen fragments, although it remains
unclear how TRAcP is initially targeted to the
RB. The transcytotic pathway might
additionally play a role in maintenance of the
RB membrane by balancing exocytic and
endocytic events (Stenbeck, 2002).
Many of the processes involved in osteoclast
polarisation and vesicular trafficking that enable
bone resorption are regulated by small GTPases
such as Rac, Rho and Rabs (Coxon and Taylor,
2008), which require post-translational
modification by isoprenylation to localise
correctly in the cell and to exert their specific
function. Indeed, disruption of the key enzymes
involved in isoprenylation in osteoclasts by
bisphosphonates and related compounds leads
to effective inhibition of bone resorption (Coxon
et al., 2006). By virtue of their mineral-binding
property, bisphosphonates specifically target to
bone, where they are released and preferentially
taken up by resorbing osteoclasts during
dissolution of the mineral, explaining their
remarkable selectivity for this cell type in vivo
(Coxon et al., 2006). Bisphosphonate drugs
have been in clinical use for several decades to
treat diseases associated with excessive bone
resorption, such as Paget’s disease of bone,
cancer-induced bone disease and for
Journal of Cell Science 124 (7)
Box 2. Wnt signalling in bone remodelling
Canonical Wnt signalling is a key pathway in bone formation. The activation of -catenin
through the Wnt co-receptors low-density lipoprotein receptor-related proteins 5 and 6
(LRP5, LRP6) and Frizzled results in the upregulation of transcription factors that are
crucial for osteoblast differentiation. Gain-of-function or loss-of-function mutations within
LRP5 are associated with high and low bone mass phenotypes in humans, respectively
(Boyden et al., 2002; Little et al., 2002; Gong et al., 2001). However, as osteoblast-specific
expression of gain-of-function mutations in mice results in a high bone mass phenotype
(Babij et al., 2003) and LRP5 also regulates synthesis of serotonin, a systemic negative
regulator of bone mass, in the duodenum (Yadav et al., 2008; Yadav et al., 2009), the
relative contribution of osteoblasts versus duodenal LRP5 to the regulation of bone mass is
Non-canonical Wnt signalling mediates the commitment of mesenchymal stem cells to
the osteoblast lineage by preventing the expression of peroxisome proliferator activated
receptor- (PPAR), which is required for adipocyte differentiation (Takada et al., 2007).
Osteoporosis and reduced levels of circulating oestrogen are associated with a switch that
favours adipocytic over osteoblastic development (Rosen et al., 2009).
There is evidence for cross-talk between PTH and Wnt signalling, because binding of
PTH to its receptor recruits and phosphorylates LRP6, which leads to stabilisation of -
catenin. In addition, the endogenous inhibitor of Wnt signalling Dickkopf 1 (DKK1) prevents
Wnt-dependent PTH effects (Wan et al., 2008; Guo et al., 2010). In differentiated
osteoblasts, canonical Wnt signalling also stimulates OPG and inhibits RANKL expression,
thereby negatively regulating osteoclast formation (Glass et al., 2005).
Journal of Cell Science
Osteocytes: formation and function
Osteocytes are the most abundant bone cell type,
accounting for 95% of all bone cells. These cells
are osteoblasts that have been spared apoptosis
at the end of a bone formation cycle and have
become incorporated into the bone matrix (see
Poster panel “Osteoblast lineage”), where they
can have a lifespan of decades. During
entombment into the bone matrix (paradoxically
also called osteocyte birth), osteoblasts
profoundly change their morphology, losing
over 70% of cell organelles and cytoplasm, and
acquiring a stellar shape with 50 or more thin
extensions (termed osteocyte processes) that
connect with other osteocytes and also remain
connected with osteoblasts on the bone surface
(Rochefort et al., 2010). The resulting osteocyte
network provides microporosity in the
mineralised bone. Osteocyte bodies are
contained within spaces referred to as lacunae,
whereas their connected processes are contained
within channels (termed canaliculi) – together
they make up the lacunar–canalicular network.
Like the neuronal network, the osteocyte
network processes and transmits signals from
their site of origin to a distant site where an effect
is required. Specifically, the osteocyte network
senses mechanical forces on bone, for example,
by compression or stretching of the bone matrix
during locomotion, and transmits this signal
through its network to ultimately influence the
activities of osteoblasts and osteoclasts on
the bone surface. It was initially thought that
osteocytes respond exclusively to mechanical
stimuli, but it is now clear that they also sense
metabolic signals. Osteocyte death (as evidenced
by the presence of empty osteocyte lacunae) is
increased after oestrogen withdrawal, unloading
of bone and during ageing (Manolagas and
Parfitt, 2010), conditions that are associated with
lower bone mass due to increased remodelling.
These observations, combined with evidence
that experimental ablation of osteocytes in young
mice leads to rapid induction of bone resorption
(Tatsumi et al., 2007), suggest strongly that
living osteocytes have an important role in
negatively regulating osteoclastic resorption,
although the precise signals they send to inhibit
osteoclasts are not yet known. The effects of
osteocytes on osteoblasts are twofold: in
response to sensing mechanical effects on bone,
they positively regulate osteoblasts through the
production of messengers, such as nitric oxide
and prostaglandin E2, and they negatively
regulate osteoblasts through secretion of
sclerostin (discussed further below) (Rochefort
et al., 2010).
Bone cells and bone matrix
Bone cells, like any connective tissue cells, live
in close contact with the abundant ECM, which
has a key role in regulating their proliferation,
differentiation and activation through a variety
of adhesion molecules, as discussed below.
Osteoblasts stably interact with matrix through
integrins. 1 integrins (11, 21 and 51)
seem the most abundant (Helfrich et al., 2008)
and have a crucial role in organising the cells on
the developing bone surface during osteoid
production (Zimmerman et al., 2000).
Osteoblasts also express a range of cell–cell
adhesion molecules, particularly cadherins,
which have a role in osteoblast differentiation
and function (Civitelli et al., 2002; Marie, 2002).
Coupling between cells in the osteoblast lineage
is further mediated by gap junctions and
hemichannels, particularly the junctions formed
by connexin 43 (Civitelli, 2008). These allow
exchange of ions and small molecules, for
example, ATP, nitric oxide and prostaglandins.
Osteoclasts migrate over mineralised trabecular
surfaces and tunnel through cortical bone, and
therefore have only an intermittent relationship
with the matrix. At times, they form tight
adhesive interactions with bone as described
above, but they are also highly motile, even
during active resorption. Osteoclasts use mainly
v3 and 21 integrins to interact with the
ECM (Helfrich et al., 2008). They bind to
collagen through 1 integrins, whereas bone-
specific or bone-enriched RGD-containing
proteins, such as bone sialoprotein and
osteopontin, are bound through 3 integrin
(Helfrich et al., 1992; Helfrich et al., 1996). As
migrating cells, mature osteoclasts do not
express cadherins, but it has been suggested
that cadherins have a role during osteoclast
differentiation to facilitate intimate contact with
Journal of Cell Science 124 (7)
Table 1. New treatments for osteoporosis that have been developed following identification of bone-disease-causing
Mutated protein Normal function
Bone disease resulting from
loss of function (phenotype)
Reference for therapy
RANKL Pro-catabolic: crucial cytokine required for
osteoclast formation, function and survival
(high bone mass)
(Rizzoli et al., 2010)
Cathepsin-K Pro-catabolic: proteolytic enzyme released
by osteoclasts that degrades collagen
(high bone mass)
(Pérez-Castrillón et al.,
V-ATPase Pro-catabolic: proton pump on ruffled
border of resorbing osteoclasts to create
acidic environment to dissolve bone
(high bone mass)
(Huss and Wieczorek,
Ostm1 Pro-catabolic: proton-chloride antiporter on
ruffled border of resorbing osteoclasts,
essential to maintain electroneutrality.
Ostm1 is a subunit of this antiporter
(high bone mass)
(Schaller et al., 2005)
LRP5 Pro-anabolic: co-receptor for canonical Wnt
signalling pathway, promotes osteoblast
differentiation (inhibited by DKK-1)
(low bone mass)
(Glantschnig et al.,
Sclerostin Anti-anabolic: secreted by osteocytes and
inhibits osteoblast differentiation
Sclerosteosis and Van Buchem
(both high bone mass)
(Li et al., 2009)
The anti-catabolic and pro-anabolic therapies described here are also licensed or in development for the treatment of other diseases. Denosumab (Amgen)
has been licensed for use in the treatment of metastatic bone disease and future indications are likely to include rheumatoid arthritis, psoriatic arthritis and
multiple myeloma. Anti-DKK antibodies (Novartis) are in phase I/II clinical trials for the treatment of multiple myeloma. The use of anti-sclerostin antibodies
as an anabolic factor to improve fracture healing is being investigated and is showing promising results in animal models (Paszty et al., 2010).
Journal of Cell Science
stromal cells that express essential growth
factors (Mbalaviele et al., 2006).
Specific interaction points between osteocyte
integrins and the matrix lining the lacunae and
canaliculi might be crucial in generating
and amplifying signals that are induced by tissue
deformation (Wang et al., 2007); roles for 1 or
3 integrins have been suggested (Litzenberger
et al., 2010; McNamara et al., 2009). Live-cell
imaging studies have shown that a population
of osteocytes near the surface of bone is
surprisingly motile, suggesting that the
formation of the osteocyte network might be
more actively controlled by the cells involved
than initially thought (Dallas and Bonewald,
Coupling bone formation to bone
During bone remodelling, bone formation is
tightly coupled to bone resorption, and direct
contacts between osteoclasts and osteoblasts
have been proposed to maintain this
relationship. Recently, the ephrin B (EphB)
receptors (the largest class of receptor tyrosine
kinases) and their ephrinB ligands have been
implicated in this coupling. These
receptor–ligand interactions activate
bidirectional signalling, where interaction
between ligand and receptor induces signalling
in both the receptor-expressing and the ligand-
expressing cells. Here, ‘forward’ signalling
from receptor EphB4 present on osteoblasts
activates a RhoA-dependent pathway to
enhance osteoblast differentiation, whereas the
‘reverse’ signalling from its ligand ephrinB2,
which is expressed by osteoclasts,
downregulates c-Fos and NFATc1 to inhibit
osteoclast function (Matsuo, 2010). The result
of this bidirectional signalling might affect the
switch from bone resorption to bone formation.
EphB4 and ephrinB2 also signal between cells
that belong to the osteoblast lineage and could
thus have additional positive effects on bone
formation during bone remodelling (Martin et
In addition, a number of soluble factors have
been implicated in the coupling between bone
formation and bone resorption, including factors
that are released from bone matrix during
resorption, such as transforming growth factor
(TGF-), factors that are secreted by osteoclasts,
including cardiotrophin-1, TRAcP and
glutamate (Walker et al., 2008; Karsdal et al.,
2007; Coxon and Taylor, 2008), and osteoblast-
derived factors, including oncostatin M (Walker
et al., 2010).
Systemic regulation of bone
Osteoclastic bone resorption is controlled
systemically by four main hormones: calcitonin,
PTH, vitamin D
(1,25 Vit D
) and oestrogen.
Secretion of the first three is driven by the need
to control the serum calcium level within precise
physiological limits (i.e. 2.22.6 mM), with
bone acting as a mineral reservoir for this
homeostasis. Calcitonin acts through its
receptors that are expressed specifically on
osteoclasts and directly inhibits osteoclastic
resorption (Zaidi et al., 2002). By contrast, PTH
binds to its receptors that are expressed on
osteoblasts and bone marrow stromal cells, in
which, through signalling by cAMP responsive
element binding protein (CREB), it activates
expression of MCSF and RANKL, thereby
indirectly stimulating osteoclastic bone
resorption. An important non-skeletal action of
PTH is to stimulate increased renal reabsorption
of calcium, which, together with increased
resorption to mobilise calcium from bone,
restores physiological serum calcium levels
(Talmage and Elliott, 1958). PTH also
stimulates the production of 1,25 Vit D
circulating inactive precursor. 1,25 Vit D
turn, facilitates calcium absorption from the gut
and the kidney, and also positively regulates
bone resorption indirectly through the 1,25 Vit
and retinoid X receptors in osteoblasts,
which increases RANKL and MCSF
The major role for oestrogen in the skeletal
system is as a bone-sparing hormone that acts
through receptors expressed by both osteoclasts
and osteoblasts. This sex hormone is crucial in
the control of osteoclast lifespan, and can cause
pre-osteoclast and osteoclast apoptosis through
Fas and Fas ligand signalling. Therefore, loss of
oestrogen in women after the menopause results
in increased osteoclast formation and survival
(Krum et al., 2008; Nakamura et al., 2007).
Oestrogen also blocks osteoclast function
indirectly through effects on the immune system
and has a role in regulating the response of bone
to mechanical stimulation (Zaman et al., 2006).
Bone as an endocrine organ
A recently emerged role for bone is that of an
endocrine organ. Firstly, the osteoblast-derived
protein osteocalcin was identified as a positive
regulator of pancreatic insulin secretion (Lee
et al., 2007). Osteocalcin expression is
upregulated by insulin signalling in osteoblasts
through downregulation of Twist, an inhibitor of
Runx2 (Fulzele et al., 2010), whereas insulin-
dependent downregulation of OPG in
osteoblasts stimulates osteoclasts and lowers the
pH of the bone ECM, which is required for
activation of osteocalcin before it enters the
circulation (Hinoi et al., 2008; Ferron et al.,
Secondly, a new role for osteocytes in
regulating bone metabolism has recently come
to light. Osteocytes synthesise fibroblast growth
factor 23 (FGF23), which plays a key role in
phosphate homeostasis by acting on the
parathyroid gland and the kidney to reduce
circulating phosphate levels. FGF23 production
is stimulated by 1,25 Vit D
and acts, in turn, by
reducing 1,25 Vit D
levels. Two other
osteocyte-expressed proteins, phosphate
regulating endopeptidase homolog, X-linked
(PHEX) and dentin matrix acidic
phosphoprotein 1 (DMP1), are thought to
negatively regulate of FGF23 in the osteocyte
Finally, through studying patients with the
rare hereditary high bone mass conditions
sclerosteosis and Van Buchem disease, the
SOST gene that encodes sclerostin, an inhibitor
of bone formation, has been discovered
(Balemans and Van Hul, 2004). Sclerostin is
synthesised exclusively by bone cells that are in
contact with mineral, that is, late-stage
osteoblasts and osteocytes. Expression of
sclerostin is inhibited by PTH and oncostatin M
(Walker et al., 2010), and by mechanical loading
of bone (Robling et al., 2008). Sclerostin acts as
an inhibitor of Wnt signalling (Box 2). Owing to
its exclusive expression in bone, sclerostin has
become a key target for development of novel
bone anabolics; anti-sclerostin antibodies are
currently in phase 2 clinical trials (Table 1).
Mechanical regulation of bone
Mechanical force is a key regulator of bone
remodelling and of bone architecture in general
(Jacobs et al., 2010). It influences bone
metabolism not only locally (e.g. resulting in a
bigger bone in the serving arm of a professional
tennis player), but also systemically (as
illustrated by the profound bone loss in
astronauts experiencing zero gravity and
in immobilised patients). Whole-animal studies
have shown dramatic responses to mechanical
stimuli at the tissue level and in vitro studies
have confirmed that individual bone cells, such
as osteocytes and osteoblasts, are able to sense
and respond to mechanical forces. Early signals
produced in response to mechanical stimuli
include nitric oxide, prostaglandins and Wnt
signalling proteins (Bonewald and Johnson,
2008). In osteocytes, -catenin rapidly
translocates to the nucleus after exposure to
fluid shear stress, suggesting activation of Wnt
signalling or cadherin-mediated signalling
(Huesa et al., 2009; Norvell et al., 2004; Santos
et al., 2010). However, the precise mechanical
stimulus that is sensed by bone cells in vivo and
Journal of Cell Science 124 (7)
Journal of Cell Science
the signal produced as a result remain unclear.
Despite this, the profound anabolic effects of
mechanical stimulation of bone have prompted
the development of mechanical therapies to
increase bone mass. The presence of
microcracks in bone affects mechanosensing
and is currently considered a crucial driver of the
remodelling response by initiating osteoclastic
resorption (Cardoso et al., 2009).
Over the past “Bone and Joint Decade”, the
progress made in understanding bone
remodelling, both through experimental
approaches and by uncovering the molecular
basis of inherited bone disease, has been truly
spectacular. The vast amount of new knowledge
has already been translated into novel
therapeutic approaches for the treatment of
common bone diseases and is leading to the
development of better biomarkers to monitor
response to treatment. There is the exciting
possibility that soon we will be able to
understand the genetic predisposition for age-
related bone loss (Box 3), develop better
screening methods to identify those most at risk
and use genetic information to decide on the
most appropriate treatment. All this will require
intricate knowledge of bone anatomy, bone cell
function and bone remodelling to inform our
understanding of the functional consequences of
such genetic differences.
Work of the authors in this field has been supported by
research grants from Arthritis Research UK, Medical
Research Council, Nuffield Foundation, National
Association for the Relief of Paget’s Disease, Chief
Scientist’s Office, European Calcified Tissue Society,
Warner Chilcott and Novartis.
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Box 3. Genetic contributors to osteoporosis
Identification of the molecules that are essential for bone cell formation or cause inherited
disorders of osteoclast function has led to a better molecular understanding of the common
bone disorder osteoporosis. This age-related low bone mass condition, most prominent in
women following the menopause, can be assessed by measuring bone mineral density
(BMD). BMD varies naturally within the population, and peak bone mass and the rate of
bone loss in later life are determined by the interplay between multiple genetic and
environmental factors. Common genetic variants associated with BMD have been reported
for components of several of the signalling pathways mentioned above, including those
activated by RANKL, Wnt-LRP, TGF- and BMPs, as well as cytoskeletal scaffolding
proteins, GAPs and endosomal transporters. In addition, genetic variation also affects
nuclear hormone receptors, for example, vitamin D receptor and oestrogen receptor, and
the ECM (Kiel et al., 2007; Rivadeneira et al., 2009).
However, the contribution of the individual genetic variants that have been identified to
date to the overall variation in BMD and bone loss is typically small (Rivadeneira et al.,
2009). Therefore, their combined effects will be the important factor in determining the risk
of osteoporotic fractures. Additionally, in the majority of cases, the functional variants of
these genes have not yet been identified, thus preventing an accurate estimation of the
actual risk or a meaningful assessment of their combined effects at a molecular or cellular
level. There is also evidence for site-specific effects of genetic variants. Such localised
effects undoubtedly occur through interactions with the local environment – for example
under different loading conditions – but also concur with the evidence for intrinsic
differences in bone cell functions between different skeletal sites (Everts et al., 2009).
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