Skip Navigation



Rheumatology Advance Access published online on October 16, 2008
Rheumatology, doi:10.1093/rheumatology/ken368
This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow All Versions of this Article:
47/12/1785    most recent
ken368v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Bonnelye, E.
Right arrow Articles by Aubin, J. E.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Bonnelye, E.
Right arrow Articles by Aubin, J. E.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?

© The Author 2008. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org

Estrogen receptor-related receptor-{alpha} (ERR-{alpha}) is dysregulated in inflammatory arthritis

E. Bonnelye1, N. Laurin1, P. Jurdic2, D. A. Hart3 and J. E. Aubin1

1Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada 2Institut de Génomique Fonctionelle de Lyon, UMR 5242, CNRS/INRA/ENS/Université Lyon1, IFR 128 Lyon Biosciences, Ecole Normale Superieure, Lyon, France and 3Department of Surgery, McCaig Institute for Bone and Joint Health, Faculty of Medicine, University of Calgary, AB, Canada.

Correspondence to: J. E. Aubin, Department of Molecular Genetics, Faculty of Medicine, University of Toronto, Room 6233, Medical Sciences Building, 1 King's College Circle, Toronto, ON M5S 1A8, Canada. E-mail: jane.aubin{at}utoronto.ca


   Abstract
Top
 Abstract
Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
Objectives. Subchondral bone loss is a characteristic feature of inflammatory arthritis. Recently, estrogen receptor-related receptor-{alpha} (ERR-{alpha}), an orphan nuclear receptor, has been found to be involved in activation of macrophages. We hypothesized that ERR-{alpha} which is expressed and also functional in articular chondrocytes, osteoblasts and osteoclasts, may be involved in rodent models of inflammatory arthritis.

Methods. Erosive arthritis was induced in DBA/1 mice by injection of type II collagen in Freund's complete adjuvant. RNA was isolated from the bone and joints and expression of ERR-{alpha} and cartilage (GDF5 and Col2a1) and bone [bone sialoprotein (BSP) and osteocalcin (OCN)] markers was analysed by semi-quantitative PCR.

Results. We report for the first time that the expression of ERR-{alpha} is dysregulated in bones and joints in a mouse model of inflammatory arthritis. Specifically, we show that ERR-{alpha} expression is down-regulated early in bone and later in joints of mice with type II CIA. Concomitantly, temporal changes were observed in GDF-5 and Col2a1 expression in joints following both initial injection and booster injection of type II collagen. Similarly, down-regulation of ERR-{alpha} mRNA expression in subchondral bone in mice with induced joint inflammation was also paralleled by down-regulation of markers of bone formation (BSP, OCN).

Conclusions. These data suggest that dysregulation of ERR-{alpha} expression may precede and contribute to the destruction of cartilage and bone accompanying inflammatory arthritis.

KEY WORDS: Joint, Bone, Rheumatoid arthritis, Nuclear receptor, Estrogen receptor-related receptor


   Introduction
Top
 Abstract
 Introduction
Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
RA is a chronic inflammatory autoimmune disease characterized by invasive synovial hyperplasia leading to progressive joint and bone destruction [1]. Bone erosion most frequently occurs at sites of inflammation and cartilage–pannus and bone–pannus interfaces, with involvement of multiple cell types including synovial cells, immune cells, osteoclasts and articular chondrocytes.

The nuclear receptor superfamily comprises both ligand-dependent molecules such as the estrogen receptors (ERs), and a large number of so-called orphan receptors for which no ligand has yet been identified [2]. Three orphan receptors, ER-related receptor {alpha} (ERR-{alpha}), ERR-β and ERR-{gamma} (NR3B1, NR3B2 and NR3B3, respectively, according to the Nuclear Receptors Nomenclature Committee, 1999), share similarity with ER-{alpha} and ER-β (NR3A1 and NR3A2, respectively) [3, 4]; however, they do not bind estrogens [5–9]. Indeed, sequence alignment of ERR-{alpha} and the ERs reveals a high similarity (68%) in the 66 amino acids of the DNA-binding domain but only a moderate similarity (36%) in the ligand-binding domain which may explain the fact that ERR-{alpha} recognizes the same DNA-binding elements as ERs but does not bind estrogen [10]. Based on its ability to recognize similar DNA sequences as the ERs and the ability of estrogens to stimulate ERR-{alpha} gene expression in several tissues (uterus, heart, bone, etc.), ERR-{alpha} has been proposed to modulate estrogen signalling [11–16]. In addition to modulation of ER-mediated transcriptional activity, ERR-{alpha} regulates fatty acid oxidation and the adaptative bioenergetic response [17–19]. Moreover, ERR-{alpha} is involved in cartilage and bone formation in vitro [20–22], and an ESRRA gene regulatory variant associated with increased ERR-{alpha} expression has been associated with increased bone mineral density in pre-menopausal women [23, 24]. Very recently, ERR-{alpha} has also been described to be involved in macrophage activation [25].

Due to ERR-{alpha} expression in bone and articular chondrocytes in adult rats and its function in bone and cartilage formation in vitro, and its function in immune cells, we hypothesized that ERR-{alpha} may be regulated in rodent models of inflammatory arthritis.


   Materials and methods
Top
 Abstract
 Introduction
 Materials and methods
Results
 Discussion
 Acknowledgements
 References
 
Mouse model of collagen-induced inflammatory arthritis
The protocol was given ethical approval by the University of Calgary Animal Care Committee. Male DBA/1 mice were obtained from Jackson Laboratories (Bar Harbor, ME, USA) at the age of 8 weeks. The animals were housed locally in the University of Calgary Animal Care Centre until 12–14 weeks of age to acclimatize and reach an age when induction of arthritis by collagen II is more uniform. This model of inflammatory arthritis is widely used [26, 27] and animals injected with collagen II in adjuvant (adjuvant + type II collagen) develop an RA-like disease based on histological analysis. Groups of animals were injected with either pyrogen-free phosphate buffered saline (PBS, controls), Freund's complete adjuvant (FCA, adjuvant controls) or 100 µg collagen II in FCA (experimentals) via the base of the tail on day 0. Emulsions of collagen II (bovine, catalogue number IMAD, Chondrex, Inc., Redmond, WA, USA) and FCA (catalogue number IMBII, Chondrex, Inc.) were prepared with a Polytron homogenizer according to the supplier's protocols (Chondrex, Inc.). In specific experiments, groups of animals were boosted with the appropriate preparation at week 5. At the time of sacrifice (1, 2, 3, 5 and 7 weeks post-treatment), animals were anaesthetized, bled for serum and then terminated by cervical dislocation.

Production of Flt3 ligand cells and macrophage or dendritic cell differentiation
Flt3 ligand cells (FL cells) were generated in vitro as previously described [28]. Briefly, total bone marrow cells from C57BL/6 mice (IFRA-CREDO, Lyon, France) were seeded at 5 x 105cells/ml in Iscove's modified Dulbecco's medium (IMDM; Life Technologies, Cergy-Pontoise, France) supplemented with 15% fetal calf serum (FCS, Dutsher, Brumath, France) and 5 ng/ml human FL (R&D Systems, Lille, France). After 6 days of culture, non-adherent cells were collected and designated as FL cells. For macrophage production, FL cells were cultured for 4 days in the presence of 20 ng/ml of M-CSF. Dendritic cells (DCs) were obtained from FL cells cultured in RPMI 1640, supplemented with 10 mM Hepes, 2 mM L-glutamine complemented with 5 ng/ml murine GM-CSF and 10 ng/ml murine TNF-{alpha} (Peprotech, Rocky Hill, NJ, USA) [28].

RT-PCR
Total RNA was extracted with Trizol reagent from mouse joints and bones, DCs, FL cells and macrophages. Briefly, the epiphyses of the mouse distal femurs and proximal tibiae (joints) were dissected from the metaphyseal and diaphyseal bones (bones) (Fig. 1); marrow was flushed quickly from bones with ice cold PBS until bones were blanched. Both joints and bones were immediately snap-frozen in liquid nitrogen, pulverized over liquid nitrogen using a mortar and then extracted with Trizol reagent. Samples of total RNA (1.5–5 µg) were reverse-transcribed using oligo dT and the first-strand synthesis kit of SuperscriptTM II (Invitrogen, Carlsbad, CA, USA). PCR was performed with primers specific for L32, a ribosomal protein used as internal control for normalization and ERR-{alpha} as described [22] (Qiagen, Hilden, Germany). Chondrocyte- and osteoblast-associated and other markers were amplified using primers specific for Col2a1 (collagen type II {alpha} chain) [21], osteocalcin (OCN) and bone sialoprotein (BSP) [22], GDF-5 [29], c-fms (MM_FMSCR), MHCII (AF387099 [GenBank] ). Amplimers were sequenced for verification. All primers used are shown in Table 1.


Figure 1
View larger version (13K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		FIG. 1. Schematic representation of the dissection protocol for isolating total RNA from bones and joints. The epiphyses of the mouse distal femurs and proximal tibiae (joints; includes pannus, dark grey) were dissected from the metaphyseal and diaphyseal bones from which marrow was removed by flushing (bones). See Materials and methods section for more details.

 

View this table:
[in this window]
[in a new window]

  		TABLE 1. Primers and conditions used for PCR

 
Statistical analysis
Results of PCR analysis were expressed as mean ± S.D., and analysed statistically by one-way analysis of variance (ANOVA) with treatment group as variance and t-test post-tests with InStat software (version 2.01, GraphPad Software, San Diego, CA, USA). Statistical significance was taken as P < 0.05.


   Results
Top
 Abstract
 Introduction
 Materials and methods
 Results
Discussion
 Acknowledgements
 References
 
ERR-{alpha} expression is decreased in the joints and subchondral bone in type II CIA in mouse
To ask whether ERR-{alpha} dysregulation may play a role in conditions in which cartilage integrity is lost, e.g. in joints undergoing destruction as seen in inflammatory arthritis, erosive arthritis was induced by injection of type II collagen in FCA (adjuvant + type II collagen) into DBA/1 mice [26, 27]. Semi-quantitative RT–PCR of RNA from joints 7 weeks after initiation of treatment revealed a dramatic decrease in ERR-{alpha} expression in adjuvant-injected (Fig. 2A and B; samples 5–8) and adjuvant + type II collagen-injected (Fig. 2A and B; samples 9–12) mice compared with control mice injected with PBS (Fig. 2A and B; samples 1–4). The decrease in ERR-{alpha} expression was even more striking when mice were boosted with the same treatments 5 weeks after the first injection (Fig. 2A and B; samples 13–16). Given our previous observations on a functional role of ERR-{alpha} in osteoblasts and bone [22] and the changes that occur in subchondral bone in erosive arthritis [30], we also assessed ERR-{alpha} gene expression in subchondral bones from the same treated mice. Although no differences were seen in ERR-{alpha} gene expression in adjuvant-treated mice ([Fig. 2C and D; samples 5–8) compared with PBS-treated control mice (Fig. 2C and D; samples 1–4), ERR-{alpha} expression was significantly inhibited in mice injected with adjuvant + type II collagen (Fig. 2C and D; samples 9–12) and in mice boosted at 5 weeks (Fig. 2C and D; samples 13–16).


Figure 2
View larger version (31K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		FIG. 2. Expression of ERR-{alpha} in joints and femur samples from mice with inflammatory arthritis induced by injection of type II collagen. RNA was extracted at 7 weeks separately from joints or bones (minus bone marrow) from each of four mice injected with PBS as a control (samples 1–4), adjuvant alone (samples 5–8) or adjuvant and collagen type II (adjuvant + type II collagen) once at day 0 (samples 9–12) or twice (booster injection at 5 weeks; samples 13–16). Semi-quantitative RT–PCR was done separately on RNA from joint samples (A) and femur samples (C) from each mouse. The mean ± S.D. of the pooled joint samples (n = 4) (B) and femur samples (n = 4) (D) is also shown (ANOVA, P < 0.05; Student's t-test post-tests *P < 0.05, **P < 0.01, PBS control vs injected).

 
ERR-{alpha} expression is down-regulated late in joints and early in bone in type II CIA
To determine whether changes in ERR-{alpha} occur acutely and are early indicators of joint destruction or reflect loss of chondrocytes in the later stages of disease, ERR-{alpha} expression was assessed over a time course beginning at 1 week post-injection. We first confirmed that an inflammatory response was present at all time-points, as seen by the robust increase in M-CSF receptor c-fms [31] expression in injected compared with non-injected joints (Fig. 3G and H). Consistent with results reported above, ERR-{alpha} expression was significantly reduced in joints 7 weeks after initial induction [mice boosted at 5 weeks; Fig 3A and B (samples 16–19)] compared with PBS controls [Fig. 3A and B; samples 1–4). There was a non-significant trend towards reduced ERR-{alpha} expression at 1, 2 and 3 weeks post-injection (Fig. 3A and B; samples 5–8, 9–12 and 13–15, respectively). We next assessed expression, in the same samples, of mRNA for articular chondrocytes (GDF5, a TGF-β/BMP superfamily member [32]) and for proliferating chondrocytes (Col2a1). Compared with PBS controls (Fig 3C and D; samples 1–4), there was a striking down-regulation of GDF5 gene expression at 1 and 2 weeks (Fig. 3C and D; samples 5–8 and 9–12) and at 7 weeks (mice boosted at 5 weeks; Fig. 3C and D (samples 16–19)] but a recovery to control levels at 3 weeks (Fig. 3C and D; samples 13–15). Col2a1 expression, on the other hand, was increased at 1, 2 (Fig. 3E and F; samples 5–8 and 9–12) and 7 weeks [mice boosted at 5 weeks; Fig. 3E and F (samples 16–19)] and inhibited at 3 weeks (Fig. 3E and F; samples 13–15) compared with PBS controls (Fig. 3E and F; samples 1–4).


Figure 3
View larger version (33K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		FIG. 3. Late down-regulation of ERR-{alpha} expression in joint cartilage destruction. ERR-{alpha} and marker expression during a time-course experiment in RNA extracted from joints from mice injected with PBS (samples 1–4) (after 4 weeks; midway between start and end points of treated mice), or adjuvant + type II collagen after 1 week (samples 5–8), 2 weeks (samples 9–12), 3 weeks (samples 13–15) and at 7 weeks (booster at 5 weeks; samples 16–19). Semi-quantitative RT–PCR was done separately on RNA from joint samples from each mouse (A, C, E and G). The mean ± S.D. of the pooled joint samples (n = 4) is also shown (B, D, F and H). Results are shown for expression of ERR-{alpha} (A and B), GDF-5 (articular chondrocyte marker) (C and D), Col2a1 (proliferating chondrocyte marker) (E and F) and c-fms (macrophages marker) (G and H). PCR products in each case were normalized to L32. There was a downward trend in ERR-{alpha} expression compared PBS control at all times but with a significant decrease seen only in joints from boosted mice (ANOVA, P < 0.05; Student t post test, **P < 0.01, PBS control vs treated). c-fms was markedly increased at all times in adjuvant + type II collagen-injected mice compared with PBS controls, but GDF5 and Col2a1 were differentially (and oppositely) regulated with time (ANOVA, P < 0.001 for GDF5, P < 0.0001 for Col2a1 and c-fms, Student's t-test post-tests **P < 0.01, ***P < 0.001, PBS control vs treated).

 
Interestingly, and in contrast to what was seen in cartilage in the same mice, ERR-{alpha} gene expression was significantly down-regulated in subchondral bone at all time-points post-injection (Fig. 4A and B). Expression of the osteoblast associated markers, BSP and OCN, in the same samples was also significantly decreased at 1, 3 and 7 (after boost at 5 weeks) weeks [Fig. 3C and D (samples 5–8); Fig. 3E and F (samples 5–8)] compared with PBS controls [Fig. 4C and D (samples 1–4); Fig. 4E and F (samples 1–4)]; both markers tended towards control levels at 2 weeks [Fig. 4C and D (samples 9–12); Fig. 4E and F (samples 9–12)].


Figure 4
View larger version (37K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		FIG. 4. Early decrease of ERR-{alpha} expression in bone destruction in type II CIA. ERR-{alpha} and marker expression during a time-course experiment in RNA extracted from femoral bones from mice injected with PBS (samples 1–4) (after 4 weeks; mid-way between start and end points of treated mice), or adjuvant + type II collagen after 1 week (samples 5–8), 2 weeks (samples 9–12), 3 weeks (samples 13–15) and at 7 weeks (booster at 5 weeks; samples 16–19). Semi-quantitative RT-PCR was done separately on RNA from bone samples from each mouse (A, C and E). The mean ± S.D. of the pooled bone samples (n = 4) is also shown (B, D and F). Results are shown for expression of ERR-{alpha} (A and B) and osteoblast markers [BSP (C and D) and OCN (E and F)]. PCR products in each case were normalized to L32. ERR-{alpha} expression was significantly decreased at all times in adjuvant + type II collagen-injected mice compared with PBS controls; BSP and OCN were also decreased at all times (except BSP at 2 weeks; ANOVA P < 0.01, P < 0.05 and P < 0.0001, respectively; Student's t-test post-tests *P < 0.05, **P < 0.01, ***P < 0.001, PBS control vs treated).

 
ERR-{alpha} mRNA is expressed in monocyte-derived cells and precursors
We have previously shown that ERR-{alpha} is expressed in monocyte-derived cells such as osteoclasts [13], suggesting that it may also be expressed by other cell types derived from the same haematopoietic lineage, including macrophages (M) or DCs, both of which are known to be crucial in the inflammation process in RA. Since RNA extracted from the joints also includes a contribution from the pannus, we therefore hypothesized that the late down-regulation of ERR-{alpha} observed in joints in type II CIA may reflect ERR-{alpha} expression in haematopoietic cells. Consistent with this, we found that ERR-{alpha} mRNAs is expressed in early monocyte precursors (FL cells) obtained from mouse bone marrow after 6 days of expansion with FL, in mature DCs and macrophages (M) (Fig. 5A and B). FL cells are haematopoietic precursors that can differentiate into macrophages, osteoclasts or DCs depending on the cytokines used during cell cultures and exhibit immature monocyte characteristics including expression of the M-CSF receptor, c-fms (Fig. 5A) [28]. As cell lineage controls, mRNA levels for the c-fms marker (expressed by FL cells and macrophages) and the immunoreceptor, MHCII (known to be very abundant in DCs) are also shown (Fig. 5B).


Figure 5
View larger version (45K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		FIG. 5. ERR-{alpha} expression in monocyte-macrophage lineage cells. Detection by PCR of ERR-{alpha} mRNA expression in (A) FL cells and in (B) macrophages (M) and DCs. Specific markers of FL cells (c-fms), DCs (MHII) and macrophages (c-fms) are also shown.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
Acknowledgements
 References
 
ERR-{alpha} expression in fetal and adult growth plate and bone suggests that it may be involved not only in the formation but also in the maintenance and integrity of cartilage/bone throughout the lifetime of the organism. In keeping with this hypothesis, we showed previously that ERR-{alpha} is expressed in osteoblasts and chondrocytes and appears to have an anabolic function in both bone and cartilage [21, 22, 24]. This is consistent with the results reported here in which we found that ERR-{alpha} is decreased in both the joints and subchondral bone, concomitant with decreases in chondrocyte and osteoblast markers respectively, in collagen-induced inflammatory arthritis in susceptible DBA/1 male mice (Fig. 6). It is also worth noting that only very small changes in ERR-{alpha} levels were seen, and only in the more susceptible male but not female mice, in parallel experiments done in C57BL6 mice, a strain not as highly susceptible to CIA [33] (data not shown).


Figure 6
View larger version (33K):
[in this window]
[in a new window]
[Download PowerPoint slide]
  		FIG. 6. Schematic representation of ERR-{alpha} dysregulation and other markers in inflammatory arthritis vs normal joints. ERR-{alpha} is decreased in mouse joints when inflammatory arthritis is induced by collagen type II injection. Concomitantly, GDF5 expression is decreased in joints and Col2a1 expression is increased, as cells undergo a repair response. A bone response is also evident with decreased expression of matrix molecules BSP and OCN.

 
In spite of some animal to animal variation in levels of molecules quantified, statistically significant differences in ERR-{alpha} and cartilage and bone markers were seen with treatment condition and time. The temporal changes observed in GDF-5 (articular chondrocyte marker) and Col2a1 (marker of articular and growth plate chondrocytes) expression following both the initial injection and the booster injection of collagen type II are similar to the changes seen in the phenotype of articular chondrocytes during osteoarthritic cartilage degeneration, i.e. transition towards a more fetal cellular phenotype [34, 35]. Although some aspects of the chondrocyte phenotype appear to ‘recover’ at intermediate times between initial and booster injections (see the 2–3 week time-points), ERR-{alpha} expression remains depressed at all times, suggesting that the chondrocyte phenotype is irreversibly modified by the inflammatory stimulus and that ERR-{alpha} may be a marker of this modification. However, it is also likely that the overall ERR-{alpha} levels reflect the complexity of the changes in phenotype of both the articular and growth plate chondrocytes in which ERR-{alpha} is expressed. Further, and as previously mentioned, the RNA extracted from the joints also includes a contribution from the pannus, which is known to contain macrophages and multinucleated tartrate-resistant acid phosphatase-positive cells (osteoclast-like cells) identified at erosive surfaces in joints of mice with CIA [36, 37]. We previously showed that ERR-{alpha} is expressed in osteoclasts in vivo and in vitro (i.e., the mouse leukaemic monocyte macrophage cell line, RAW, before and after differentiation; [13]). Although a functional role for ERR-{alpha} in osteoclasts has not yet been elucidated, we have shown that ERR-{alpha} is a potent regulator of the osteoprotegerin (OPG)/receptor activator of nuclear factor-kappa B ligand (RANKL) ratio [38–40] in osteoblasts in vitro [13], indicating that ERR-{alpha} can regulate osteoclast development at least indirectly and may therefore predict a parallel role in joint destruction in RA. Moreover, recently, ERR-{alpha} had been associated with activation of macrophages [25]. Concomitant to these results, we confirm ERR-{alpha} expression in macrophages and found its presence in DCs suggesting that ERR-{alpha} may also play a role in the inflammation in RA. It should be noted that ERR-{alpha} mRNA levels were also decreased in the joints following exposure of the mice to Freund's adjuvant alone. As adjuvant alone does not lead to a T-lymphocyte-dependent, progressive and erosive arthritis [41, 42], other mechanisms must be operative. These may relate to macrophage activation and down-regulation of ERR-{alpha} and/or release of soluble factors from such activated macrophages which could contribute to the apparent down-regulation of ERR-{alpha} expression by other cell types. This is a potentially important point since the mice receiving adjuvant + type II collagen would have the induction of the T-lymphocyte arthritis superimposed on a background of adjuvant-induced changes. However, while the changes in expression of ERR-{alpha} mRNA in joint tissues was similar in adjuvant and adjuvant + type II collagen-treated mice, only in the latter were changes detected in ERR-{alpha} mRNA levels in the subchondral bone.

The down-regulation of ERR-{alpha} mRNA expression in subchondral bone (bone from which marrow had been removed; see Materials and methods section) in mice with induced joint inflammation, paralleled by down-regulation of markers of bone formation (BSP, OCN), are notable for the rapidity of response of subchondral osteoblasts to joint inflammation. These data suggest that dysregulation of ERR-{alpha} expression may precede and contribute to the development of the osteopenia accompanying RA. Our data on bone after initial and booster injections also suggest that osteoblast function parallels closely the presence or loss of the inflammatory stimulus. The fact that osteoblast markers, but not ERR-{alpha} itself, recovers in bone again suggests the interesting possibility that ERR-{alpha} is a useful marker of early changes in subchondral bone in inflamed joints. Moreover, the acute decrease in ERR-{alpha} expression in bone after induction of joint inflammation may participate in stimulation of osteoclast development and, consequently, the destruction of joint cartilage and bone. This is consistent with the fact that RANKL is now considered a therapeutic target in arthritis, as OPG treatment prevents bone loss at inflamed joints and has partially beneficial effects on cartilage destruction in arthritis models [43].

ERR-{alpha} has recently been shown to be activated by IFN-{gamma} and to be a key effector of IFN-{gamma} in innate response to bacterial pathogenesis [25]. As a result, mice lacking ERR-{alpha} are susceptible to infection by intracellular pathogens. IFN-{gamma} is also known to play a critical role in RA as an anti-inflammatory molecule, especially during the initial phase of RA [44], and both inflammatory and degenerative arthritis parameters (cartilage and bone destruction) are significantly exacerbated in the absence of IFN-{gamma} [45]. The role of IFN-{gamma}—as an anti-inflammatory, chondroprotective and anti-osteoclastogenic molecule—support the importance of ERR-{alpha} activation in immune cells, articular chondrocytes and osteoclasts in RA.

In conclusion, our data suggest that ERR-{alpha} expression level may be useful as a diagnostic marker for diseases characterized by destruction of cartilage and bone such as that seen in inflammatory arthritic conditions. Together with our analyses of ERR-{alpha} function in osteogenesis and chondrogenesis, they also point towards modulation of ERR-{alpha} activity as a therapeutic strategy in skeletal diseases.

Formula


   Acknowledgements
Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
References
 
We are very grateful to Usha Bhargava, Carol Reno, and Helen Tsao for their technical help. This work was supported by grants from The Arthritis Society of Canada (TAS 04/0031), the Canadian Institutes of Health Research (CIHR; IMHA-67519; MOP-88104) and the Canadian Arthritis Network to J.E.A., by fellowship support from the Association Jacques Cartier, the Association pour la Recherche sur la Polyarthrite (ARP, France), The Arthritis Society of Canada and CNRS to E.B., and by grants from the Institute of Gender and Health (CIHR) and The Arthritis Society of Canada to D.A.H. D.A.H. is the Calgary Foundation-Grace Glaum Professor in Arthritis Research.

Funding: This work was supported by grants from The Arthritis Society of Canada (TAS 04/0031) and the Canadian Arthritis Network to J.E.A., by fellowship support from the Association Jacques Cartier and the Association pour la Recherche sur la Polyarthrite (ARP, France) and the Arthritis Society of Canada and CNRS to E.B., and by grants from the Institute of Gender and Health (CIHR) and The Arthritis Society to D.A.H. D.A.H. is the Calgary Foundation-Grace Glaum Professor in Arthritis Research.

Disclosure statement: The authors have declared no conflicts of interest.


   Notes
 
Present addresses: E. Bonnelye, INSERM UMR 664, Faculté de Médecine RTH Laennec, Rue Guillaume Paradin. 69372 Lyon Cedex 08, France; N. Laurin, Department of Biochemistry, Microbiology, and Immunology, University of Ottawa, Ottawa, ON, Canada.


   References
Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 

  1. Firestein GS. Evolving concepts of rheumatoid arthritis. Nature (2003) 423:356–61.[CrossRef][Web of Science][Medline]
  2. Benoit G, Cooney A, Giguere V, et al. International union of pharmacology. LXVI. Orphan nuclear receptors. Pharmacol Rev (2006) 58:798–836.[Abstract/Free Full Text]
  3. Kuiper GG, Enmark E, Pelto-Huikko M, Nilsson S, Gustafsson JA. Cloning of a novel receptor expressed in rat prostate and ovary. Proc Natl Acad Sci USA (1996) 93:5925–30.[Abstract/Free Full Text]
  4. Green S, Walter P, Kumar V, et al. Human oestrogen receptor cDNA: sequence, expression and homology to v-erb-A. Nature (1986) 320:134–9.[CrossRef][Web of Science][Medline]
  5. Kallen J, Schlaeppi JM, Bitsch F, et al. Evidence for ligand-independent transcriptional activation of the human estrogen-related receptor alpha (ERRalpha): crystal structure of ERRalpha ligand binding domain in complex with peroxisome proliferator-activated receptor coactivator-1alpha. J Biol Chem (2004) 279:49330–7.[Abstract/Free Full Text]
  6. Greschik H, Wurtz JM, Sanglier S, et al. Structural and functional evidence for ligand-independent transcriptional activation by the estrogen-related receptor 3. Mol Cell (2002) 9:303–13.[CrossRef][Web of Science][Medline]
  7. Greschik H, Flaig R, Renaud JP, Moras D. Structural basis for the deactivation of the estrogen-related receptor gamma by diethylstilbestrol or 4-hydroxytamoxifen and determinants of selectivity. J Biol Chem (2004) 279:33639–46.[Abstract/Free Full Text]
  8. Hong H, Yang L, Stallcup MR. Hormone-independent transcriptional activation and coactivator binding by novel orphan nuclear receptor ERR3. J Biol Chem (1999) 274:22618–26.[Abstract/Free Full Text]
  9. Coward P, Lee D, Hull MV, Lehmann JM. 4-hydrotamoxifen bind to and deactivates the estrogen-related receptor gamma. Proc Natl Acad Sci USA (2001) 98:8880–4.[Abstract/Free Full Text]
  10. Giguere V, Yang N, Segui P, Evans RM. Identification of a new class of steroid hormone receptors. Nature (1988) 331:91–4.[CrossRef][Web of Science][Medline]
  11. Giguere V. To ERR in the estrogen pathway. Trends Endocrinol Metab (2002) 13:220–5.[CrossRef][Web of Science][Medline]
  12. Shigeta H, Zuo W, Yang N, DiAugustine R, Teng CT. The mouse estrogen receptor-related orphan receptor alpha 1: molecular cloning and estrogen responsiveness. J Mol Endocrinol (1997) 19:299–309.[Abstract/Free Full Text]
  13. Bonnelye E, Kung V, Laplace C, Galson DL, Aubin JE. Estrogen receptor-related receptor alpha impinges on the estrogen axis in bone: potential function in osteoporosis. Endocrinology (2002) 143:3658–70.[Abstract/Free Full Text]
  14. Liu D, Zhang Z, Gladwell W, Teng CT. Estrogen stimulates estrogen-related receptor alpha gene expression through conserved hormone response elements. Endocrinology (2003) 144:4894–904.[CrossRef][Web of Science][Medline]
  15. Vanacker JM, Pettersson K, Gustafsson JA, Laudet V. Transcriptional targets shared by estrogen receptor- related receptors (ERRs) and estrogen receptor (ER) alpha, but not by ERbeta. EMBO J (1999) 18:4270–9.[CrossRef][Web of Science][Medline]
  16. Johnston SD, Liu X, Zuo F, et al. Estrogen-related receptor alpha 1 functionally binds as a monomer to extended half-site sequences including ones contained within estrogen-response elements. Mol Endocrinol (1997) 11:342–52.[Abstract/Free Full Text]
  17. Dufour CR, Wilson BJ, Huss JM, et al. Genome-wide orchestration of cardiac functions by the orphan nuclear receptors ERRalpha and gamma. Cell Metab (2007) 5:345–56.[CrossRef][Medline]
  18. Luo J, Sladek R, Carrier J, Bader JA, Richard D, Giguere V. Reduced fat mass in mice lacking orphan nuclear receptor estrogen-related receptor alpha. Mol Cell Biol (2003) 23:7947–56.[Abstract/Free Full Text]
  19. Huss JM, Imahashi K, Dufour CR, et al. The nuclear receptor ERRalpha is required for the bioenergetic and functional adaptation to cardiac pressure overload. Cell Metab (2007) 6:25–37.[CrossRef][Web of Science][Medline]
  20. Bonnelye E, Aubin JE. Estrogen receptor-related receptor alpha: a mediator of estrogen response in bone. J Clin Endocrinol Metab (2005) 90:3115–21.[Abstract/Free Full Text]
  21. Bonnelye E, Zirngibl RA, Jurdic P, Aubin JE. The orphan nuclear estrogen receptor-related receptor-{alpha} regulates cartilage formation in vitro: implication of Sox9. Endocrinology (2007) 148:1195–205.[Abstract/Free Full Text]
  22. Bonnelye E, Merdad L, Kung V, Aubin JE. The orphan nuclear estrogen receptor-related receptor (ERR) is expressed throughout osteoblast differentiation and regulates bone formation in vitro. J Cell Biol (2001) 153:971–83.[Abstract/Free Full Text]
  23. Kamei Y, Lwin H, Saito K, et al. The 2.3 genotype of ESRRA23 of the ERR alpha gene is associated with a higher BMI than the 2.2 genotype. Obes Res (2005) 13:1843–4.[Web of Science][Medline]
  24. Laflamme N, Giroux S, Loredo-Osti JC, et al. A frequent regulatory variant of the estrogen-related receptor alpha gene associated with BMD in French-Canadian premenopausal women. J Bone Miner Res (2005) 20:938–44.[CrossRef][Web of Science][Medline]
  25. Sonoda J, Laganiere J, Mehl IR, et al. Nuclear receptor ERR alpha and coactivator PGC-1 beta are effectors of IFN-gamma-induced host defense. Genes Dev (2007) 21:1909–20.[Abstract/Free Full Text]
  26. Holmdahl R, Bockermann R, Backlund J, Yamada H. The molecular pathogenesis of collagen-induced arthritis in mice – a model for rheumatoid arthritis. Ageing Res Rev (2002) 1:135–47.[CrossRef][Web of Science][Medline]
  27. Wooley PH. Immunotherapy in collagen-induced arthritis: past, present, and future. Am J Med Sci (2004) 327:217–26.[CrossRef][Web of Science][Medline]
  28. Servet-Delprat C, Arnaud S, Jurdic P, et al. Flt3 + macrophage precursors commit sequentially to osteoclasts, dendritic cells and microglia. BMC Immunol (2002) 3:15.[CrossRef][Medline]
  29. Sena K, Morotome Y, Baba O, Terashima T, Takano Y, Ishikawa I. Gene expression of growth differentiation factors in the developing periodontium of rat molars. J Dent Res (2003) 82:166–71.[Abstract/Free Full Text]
  30. Gravallese EM. Bone destruction in arthritis. Ann Rheum Dis (2002) 61(Suppl. 2):ii84–6.[Abstract/Free Full Text]
  31. Scheijen B, Griffin JD. Tyrosine kinase oncogenes in normal hematopoiesis and hematological disease. Oncogene (2002) 21:3314–33.[CrossRef][Web of Science][Medline]
  32. Storm EE, Huynh TV, Copeland NG, Jenkins NA, Kingsley DM, Lee SJ. Limb alterations in brachypodism mice due to mutations in a new member of the TGF beta-superfamily. Nature (1994) 368:639–43.[CrossRef][Web of Science][Medline]
  33. Williams RO. Collagen-induced arthritis in mice. Methods Mol Med (2007) 136:191–9.[CrossRef][Medline]
  34. Gebhard PM, Gehrsitz A, Bau B, Soder S, Eger W, Aigner T. Quantification of expression levels of cellular differentiation markers does not support a general shift in the cellular phenotype of osteoarthritic chondrocytes. J Orthop Res (2003) 21:96–101.[CrossRef][Web of Science][Medline]
  35. Karsenty G, Wagner EF. Reaching a genetic and molecular understanding of skeletal development. Dev Cell (2002) 2:389–406.[CrossRef][Web of Science][Medline]
  36. Romas E, Bakharevski O, Hards DK, et al. Expression of osteoclast differentiation factor at sites of bone erosion in collagen-induced arthritis. Arthritis Rheum (2000) 43:821–6.[CrossRef][Web of Science][Medline]
  37. Suzuki M, Uetsuka K, Shinozuka J, et al. Changes in location and number of tartrate-resistant acid phosphatase (TRAP)-positive cells during the development of type II collagen-induced arthritis in DBA/1J mice. Exp Anim (1998) 47:211–4.[CrossRef][Web of Science][Medline]
  38. Kong YY, Yoshida H, Sarosi I, et al. OPGL is a key regulator of osteoclastogenesis, lymphocyte development and lymph-node organogenesis. Nature (1999) 397:315–23.[CrossRef][Web of Science][Medline]
  39. Lacey DL, Timms E, Tan HL, et al. Osteoprotegerin ligand is a cytokine that regulates osteoclast differentiation and activation. Cell (1998) 93:165–76.[CrossRef][Web of Science][Medline]
  40. Yasuda H, Shima N, Nakagawa N, et al. Osteoclast differentiation factor is a ligand for osteoprotegerin/osteoclastogenesis-inhibitory factor and is identical to TRANCE/RANKL. Proc Natl Acad Sci USA (1998) 95:3597–602.[Abstract/Free Full Text]
  41. Campbell IK, Hamilton JA, Wicks IP. Collagen-induced arthritis in C57BL/6 (H-2b) mice: new insights into an important disease model of rheumatoid arthritis. Eur J Immunol (2000) 30:1568–75.[CrossRef][Web of Science][Medline]
  42. Courtenay JS, Dallman MJ, Dayan AD, Martin A, Mosedale B. Immunisation against heterologous type II collagen induces arthritis in mice. Nature (1980) 283:666–8.[CrossRef][Web of Science][Medline]
  43. Nakashima T, Wada T, Penninger JM. RANKL and RANK as novel therapeutic targets for arthritis. Curr Opin Rheumatol (2003) 15:280–7.[CrossRef][Web of Science][Medline]
  44. Irmler IM, Gajda M, Brauer R. Exacerbation of antigen-induced arthritis in IFN-gamma-deficient mice as a result of unrestricted IL-17 response. J Immunol (2007) 179:6228–36.[Abstract/Free Full Text]
  45. Williams AS, Richards PJ, Thomas E, et al. Interferon-gamma protects against the development of structural damage in experimental arthritis by regulating polymorphonuclear neutrophil influx into diseased joints. Arthritis Rheum (2007) 56:2244–54.[CrossRef][Web of Science][Medline]
  46. Valcourt U, Gouttenoire J, Moustakas A, Herbage D, Mallein-Gerin F. Functions of transforming growth factor-beta family type I receptors and Smad proteins in the hypertrophic maturation and osteoblastic differentiation of chondrocytes. J Biol Chem (2002) 277:33545–58.[Abstract/Free Full Text]
Submitted 7 March 2008; revised version accepted 11 August 2008.
Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us    What's this?



This Article
Right arrow Abstract Freely available
Right arrow FREE Full Text (PDF) Freely available
Right arrow All Versions of this Article:
47/12/1785    most recent
ken368v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to My Personal Archive
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Disclaimer
Google Scholar
Right arrow Articles by Bonnelye, E.
Right arrow Articles by Aubin, J. E.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Bonnelye, E.
Right arrow Articles by Aubin, J. E.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us  
What's this?