UBC Faculty Research and Publications

The retinoic acid binding protein CRABP2 is increased in murine models of degenerative joint disease Welch, Ian D; Cowan, Matthew F; Beier, Frank; Underhill, Tully M Jan 28, 2009

Your browser doesn't seem to have a PDF viewer, please download the PDF to view this item.

Item Metadata


52383-13075_2008_Article_2452.pdf [ 1.71MB ]
JSON: 52383-1.0223179.json
JSON-LD: 52383-1.0223179-ld.json
RDF/XML (Pretty): 52383-1.0223179-rdf.xml
RDF/JSON: 52383-1.0223179-rdf.json
Turtle: 52383-1.0223179-turtle.txt
N-Triples: 52383-1.0223179-rdf-ntriples.txt
Original Record: 52383-1.0223179-source.json
Full Text

Full Text

Available online http://arthritis-research.com/content/11/1/R14Open AccessVol 11 No 1Research articleThe retinoic acid binding protein CRABP2 is increased in murine models of degenerative joint diseaseIan D Welch1, Matthew F Cowan2, Frank Beier3 and Tully M Underhill21Department of Animal Care and Veterinary Services, University of Western Ontario, London, Ontario, N6A 5C1, Canada2Department of Cellular and Physiological Sciences, The University of British Columbia, Vancouver, British Columbia, V6T 1Z3, Canada3Department of Physiology and Pharmacology, CIHR Group in Skeletal Development and Remodeling, The University of Western Ontario, London, Ontario, N6A 5C1, CanadaCorresponding author: Tully M Underhill, tunderhi@brc.ubc.caReceived: 12 Sep 2008 Revisions requested: 12 Nov 2008 Revisions received: 4 Dec 2008 Accepted: 28 Jan 2009 Published: 28 Jan 2009Arthritis Research & Therapy 2009, 11:R14 (doi:10.1186/ar2604)This article is online at: http://arthritis-research.com/content/11/1/R14© 2009 Welch et al.; licensee BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.AbstractIntroduction Osteoarthritis (OA) is a debilitating disease withpoorly defined aetiology. Multiple signals are involved indirecting the formation of cartilage during development and thevitamin A derivatives, the retinoids, figure prominently inembryonic cartilage formation. In the present study, weexamined the expression of a retinoid-regulated gene in murinemodels of OA.Methods Mild and moderate forms of an OA-like degenerativedisease were created in the mouse stifle joint by meniscotibialtransection (MTX) and partial meniscectomy (PMX),respectively. Joint histopathology was scored using anOsteoarthritis Research Society International (OARSI) systemand gene expression (Col1a1, Col10a1, Sox9 and Crabp2) inindividual joints was determined using TaqMan quantitative PCRon RNA from microdissected articular knee cartilage.Results For MTX, there was a significant increase in the jointscore at 10 weeks (n = 4, p < 0.001) in comparison to shamsurgeries. PMX surgery was slightly more severe and producedsignificant changes in joint score at six (n = 4, p < 0.01), eight(n = 4, p < 0.001) and 10 (n = 4, p < 0.001) weeks. Theexpression of Col1a1 was increased in both surgical models attwo, four and six weeks post-surgery. In contrast, Col10a1 andSox9 for the most part showed no significant difference inexpression from two to six weeks post-surgery. Crabp2expression is induced upon activation of the retinoid signallingpathway. At two weeks after surgery in the MTX and PMXanimals, Crabp2 expression was increased about 18-fold andabout 10-fold over the sham control, respectively. By 10 weeks,Crabp2 expression was increased about three-fold (n = 7, notsignificant) in the MTX animals and about five-fold (n = 7, p <0.05) in the PMX animals in comparison to the contralateralcontrol joint.Conclusions Together, these findings suggest that the retinoidsignalling pathway is activated early in the osteoarthritic processand is sustained during the course of the disease.IntroductionOsteoarthritis (OA) is a degenerative joint disease (DJD) thatimpacts multiple joint tissues (i.e. subchondral bone, syn-ovium), but is typically associated with a deterioration of artic-ular cartilage. Although numerous factors have beensuggested to be important contributors to the developmentand progression of this disease, very few, with the possibleexception of FRZB or GDF5, have been confirmed to havecausal roles [1]. OA is considered in many instances to resultfrom years of wear and tear on the joint. In this scenario, aswith many other structures and organs within the body, thecartilage is considered to wear out as a result of ageing. There-fore, OA usually develops over a protracted period, which canbe accelerated in certain individuals because of an underlyinggenetic predisposition or various environmental factors.In the past few years, genetic links to OA have been estab-lished, and the first mutations in the collagen type II geneinvolved in the disease described [2,3]. More recently, othergenes associated with the WNT and GDF signalling pathwaysPage 1 of 9(page number not for citation purposes)ACAN: aggrecan; ACL: anterior cruciate ligament; ANOVA: analysis of variance; DMM: destabilisation of the medial meniscus; DJD: degenerative joint disease; ECM: extracellular matrix; MCL: medial collateral ligament; MTX: meniscotibial transection; OA: osteoarthritis; PMX: partial meniscoec-tomy; RT-qPCR: reverse transcription quantitative polymerase chain reaction.Arthritis Research & Therapy    Vol 11 No 1    Welch et al.have been implicated in OA susceptibility [4-6]. With regard toenvironmental factors, the biggest contributor is most likely tobe physical activity/trauma and underlying medical conditionsthat place a greater mechanical burden on articular cartilage.The net result of these various factors is the loss of the integrityof the cartilaginous extracellular matrix (ECM), leading to adecrease in mechanical strength. This increases the suscepti-bility of the articular cartilage to further damage, and becauseof its limited ability to repair itself the disease worsens. In OA,the structural integrity of the matrix is irreversibly lost, leadingto joint dysfunction [7].One class of molecules that is important in development andhomeostasis are the metabolites of vitamin A, the retinoids[8,9]. In the developing mammalian limb, retinoic acid has longbeen known to affect cells of mesenchymal and chondrogenicorigin [10-13]. The addition of retinoic acid to high-density cul-tures of limb bud mesenchymal cells (which form cartilagenodules in vitro) has been shown to decrease the number andsize of cartilage nodules formed. More interestingly, treatmentof mature chondrocytes with retinoic acid causes them toassume an immature phenotype [14-17]. This is accompaniedby a decrease in Col2a1 expression [14] and an increase inmetalloproteinase expression [18] that leads to degradation ofthe ECM. In this regard, retinoic acid treatment of cartilage iscommonly used to study cartilage degeneration [19,20]. Invivo, intra-articular injection of retinoic acid leads to chondro-cyte dedifferentiation and DJD [21]. More recently, antago-nists of the retinoic acid receptors have been tested in arheumatoid arthritis model in mice and rats and found toimprove histological scores, and this was associated withdecreased expression of Mmp13 [22].The changes in aggrecan metabolism seen in OA are similarto those produced by treatment of cartilage with retinoic acid.Bovine cartilage explant cultures treated with retinoic acidexhibit increased degradation of proteoglycans [23]. In ratosteosarcoma cells and primary bovine chondrocytes, treat-ment with retinoic acid produces cleavage of aggrecan(ACAN) at the E373-A374 peptide bond that is also cleavedin OA [24]. The retinoic acid-mediated degradation of ACANis inhibited by metalloprotease inhibitors, but not by inhibitorsof cathepsin B [23]. Others have shown that the addition ofretinoic acid to chondrocytes stimulates maturation and hyper-trophy consistent with the effects observed in vivo [25,26]. Aswitch from type II expression to type X and a decrease inACAN expression accompanied by an increase in catabolismof collagen type II and ACAN was observed. In this regard,retinoic acid has been shown to enhance chondrocyte hyper-trophy both in vitro and in vivo, where retinoic acid wasobserved to promote premature closure of the growth plate[27,28].To examine the status of the retinoic acid signalling pathway inexpression of a retinoic acid-regulated gene, including Crabp2in the articular cartilage. In a recent study, Crabp2 was foundto be elevated in DJD in a rat model of OA [29,30]. We foundCrapb2 to be significantly increased in early OA, indicatingthat the retinoic acid pathway may play a role in OA patho-physiology.Materials and methodsSurgerySurgeries were performed on 10-week-old male C57BL/6NCr1 mice (Charles River Laboratories, St. Constant, Que-bec, Canada). After a one-week period of acclimation afterarrival, mice were sorted into random groups using a lotterysystem. For each experimental time point a minimum of fivemice were evaluated. Mice were induced with 4% isofluraneand 1 L/minute oxygen. Once mice were anaesthetised theywere transferred to a mask and maintained on 2% isofluraneand 0.8 L/minute oxygen. The surgical area was shaved and athree-part preparation was applied containing hibitane soap(Ayerst, Montreal, Quebec, Canada), isopropyl alcohol andbetadine solution (Purdue Pharma, Pickering, Ontario, Can-ada).During surgery, the body temperature of the animals was main-tained by placing them on a warm circulating water pad.Standard sterile techniques were used throughout the sur-gery. A small sterile drape was fitted over each mouse toexpose only the area of interest, the medial side of the leftknee. The surgery began with a small skin incision startingfrom the distal femur and extending to the proximal tibia on themedial side of the knee. The subcutaneous tissue was dis-sected throughout the length of the skin incision. The deepfascia that connects the parapatellar fascia to the biceps fem-oris was separated exposing the medial side of the joint includ-ing the medial collateral ligament (MCL) and the joint capsule.In the partial meniscectomy (PMX) surgical paradigm both theMCL and the joint capsule were incised to reveal the medialmeniscus. The medial meniscus was gently held and retractedin a way that allowed for identification and transection of themeniscotibial ligament (Figure 1). The medial meniscus cansubsequently be observed to be 'free' at its dorsal border. Thejoint capsule incision was continued medially progressing tothe caudomedial edge of the tibial plateau. Gentle traction onthe medial meniscus allowed the meniscus to be isolated andtransected (Figure 1).In the meniscotibial transection (MTX) surgical paradigm therewas no medial dissection of the joint capsule or transection ofthe MCL. In this regard, the MTX model is similar to the desta-bilisation of the medial meniscus (DMM) model recentlydescribed by Glasson and colleagues [31]. Once the proce-dure was completed the deep fascia was closed using aninterrupted suturing pattern with 5-0 vicryl (Ethicon Inc.,Markham, Ontario, Canada). The subcutaneous tissue andPage 2 of 9(page number not for citation purposes)OA, we have used two murine DJD models and quantified the skin were also closed together in a continuous subcuticularAvailable online http://arthritis-research.com/content/11/1/R14pattern using 5-0 vicryl. Any loose skin edges were apposedwith sterile surgical glue. On completion of the surgery theinhalation anaesthesia was turned off and to control pain andinfection, the mice were injected subcutaneously withbuprenorphine (Schering-Plough, Hertfordshire, UK) and amp-icillin (Novopharm, Toronto, Ontario, Canada) in normal physi-ological saline. The sham operation involved a similar incisionto the left knee without compromising the joint capsule. All ani-mal experiments were sanctioned by The University of West-ern Ontario's Animal Care Committee and conducted in fullcompliance with the Canadian Council on Animal Care.Joint histologyAt the time of processing, mice were euthanased by carbondioxide inhalation and both stifle joints were harvested andfixed in 4% paraformaldehyde. After a minimum of 48 hours fix-ation the joints were decalcified for 96 hours in 26% formicacid (TBD-2, Thermo Inc., Pittsburgh, PA, USA). Once decal-cified the knees were paraffin embedded and serially sec-tioned in a sagital plane starting on the medial edge of thejoint. Slides were stained with safranin-O and the medial sideker and colleagues from a minimum of three slides [32]. Thesection with the highest score was recorded. Briefly, this scor-ing system, ranging from 1 to 24, involves the product of thehorizontal extent of the OA by the vertical severity of anylesions present.RNA isolation and quantitative PCRAt predetermined endpoints the mice were euthanased andthe knees were carefully dissected to expose the cartilage sur-face of both tibial plateau and the femoral chondyles. Under anoperating microscope, using a pair of micro-rongeurs, thearticular cartilage was gently scraped away from the underly-ing subchondral bone and transferred into Qiazol (Qiagen,Mississauga, Ontario, Canada). The cartilage was subse-quently homogenised in a microcentrifuge tube using a dis-posable plastic pestle and stored at -80°C. RNA was isolatedfrom the samples according to the manufacturer's guidelinesand for real-time quantitative PCR (RT-qPCR) the RNA wasreverse-transcribed using the High Capacity cDNA ReverseTranscription Kit (Applied Biosystems, Foster City, CA, USA).Gene expression was quantitated using qPCR on an ABI7500 Fast system with either custom TaqMan MGB probe orprimer sets (Col1a1, Sox9) or off the shelf TaqMan GeneExpression Assays (Applied Biosystems, Foster City, CA,USA). Relative expression was determined using the relativequantitation method with a standard curve and gene expres-sion was normalised to 18S abundance.Statistical analysesFor multiple comparisons, significance was determined usinganalysis of variance (ANOVA) with Bonferroni's or Tukey'spost-hoc tests as indicated. With the exception of the analysisof Crabp2 in the 10-week-old mice, all tests in the geneexpression studies were made between the operated left kneeand the contralateral unoperated right knee and a sham. Forcomparison between operated and the contralateral knee inthe 10-week-old mice significance was determined using two-tailed t-test. Significance is represented as follows: * p < 0.05;** p < 0.01; *** p < 0.001.Results and discussionIn an earlier study of a rat surgically induced model of DJD,Appleton and colleagues reported that Crabp2 expression asdetermined from microarray analysis was elevated in the OAjoint four weeks post-surgery [30]. Crabp2 is regulated byretinoic acid and is often induced upon activation of this path-way [33]. We were interested in confirming these findings anddetermining the kinetics of Crabp2 induction during OA. Fur-ther, we desired to develop and validate murine cartilage-spar-ing models of OA to facilitate the use of genetically modifiedlines and enable molecular analysis of gene expression in thearticular cartilage. For these purposes, surgeries were tailoredto produce less joint instability with the intent of producing amore slowly progressing disease than the standard anteriorFigure 1Overview of the surgical procedures used to generate meniscotibial transection (MTX) and partial meniscoectomy (PMX) modeltransection (MTX) and partial meniscoectomy (PMX) models. (a) Sche-matic representation of the joint (Adapted from Kamekura and col-leagues [34]). (b) An incision through the skin and subcutaneous tissue exposes the stifle joint. Prominent features include the medial collateral ligament (MCL), the patellar ligament and the tibial tuberosity. (c) The joint capsule has been cut on the medial side of the joint and the patella has been reflected laterally. This exposes the internal struc-tures of the joint. Prominent features include the medial femoral chondyle (MFC), the medial meniscus (MM), the meniscotibial ligament (MTL) and the anterior cruciate ligament (ACL). (d) The MTL has been cut to release the MM. (e) The partial meniscectomy has been com-pleted and the cut edge of the MM can be seen. Note that in order to visualise the cut edge of the meniscus, for pictorial purposes, the ante-rior cruciate ligament (ACL) has been transected. LM, lateral meniscus.Page 3 of 9(page number not for citation purposes)of tibial plateaus were subsequently scored according to Pritz-Arthritis Research & Therapy    Vol 11 No 1    Welch et al.cruciate ligament (ACL) models as has been recentlydescribed for the DMM and other models [31,34].MTX and PMX surgeries were performed and joint histologywas scored at various times up to 10 weeks post surgery. Inthe MTX group there was a significant increase in the jointscore by 10 weeks (n = 7, p < 0.001), although increases attwo and six weeks were not significant (Figure 2). In therecently reported DMM model, significant joint deteriorationwas observed at four weeks, and the differences between thismodel and the related MTX model may be a consequence ofthe different mouse strains used (129/SvEv versus theC57BL/6NCr1 strain utilised herein) and/or the different scor-ing methodologies employed [31]. Consistent with the moreaggressive nature of the surgery, the PMX mice displayed asignificant increase in joint score by six weeks after surgery (n= 7, p < 0.01); at 10 weeks these animals had a joint score of8.4 (n = 5) out of a possible total of 24 (Figure 2). The earlyhistopathological findings are focal, superficial to deep fibrilla-tion sometimes associated with variable degrees of matrixdepletion in these early stages. In the more aggressive PMXsurgical paradigm the lesions become more diffuse and aremore likely to have vertical fissures through the midzone withless common delamination of the superficial layer (Figure 3).Together, these results demonstrate that the PMX and MTXsurgeries give rise to moderate and mild forms of DJD that areassociated with a slowly progressing joint disease.To quantify gene expression in single knee joints we devel-oped efficient RNA isolation methods that yielded about 100to 150 ng of total RNA from individual stifle joints sufficient foranalysis of five to seven genes (20 ng of RNA per gene). Pre-vious studies have shown that Col1a1 and Col10a1 are bothelevated in OA [35-38]. We also examined the expression ofSox9, a transcription factor important in chondrocyte differen-tiation and matrix production [39]. With the exception of thefour-week time point, the expression of Col1a1 was found tobe elevated more than three-fold at all times in the MTX/PMXjoint in comparison to the sham control (Figure 4). In contrast,there were only slight changes in the expression of Col10a1or Sox9 during the first six weeks after surgery (Figure 4), witha significant (n = 5, p < 0.05) two-fold increase in Col10a1being observed in six-week PMX samples in comparison to thesham control (Figure 4). Interestingly, examination of Crabp2expression revealed a large increase in expression at twoweeks after surgery in both MTX and PMX of about 18-foldand 10-fold, respectively (n = 5; MTX, p < 0.05; PMX, p <0.01; Figure 5a). This magnitude of induction declined overtime; however, by six weeks in the PMX mice there was abouta three-fold increase in Crabp2 expression in PMX knees incomparison to the sham or contralateral knee (n = 5, p < 0.05;Figure 5a), whereas at 10 weeks, there was still about a five-fold increase in Crabp2 expression in PMX knees in compari-son to the contralateral control (n = 7, p < 0.05; Figure 5b).Together these results show that PMX and MTX surgeries leadto a slowly progressing arthrosis in mice and that Crabp2 rep-resents an early and sustained marker of DJD.Crabp2 expression is regulated by retinoic acid and itsincrease in expression is consistent with activation of this path-way. CRABP2 has been shown to function both to suppressretinoic acid receptor activity by sequestering ligands and alsoas a vehicle to deliver ligands to the retinoic acid receptors,thereby enhancing ligand-mediated retinoic acid receptor tran-scriptional activity [40-45]. More recent reports favour the lat-ter function, indicating that increased expression of Crabp2Figure 2Meniscotibial transection (MTX) and partial meniscoectomy (PMX) surgeries lead to degenerative changes in the joint as evaluated by joint scoringj i  i . Joints were scored based on the Osteoarthritis Research Society International (OARSI) system. The PMX surgery was associated with a higher his-Page 4 of 9(page number not for citation purposes)tological score than that of the MTX surgery. The sham surgeries at week 10 had a 0 score. ** p < 0.01; *** p < 0.001.Available online http://arthritis-research.com/content/11/1/R14enhances retinoic acid receptor transcriptional activity[41,42,44]. Further, Crabp2 null animals present with minorlimb defects including an extra post-axial digit that (based onmore recent reports) would suggest decreased retinoid sig-nalling in the absence of CRABP2 [9,46,47]. In aggregate,increased expression of Crabp2 is generally linked to activa-tion of the retinoid pathway either directly, because expressionof Crabp2 is regulated by retinoic acid, or indirectly, becauseincreased CRABP2 would increase retinoic acid receptortranscriptional activity. Inappropriate activation of the retinoicacid signalling pathway is expected to enhance cartilage deg-radation and/or chondrocyte dedifferentiation, both of whichare observed in the osteoarthritic process.As mentioned above, retinoic acid has been commonly usedto promote degeneration in cartilage explants, and this hasbeen associated with increased activity of various aggreca-nases. Knockout animals of the gene encoding the aggreca-nase ADAMTS5 are protected to a great extent from OA injoint instability models, indicating that this enzyme may play amajor role in cartilage catabolism, at least in mouse models ofOA [48-50]. Interestingly, in explants derived from doublemutants of Adamts5 and another major aggrecanaseAdamts4, addition of retinoic acid was still found to promoterelease of aggrecan through cleavage in the CS-2 domain[51]. Retinoic acid has been shown to increase Adamts5 andson and colleagues [51] suggest that retinoic acid may also bepromoting cartilage degradation through additional and as yetundefined aggrecanase(s). Further, in collagen-induced arthri-tis in the mouse and streptococcal cell wall-induced arthritis inrats, small molecule antagonists of the retinoic acid receptorswere found to ameliorate pain and decrease cartilage loss[22].In addition to Crabp2, other components of the retinoid signal-ling pathway were found to be significantly elevated in thejoints of the aforementioned rat OA joint instability model,including genes encoding proteins involved in retinoic acidsynthesis (Aldh1a3) and retinol transport and delivery (Lratand retinol dehydrogenase) and a putative retinoic acid targetgene (Stra3) [30]. Together, these findings along with ourobservations of elevated Crabp2 expression in mouse modelsof DJD suggest that retinoic acid may play a fundamental andperhaps unappreciated role in the osteoarthritic process.As Crabp2 is robustly expressed in early OA, it may representa marker for detection of early OA. Further, polymorphisms inFRZB and GDF5 have been linked to OA, and similar to theretinoid signalling pathway, they all play a role in endochondralossification and modulation of their activity may impact main-tenance of the articular chondrocyte [1]. In this regard, as hasbeen previously suggested, antagonists of the retinoic acidFigure 3Meniscotibial transection (MTX) and partial meniscoectomy (PMX) surgeries lead to degenerative joint disease in mouse knees. Histological sec-tions were collected at the indicated time points and stained with safranin O. Note in both the six-week and 10-week PMX and 10-week MTX sur-gery groups the loss of proteoglycan staining (arrowhead) in the superficial layers and in the 10-week PMX there is delamination of the superficial layer (arrow). WT, wild-type.Page 5 of 9(page number not for citation purposes)Mmp13 expression [19,22], and these new findings by Roger-Arthritis Research & Therapy    Vol 11 No 1    Welch et al.signalling pathway may prove useful for maintaining thechondrocyte phenotype [8].ConclusionThe joint instability models presented herein in contrast tomore aggressive models involving for instance ACL transec-tion, present with OA-like pathology but still appreciable artic-ular cartilage 10 weeks after surgery, thereby enabling the useof molecular approaches to quantify gene expression changesin early OA [34].The expression of the retinoic acid-regulated gene Crabp2 issignificantly elevated in early DJD, and may be a useful markerto follow early changes in cartilage in response to joint insta-bility or in OA. Manipulation of the retinoic acid signalling path-way may prove useful in modifying the clinical course of OA.Figure 4Analysis of Col1a1, Col10a1 and Sox9 expression in knee cartilage from meniscotibial transection (MTX), partial meniscoectomy (PMX) and sham surgeriessurgeries. Genes analysed are shown on the left and significance was determined by analysis of variance (ANOVA) with Tukey's post-hoc tests for multiple comparisons. In comparison to Sox9 and Col10a1, Col1a1 is significantly increased at multiple time points in the different surgeries. Gene expression was normalised to an 18S internal control and the normalised expression (arbitrary units) for each gene is shown for the contralateral unoperated right knee, a sham control (left) knee (n = 5) and the operated left knee. Significance to the contralateral and sham controls is indicated on the top and bottom, respectively. * p < 0.05; ** p < 0.01; *** p < 0.001; ns = not significant.Page 6 of 9(page number not for citation purposes)Available online http://arthritis-research.com/content/11/1/R14Page 7 of 9(page number not for citation purposes)Figure 5Crabp2 expression is increased in joint cartilage from meniscotibial transection (MTX) and partial meniscoectomy (PMX) operated knees. (a) Crabp2 expression was analysed at two, four and six weeks post-surgery and its expression was significantly changed in MTX/PMX-operated knees (n = 5 to 7) in comparison to either the contralateral knee or a sham control. Significance was determined by analysis of variance (ANOVA) with Tukey's post-hoc tests for multiple comparisons. Gene expression was normalised to an 18S internal control and the normalised expression for each gene is shown for the contralateral unoperated right knee, a sham control (left) knee and the operated left knee. Significance to the contralateral and sham controls is indicated on the top and bottom, respectively. (b) The expression of Crabp2 is still elevated 10 weeks post-surgery. Significance in gene expression at 10 weeks post-surgery was determined by two-tailed t-test analysis of the left (L, operated) versus right (R, contralateral control) knee. No sham was included in the 10-week group. * p < 0.05; ** p < 0.01; ns = not significant.Arthritis Research & Therapy    Vol 11 No 1    Welch et al.Competing interestsThe authors declare that they have no competing interests.Authors' contributionsIW performed experiments, contributed to experimentaldesign, writing of the manuscript and data interpretation. MCcontributed to experimental design and carried out the experi-ments. FM and TMU were involved in experimental design,data interpretation and writing of the manuscript.AcknowledgementsThe authors would like to thank Tracy Hill for technical assistance. This grant was supported by Canadian Arthritis Networks grants to FB and TMU. FB holds a Canada Research Chair and TMU is an Investigator of the Arthritis Society.References1. Bos SD, Slagboom PE, Meulenbelt I: New insights into osteoar-thritis: early developmental features of an ageing-related dis-ease.  Curr Opin Rheumatol 2008, 20:553-559.2. Williams CJ, Jimenez SA: Heritable diseases of cartilage causedby mutations in collagen genes.  J Rheumatol Suppl 1995,43:28-33.3. Sahlman J, Pitkanen MT, Prockop DJ, Arita M, Li SW, Helminen HJ,Langsjo TK, Puustjarvi K, Lammi MJ: A human COL2A1 gene withan Arg519Cys mutation causes osteochondrodysplasia intransgenic mice.  Arthritis Rheum 2004, 50:3153-3160.4. Ikegawa S: New gene associations in osteoarthritis: what dothey provide, and where are we going?  Curr Opin Rheumatol2007, 19:429-434.5. Rodriguez-Lopez J, Pombo-Suarez M, Liz M, Gomez-Reino JJ,Gonzalez A: Further evidence of the role of frizzled-related pro-tein gene polymorphisms in osteoarthritis.  Ann Rheum Dis2007, 66:1052-1055.6. Loughlin J: Polymorphism in signal transduction is a majorroute through which osteoarthritis susceptibility is acting.Curr Opin Rheumatol 2005, 17:629-633.7. Aigner T, McKenna L: Molecular pathology and pathobiology ofosteoarthritic cartilage.  Cell Mol Life Sci 2002, 59:5-18.8. Hoffman LM, Weston AD, Underhill TM: Molecular mechanismsregulating chondroblast differentiation.  J Bone Joint Surg Am2003, 85-A(Suppl 2):124-132.9. Weston AD, Hoffman LM, Underhill TM: Revisiting the role ofretinoid signaling in skeletal development.  Birth Defects Res CEmbryo Today 2003, 69:156-173.10. Amos B, Lotan R: Retinoid-sensitive cells and cell lines.  Meth-ods Enzymol 1990, 190:217-225.11. Lewis CA, Pratt RM, Pennypacker JP, Hassell JR: Inhibition oflimb chondrogenesis in vitro by vitamin A.  Dev Biol 1978,64:31-47.12. Solursh M, Meier S: The selective inhibition of mucopolysac-charide synthesis by vitamin A treatment of cultured chickembryo chondrocytes.  Calcif Tissue Res 1973, 13:131-142.13. Zimmerman B, Tsambos D: Evaluation of the sensitive step ofinhibition of chondrogenesis by retinoids in limb mesenchy-mal cells in vitro.  Cell Differentiation 1985, 17:95-103.14. Benya PD, Padilla SR: Modulation of the rabbit chondrocytephenotype by retinoic acid terminates type II collagen synthe-sis without inducing type I collagen: the modulated phenotypediffers from that produced by subculture.  Dev Biol 1986,118:296-305.15. Takishita Y, Hiraiwa K, Nagayama M: Effect of retinoic acid onproliferation and differentiation of cultured chondrocytes interminal differentiation.  J Biochem 1990, 107:592-596.16. Lee KK, Li FC, Yung WT, Kung JLS, Ng JN, Cheah KS: Influenceof digits, ectoderm, and retinoic acid on chondrogenesis bymouse interdigital mesoderm in culture.  Dev Dyn 1994,201:297-309.18. Ballock RT, Heydemann A, Wakefield LM, Flanders KC, RobertsAB, Sporn MB: Inhibition of the chondrocyte phenotype byretinoic acid involves upregulation of metalloproteinase genesindependent of TGF-b.  J Cell Physiol 1994, 159:340-346.19. East CJ, Stanton H, Golub SB, Rogerson FM, Fosang AJ:ADAMTS-5 deficiency does not block aggrecanolysis at pre-ferred cleavage sites in the chondroitin sulfate-rich region ofaggrecan.  J Biol Chem 2007, 282:8632-8640.20. Glasson SS, Askew R, Sheppard B, Carito BA, Blanchet T, Ma HL,Flannery CR, Kanki K, Wang E, Peluso D, Yang Z, Majumdar MK,Morris EA: Characterization of and osteoarthritis susceptibilityin ADAMTS-4-knockout mice.  Arthritis Rheum 2004,50:2547-2558.21. Lapadula G, Nico B, Cantatore FP, La Canna R, Roncali L, PipitoneV: Early ultrastructural changes of articular cartilage and syn-ovial membrane in experimental vitamin A-induced osteoar-thritis.  J Rheumatol 1995, 22:1913-1921.22. Beehler BC, Hei YJ, Chen S, Lupisella JA, Ostrowski J, Starrett JE,Tortolani D, Tramposch KM, Reczek PR: Inhibition of diseaseprogression by a novel retinoid antagonist in animal models ofarthritis.  J Rheumatol 2003, 30:355-363.23. Buttle DJ, Handley CJ, Ilic MZ, Saklatvala J, Murata M, Barrett AJ:Inhibition of cartilage proteoglycan release by a specific inac-tivator of cathepsin B and an inhibitor of matrix metalloprotei-nases. Evidence for two converging pathways of chondrocyte-mediated proteoglycan degradation.  Arthritis Rheum 1993,36:1709-1717.24. Lark MW, Gordy JT, Weidner JR, Ayala J, Kimura JH, Williams HR,Mumford RA, Flannery CR, Carlson SS, Iwata M, Sandy JD: Cell-mediated catabolism of aggrecan. Evidence that cleavage atthe "aggrecanase" site (Glu373-Ala374) is a primary event inproteolysis of the interglobular domain.  J Biol Chem 1995,270:2550-2556.25. Iwamoto M, Golden EB, Adams SL, Noji S, Pacifici M: Respon-siveness to retinoic acid changes during chondrocyte matura-tion.  Exp Cell Res 1993, 205:213-224.26. Iwamoto M, Shapiro IM, Yagami K, Boskey AL, Leboy PS, AdamsSL, Pacifici M: Retinoic acid induces rapid mineralization andexpression of mineralization-related genes in chondrocytes.Exp Cell Res 1993, 207:413-420.27. De Luca F, Uyeda JA, Mericq V, Mancilla EE, Yanovski JA, BarnesKM, Zile MH, Baron J: Retinoic acid is a potent regulator ofgrowth plate chondrogenesis.  Endocrinology 2000,141:346-353.28. Koyama E, Golden EB, Kirsch T, Adams SL, Chandraratna RA,Michaille JJ, Pacifici M: Retinoid signaling is required forchondrocyte maturation and endochondral bone formationduring limb skeletogenesis.  Dev Biol 1999, 208:375-391.29. Appleton CT, McErlain DD, Henry JL, Holdsworth DW, Beier F:Molecular and histological analysis of a new rat model ofexperimental knee osteoarthritis.  Ann N Y Acad Sci 2007,1117:165-174.30. Appleton CT, Pitelka V, Henry J, Beier F: Global analyses of geneexpression in early experimental osteoarthritis.  ArthritisRheum 2007, 56:1854-1868.31. Glasson SS, Blanchet TJ, Morris EA: The surgical destabilizationof the medial meniscus (DMM) model of osteoarthritis in the129/SvEv mouse.  Osteoarthr Cartil 2007, 15:1061-1069.32. Pritzker KP, Gay S, Jimenez SA, Ostergaard K, Pelletier JP, RevellPA, Salter D, Berg WB van den: Osteoarthritis cartilage histopa-thology: grading and staging.  Osteoarthr Cartil 2006, 14:13-29.33. Durand B, Saunders M, Leroy P, Leid M, Chambon P: All-transand 9-cis retinoic acid induction of CRABPII transcription ismediated by RAR-RXR heterodimers bound to DR1 and DR2repeated motifs.  Cell 1992, 71:73-85.34. Kamekura S, Hoshi K, Shimoaka T, Chung U, Chikuda H, YamadaT, Uchida M, Ogata N, Seichi A, Nakamura K, Kawaguchi H: Oste-oarthritis development in novel experimental mouse modelsinduced by knee joint instability.  Osteoarthr Cartil 2005,13:632-641.35. Aigner T, Reichenberger E, Bertling W, Kirsch T, Stoss H, Mark Kvon der: Type × collagen expression in osteoarthritic and rheu-matoid articular cartilage.  Virchows Arch B Cell Pathol Incl MolPathol 1993, 63:205-211.Page 8 of 9(page number not for citation purposes)17. Horton WE, Yamada Y, Hassell JR: Retinoic acid rapidly reducescartilage matrix synthesis by altering gene transcription inchondrocytes.  Dev Biol 1987, 123:508-516.36. Girkontaite I, Frischholz S, Lammi P, Wagner K, Swoboda B,Aigner T, Mark K Von der: Immunolocalization of type × colla-Available online http://arthritis-research.com/content/11/1/R14gen in normal fetal and adult osteoarthritic cartilage with mon-oclonal antibodies.  Matrix Biol 1996, 15:231-238.37. Nerlich AG, Wiest I, Mark K von der: Immunohistochemical anal-ysis of interstitial collagens in cartilage of different stages ofosteoarthrosis.  Virchows Arch B Cell Pathol Incl Mol Pathol1993, 63:249-255.38. Mark K von der, Frischholz S, Aigner T, Beier F, Belke J, ErdmannS, Burkhardt H: Upregulation of type × collagen expression inosteoarthritic cartilage.  Acta Orthop Scand Suppl 1995,266:125-129.39. Lefebvre V, Smits P: Transcriptional control of chondrocyte fateand differentiation.  Birth Defects Res C Embryo Today 2005,75:200-212.40. Boylan JF, Gudas LJ: Overexpression of the cellular retinoicacid binding protein-I (CRABP-I) results in a reduction in dif-ferentiation-specific gene expression in F9 teratocarcinomacells.  J Cell Biol 1991, 112:965-979.41. Schug TT, Berry DC, Shaw NS, Travis SN, Noy N: Opposingeffects of retinoic acid on cell growth result from alternate acti-vation of two different nuclear receptors.  Cell 2007,129:723-733.42. Sessler RJ, Noy N: A ligand-activated nuclear localization signalin cellular retinoic acid binding protein-II.  Mol Cell 2005,18:343-353.43. Wu Z, Yang Y, Shaw N, Bhattacharya S, Yan L, West K, Roth K,Noy N, Qin J, Crabb JW: Mapping the ligand binding pocket inthe cellular retinaldehyde binding protein.  J Biol Chem 2003,278:12390-12396.44. Budhu AS, Noy N: Direct channeling of retinoic acid betweencellular retinoic acid-binding protein II and retinoic acid recep-tor sensitizes mammary carcinoma cells to retinoic acid-induced growth arrest.  Mol Cell Biol 2002, 22:2632-2641.45. Noy N: Retinoid-binding proteins: mediators of retinoid action.Biochem J 2000, 348:481-495.46. Fawcett D, Pasceri P, Fraser R, Colbert M, Rossant J, Giguere V:Postaxial polydactyly in forelimbs of CRABP-II mutant mice.Development 1995, 121:671-679.47. Hoffman LM, Garcha K, Karamboulas K, Cowan MF, Drysdale LM,Horton WA, Underhill TM: BMP action in skeletogenesisinvolves attenuation of retinoid signaling.  J Cell Biol 2006,174:101-113.48. Glasson SS, Askew R, Sheppard B, Carito B, Blanchet T, Ma HL,Flannery CR, Peluso D, Kanki K, Yang Z, Majumdar MK, Morris EA:Deletion of active ADAMTS5 prevents cartilage degradation ina murine model of osteoarthritis.  Nature 2005, 434:644-648.49. Majumdar MK, Askew R, Schelling S, Stedman N, Blanchet T,Hopkins B, Morris EA, Glasson SS: Double-knockout ofADAMTS-4 and ADAMTS-5 in mice results in physiologicallynormal animals and prevents the progression of osteoarthritis.Arthritis Rheum 2007, 56:3670-3674.50. Stanton H, Rogerson FM, East CJ, Golub SB, Lawlor KE, MeekerCT, Little CB, Last K, Farmer PJ, Campbell IK, Fourie AM, FosangAJ: ADAMTS5 is the major aggrecanase in mouse cartilage invivo and in vitro.  Nature 2005, 434:648-652.51. Rogerson FM, Stanton H, East CJ, Golub SB, Tutolo L, Farmer PJ,Fosang AJ: Evidence of a novel aggrecan-degrading activity incartilage: Studies of mice deficient in both ADAMTS-4 andADAMTS-5.  Arthritis Rheum 2008, 58:1664-1673.Page 9 of 9(page number not for citation purposes)


Citation Scheme:


Citations by CSL (citeproc-js)

Usage Statistics



Customize your widget with the following options, then copy and paste the code below into the HTML of your page to embed this item in your website.
                            <div id="ubcOpenCollectionsWidgetDisplay">
                            <script id="ubcOpenCollectionsWidget"
                            async >
IIIF logo Our image viewer uses the IIIF 2.0 standard. To load this item in other compatible viewers, use this url:


Related Items