The Role And Mechanism Of FGFR3in The Occurrence And Development Of Osteoarthritis

Osteoarthritis (OA) is a common degenerative joint disease in the elderly, and causesolder people walking inconvenience. Osteoarthritis is a chronic progressive disease whichinvolving weight-bearing joints, limited clinical characteristics of joint pain, joint activities,joint deformity. Although more research focused on OA, but still lack of effective safetyprevention and treatment of osteoarthritis. So a better understanding of the molecularmechanism of osteoarthritis progression, looking for drugs which can slow the diseaseprocess in osteoarthritis, may allow for the development of new biologic therapies aimed atslowing the disease process.FGFR3(fibroblast growth factor3) which belongs to the FGFR family is tyrosinekinase receptors. In the23kinds of FGFs, FGF9and FGF18are relatively specific ligandsof FGFR3. In addition to FGFR3plays an important role in bone development, but it playsan important role in cartilage development and homeostasis in articular cartilage. FGFR3conventional KO mice appear the phenotype of osteoarthritis, increased expression ofMMP13in the articular cartilage, suggesting protective effects of FGFR3in articularcartilage, but the emerging of OA phenotype in FGFR3conventional KO mice does notexclude the mechanical impact of its skeletal malformations. FGF9is relatively specificligand of FGFR3, FGF9mutation causes Elbow knee synostosis (EKS) in mice, canMultiple synostosis syndrome (SYNS) in human, joint fusion is an important feature ofEKS and SYNS in mice or patients. These results suggest that FGFR3plays an importantrole in regulating osteoarthritis cartilage repair, but direct evidence of the specificmechanisms of FGFR3in cartilage repair is still lacking.To investigate the role and mechanism of FGFR3in the development of osteoarthritis,we use the inducible FGFR3cartilage-specific KO mice and build the OA model mice toexplore effects of FGFR3in the process of osteoarthritis and cartilage repair.Methods: 1. Create three mouse models, Fgfr3G369C/+is enhanced FGFR3mutation mouse(simulate human ACH disease, referred to as ACH mice), Col2a1-CreERT2;Fgfr3flox/floxisFGFR3cartilage-specific induced knockout mouse (referred to as Fgfr3cKO mice),Col2a1-CreERT2;Fgfr3+/K644Eneois FGFR3cartilage-specific inducedconstitutive activationmouse(referred to as the Fgfr3cActivation mouse). PCR assay was used to identify the genotypeof mice.2-month-old Fgfr3cKO mice and Fgfr3cActivation mice were injectedTamoxifenintraperitoneally;2. Establish two mouse models of arthritis, including aging-associated spontaneous OAmodel and DMM (Destablilization of the medial meniscus) surgery-induced OAmodel.DMM surgery was perfomed after Fgfr3cKO mice and Fgfr3cActivation have beeninjected Tamoxifen.3. ACH mice were sacrificed at12months and20months of age, Fgfr3cKO and Fgfr3cActivation mice were sacrificed at3months and4months of age. X-ray was used toobserveFgfr3cKO mice in length and joint general situation. Safranin-o/Fast green stainingwas used to observearticular cartilage situation and scorearticular cartilage matrixloss andinjuryreference to OARSI (Osteoarthrisits Research Society International) recommendedmethods. Immunohistochemical staining was used to observeexpression ofCollagen II,Collagen X,MMP13and FGFR1in articular cartilage.4. In vitro, Fgfr3cKO mouse primary articular chondrocytes treated with4-hydroxy-Tamoxifen, then real-time quantitative PCR was used to examine RNA expression foldchange of Mmp13, Collagen X, Collagen II, Adamts-5and Ihh.5. Overexpressing Fgfr3in ATDC5cells carried out by transfection experiments, thenreal-time quantitative PCR was used to detect expression change of Mmp13, Collagen X.Results:1. Abnormal joint cartilage hypertrophy in Fgfr3cKO mouse and articular cartilagedamage is more serious after DMM surgery.Increased expressionof MMP13, Collagen Xand decreased expression of Collagen II in articular cartilage of Fgfr3cKOmouse.Decreased expression of FGFR3and increased expression of FGFR1in articularcartilage of Fgfr3cKO mouse.RNA levels of Mmp13, Collagen X, Adatms-5and Ihh wasupregulated inFgfr3cKO mouse primary articularchondrocytes.2. In aging-associated spontaneous OA model, articular cartilage extracellular matrix degradation delayed in ACH mouse, and expressionof MMP13, Collagen X expressiondecreased in articular cartilage of ACH mouse.3. Articular cartilage damage process delayed after DMM surgery in Fgfr3cActivationmouse.Decreased expressing of MMP13and Collagen X and increased expressionofCollagen II in articular cartilage of Fgfr3cActivation mouse after DMM surgery.4. Overexpressing of Fgfr3causesdownregulation of Mmp13, Collagen X in ATDC5cells.Conclusions:1. FGFR3signaling deficiency leads to abnormal cell hypertrophy and increase OAphenotype after DMM surgery.2. Deficiency of FGFR3signalscan promote increasing of cartilage catabolism,probably because of lack of FGFR3signals due to activation of IHHsignals and FGFR1signals, leading to metabolic disorder of articular cartilage.3. FGFR3signals can slow down the process of articular cartilage injuryin OA.