Extracellular Matrix Proteoglycans Regulate Condylar Chondrocyte Activity

Objective:  Biglycan (Bgn) and fibromodulin (Fmod) are extracellular matrix (ECM) proteins highly expressed in temporomandibular joint (TMJ) condylar cartilage and may regulate the activity of growth factors like TGF-β1.  Mice deficient in both Bgn and Fmod (Bgn^-/0Fmod ^-/-) develop TMJ osteoarthritis (OA). Little is known about the role of the ECM micro-environment in condylar cartilage degradation.  The goal of this study was to utilize Bgn^-/0Fmod^-/- mice to elucidate the mechanistic function of the ECM and TGF-β1 in TMJ OA pathology.  Methods: Primary chondrocytes were isolated from WT and Bgn^-/0Fmod^-/-condyles.  TGF-β1 sequestration within the ECM was examined by TGF-β1 ELISA.  TGF-β1 signal transduction was investigated by Western blot analysis.  Chondrocytes were transfected with a TGF-β1 responsive luciferase reporter construct (pID-lux) in order to determine TGF-β1 transcriptional activity.  To assess aggrecan degradation, WT and Bgn^-/0Fmod^-/- condyles were isolated and cultured ex-vivo for 48 hours in 2ng/ml TGF- β1.  Explants were prepared for histology and aggrecan levels were examined by immunohistochemistry.  Aggrecan degradation was examined using an ELISA that measured aggrecan neoepitopes released from the explants in conditioned media.  Results: TGF-β1 ELISA revealed less TGF-β1 sequestered in the ECM of Bgn^-/0Fmod^-/- chondrocytes. Western blot showed increased pSmad2 levels in Bgn^-/0Fmod^-/- chondrocytes and luciferase assays showed increased reporter activity.  The addition of TGF-β1 to explants cultures decreased aggrecan immunostaining in Bgn^-/0Fmod^-/- explants and significantly increased the generation of aggrecan neoepitopes in conditioned media from Bgn^-/0Fmod^-/- explants. Conclusion: We identified Bgn and Fmod as critical components of the ECM, which control chondrocyte function by modulating TGF-β1 activity. Our studies showed the absence of Bgn and Fmod prevented proper TGF-β1 sequestration within the ECM, leading to increased TGF-β1 signal transduction and aggrecan degradation.  These findings demonstrate the ECM micro-environment is imperative for maintaining normal chondrocyte activities and reveal a novel cellular mechanism underlying TMJ OA pathology.