L Enochson1, J Stenberg2, M Brittberg3, A Lindahl4. 1. Department of Clinical Chemistry and Transfusion Medicine, The Sahlgrenska Academy at Gothenburg University, Gothenburg, Sweden. Electronic address: lars.enochson@gu.se. 2. Department of Clinical Chemistry and Transfusion Medicine, The Sahlgrenska Academy at Gothenburg University, Gothenburg, Sweden. Electronic address: johan.stenberg@gu.se. 3. Cartilage Research Unit, Gothenburg University, Department of Orthopaedics, Kungsbacka Hospital, Kungsbacka, Sweden. Electronic address: mats.brittberg@regionhalland.se. 4. Department of Clinical Chemistry and Transfusion Medicine, The Sahlgrenska Academy at Gothenburg University, Gothenburg, Sweden. Electronic address: anders.lindahl@clinchem.gu.se.
Abstract
OBJECTIVE: Growth differentiation factor 5 (GDF5) is important for joint formation and associated with osteoarthritis (OA). Its role for the homeostasis of cartilage extracellular matrix (ECM) is, however, unknown. The canonical Wnt signaling pathway is also implemented in OA and activation of the pathway has detrimental effects on the cartilage ECM. The objective of this study was to investigate the effect of GDF5 stimulation on the Wnt signaling pathway and on the expression of known modulators of cartilage ECM. DESIGN: Human chondrocytes were cultured in the pellet mass system and stimulated with increasing concentrations of GDF5. Expression of matrix modulating enzymes and canonical Wnt inhibitors dickkopf 1 (DKK1) and frizzled related protein (FRZB) were measured with quantitative PCR (qPCR). Protein levels of matrix metalloprotease 13 (MMP13), DKK1 and β-catenin were measured with enzyme-linked immunosorbent assay (ELISA). Canonical Wnt signaling was stimulated with Wnt3a and small molecule CHIR-99021 and DKK1 was blocked with small molecule WAY-262611. RESULTS: In this study, we show that GDF5 stimulation of human chondrocytes inhibits expression of the cartilage ECM degrading enzymes MMP13 and ADAMTS4 and stimulates the expression of cartilage anabolic genes ACAN and SOX9. We further show that the stimulation inhibits the canonical Wnt signaling pathway through expression of the canonical Wnt inhibitors DKK1 and FRZB. Finally we show that inhibition of MMP13 expression through GDF5 stimulation is mediated by DKK1. CONCLUSION: Herein, we provide evidence of a previously unknown link between GDF5 signaling and canonical Wnt signaling that may contribute to the understanding of the molecular mechanisms of OA.
OBJECTIVE:Growth differentiation factor 5 (GDF5) is important for joint formation and associated with osteoarthritis (OA). Its role for the homeostasis of cartilage extracellular matrix (ECM) is, however, unknown. The canonical Wnt signaling pathway is also implemented in OA and activation of the pathway has detrimental effects on the cartilage ECM. The objective of this study was to investigate the effect of GDF5 stimulation on the Wnt signaling pathway and on the expression of known modulators of cartilage ECM. DESIGN:Human chondrocytes were cultured in the pellet mass system and stimulated with increasing concentrations of GDF5. Expression of matrix modulating enzymes and canonical Wnt inhibitors dickkopf 1 (DKK1) and frizzled related protein (FRZB) were measured with quantitative PCR (qPCR). Protein levels of matrix metalloprotease 13 (MMP13), DKK1 and β-catenin were measured with enzyme-linked immunosorbent assay (ELISA). Canonical Wnt signaling was stimulated with Wnt3a and small molecule CHIR-99021 and DKK1 was blocked with small molecule WAY-262611. RESULTS: In this study, we show that GDF5 stimulation of human chondrocytes inhibits expression of the cartilage ECM degrading enzymes MMP13 and ADAMTS4 and stimulates the expression of cartilage anabolic genes ACAN and SOX9. We further show that the stimulation inhibits the canonical Wnt signaling pathway through expression of the canonical Wnt inhibitors DKK1 and FRZB. Finally we show that inhibition of MMP13 expression through GDF5 stimulation is mediated by DKK1. CONCLUSION: Herein, we provide evidence of a previously unknown link between GDF5 signaling and canonical Wnt signaling that may contribute to the understanding of the molecular mechanisms of OA.
Authors: Ming-Song Lee; Matthew J Stebbins; Hongli Jiao; Hui-Ching Huang; Ellen M Leiferman; Brian E Walczak; Sean P Palecek; Eric V Shusta; Wan-Ju Li Journal: Sci Adv Date: 2021-05-19 Impact factor: 14.136