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Role of sox9 in growth factor regulation of articular chondrocytes.

ABSTRACT: Chondrogenic polypeptide growth factors influence articular chondrocyte functions that are required for articular cartilage repair. Sox9 is a transcription factor that regulates chondrogenesis, but its role in the growth factor regulation of chondrocyte proliferation and matrix synthesis is poorly understood. We tested the hypotheses that selected chondrogenic growth factors regulate sox9 gene expression and protein production by adult articular chondrocytes and that sox9 modulates the actions of these growth factors. To test these hypotheses, we delivered insulin-like growth factor-I (IGF-I), fibroblast growth factor-2 (FGF-2), bone morphogenetic protein-2 (BMP-2) and/or bone morphogenetic protein-7 (BMP-7), or their respective transgenes to adult bovine articular chondrocytes, and measured changes in sox9 gene expression and protein production. We then knocked down sox9 gene expression with sox9 siRNA, and measured changes in the expression of the genes encoding aggrecan and types I and II collagen, and in the production of glycosaminoglycan, collagen and DNA. We found that FGF-2 or the combination of IGF-I, BMP-2, and BMP-7 increased sox9 gene expression and protein production and that sox9 knockdown modulated growth factor actions in a complex fashion that differed both with growth factors and with chondrocyte function. The data suggest that sox9 mediates the stimulation of matrix production by the combined growth factors and the stimulation of chondrocyte proliferation by FGF-2. The mitogenic effect of the combined growth factors and the catabolic effect of FGF-2 appear to involve sox9-independent mechanisms. Control of these molecular mechanisms may contribute to the treatment of cartilage damage.

SUBMITTER: Shi S

PROVIDER: S-EPMC5210171 | biostudies-literature | 2015 Jul

REPOSITORIES: biostudies-literature

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Role of sox9 in growth factor regulation of articular chondrocytes.

Shi Shuiliang S Wang Congrong C Acton Anthony J AJ Eckert George J GJ Trippel Stephen B SB

Journal of cellular biochemistry 20150701 7

Chondrogenic polypeptide growth factors influence articular chondrocyte functions that are required for articular cartilage repair. Sox9 is a transcription factor that regulates chondrogenesis, but its role in the growth factor regulation of chondrocyte proliferation and matrix synthesis is poorly understood. We tested the hypotheses that selected chondrogenic growth factors regulate sox9 gene expression and protein production by adult articular chondrocytes and that sox9 modulates the actions o ...[more]

PMID: 25708223

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Project description:Cartilage degeneration driven by catabolic stimuli is a critical pathophysiological process in osteoarthritis (OA). We have defined fibroblast growth factor 2 (FGF-2) as a degenerative mediator in adult human articular chondrocytes. Biological effects mediated by FGF-2 include inhibition of proteoglycan production, up-regulation of matrix metalloproteinase-13 (MMP-13), and stimulation of other catabolic factors. In this study, we identified the specific receptor responsible for the catabolic functions of FGF-2, and established a pathophysiological connection between the FGF-2 receptor and OA.Primary human articular chondrocytes were cultured in monolayer (24 hours) or alginate beads (21 days), and stimulated with FGF-2 or FGF18, in the presence or absence of FGFR1 (FGF receptor 1) inhibitor. Proteoglycan accumulation and chondrocyte proliferation were assessed by dimethylmethylene blue (DMMB) assay and DNA assay, respectively. Expression of FGFRs (FGFR1 to FGFR4) was assessed by flow cytometry, immunoblotting, and quantitative real-time PCR (qPCR). The distinctive roles of FGFR1 and FGFR3 after stimulation with FGF-2 were evaluated using either pharmacological inhibitors or FGFR small interfering RNA (siRNA). Luciferase reporter gene assays were used to quantify the effects of FGF-2 and FGFR1 inhibitor on MMP-13 promoter activity.Chondrocyte proliferation was significantly enhanced in the presence of FGF-2 stimulation, which was inhibited by the pharmacological inhibitor of FGFR1. Proteoglycan accumulation was reduced by 50% in the presence of FGF-2, and this reduction was successfully rescued by FGFR1 inhibitor. FGFR1 inhibitors also fully reversed the up-regulation of MMP-13 expression and promoter activity stimulated by FGF-2. Blockade of FGFR1 signaling by either chemical inhibitors or siRNA targeting FGFR1 rather than FGFR3 abrogated the up-regulation of matrix metalloproteinases 13 (MMP-13) and a disintegrin and metalloproteinase with a thrombospondin type 1 motif 5 (ADAMTS5), as well as down-regulation of aggrecan after FGF-2 stimulation. Flow cytometry, qPCR and immunoblotting analyses suggested that FGFR1 and FGFR3 were the major FGFR isoforms expressed in human articular chondrocytes. FGFR1 was activated more potently than FGFR3 upon FGF-2 stimulation. In osteoarthritic chondrocytes, FGFR3 was significantly down regulated (P < 0.05) with a concomitant increase in the FGFR1 to FGFR3 expression ratio (P < 0.05), compared to normal chondrocytes. Our results also demonstrate that FGFR3 was negatively regulated by FGF-2 at the transcriptional level through the FGFR1-ERK (extracellular signal-regulated kinase) signaling pathway in human articular chondrocytes.FGFR1 is the major mediator with the degenerative potential in the presence of FGF-2 in human adult articular chondrocytes. FGFR1 activation by FGF-2 promotes catabolism and impedes anabolism. Disruption of the balance between FGFR1 and FGFR3 signaling ratio may contribute to the pathophysiology of OA.

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Role of sox9 in growth factor regulation of articular chondrocytes.

Publications

Role of sox9 in growth factor regulation of articular chondrocytes.

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