Fibroblast growth factor receptor 1 is principally responsible for fibroblast growth factor 2-induced catabolic activities in human articular chondrocytesReport as inadecuate




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Arthritis Research and Therapy

, 13:R130

First Online: 11 August 2011Received: 22 December 2010Revised: 06 June 2011Accepted: 11 August 2011

Abstract

IntroductionCartilage 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.

MethodsPrimary 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.

ResultsChondrocyte 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.

ConclusionsFGFR1 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.

AbbreviationsADAMTSa disintegrin and metalloproteinase with a thrombospondin type 1 motif

BCAbicinchoninic acid

DMEMDulbecco-s modified Eagle-s medium

DMMBdimethylmethylene blue

ERKextracellular signal-regulated kinase

FGF-2fibroblast growth factor 2

FGFRFGF receptor

IPimmunoprecipitation

MMPmatrix metalloproteinase

OAosteoarthritis

qPCRquantitative real-time polymerase chain reaction

RTreverse transcription

siRNAsmall interfering RNA.

Electronic supplementary materialThe online version of this article doi:10.1186-ar3441 contains supplementary material, which is available to authorized users.

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Author: Dongyao Yan - Di Chen - Simon M Cool - Andre J van Wijnen - Katalin Mikecz - Gillian Murphy - Hee-Jeong Im

Source: https://link.springer.com/



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