Background/Purpose:

Initiation of protein synthesis is regulated by a repressor of cap-dependent translation, eukaryotic initiation factor 4E-binding protein 1 (4E-BP1), which prevents eukaryotic initiation factor 4E (eIF4E) from participating in the eukaryotic translation apparatus through its binding to eIF4E. In this study we determined whether the control of translational apparatus activity could have an effect on the expression of transforming growth factor-beta 1 (TGF-β1)-induced catabolic or anabolic genes in primary chondrocytes and be responsible for the pathogenesis of osteoarthritis (OA).

Methods:

Human articular chondrocytes were enzymatically isolated from articular cartilage and cultured in monolayer. The relative levels of mRNA and protein were analyzed by real-time quantitative reverse transcription-polymerase chain reaction and Western blot analysis, respectively. Immunoprecipitation (IP) assay was performed to investigate the binding of 4E-BP1 to eIF4E. The total protein synthesis was measured by flow cytometry using protein synthesis assay kit. 4E-BP1 and mTOR genes were knocked down by transfection with small interfering RNAs (siRNAs).

Results:

The level of 4EBP-1 was significantly higher in human OA cartilage than normal cartilage. TGF-β1 increased total protein synthesis, including aggrecan (ACAN) and collagen type II (Col II), together with activation of Akt/mTOR signaling pathway. IP data demonstrated that TGF-β1 treatment suppressed the interaction of 4E-BP1 and eIF4E. mTOR silencing significantly suppressed ACAN and Col II expressions through decreased level of phosphorylated 4E-BP1 in TGF-β1-treated chondrocytes. On the other hand, 4E-BP1 knockdown promoted total protein synthesis but suppressed Col II and ACAN expressions with decreased expression of Smad2/3 and Smad4 and increased expression of inhibitory Smad6 and Smad7. Furthermore, knockdown of Smad4 suppressed Col II and ACAN expression whereas increased expressions of Col II and ACAN were observed in Smad6- or Smad7-knocked down chondrocytes

Conclusion:

These results demonstrated that TGF-β1-modulated phosphorylation of 4EBP1 plays a role in the expression of Col II and ACAN through alteration of Smad signaling pathway.

« Back to 2018 ACR/ARHP Annual Meeting

ACR Meeting Abstracts - https://acrabstracts.org/abstract/the-effect-of-4e-bp1-on-osteoarthritis-related-gene-expression-in-tgf-%ce%b21-stimulated-chondrocytes/