- Meeting abstract
Reporter cell lines for real-time imaging of NF-κB activation in the arthritic joint
Arthritis Res Ther volume 6, Article number: 70 (2004)
Inadequate functioning of NF-κB is implicated in many diseases, such as rheumatoid arthritis (RA), and increased activation of NF-κB in synovial tissue precedes the clinical manifestation of experimental arthritis.
The aim of the study was to develop NF-κB reporter cell-lines that can be used for real-time evaluation of NF-κB activation in knee joints during experimental arthritis in living animals.
For determination of NF-κB transcription activity, an eukaryotic expression vector was created containing five tandemly arranged NF-κB binding sites (5'-GGGACTTTCC-3') regulating luciferase expression. Cell lines were selected on stable incorporation of 5 × NF-κB luciferase and responsiveness was tested toward NF-κB inducing stimuli.
These cell-lines, derived from fibroblasts, chondrocytes and thymocytes, showed good cytokine responsiveness toward IL-1, IL-17, IL-18, and TNF. We showed that these cells could also be used in in vitro inhibition assays using cytokine protein antagonist (e.g. IL-1Ra and IL-18BPc) or RNA interference. By transplanting these cells into HLA matched recipients we were able to image in real time NF-κB activation in knee joints of mice with IL-1- or SCW-induced inflammation using a cooled-charge-coupled device (CCCD) camera. Interestingly, NF-κB activation was also activated at sites remote from the monokine or SCW injection sites. Synovial mRNA revealed a striking upregulation of IL-1 in both SCW challenged and contralateral knee joint. This effect was confirmed by pretreatment of mice with neutralizing anti-IL-1 Abs, which markedly prevented the NF-κB activation in the contralateral naive joints.
These reporter cell-lines can be a valuable tool to study the role of NF-κB activation in arthritis and to evaluate cytokine/NF-κB based inhibition therapies. Furthermore, we show that local induction of experimental arthritis results in contralateral NF-κB activation, which is dependent on IL-1.
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Smeets, R., Joosten, L., van den Berg, W. et al. Reporter cell lines for real-time imaging of NF-κB activation in the arthritic joint. Arthritis Res Ther 6, 70 (2004). https://doi.org/10.1186/ar1112
- Knee Joint
- Eukaryotic Expression Vector
- Experimental Arthritis
- Stable Incorporation
- Cytokine Responsiveness