Abstract
The synovium secretes synovial fluid, but is also richly innervated with nociceptors and acts as a gateway between avascular joint tissues and the circulatory system. Resident fibroblast-like synoviocytes’ (FLS) calcium-activated potassium channels (KCa) change in activity in arthritis models and this correlates with FLS activation.ObjectiveTo investigate this activation in an in vitro model of inflammatory arthritis; 72 h treatment with cytokines TNFα and IL1β.MethodsFLS cells were isolated from rat synovial membranes. We analyzed global changes in FLS mRNA by RNA-sequencing, then focused on FLS ion channel genes and the corresponding FLS electrophysiological phenotype and finally modeling data with ingenuity pathway analysis (IPA) and MATLAB.ResultsIPA showed significant activation of inflammatory, osteoarthritic and calcium signaling canonical pathways by cytokines, and we identified ∼200 channel gene transcripts. The large KCa (BK) channel consists of the pore forming Kcnma1 together with β-subunits. Following cytokine treatment, a significant increase in Kcnma1 RNA abundance was detected by qPCR and changes in several ion channels were detected by RNA-sequencing, including a loss of BK channel β-subunit expression Kcnmb1/2 and an increase in Kcnmb3. In electrophysiological experiments, there was a decrease in over-all current density at 20 mV without change in chord conductance at this potential.ConclusionTNFα and IL1β treatment of FLS in vitro recapitulated several common features of inflammatory arthritis at the transcriptomic level, including increase in Kcnma1 and Kcnmb3 gene expression.
Highlights
Rheumatoid arthritis (RA) and osteoarthritis (OA) are degenerative diseases that target articular joint structures resulting in pain, loss of function and frequent disability
We investigate whether the TNFα and IL1β cytokine in vitro model of inflammation leads to a significant change of the BK ion channel and quantify the mechanism of this change
Our first bioinformatic analysis tested the reproducibility of the 72 h, 10 ng/ml TNFα + IL1β treatment
Summary
Rheumatoid arthritis (RA) and osteoarthritis (OA) are degenerative diseases that target articular joint structures resulting in pain, loss of function and frequent disability. Whilst RA is an established inflammatory condition, the contribution of inflammatory processes to OA was less well known until recently. Mediators of inflammation (i.e., including pro-inflammatory cytokines) contribute to the development of synovitis, which is known to drive disease progression in RA. How inflammation drives joint destruction is not fully known. One feature of synovitis is the presence of major pro-inflammatory cytokines, such as interleukin-1β (IL-1β), tumor necrosis factor alpha (TNFα), and interleukin 6 (IL-6), resulting in suppression of collagen and proteoglycan synthesis, increased inflammatory signaling, and protease expression and activation (Martel-Pelletier et al, 1999)
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