Session Title: Rheumatoid Arthritis – Human Etiology and Pathogenesis I
Session Type: ACR Concurrent Abstract Session
Session Time: 2:30PM-4:00PM
Aberrant epigenetic marks in RA FLS contribute to disease pathogenesis and aggressive FLS behavior. Computational data also suggest that RA FLS isolated from different joint locations display distinct epigenetic states and transcriptomes that implicate joint-specific pathogenic pathways. For example, IL-6 signaling and JAK-STAT epigenetic marks differ in RA hips and knees. This suggests that hip and knee FLS might have differential responses to IL-6 stimulation and JAK inhibitors like tofacitinib, which could contribute to asynchronous clinical responses between joints in RA patients treated with targeted agents. In this study, the joint-specific biology of IL-6 and JAK in RA hip and knee FLS were explored to biologically validate the computational data using IL-6-induced monocyte chemoattractant protein 1 (MCP1) gene expression as a primary differential function.
FLS lines were established from synovial tissue of RA patients undergoing total hip or knee replacement surgery. The cells were expanded in 10% fetal calf serum/DMEM and used at passages 3 – 7. FLS were plated, serum starved for 18 hours and then stimulated with various concentrations of IL-6 for 2h. In some experiments, the cells were pre-treated with tofacitinib prior to IL-6 stimulation. RNA was isolated and GAPDH-normalized gene transcripts were quantified by qPCR.
Under basal conditions, hip FLS expressed significantly less MCP1 than knee FLS (1.37±0.22 and 2.5±0.27, respectively; n=15/group, p=0.003). At low doses of IL-6 (<20ng/ml), MCP1 expression for hip FLS remained lower than for knee FLS, but the fold increase was greater for hip. For example, at 7.5 ng/ml of IL-6, MCP1 increased 2.4±0.4 fold for hip FLS and 1.2±0.07 fold for knee FLS). At high concentrations of IL-6 (40-100ng/ml) hip and knee MCP1 gene expression were similar, suggesting that the differences under basal conditions could be overcome. Because JAK-STAT signaling in RA hips and knee FLS were predicted to be different based on our computational data, we performed dose responses with the JAK inhibitor tofacitinib. Pre-incubation of FLS with various concentrations of tofacitinib prior to IL-6 stimulation reduced IL-6-induced MCP1 expression for both hip and knee FLS, but with greater sensitivity in hip FLS (EC50 of 30 nM for hip FLS and 80 nM for knee FLS). These results suggest that knee FLS required higher concentrations of tofacitinib for the same degree of MCP1 inhibition as hip FLS. Inhibition of basal expression of MCP1 in the absence of exogenous IL-6 was greater for hip compared with knee FLS (50% and 13% inhibition, respectively, n=4/group) suggesting that JAK-STAT-related signaling differs in the joint-specific FLS.
These studies biologically validate the prediction that RA hip and knee FLS are imprinted with distinct functions related to IL-6 signaling and the JAK-STAT pathway. Differential sensitivity to JAK inhibition could be related to the unique epigenetic imprinting due to joint location and could contribute to variable clinical responses to tofacitinib. Additional studies that profile the epigenome and transcriptome in individual joints could create opportunities for personalized therapy.
To cite this abstract in AMA style:Hammaker D, Nygaard G, Boyle DL, Ai R, Wang W, Firestein GS. Joint Location-Specific JAK-STAT Signaling in Rheumatoid Arthritis (RA) Fibroblast-like Synoviocytes (FLS) [abstract]. Arthritis Rheumatol. 2017; 69 (suppl 10). https://acrabstracts.org/abstract/joint-location-specific-jak-stat-signaling-in-rheumatoid-arthritis-ra-fibroblast-like-synoviocytes-fls/. Accessed August 3, 2021.
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