Session Information
Session Type: Abstract Submissions (ACR)
Background/Purpose:
Rheumatoid arthritis (RA) is characterized by hyperplasia of the synovial tissue due to local proliferation of stromal and recruitment of inflammatory immune cells. Accumulation of synovial fibroblasts likely is due to an imbalance between cell proliferation, survival and death. Both cytosolic dsRNA receptors, MDA5 and RIG-I, can initiate pro-apoptotic signalling in a variety of cell types. In animal models of cancer and multiple sclerosis, treatment with dsRNA ligands causes apoptosis of effector cells and ameliorates disease. We showed that fibroblast-like synoviocytes (FLS) from RA synovial tissue are equipped with functional cytosolic dsRNA receptors. We have studied the consequences of stimulation with dsRNA on the survival of FLS.
Methods:
FLS from arthritis patients were stimulated with poly(I:C) or 3pRNA complexed with fugene (FG) for intracellular uptake. Stimulation was confirmed by RT-PCR of dsRNA response genes (MDA5, RIG-I and CXCL10). Cell death of FLS after triggering of dsRNA sensors was detected by flow-cytometric analysis using annexin V/PI staining. Changes in protein expression of anti- and pro-apoptotic genes were analyzed by Western blot. Knockdown of MDA5, RIG-I and their adaptor proteins was performed using SMART pool siRNAs.
Results:
dsRNA response genes, including MDA5, RIG-I and CXCL10 were similarly induced after 16 h stimulation of RA FLS with either 3pRNA or poly(I:C), complexed with FG. However, 3pRNA + FG treatment of FLS caused about 15% of cell death after 48 h, whereas poly(I:C) + FG treatment induced up to 75% cell death. Surprisingly, we did not find differences in the expression of pro- and anti-apoptotic proteins between cells stimulated with 3pRNA + FG or poly(I:C) + FG. siRNA knockdown of MDA5, a known receptor for poly(I:C), did not effect poly(I:C) + FG stimulation on dsRNA response genes, whereas knockdown of RIG-I, a known receptor for 3pRNA, completely abrogated the response to 3pRNA + FG. Knockdown of downstream adaptor proteins IPS, STING and TRIF did not have any effect on dsRNA response genes after poly(I:C) + FG stimulation.
Conclusion:
Stimulation of RA FLS with artificial dsRNA poly(I:C), but not 3pRNA induced apoptosis of FLS. We confirmed that 3pRNA is recognized by the dsRNA sensor RIG-I. In contrast, intracellular recognition of poly(I:C) in FLS does not involve MDA5 and the adapter proteins IPS, STING and TRIF, suggesting the existence of a different recognition mechanism.
Disclosure:
J. Hamann,
None;
P. P. Tak,
None;
O. N. Karpus,
None.
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/polyic-induced-cell-death-of-synovial-fibroblasts-via-an-unknown-dsrna-receptor/