Background Synovitis-associated pain is an important aspect of arthritis pathology. Several inflammatory mediators released by the synovium have been implicated in the regulation of pain, including S100A8 and S100A9 which may regulate pain either via direct stimulation of TLR4 on the nerve endings in the synovium or via stimulation of the dorsal root ganglia (DRG), thereby enabling an increased phagocyte infiltration.
Objectives To investigate the role of S100A9 in the pain response after induction of an acute synovitis using streptococcal cell walls (SCW) as a trigger, comparing S100A9-/- mice and their WT controls.
Methods Acute synovitis was induced by a single i.a. injection of SCW in the knee joint of C57Bl6 (WT) mice and S100A9-/- mice, control mice received an i.a. saline injection. Serum S100A8/A9 levels were investigated by ELISA and expression of S100A8 and S100A9 in synovium and DRG by immunohistochemistry. Joint swelling and cell influx was assessed by 99mTc accumulaiton and histology, respectively. Pain response were investigated Incapacitance Tester (weight bearing), Catwalk (gait analysis) and von Frey's filaments (mechanical allodynia). Gene expression of inflammatory mediators and neuron activation markers in DRG were determined by q-PCR.
Results A single i.a. injection of SCW resulted in increased synovial expression of S100A8 and S100A9 and subsequent increased serum S100A8/A9 levels (2.6-fold, P<0.001) 1 day p.i., which returned to basal levels at 7 days p.i. The increased expression of S100A9 did not contribute to the development of inflammation since joint swelling and cell influx were similar in WT and S100A9-/- mice 1 day p.i. Using the Incapacitance Tester, WT mice showed a marked and significant decrease in percentage of weight bearing on the SCW injected hindpaw (28%) compared to saline injection (47%, P <0.001) 1 day p.i., whereas S100A9-/- mice did not. In addition, gait analysis showed that the stand-phase of the unaffected paws were significantly increased in WT mice 1 day p.i., which will reduce the load on the inflamed paw, while in S100A9-/- mice these parameters were not altered. No difference in mechanical allodynia was observed, both mouse strains showed a similar reduction of paw withdrawal threshold (4.2 and 4.5 fold decrease respectively). Analysis of DRG showed no increased phagocyte infiltration after SCW injection as determined by S100A8 and S100A9 immunohistochemistry and no change in gene expression of MCP-1, KC, IL-1β or TNF was measured. However, expression of neuron activation markers NAV1.7, ATF3 and GAP43 were significantly increased at 1 day after SCW injection in WT mice as compared to saline injected mice (P =0.022, 0.004 and 0.030, respectively) while SCW injection in S100A9-/- mice did not show increased expression, which is in line in with the reduced pain response observed earlier in S100A9-/- mice.
Conclusions These findings show that S100A9, which is released from the synovium upon inflammation, is an important mediator of inflammatory pain response in the knee, and that during the acute phase of inflammation is likely regulated via direct activation of TLR4 on nerve endings in the synovium and not via increased infiltration of phagocytes in the DRG.
Disclosure of Interest None declared
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