Background Contribution of Toll-Like Receptor 4 (TLR4) to pain sensitisation has been demonstrated to occur under chronic pain conditions. We previously described an antinociceptive effect of TLR4-A1, a TLR4 inhibitor, in two chronic pain conditions, peripheral neuropathic pain and osteoarthritis (OA).
Objectives The aim of this study was to evaluate TLR4-A1 effect on allodynia and hyperalgesia in OA model, and to evaluate whether this effect is correlated with changes in spinal glial activation.
Methods Wistar rats weighing 200–250g were used. OA was induced by a single intraarticular injection of 2mg of monosodium iodoacetate (MIA) into the right knee joint of anaesthetised rats. TLR4-A1, 10 mgkg-1, was intraperitoneally administered during the first five days post-MIA injection. TLR4-A1 was synthesised by Dr Quesada. Vehicle-treatment (ethanol:saline, 1:9) was used as control. Each group was composed of 6 animals. After three weeks (day 22 post-MIA injection), animals were sacrificed for tissue collection. L3-L5 spinal segments were collected and embedded in paraffin wax. Eventually, samples were immune-stained with anti-GFAP or Iba-1 antibodies. Photomicrographs were recorded to make montages of the entire spinal cord at a final magnification of 20x (n=3 per lumbar section). Total number of GFAP or Iba-1 positive cells were counted separately in laminas I-II, III-IV and V-VI.
Results Intraarticular injection of MIA increased microglial expression (Iba-1 labelling) in the ipsilateral spinal cord compared to the contralateral side, being the difference statistically significant for the superficial (I-II, +72.25%; P<0.01) and deeper (V-VI, +95.31%; P<0.001) laminae of L3 and for the superficial laminae of L4 (+87.5%; P<0.01). In animals treated with TLR4-A1, Iba-1 labelling in the ipsilateral dorsal horn showed a similar pattern to the contralateral dorsal horn. Pre-treatment with TLR4 blocker prevented microglia activation after MIA-injection in L3 and L4 segments.
Intraarticular injection of MIA also increased the number of GFAP-positive activated astrocytes in the ipsilateral spinal cord compared to the contralateral side; in this case, statistically significant differences were found for the superficial (I-II; +41.62%; p<0.01) and middle (III-IV; +64.35%, p<0.001) laminae of L3 sections. GFAP in TLR4-A1-treated rats showed a similar pattern for the ipsi- and the contra-lateral sides. That is, TLR4-A1 prevented L3 increased activated astroglia following MIA-injection.
Conclusions Early toll-like receptor 4 blockade hampers spinal glial activation, which correlates with diminished allodynia and hyperalgesia observed in TLR4-A1-treated animals in a model of MIA-induced OA. Although further studies are needed, TLR4 blockade could be a good option in the treatment of osteoarthritis.
Acknowledgements Franco R and Marquez A for technical support.
Granted: Ministerio de Economía y Competitividad. SAF2012–40075-C02–01.
Disclosure of Interest None declared