INSERM UMR_S 938, Centre de Recherche de Saint-Antoine, Hôpital Saint-Antoine, Paris F-75012, France.
J Neuroinflammation. 2012 Feb 21;9:36. doi: 10.1186/1742-2094-9-36.
Sustained neuroinflammation strongly contributes to the pathogenesis of pain. The clinical challenge of chronic pain relief led to the identification of molecules such as cytokines, chemokines and more recently matrix metalloproteinases (MMPs) as putative therapeutic targets. Evidence points to a founder member of the matricial CCN family, NOV/CCN3, as a modulator of these inflammatory mediators. We thus investigated the possible involvement of NOV in a preclinical model of persistent inflammatory pain.
We used the complete Freund's adjuvant (CFA)-induced model of persistent inflammatory pain and cultured primary sensory neurons for in vitro experiments. The mRNA expression of NOV and pro-inflammatory factors were measured with real-time quantitative PCR, CCL2 protein expression was assessed using ELISA, MMP-2 and -9 activities using zymography. The effect of drugs on tactile allodynia was evaluated by the von Frey test.
NOV was expressed in neurons of both dorsal root ganglia (DRG) and dorsal horn of the spinal cord (DHSC). After intraplantar CFA injection, NOV levels were transiently and persistently down-regulated in the DRG and DHSC, respectively, occurring at the maintenance phase of pain (15 days). NOV-reduced expression was restored after treatment of CFA rats with dexamethasone. In vitro, results based on cultured DRG neurons showed that siRNA-mediated inhibition of NOV enhanced IL-1β- and TNF-α-induced MMP-2, MMP-9 and CCL2 expression whereas NOV addition inhibited TNF-α-induced MMP-9 expression through β1 integrin engagement. In vivo, the intrathecal delivery of MMP-9 inhibitor attenuated mechanical allodynia of CFA rats. Importantly, intrathecal administration of NOV siRNA specifically led to an up-regulation of MMP-9 in the DRG and MMP-2 in the DHSC concomitant with increased mechanical allodynia. Finally, NOV intrathecal treatment specifically abolished the induction of MMP-9 in the DRG and, MMP-9 and MMP-2 in the DHSC of CFA rats. This inhibitory effect on MMP is associated with reduced mechanical allodynia.
This study identifies NOV as a new actor against inflammatory pain through regulation of MMPs thus uncovering NOV as an attractive candidate for therapeutic improvement in pain relief.
持续的神经炎症强烈促进疼痛的发病机制。慢性疼痛缓解的临床挑战导致鉴定出细胞因子、趋化因子和最近的基质金属蛋白酶 (MMPs) 等作为潜在的治疗靶点。有证据表明，基质细胞周期素 (CCN) 家族的创始人 NOV/CCN3 是这些炎症介质的调节剂。因此，我们研究了 NOV 在持续性炎症性疼痛的临床前模型中的可能参与。
我们使用完全弗氏佐剂 (CFA) 诱导的持续性炎症性疼痛模型和原代感觉神经元进行体外实验。使用实时定量 PCR 测量 NOV 和促炎因子的 mRNA 表达，使用 ELISA 评估 CCL2 蛋白表达，使用明胶酶谱法评估 MMP-2 和 -9 活性。通过 von Frey 试验评估药物对触觉过敏的影响。
NOV 在背根神经节 (DRG) 和脊髓背角 (DHSC) 的神经元中表达。足底注射 CFA 后，NOV 水平在 DRG 和 DHSC 中分别短暂和持续下调，发生在疼痛维持阶段 (15 天)。用地塞米松治疗 CFA 大鼠后，NOV 表达减少得到恢复。在体外，基于培养的 DRG 神经元的结果表明，siRNA 介导的 NOV 抑制增强了 IL-1β 和 TNF-α诱导的 MMP-2、MMP-9 和 CCL2 表达，而 NOV 添加通过β1 整合素结合抑制了 TNF-α诱导的 MMP-9 表达。在体内，鞘内给予 MMP-9 抑制剂可减轻 CFA 大鼠的机械性痛觉过敏。重要的是，鞘内给予 NOV siRNA 特异性导致 DRG 中 MMP-9 和 DHSC 中 MMP-2 的上调，同时伴有机械性痛觉过敏增加。最后，NOV 鞘内给药特异性地抑制了 CFA 大鼠 DRG 中 MMP-9 的诱导，以及 DHSC 中 MMP-9 和 MMP-2 的诱导。这种对 MMP 的抑制作用与机械性痛觉过敏的减少有关。
本研究通过调节 MMP 将 NOV 鉴定为一种新的抗炎痛因子，从而揭示 NOV 作为治疗疼痛缓解的有吸引力的候选药物。
