Institute of Pharmacology, Polish Academy of Sciences, Department of Pain Pharmacology, Krakow, Poland.
Institute of Pharmacology, Polish Academy of Sciences, Department of Pain Pharmacology, Krakow, Poland.
Cytokine. 2019 Jul;119:202-213. doi: 10.1016/j.cyto.2019.03.007. Epub 2019 Apr 16.
The complex neuroimmunological interactions mediated by chemokines are suggested to be responsible for the development of neuropathic pain. The lack of knowledge regarding the detailed pathomechanism of neuropathy is one reason for the lack of optimally efficient therapies. Recently, several lines of evidence indicated that expression of CCR2 is increased in spinal cord neurons and microglial cells after peripheral nerve injury. It was previously shown that administration of CCR2 antagonists induces analgesic effects; however, the role of CCR2 ligands in neuropathic pain still needs to be explained. Thus, the goal of our studies was to investigate the roles of CCL2, CCL7, and CCL12 in neuropathic pain development and opioid effectiveness. The experiments were conducted on primary glial cell cultures and two groups of mice: naive and neuropathic. We used chronic constriction injury (CCI) of the sciatic nerve as a neuropathic pain model. Mice intrathecally received chemokines (CCL2, CCL7, CCL12) at a dose of 10, 100 or 500 ng, neutralizing antibodies (anti-CCL2, anti-CCL7) at a dose of 1, 4 or 8 μg, and opioids (morphine, buprenorphine) at a dose of 1 μg. The pain-related behaviors were assessed using the von Frey and cold plate tests. The biochemical analysis of mRNA expression of glial markers, CCL2, CCL7 and CCL12 was performed using quantitative reverse transcriptase real-time PCR. We demonstrated that CCI of the sciatic nerve elevated spinal expression of CCL2, CCL7 and CCL12 in mice, in parallel with microglia and astroglial activation markers. Moreover, intrathecal injection of CCL2 and CCL7 induced pain-related behavior in naive mice in a dose-dependent manner. Surprisingly, intrathecal injection of CCL12 did not influence nociceptive transmission in naive or neuropathic mice. Additionally, we showed for the first time that intrathecal injection of CCL2 and CCL7 neutralizing antibodies not only attenuated CCI-induced pain-related behaviors in mice but also augmented the analgesia induced by morphine and buprenorphine. In vitro studies suggest that both microglia and astrocytes are an important cellular sources of the examined chemokines. Our results revealed the crucial roles of CCL2 and CCL7, but not CCL12, in neuropathic pain development and indicated that pharmacological modulation of these factors may serve as a potential therapeutic target for new (co)analgesics.
趋化因子介导的复杂神经免疫相互作用被认为是导致神经性疼痛的原因。对神经病变详细发病机制缺乏了解是缺乏最佳有效治疗方法的原因之一。最近，有几条证据表明，在外周神经损伤后，脊髓神经元和小胶质细胞中的 CCR2 表达增加。先前的研究表明，CCR2 拮抗剂的给药可诱导镇痛作用；然而，趋化因子配体在神经性疼痛中的作用仍需要解释。因此，我们的研究目标是研究 CCL2、CCL7 和 CCL12 在神经性疼痛发展和阿片类药物有效性中的作用。该实验在原代神经胶质细胞培养物和两组小鼠中进行：未损伤和神经病变。我们使用坐骨神经慢性缩窄性损伤 (CCI) 作为神经性疼痛模型。将趋化因子（CCL2、CCL7、CCL12）以 10、100 或 500ng 的剂量、中和抗体（抗 CCL2、抗 CCL7）以 1、4 或 8μg 的剂量、阿片类药物（吗啡、丁丙诺啡）以 1μg 的剂量鞘内给药。使用 von Frey 和冷板测试评估与疼痛相关的行为。使用定量逆转录实时 PCR 进行神经胶质标志物、CCL2、CCL7 和 CCL12 的 mRNA 表达的生化分析。我们证明，坐骨神经 CCI 会导致小鼠脊髓中 CCL2、CCL7 和 CCL12 的表达升高，同时还伴有小胶质细胞和星形胶质细胞激活标志物。此外，CCL2 和 CCL7 的鞘内注射以剂量依赖性方式诱导未损伤小鼠的疼痛相关行为。令人惊讶的是，CCL12 的鞘内注射不会影响未损伤或神经病变小鼠的伤害性传递。此外，我们首次表明，CCL2 和 CCL7 中和抗体的鞘内注射不仅减弱了 CCI 诱导的小鼠疼痛相关行为，还增强了吗啡和丁丙诺啡诱导的镇痛作用。体外研究表明，小胶质细胞和星形胶质细胞都是这些趋化因子的重要细胞来源。我们的结果揭示了 CCL2 和 CCL7 而不是 CCL12 在神经性疼痛发展中的关键作用，并表明这些因素的药理学调节可能成为新的（共同）镇痛药的潜在治疗靶点。
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