Evaluation of Trigeminal Ganglia Sensory Neuronal Population/s Mediating MIF-Induced Anti-Nociception in a Model of Apical Periodontitis.

摘要 尽管根管治疗在治疗根尖周炎等疾病方面取得了成功， 可能会在治疗后六个月出现持续性疼痛。根尖牙周炎是由牙周组织感染引起的。 牙髓导致机械性异常性疼痛和炎症。持续性牙痛增加负担成本 以及服用止痛药。此外，非甾体抗炎药（NSAID）和阿片类药物提供了 不完全的疼痛缓解和长期服用时的不良反应，突出了对新治疗的需求 可以缓解数百万持续牙痛的患者。干细胞的研究 镇痛性能正在迅速增长，希望开发出一类新型镇痛药。疗效 在神经性疼痛的临床前模型和患者中， 被诊断为偏头痛和骨关节炎初步数据表明，静脉注射人 根尖乳头干细胞（hSCAP）完全逆转与根尖牙周炎相关的超敏反应， 小鼠对感染牙齿的hSCAP的RNA测序显示， 巨噬细胞移动抑制因子（MIF）。此外，MIF受体CD 74和CXCR 4 共定位于三叉神经节中的TRPV 1+神经元上。此外，局部注射重组MIF可逆转 与根尖牙周炎相关的超敏反应。最后，将来自小鼠共培养物的条件培养基 根尖周肉芽肿（感染的牙齿）和hSCAP减弱辣椒素诱发的Ca 2+反应， 三叉神经节神经元，并且用MIF抗体的预处理抑制。这些数据支持了一种新的 通过可能由MIF介导的干细胞的直接作用的干细胞抗伤害感受的机制。 然而，尽管原始数据表明MIF直接抑制三叉神经节神经元的活动，但神经元的活动并不受影响。 MIF诱导的抗伤害感受的亚型是未知的。根尖周炎引起的疼痛通常 如所提及的那样报道，表明由于中枢致敏而涉及非伤害性感受纤维。 了解这些亚群中哪些介导了MIF诱导的抗伤害感受，将使我们能够开发 针对根尖周炎相关的持续性牙痛的治疗。我们将测试中央 假设MIF诱导的三叉神经节神经元活性抑制是由伤害性和非伤害性介导的， 伤害感受神经元为了验证这一假设，我们将1）进行单细胞RT-PCR和免疫组织化学 确定MIF受体CD 74和CXCR 4与不同神经元标记物的共表达，和2）使用Cre- 重组酶从特定的神经元群体中有条件地敲除MIF受体CD 74和CXCR 4， 在根尖牙周炎模型中评价它们对MIF抗伤害感受的贡献。这些研究将 不仅为介导MIF诱导的抗伤害感受的神经元群体提供了新的见解， 作为一个优秀的培训工具，我的职业生涯作为一个临床科学家。

Abstract Despite the success of root canal treatments at treating diseases such as apical periodontitis, 1.8 million patients may experience persistent pain six months after treatment. Apical periodontitis is caused by an infection of the dental pulp leading to mechanical allodynia and inflammation. Persistent dental pain increases cost of burden and intake of analgesic drugs. Furthermore, non-steroidal anti-inflammatory drugs (NSAIDs) and opioids provide incomplete pain relief and adverse effects when taken chronically, highlighting the need for a novel treatment that can provide relief to millions of patients experiencing persistent dental pain. The study of stem cells for their analgesic properties has been rapidly growing in hopes of developing a novel class of analgesics. The efficacy of mesenchymal stem cells has been demonstrated in pre-clinical models of neuropathic pain and in patients diagnosed with migraines and osteoarthritis. Preliminary data demonstrates that intravenous injections of human stem cells of the apical papilla (hSCAP) fully reverses hypersensitivity associated with apical periodontitis in mice. RNA sequencing of hSCAP homed to the infected tooth shows a 133-fold increase in the expression of the cytokine Macrophage Migratory Inhibitory Factor (MIF). Additionally, MIF receptors CD74 and CXCR4 colocalize on TRPV1+ neurons in the trigeminal ganglia. Moreover, a local injection of recombinant MIF reverses hypersensitivity associated with apical periodontitis. Lastly, conditioned media from co-cultures of mouse periapical granulomas (the infected tooth) and hSCAP attenuates capsaicin evoked Ca2+ response from trigeminal ganglia neurons and is inhibited with pre-treatment of a MIF antibody. This data supports a novel mechanism for stem cell anti-nociception through the direct effect of stem cells that is possibly mediated by MIF. However, despite primary data suggesting MIF directly inhibits trigeminal ganglia neuronal activity, the neuronal subtypes that MIF-induced anti-nociception are unknown. The pain induced by apical periodontitis is often reported as referred, suggesting the involvement of non-nociceptive fibers due to central sensitization. Understanding which of these subpopulations mediate MIF-induced anti-nociception will allow us to develop targeted treatments for persistent dental pain associated with apical periodontitis. We will test the central hypothesis that MIF-induced inhibition of trigeminal ganglia neuronal activity is mediated by nociceptive and non- nociceptive neurons. To test this hypothesis, we will 1) conduct single cell RT-PCR and immunohistochemistry to determine co-expression of MIF receptors CD74 and CXCR4 with different neuronal markers and 2) use Cre- recombinase to conditionally knockout MIF receptors CD74 and CXCR4 from specific neuronal populations in vivo and evaluate their contributions to MIF anti-nociception in a model of apical periodontitis. These studies will not only provide novel insight into the neuronal populations that mediate MIF-induced anti-nociception but also serve as an excellent training vehicle for my career as a clinician-scientist.