A population of Regulatory T cells controls nociception through enkephalins to restrain skin inflammation [plate scRNA-seq]

The skin, as a principal barrier tissue, integrates diverse signals discerned by sensory neurons and by immune cells to elicit adaptive responses to a wide range of stresses. While nociceptors, sensory neurons that initiate the sensation of pain, can mobilize and amplify the function of innate and adaptive immune effector cells, regulatory T (Treg) cells, a T cell subset expressing the transcription factor Foxp3, suppress systemic and local inflammatory responses. Considering the connectivity between nervous and immune systems for sensing and responding to damaging insults, we questioned whether Treg cells can modulate proinflammatory neuronal activity by acting directly on peripheral neurons. We found that Treg cells localized near axons innervating the skin and that short-term inactivation or ablation of Treg cells increased neuronal activation to noxious stimuli in a manner separable from their immunosuppressive function. Furthermore, a population of skin Treg cells was highly enriched for expression of Penk, the gene encoding the neuropeptide precursor for endogenous opioids enkephalins, which inhibit pain perception by inducing analgesia. Acute depletion of Penk expressing Treg cells, or inducible ablation of Penk in Treg cells increased neuronal activation in response to noxious stimuli and associated inflammation. Our study suggests that a population of Treg cells residing at barrier tissues exhibit a direct neuromodulatory activity opposing exacerbated activation of sensory neurons, at least in part through the production of an endogenous opioids, and by acting as an immune component of the neuro-immune axis, dampen responses to noxious pain stimuli and local inflammation. Mice were anesthetized with 2.5% isofluorane and treated with 5% Imiquimod (Perrigo) or vehicle control, applied topically to dorsal and ventral sides of ear skin. Mice (2 IMQ, 2 VEH) were treated starting on day 0 three times and euthanized for sorting on day 3.

皮肤作为人体主要的屏障组织，可整合感觉神经元与免疫细胞所感知的各类信号，从而针对多种应激源触发适应性应答。尽管作为引发疼痛感知的感觉神经元——伤害性感受器（nociceptors）——能够动员并增强固有免疫与适应性免疫效应细胞的功能，表达转录因子Foxp3的T细胞亚群即调节性T（Treg）细胞，可抑制全身性与局部性炎症应答。鉴于神经系统与免疫系统在感知并应对损伤性刺激时存在交互联系，我们提出疑问：调节性T细胞能否通过直接作用于外周神经元，来调控促炎性神经元活动？我们发现，调节性T细胞定位于皮肤神经支配轴突附近；且短期灭活或消融调节性T细胞，会以独立于其免疫抑制功能的方式，增强神经元对伤害性刺激的激活反应。此外，皮肤调节性T细胞群体中，高度富集编码内源性阿片肽（endogenous opioids）前体脑啡肽（enkephalins）的基因Penk的表达；脑啡肽可通过诱导镇痛作用来抑制疼痛感知。急性清除表达Penk的调节性T细胞，或诱导性消融调节性T细胞中的Penk基因，均会增强神经元对伤害性刺激及相关炎症的激活反应。本研究表明，定居于屏障组织的一类调节性T细胞，可通过直接发挥神经调节活性来对抗感觉神经元的过度激活——这一作用至少部分依赖于内源性阿片肽的产生；同时作为神经-免疫轴的免疫组分，这类细胞可减弱机体对伤害性疼痛刺激与局部炎症的应答。实验中，小鼠以2.5%异氟烷（isofluorane）麻醉，随后将5%咪喹莫特（Imiquimod，Perrigo公司）或溶剂对照涂抹于耳部皮肤的背侧与腹侧。于第0日开始对小鼠（2只咪喹莫特组、2只溶剂对照组）进行连续3次处理，并于第3日处死小鼠以进行细胞分选。