Stress Induces Skin Mast Cell Activation & Vasodilation

描述（申请人摘要）：肥大细胞位于血管周围， 神经过程，并能分泌许多血管活性和促炎 分子。除了过敏的触发因素，肥大细胞也可以被激活 通过直接神经刺激、神经肽和急性制动 通过促肾上腺皮质激素释放激素（CRH）的局部释放产生应激，或 尿皮质素（UCN），比CRH更有效。CRH皮内注射 或Ucn诱导大鼠皮肤肥大细胞活化， 渗透性，这两种抑制预处理与中和 CRH或CRH受体拮抗剂antalarmin的抗血清。CRH或急性 伊文思蓝法测定应激性皮肤血管通透性 在W/W（v）肥大细胞缺陷小鼠中不存在外渗，但存在 在他们的野生型对照中，表明它是肥大细胞依赖性的;这一发现是 支持这一点的事实是，血管渗透性也被“肥大细胞”阻断， 细胞稳定剂”色甘酸二钠（croglycate）。了类似的结果 在大鼠和小鼠对急性制动应激的反应中获得。的in 在用辣椒素处理的啮齿类动物中，CRH在体内没有作用，但在原位没有作用 耗尽感觉神经纤维的P物质（SP）含量， 被神经降压素（NT）受体拮抗剂SR 48692抑制。我们 假设急性应激在皮肤中释放CRH和/或Ucn， 直接或通过SP、NT等神经肽参与肥大细胞的活化 和血管通透性增加;白细胞浸润，然后可能有助于 局部炎症和可能的迟发型超敏反应（DTH）。 我们建议使用正常和遗传缺陷小鼠来研究：（1） 急性应激对（a）通过成像确定皮肤肥大细胞活化的影响 分析，以及残留的皮肤组胺和小鼠肥大细胞蛋白酶 CRH敲除小鼠中的（MMCP）-6含量，和（B）血管通透性 通过CRH敲除中的99锝-葡庚糖酸盐（99 Tc）外渗定量 和W/W（v）小鼠及其+/+对照;（2）皮肤背根的任何变化 神经节（DRG）或脊髓CRH，应激后Ucn或CRH受体表达， 采用免疫组化、Western和北方分析或RT-PCR;（3） SP参与应激诱导肥大细胞活化和血管生成 SP和NK-1受体敲除小鼠的通透性，以及可能的 使用敲除动物和CRH的组合的CRH或SP的作用顺序 或SP受体拮抗剂;（4）CRH和Ucn对分泌的影响， 分别来自小鼠纯化皮肤组胺、IL-6和MMCP-6或类胰蛋白酶 和人脐带来源的肥大细胞的免疫刺激或通过 这些研究将帮助我们了解急性应激如何触发皮肤肥大 细胞活化、血管通透性增加和可能的DTH。他们将 也有助于研究皮肤综合征的病理生理学， 应激，如特应性皮炎或银屑病，特别是由于CRH和CRH 已经在人皮肤中鉴定出受体。

DESCRIPTION (applicant's abstract): Mast cells are located perivascularly close to nerve processes and can secrete many vasoactive and proinflammatory molecules. In addition to allergic triggers, mast cells can also be activated by direct nerve stimulation, by neuropeptides, and by acute immobilization stress through the local release of corticotropin releasing hormone (CRH) or urocortin (Ucn), which is more potent than CRH. Intradermal injections of CRH or Ucn induced rat skin mast cell activation and increased vascular permeability, both of which were inhibited by pretreatment with neutralizing antiserum to CRH or the CRH-receptor antagonist antalarmin. CRH or acute stress-induced skin vascular permeability measured with Evans blue extravasation was absent in W/W (v) mast cell deficient mice, but was present in their wt controls indicating it is mast cell dependent; this finding was supported by the fact that vascular permeability was also blocked by the "mast cell stabilizer" disodium cromoglycate (cromolyn). Similar results were obtained in rats and mice in response to acute immobilization stress. The in vivo, but not in situ, CRH action was absent in rodents treated with capsaicin to deplete sensory nerve fibers of their substance P (SP) content and was also inhibited by the neurotensin (NT) receptor antagonist SR48692. We are hypothesizing that acute stress releases CRH and/or Ucn in the skin leading, directly or through neuropeptides such as SP and NT, to mast cell activation and increased vascular permeability; leukocyte infiltration may then contribute to local inflammation and possibly to delayed-hypersensitivity (DTH) reactions. We propose to use normal and genetically deficient mice to investigate: (1) the effect of acute stress on (a) skin mast cell activation determined by image analysis, as well as residual skin histamine and mouse mast cell protease (MMCP)-6 content in CRH knock-out mice, and (b) vascular permeability quantitated by 99Technicium-gluceptate (99Tc) extravasation in CRH knock-out and W/W (v) mice and their +/+ controls; (2) any change in skin dorsal root ganglia (DRG) or spinal cord CRH, Ucn or CRH receptor expression after stress, using immunohistochemistry, Western and Northern analysis or RT-PCR; (3) the involvement of SP on stress-induced mast cell activation and vascular permeability in SP and NK-1 receptor knock-out mice, as well as the possible sequence of action of CRH or SP using combination of knock-out animals and CRH or SP-receptor antagonists; (4) the effect of CRH and Ucn on secretion of histamine, IL-6 and MMCP-6 or tryptase, respectively, from mouse purified skin and human umbilical cord-derived mast cells stimulated immunologically or by SP. These studies will help us understand how acute stress triggers skin mast cell activation, increased vascular permeability and possibly DTH. They will also help investigate the pathophysiology of skin syndromes exacerbated by stress, such as atopic dermatitis or psoriasis, especially since CRH and CRH receptors have been identified in human skin.