Stress Induces Skin Mast Cell Activation & Vasodilation

描述(申请人摘要)：肥大细胞位于血管周围较近的位置 神经突起，可以分泌许多血管活性物质和促炎性物质 分子。除了过敏的诱因，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 通过局部释放促肾上腺皮质激素(CRH)或 尿皮质激素(Ucn)，比CRH更有效。促肾上腺皮质激素释放激素皮内注射 UCN诱导大鼠皮肤肥大细胞活化和血管增多 渗透性，这两者都被中和预处理所抑制 CRH或CRH受体拮抗剂antalarmin的抗血清。CRH或急性 伊文思蓝检测应激诱导的皮肤血管通透性 W/W(V)肥大细胞缺陷小鼠无外渗，但存在外渗 在他们的wt对照中，表明它是肥大细胞依赖的；这一发现是 支持这一事实的是，血管的通透性也被“肥大 细胞稳定剂“色甘酸二钠(色甘宁)。类似的结果是 在大鼠和小鼠身上获得对急性制动应激的反应。入内 辣椒素处理的啮齿动物体内没有CRH作用，但不是在原位 耗尽感觉神经纤维中P物质(SP)的含量，也 被神经降压素(NT)受体拮抗剂SR48692抑制。我们是 假设急性应激导致皮肤释放CRH和/或UCN， 直接或通过神经肽(如SP和NT)来激活肥大细胞 并增加血管通透性；白细胞的渗透可能会导致 可能与局部炎症和迟发型超敏反应(DTH)有关。 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)皮肤肥大细胞激活的影响 分析，以及残留的皮肤组胺和小鼠肥大细胞蛋白酶 CRH基因敲除小鼠的(MMCP)-6含量，以及(B)血管通透性 CRH基因敲除中~(99)Tc外渗法测定CRH 和W/W(V)小鼠及其+/+对照；(2)皮肤背根的任何变化 应激后神经节(DRG)或脊髓CRH、UCN或CRH受体表达； 应用免疫组织化学、Western和Northern分析或RT-PCR；(3) P物质参与应激诱导的肥大细胞激活和血管生成 SP和NK-1受体基因敲除小鼠的通透性以及可能的 利用基因敲除动物和CRH结合的CRH或SP的作用顺序 或SP受体拮抗剂；(4)CRH和UCN对血管内皮细胞分泌的影响 从小鼠纯化的皮肤中分别提取组胺、IL-6和MMCP-6或类胰蛋白酶 和免疫刺激的人脐带来源的肥大细胞 沙棘属(SP.)这些研究将帮助我们理解急性压力如何触发皮肤肥大。 细胞活化，血管通透性增加，可能还有迟发性高血压。他们会 还有助于研究皮肤综合征的病理生理学，这些症状因 应激，如特应性皮炎或牛皮癣，特别是自CRH和CRH以来 已经在人类皮肤中发现了受体。

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.