2013-08-012024-05-13https://scholars.lib.ntu.edu.tw/handle/123456789/644123摘要：異位性皮膚炎是一種常見的慢性發炎性皮膚疾病。此病和氣喘、過敏性鼻炎，均為具有異位性體質者，才能引發的免疫疾病。異位性疾病的過敏原皆為環境中普遍存在的蛋白質抗原，其中有許多過敏原具有酵素活性。蛋白質抗原經皮致敏，已被証實為異位性皮膚炎重要的致敏途徑之ㄧ。我們於十幾年前建立小鼠持續性蛋白質貼膚模型(貼膚五天)，來研究”蛋白質抗原經皮致敏的免疫反應及機制”。發現在 BALB/c 和 B6 小鼠，蛋白質抗原持續經皮致敏引發顯著的第二型 T 細胞反應，但所引發的第一型 T 細胞反應很弱。最近，本研究室發展出小鼠非持續性蛋白質貼膚模型 (貼膚一天)，發現在 BALB/c小鼠中，仍引發顯著的 Th2 型反應，但在 B6 小鼠中，可引發強烈的Ｔh1 反應，且其Th2 反應較 BALB/c 小鼠更高。 直接主導 T 細胞亞型分化方向的是樹突狀細胞。樹突狀細胞的性狀 (成熟度、共同刺激因子細胞素)，和其主導 Th1/Th2/Th17 的分化，直接相關。對第一型 T 細胞的分化而言，樹突狀細胞所產生的 IL-12，是最重要的決定因子。對第二型 T 細胞的分化而言，因為樹突狀細胞無法產生 IL-4，故長久以來，學者們一直在尋找可以快速供給 IL-4 的固有性免疫細胞。嗜鹼性淋巴球，因可以在被刺激後，快速釋出大量 IL-4，而一直被懷疑可能是第二型 T 細胞反應的早期 IL-4 供應者。近幾年來，關於嗜鹼性淋巴球的了解大有進步，目前已知在某些第二型 T 細胞反應的小鼠模型中，尤其是蛋白質抗原具酵素活性者，嗜鹼性淋巴球會被召入淋巴結，釋放出 IL-4 和 TSLP 來幫助第二型 T 細胞的分化。甚至在某些小鼠模型中，嗜鹼性淋巴球可以做為抗原呈獻細胞。在常用的實驗小鼠中，學者早就發現 BALB/c 和 B6 小鼠，分別易於引發第二型和第一型 T 細胞反應。但是嗜鹼性淋巴球的質和量，對引發這兩種小鼠的 Th2 反應的影響，則尚未有報告。 本研究將利用非持續性小鼠貼膚模型，來研究嗜鹼性淋巴球，在 BALB/c 和 B6 小鼠的蛋白質抗原經皮致敏，所引發的 Th2 反應中的角色。我們將使用三種抗原: ovalbumin, papain (具酵素活性)和 heat-inactivated papain (不具酵素活性)。之前的實驗結果已知，ovalbumin, papain 和 heat-inactivated papain 貼膚致敏後，其局部淋巴結內 basophil 的數目會增加，在本實驗計畫中，我們將利用生物體內去除或增加嗜鹼性淋巴球的方法，以及測量淋巴結內樹突狀細胞的性狀及細胞激素和細胞素的變化，來達成兩項特定目標: 特定目標一: 確立 basophil 有助於蛋白質抗原經皮致敏所引發的 Th2 免疫反應。特定目標二: 釐清 basophil 幫助引發 Th2 免疫反應的機制。 計劃主持人長期致力於「異位性皮膚炎致病機轉」的研究，尤其著重在「蛋白質抗原經皮致敏的機制」，此計劃為此長期研究的一部分，我們期待這些研究所得到的資料，最終能幫助我們發展出避免過敏原致敏的方法或疫苗，使這些高危險群的兒童，能免受異位性疾病之苦。<br> Abstract: Atopic diseases are clinically common and important disorders. The known allergens of atopic diseases are all universally existing environmental protein antigens. Many of them posses protease activity. Epicutaneous sensitization has been proven to be one of the most important sensitization routes for atopic dermatitis. Using a murine continuous protein-patch model (five-day patch application), which we established over one decade ago, we and other investigators have demonstrated that epicutaneous sensitization with protein antigen induces predominant Th2, low Th17, and weak Th1 immune responses in BALB/c and B6 mice. Recently, our lab developed a murine non-continuous protein-patch model (one-day patch application) and observed that although BALB/c mice showed a predominant Th2 and weak Th1 response, B6 mice showed both strong Th1 and Th2 response. During priming, dendritic cells instruct naïve T cells to differentiate into Th1, Th2 or Th17 cells. Characters of DCs (maturation status, costimulator expression and cytokine production) have been proved to be crucial for their Th instructing capabilities. For Th1 differentiation, IL-12 produced by dendritic cells is a key determinator. For Th2 differentiation, due to the inability of dendritic cells to produce IL-4, it has been a long-term pursuit to search for early IL-4 provider in immune system. Basophils are considered to be candidates due to their capability to release large amount of IL-4 rapidly after activation. Recently, the role of basophils in initiating Th2 responses has been emphasized. Studies showed that basophils could be recruited into draining lymph nodes and secrete IL-4 and TSLP to initiate Th2 differentiation in some experimental systems especially when using protease as an antigen. Moreover, in some model systems, basophils function as antigen presenting cells to initiate Th2 differentiation. In murine experimental system, BALB/c and B6 mice have long been demonstrated to be Th2- and Th1-prone mice respectively. However, the contributions of basophils to the development of Th2 response in these two kinds of mice have never been explored. In this project, we will use murine non-continuous protein-patch model (one-day patch application) to investigative the roles of basophil in development of Th2 response by epicutaneous sensitization with protein antigen in BALB/c and B6 mice. We will use three kinds of antigen: ovalbumin, papain (possessing enzyme activity), heat–inactivated papain (no enzyme activity). In the preview study, it has been demonstrated that after epicutaneous sensitization with ovalbumin, papain or heat-inactivated papain , the numbers of basophils in draining lymph nodes increased. In this study, in vivo depletion or promotion of basophils as well as measurement of change of characters of DC subsets and expression levels of cytokine and chemokine in regional lymph nodes will be performed to achieve the following specific aims: 1. To confirm the role of basophil in epicutaneously-induced Th2 response. 2. To elucidate the mechanisms of initiating Th2 response with the help of basophil. We have focused on the research of “pathogenesis of atopic dermatitis “especially on” mechanisms of epicutaneous sensitization with protein antigens” for many years. This project is a part of our long-term pursuit. We hope that the results from this project will help us to understand the mechanisms of allergen sensitization and finally we could develop useful strategy or vaccines to prevent atopic children from developing atopic diseases.蛋白質抗原經皮致敏嗜鹼性淋巴球第二型 T 細胞反應protein antigenepicutaneous sensitizationbasophilTh2 response