The role of IL-4 and IL-4Ralpha in the multi-step process of Th2 development

描述(申请人提供)：土壤传播的寄生线虫是世界范围内最常见的多细胞寄生虫感染之一。人类和小鼠的适应性宿主反应的特点是存在CD4+Th2细胞及其标志性细胞因子IL-4。Th2细胞和IL-4与针对寄生虫的保护性免疫反应有关，它们在某些免疫病理过程中是有害的，如抗原诱导的哮喘反应、特应性和变态反应。虽然Th2细胞与线虫感染有关，但目前尚不清楚Th2细胞是如何从原始的CD4+T细胞发展成对感染的反应的。大量研究表明，IL-4ra介导的信号和IL-4在体外对初始的CD4+T细胞的Th2发育是必需的，但在体内对这些过程知之甚少。这在很大程度上是因为很难识别和跟踪由IL-4表达定义的Th2细胞。为了克服这些限制，我们开发了双顺反子IL-4报告(4get)小鼠。与体外研究相比，我们的初步数据显示，Th2启动在感染小鼠蠕虫H.多回的IL4Ra-/-4get小鼠中发生了令人惊讶的有效。因此，IL-4R和IL-4在Th2形成中的作用是有争议的。最近，我们创造了新的IL-4双报告小鼠(4get/kn2)，并揭示了IL-4的活性和IL-4的产生是不同的步骤。在这里，我们提出了一个新的模型，在这个模型中，Th2的发展和IL-4的产生分几个不同的、可识别的步骤进行：1.激活，2.分化，3.扩张，4.IL-4产生，5.传播。在目前的应用中，我们将使用我们独特的双报告小鼠模型重新研究IL-4和IL-4ra介导的信号在Th2分化和IL-4产生中的作用。我们假设，IL-4R功能不是启动CD4+T细胞Th2启动所必需的，而是在该模型的其他多个步骤中发挥作用。在目标1中，我们将分析内源性T细胞群Th2分化的哪一步(S)受到IL-4R信号的调节，以应对感染。在目标2中，我们将确定Th2分化的哪一步(S)受IL-4R信号直接介导到初始的、抗原特异性的CD_4+T细胞上调节。我们将过继转移apTCR转基因的CD4+T细胞，并以同源抗原作为Th2佐剂免疫多回幼虫。在目标3中，我们将通过选择性谱系分析IL-4和IL-4ra的表达在Th2多步分化过程中的作用。我们将产生各种骨髓嵌合体，这些嵌合体在选择性细胞谱系中缺乏各自的成分，并跟踪内源性T辅助细胞对感染的反应的发展。总之，提出的目标将揭示IL-4R和IL-4在体内调节Th2发育的多个步骤的机制。这些见解对于设计针对IL-4R和IL-4的激动型和拮抗型干预策略是有价值的。

DESCRIPTION (provided by applicant): Soil transmitted parasitic nematodes are world wide one of the most commonly acquired infections with multi-cellular parasites. The adaptive host response in humans and mice is characterized by the presence of CD4+ Th2 cells and their signature cytokine IL-4. Th2 cells and IL-4 are associated with protective immune responses against helminth parasites they are detrimental in certain immunopathologies such as antigen-induced asthmatic reactions, atopy and allergy. While Th2 cells are associated with nematode infections; however, it is not clear how Th2 cells develop from naive CD4+ T cells in response to infection. Numerous studies have shown that IL-4Ra-mediated signals and IL-4 are required for the Th2 development of naive CD4+ T cells in vitro, however, very little is known about these processes in vivo. This is largely due to the difficulty to identify and track Th2 cells defined by IL-4 expression. To overcome these limitations we have developed bicistronic IL-4 reporter (4get) mice. In contrast to in vitro studies, our preliminary data show that Th2 priming occurs surprisingly efficient in IL4Ra-/- 4get mice infected with the murine helminth H. polygyrus. Thus, the role of IL-4R and IL-4 for Th2 development is controversial. Recently we have generated novel IL-4 dual-reporter mice (4get/KN2) and revealed that IL-4 competence and IL-4 production are distinct steps. Here we propose a novel model whereby Th2 development and IL-4 production occur in several distinct, identifiable steps: 1. activation, 2. differentiation, 3. expansion, 4. IL-4 production, 5. dissemination. In the current application we will revisit the role of IL-4 and IL-4Ra-mediated signals for Th2 differentiation and IL-4 production using our unique dual-reporter mouse model. We hypothesize that IL-4R functions are not required to nucleate the Th2 priming of CD4+ T cells but play a role in multiple other steps of this model. In Aim 1 we will analyze which step(s) in Th2 differentiation of the endogenous T cell population are regulated by IL-4R signals in response to infection. In Aim 2 we will determine which step(s) in Th2 differentiation are regulated by IL-4R signals mediated directly on naive, antigen-specific CD4+ T cells. We will adoptively transfer apTCR transgenic CD4+ T cells and immunize with the cognate antigen using H. polygyrus larvae as a Th2 adjuvant. In Aim 3 we will dissect the role of IL-4 and IL-4Ra expression by selective lineages in the multi-step process of Th2 differentiation. We will generate various bone marrow chimeras which lack the respective components in selective cellular lineages and follow the development of the endogenous T helper response to infection. Collectively the proposed Aims will reveal the mechanism by which IL-4R and IL-4 regulate the multiple steps of Th2 development in vivo. These insights are valuable for designing agonistic and antagonistic intervention strategies targeting the IL-4R and IL-4.