REGULATION OF IL-4 PRODUCTION AND IL-4-MEDIATED IMMUNITY

IL-4 产生和 IL-4 介导的免疫的调节

• 批准号: 2063655
• 负责人: DAVID BRAM LEWIS
• 金额: $ 15.33万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-07-01 至 1997-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2063655
• 关键词: B lymphocyte; DNA footprinting; Leishmania major; Nematoda; cellular immunity; delayed hypersensitivity; gel mobility shift assay; gene expression; genetic regulatory element; genetically modified animals; helminthiasis; helper T lymphocyte; human genetic material tag; human tissue; immunoglobulin E; immunologic memory; interleukin 2; interleukin 4; laboratory mouse; leishmaniasis; leukocyte activation /transformation; molecular cloning; protein biosynthesis; transcription factor

Differential cytokine production by CD4+ T cells is an important mechanism for selectively regulating immune effector function. The expression f the interleukin-4 (IL-4) and interleukin-2 (IL-2) genes is differentially regulated during the maturation of antigenically-naive, virgin T cells into memory cells. Virgin cells produce IL-2 but low or undetectable amounts of IL-4, while memory T cells may produce high levels of both cytokines. The first aim of this project is to determine the ability of increased IL-4 production by activated T cells in vivo to influence the outcome of the immune response. For this aim, PIL-2/IL-4 transgenic mice have been generated in which the IL-2 promoter drives transcription of an IL-4 transgene. This dramatically increases IL-4 expression by T cells, presumably by virgin an memory cells, and increases serum IgE levels, a hallmark of IL-4 activity, in an activation-dependent fashion. It is predicted that the transgene will also: 1) increase the bias of CD4+ T cell cytokine production, in bulk and at the clonal level, towards a Th2 pattern (increased production of endogenous-gene derived IL-4 and IL-5, decreased production of IL-2 and interferon-gamma), 2) decrease cell- mediated immunity, including delayed-type hypersensitivity and 3) enhance expression of IgE but not other isotypes. To test these predictions, PIL- 2/IL-4 mice will be challenged with antigens and the effect of the transgene on modifying isotype expression by B cells, antigen-specific delayed-type hypersensitivity, and endogenous cytokine gene expression by CD4+ T cells will be determined. In subsequent experiments, PIL-2/IL-4 mice and littermate controls will be challenged with pathogens that normally result in either protective Th2-type (H. polygyrus) or Th1-type (Leishmania major) responses. We will determine whether the transgene will alter cytokine production in vivo, IgE expression, and the burden of infection. The second aim of this project is to identify cis-elements in the promoter of the IL-4 gene that confer increased IL-4 gene expression by memory cells and 2) to identify selective differences in potential transcription factors binding to these elements, that differ between virgin and memory T cells, using gel-shift assays and in vivo footprinting. The in vivo relevance of cis-regulatory elements defined by in vitro assays will then be tested by generating mice transgenic for human IL-4 genomic constructs. We will test whether the cis-elements of the transgene will confer, in a manner analogous to the endogenous IL-4 gene, enhanced expression by in vitro-primed T cells, memory T cells, and by T cells in vivo after helminth challenge. These studies will provide substantial insight into the biologic significance of the differential regulation of IL-4 during in vivo immune responses, as well as the molecular mechanisms responsible for this regulation. They may also point out means by which endogenous IL-4 production might be inhibited, such as in IgE-mediated allergic disorders, or, alternatively, increased, as in cytokine therapy for cancer. These studies may also point out potential pitfalls of these alterations.

CD4+ T细胞产生差异细胞因子是重要的机制 用于选择性调节免疫效应子功能。 表达f 白介素-4（IL-4）和白介素-2（IL-2）基因是差异的 在抗原侵蚀性的，处女t细胞成熟期间受到调节 记忆单元。 维珍细胞产生IL-2，但低或不可检测的数量 IL-4，而记忆T细胞可能会产生高水平的两种细胞因子。 这 该项目的第一个目的是确定IL-4增加的能力 通过体内激活的T细胞生产以影响 免疫反应。 为此，PIL-2/IL-4转基因小鼠已经 IL-2启动子驱动IL-4的转录产生 转基因。 这大大增加了T细胞的IL-4表达， 大概是通过处女的记忆细胞，并增加了血清IgE水平，a IL-4活动的标志，以激活依赖性方式。 这是 预测转基因还将：1）增加CD4+ T的偏差 细胞细胞因子的产生，散装和克隆水平，朝向Th2 模式（内源性基因衍生的IL-4和IL-5的产生增加， IL-2和干扰素伽马的产生降低），2）降低细胞 - 介导的免疫力，包括延迟型超敏反应和3）增强 IgE的表达，而不是其他同种型。 为了测试这些预测， 2/IL-4小鼠将受到抗原的挑战和效果 通过B细胞修饰同种型表达的转基因，抗原特异性 延迟型高敏性和内源性细胞因子基因表达 CD4+ T细胞将被确定。 在随后的实验中，PIL-2/IL-4 小鼠和同窝窝控制将受到病原体的挑战 通常导致保护性Th2型（H. polygyrus）或Th1型 （利什曼尼亚专业）的回应。 我们将确定转基因是否会 改变体内细胞因子的产生，IgE表达和负担 感染。 该项目的第二个目的是确定在 IL-4基因的启动子通过赋予IL-4基因表达的启动子。 记忆单元和2）确定潜在的选择性差异 与这些元素结合的转录因子，而处女之间有所不同 和记忆T细胞，使用凝胶换档测定和体内足迹。 这 由体外测定定义的顺式调节元件的体内相关性 然后，将通过为人IL-4基因组生成小鼠转基因来测试 构造。 我们将测试转基因的顺式元素是否会 以类似于内源性IL-4基因的方式授予 通过体外提出的T细胞，记忆T细胞和T细胞在体外表达 蠕虫挑战之后的体内。 这些研究将提供大量 深入了解差异调节的生物学意义 体内免疫反应期间的IL-4以及分子机制 负责此法规。 他们也可能指出手段 内源性IL-4的产生可能被抑制，例如在IgE介导的 过敏性疾病，或替代增加，如细胞因子疗法 用于癌症。 这些研究也可能指出了这些研究的潜在陷阱 改变。