Generation of a Cre-LoxP mouse line expressing hCXCR1

表达 hCXCR1 的 Cre-LoxP 小鼠系的生成

• 批准号: 7048767
• 负责人: MANUELA M. MARTINS-GREEN
• 金额: $ 7.49万
• 依托单位: UNIVERSITY OF CALIFORNIA RIVERSIDE
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-02-15 至 2008-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7048767
• 关键词: chemokine; genetically modified animals; human; inflammation; laboratory mouse; receptor; recombinase; role; tissues

DESCRIPTION (provided by applicant): The overall objective of this project is to generate a transgenic mouse line that expresses the human CXCR1 gene in a Cre-recombinase-driven tissue-specific manner as a tool to study CXCR1-dependent chemokine functions. Chemokines are important molecules playing many roles in physiological and pathological conditions, lnterteukine-8 is one of the chemokines that has been intensively studied because it functions in inflammation, wound healing, angiogenesis and tumorigenesis. Much work has been performed to elucidate the functions of this chemokine but progress has been impaired by the lack of an animal model system that can be genetically manipulated. In humans, IL-8 interacts with two receptors, CXCR1 and CXCR2, hence in order to determine how IL-8 functions it is necessary to study the role of each of these receptors in response to activation by hlL-8. These receptors and r|L-8 are highly homologous to the human proteins and rabbits express both receptors. However, it is very difficult (if at all possible) and expensive to perform gene manipulations in rabbits. Mice, on the other hand, do not have the CXCR1 gene or IL-8, hence cannot be used to fully study the functions of IL-8 in humans. We propose here a plan to create a human CXCR1 transgenic mouse line that can serve as an in vivo experimental system to elucidate the functions of IL-8. The specific aims of the work are: 1) To build and test the DMA construct for the transgenic animal. 2) To produce the founder transgenic mice. 3) To test the transgenic mouse line for tissue-specific Cre- recombinase activation. We will first generate a LoxP-based conditional transgenic founder line in which hCXCRI expression is suppressed in all tissues of the mouse. To do so, a DNA transgenic construct will be prepared by molecular cloning methods, tested in a cell culture system to confirm its functionality, including the transgene hCXCRI expression, and Cre-recombinase mediated recombination. We will then supply the DNA transgenic construct to Xenogen Biological that will create the founder transgenic mice. Finally, we will acquire specific Cre-recombinase expressing mice and cross them with our founder mice to achieve hCXCRI transgene activation in the skin to test the functionality of the mouse transgenic line in vivo. The mice expressing hCXCRI in specific tissues will serve as powerful tools to perform research related to human interieukin-8 because delivered hlL-8 will be able to interact with hCXCRI and mCXCR2 (already shown to occur) in the pertinent tissues, providing insight into the molecular mechanisms of the functions of IL-8 in humans. In general, the transgenic animal we are proposing to build will be a useful tool that brings versatility to our research and those of others for understanding the functions of hlL-8.

描述（由申请人提供）：该项目的总体目的是生成一种转基因小鼠系，该系列以CRE聚合酶驱动的组织特异性方式表达人CXCR1基因，作为研究CXCR1依赖性趋化因子函数的一种工具。趋化因子是重要的分子在生理和病理条件下起许多作用，lnteukine-8是经过深入研究的趋化因子之一，因为它在炎症，伤口愈合，血管生成和肿瘤发生中起作用。已经进行了许多工作来阐明这种趋化因子的功能，但是由于缺乏可以通过遗传操纵的动物模型系统而损害了进步。在人类中，IL-8与两个受体CXCR1和CXCR2相互作用，因此为了确定IL-8功能如何研究这些受体在HLL-8激活中的作用。这些受体和R | L-8与人蛋白质高度同源，而兔子表达这两个受体。但是，在兔子中进行基因操作非常困难（如果可能的话），并且昂贵。另一方面，小鼠没有CXCR1基因或IL-8，因此不能用来充分研究人类IL-8的功能。我们在这里提出了一个计划，以创建人类CXCR1转基因小鼠系，该系列可以用作体内实验系统，以阐明IL-8的功能。该作品的具体目的是：1）为转基因动物构建和测试DMA构建体。 2）产生创始人的转基因小鼠。 3）测试转基因小鼠系的组织特异性重组酶活性。我们将首先生成一个基于LOXP的条件转基因创始人线，其中HCXCRI表达在小鼠的所有组织中都受到抑制。为此，将通过分子克隆方法制备DNA转基因构建体，该方法在细胞培养系统中进行了测试，以确认其功能，包括转基因HCXCRI表达和Cre-Recombinase介导的重组。然后，我们将提供DNA转基因构建体，以创建创始人转基因小鼠。最后，我们将获取表达小鼠的特定CRE聚合酶，并与我们的创始人小鼠交叉，以实现皮肤中的HCXCRI转基因激活，以测试体内小鼠转基因线的功能。在特定组织中表达HCXCRI的小鼠将用作执行与人Interieukin-8相关的研究的强大工具，因为递送的HLL-8将能够与HCXCRI和MCXCR2相互作用（已经显示出在相关的组织中发生），从而洞悉IL-8 IN-8 INSMANS INSMANS IL-8功能的分子机制。通常，我们建议建造的转基因动物将是一个有用的工具，它为我们的研究和其他人的研究带来了多功能性，以理解HLL-8的功能。