Natural Killer Dendritic Cells in Listeria Infection

描述(申请人提供)：虽然人们普遍认为干扰素-伽马(干扰素-伽马)对清除单核细胞增多性李斯特菌至关重要，但干扰素-伽马的细胞来源一直存在争议。一些研究人员得出结论，NK细胞是罪魁祸首，而另一些人则与T细胞有关。最近，我们发现了一种新的细胞类型，它可以分泌大量的干扰素-γ，并且存在于正常小鼠的脾和其他器官中。由于这些独特的细胞直接杀伤靶细胞，在体外和体内激活初始T细胞，并同时表达自然杀伤细胞标志NK1.1和树突状细胞标志CD11c，我们将其命名为自然杀伤树突状细胞(NKDC)。其他一些研究人员在啮齿动物和人类中发现了NKDC样细胞。我们已有初步数据表明，在小鼠感染李斯特氏菌的过程中，NKDC会产生干扰素-γ。因此，我们推测NKDC在李斯特菌的免疫应答中起关键作用。虽然NKDC的裂解和抗原提呈能力可能很重要，但我们选择只关注他们在这两年的应用中产生的干扰素-γ。我们建议评估NKDC在小鼠感染李斯特氏菌过程中产生干扰素-γ的生物学意义，并剖析其调节机制。在目标1中，我们将确定NKDC在李斯特菌感染过程中参与干扰素-γ产生的程度。为了排除其他细胞在体内产生干扰素-γ的可能性，但一旦它们被从脾内的局部环境中移除，就无法做到这一点，我们将利用最近描述的一种体内脾细胞细胞内细胞因子分析技术。在目标2中，我们将建立NKDC在李斯特菌感染过程中产生干扰素-γ的机制。具体地说，我们将测试IL-12、IL-18和Toll样受体9，因为我们已经发现它们可以调节NKDC对细菌CpG的干扰素-γ的产生。在目标3中，我们将确定NKDC产生的干扰素-γ是否足以诱导对李斯特氏菌的免疫反应。我们将过继地将NKDC从野生型小鼠转移到干扰素-γ缺陷小鼠，然后评估它们对李斯特菌感染的反应。NKDC将与NK细胞和目标1中确定的产生干扰素-伽马的其他脾细胞亚群进行比较。我们拟议的实验将开始确定NKDC在李斯特菌免疫反应中的作用，李斯特菌是一种导致人类死亡的病原体，也是潜在的生物恐怖主义因素，并提供对这种新细胞类型如何参与其他类型免疫的更多了解。

DESCRIPTION (provided by applicant): While there is general consensus that interferon-gamma (IFN-gamma) is vital to the clearance of Listeria monocytogenes, the cellular source of IFN-gamma has been controversial. Some investigators have concluded that NK cells are responsible, while others have implicated T cells. Recently, we have identified a novel cell type that secretes large amounts of IFN-gamma and exists in the spleen and other organs of normal mice. Because these unique cells lyse targets directly, activate naive T cells in vitro and in vivo, and express both the natural killer cell marker NK1.1 and the dendritic cell marker CD11c, we have labeled them as natural killer dendritic cells (NKDC). A few other investigators have recognized NKDC-like cells in rodents and humans. We have preliminary data that splenic NKDC produce IFN-gamma during Listeria infection in mice. Therefore, we hypothesize that NKDC are critical to the immune response against Listeria. While the lytic and antigen-presenting abilities of NKDC may be important, we have chosen to focus exclusively on their IFN-gamma production for this 2-year application. We propose to assess the biologic significance of IFN-gamma production by NKDC during Listeria infection in mice and dissect the mechanisms by which it is regulated. In Aim 1, we will determine the extent of NKDC involvement in IFN-gamma production during Listeria infection. To exclude the possibility that other cells make IFN-gamma in vivo, but are unable to do so once they are removed from their local environment in the spleen, we will make use of a recently described technique of in vivo intracellular cytokine analysis of splenocytes. In Aim 2, we will establish the mechanism of IFN-gamma production by NKDC during Listeria infection. Specifically, we will test IL-12, IL-18, and Toll-like receptor 9 as we have found these to regulate NKDC IFN-gamma production in response to bacterial CpG. In Aim 3, we will determine whether NKDC production of IFN-gamma is sufficient to induce an immune response against Listeria. We will adoptively transfer NKDC from wild-type mice into IFN-gamma deficient mice and then assess their response to Listeria infection. NKDC will be compared to NK cells and other splenic cell subsets that make IFN-gamma as identified in Aim 1. Our proposed experiments will begin to define the role of NKDC in the immune response to Listeria, a pathogen that is a cause of human mortality and a potential agent for bioterrorism, and provide a greater understanding of how this novel cell type may participate in other types of immunity.