Innate Immunity to Influenza in Caloric Restricted Aged Mice

描述(申请人提供)：流感及其继发性肺炎是美国65岁及以上人群的第四大死因。与随意喂养(AL)相比，限制卡路里(CR)可延长健康啮齿动物的中位寿命和最长寿命。老年CR啮齿动物的肿瘤和癌症发生率降低，抗体滴度和T细胞增殖增加，提示CR延缓了年龄相关性免疫功能下降的发生。我们使用了一种小鼠CR模型来研究与年龄相关的对流感的主要反应下降。尽管CR延缓了与年龄相关的T细胞增殖的下降，但与之形成鲜明对比的是，老年CR小鼠在初次流感感染后4-6天死亡，表现出肺病毒滴度增加和肺NK活性降低。重要的是，感染时CR小鼠的体重比AL小鼠轻30%，并且在感染后体重显著下降。由于早期的时间进程，我们假设老年CR小鼠由于先天免疫改变而不能控制原发流感感染。这可能反映了NK细胞本身的内在缺陷，也可能继发于涉及巨噬细胞和/或树突状细胞早期产生的信号的外在缺陷。具体来说，我们将：1)确定CR小鼠对初级流感感染的敏感性增强是否与年龄有关。年轻的AL和CR C57BL/6小鼠将感染流感，并对存活、体重减轻和恢复、肺和纵隔淋巴结的细胞类型以及肺病毒滴度进行评估。2)探讨初发流感时NK细胞活性降低的机制(S)。我们将通过细胞毒性、穿孔素和颗粒酶的产生、干扰素对NK细胞的体外刺激以及细胞因子的产生来评估NK功能。我们将研究NK细胞活性降低是否与NK细胞数量减少或NK细胞占肺内淋巴细胞总数的百分比有关。我们将进一步确定NK细胞数量或功能的缺陷是否是由于巨噬细胞/树突状细胞产生的细胞因子损害了NK细胞的募集和/或激活。我们将对肺内巨噬细胞和树突状细胞的活化标志物及其细胞因子和趋化因子的产生(干扰素-α、IL-12、IL-15、IL-18、MIP-1a/β、MCP-1、IL-1、IL-6、TNF-α)进行定量检测。3)确定CR小鼠对流感的易感性是否与体重减轻有关，以及在感染前重新喂养是否恢复了对流感感染的免疫反应。在感染流感之前，将喂食AL饮食，使AL小鼠的体重恢复到AL小鼠的50%和100%，并评估特定目标1和2的结果。

DESCRIPTION (provided by applicant): Influenza and its secondary pneumonias are the fourth leading cause of death in persons 65 years and older in the United States. Caloric restriction (CR) extends median and maximal life span in healthy rodents, compared to those fed ad-libitum (AL). Aged CR rodents show decreases in tumors and cancers, and increases in antibody titers and T cell proliferation suggesting CR delays the onset of age-related decreased immune function. We have employed a mouse model of CR to examine the age-related decline in primary response to influenza. Although CR delayed the age-related decline in T cell proliferation, in stark contrast, aged CR mice died 4-6 days after primary influenza infection, exhibiting increased lung virus titers and reduced pulmonary NK activity. Importantly, CR mice weighed 30% less than AL mice at the time of infection and had dramatic weight loss following infection. Due to the early time course, we hypothesize that aged CR mice cannot control primary influenza infection because of altered innate immunity. This may reflect an intrinsic defect in NK cells themselves or may be secondary to an extrinsic defect involving signals produced early on by macrophages and/or dendritic cells. Specifically, we will: 1) Determine if enhanced susceptibility of CR mice to primary influenza infection is age-dependent. Young AL and CR C57BL/6 mice will be infected with influenza and survival, weight loss and recovery, cell types in lung and mediastinal lymph nodes, and lung virus titers will be evaluated. 2) Investigate mechanism(s) for reduced NK activity during primary influenza infection. We will assess NK function by cytotoxicity, perforin and granzyme production, in vitro stimulation of NK cells with IFN-a/¿, and cytokine production. We will investigate whether decreased NK cell activity is related to decreased NK cell number or NK cells as a percentage of total lymphocytes in lung. We will further determine if the defect in NK number or function results from impaired recruitment and/or activation of NK cells by cytokines produced by macrophages/dendritic cells. We will quantitate activation markers on macrophage and dendritic cells in lung and their cytokine and chemokine production (IFN-a/¿?, IL-12, IL-15, IL-18, MIP-1a/¿, MCP-1, IL-1¿, IL-6, TNF-a). 3) Determine if the susceptibility of CR mice to influenza is related to reduced body weight and if refeeding prior to infection restores the immune response to influenza infection. CR mice will be fed AL diets to restore weight to 50% and 100% of AL mice before infection with influenza and outcomes in Specific Aims 1 and 2 will be assessed.