Vulnerability to Smallpox Due to Declining CTL Immunity

描述（由申请人提供）：自1971年以来，停止接种天花疫苗导致美国普通人群获得性免疫力下降，从而增加了恐怖分子或不友好政府故意释放天花的重大流行病的风险。美国疾病控制与预防中心已经表明，牛痘免疫导致了世界范围内天花的根除，但在老年人中造成严重并发症和死亡率过高。由于老年人中许多其他病毒感染的较高发病率和死亡率归因于缺乏病毒特异性hla限制性CTL反应，因此预计老年人具有较低的牛痘特异性CTL活性，因此更容易患天花疾病以及牛痘免疫引起的并发症。本研究的目的是：1)描述具有代表性的美国年轻人和老年人人群中抗牛痘的血清患病率和记忆CD8+ CTL活性状态；2)确定易于检测和量化的CTL抗天花感染能力的替代物；3)探索牛痘疫苗接种并发症的免疫学基础。在美国出生的健康成人（35-49岁和50-64岁）和以前接种过牛痘的老年人（~65岁）的外周血淋巴细胞中，将测定牛痘特异性记忆CD8 CTL活性水平，并与最近（<1年）接种牛痘或未接种牛痘的年轻人（18-32岁）进行比较。测定病毒血清中和抗体和病毒特异性IgG抗体滴度，并与CTL活性相关。CD8+ T细胞表达干扰素(IFN)- γ （CTL替代品）和白细胞介素(IL)-4 （CTL拮抗剂）的频率将使用ELISPOT和流式细胞术进行检测。ifn - γ和IL-4的功能将使用反义寡核苷酸和/或抗体进行评估。将分析选定的促炎细胞因子和趋化因子（IL-1¿，TNF-a, IL-6， IL-8和il -13）。据推测，在先前接种过疫苗的人群中：a)与年轻人相比，老年人的CD8+ CTL记忆显著降低；b)牛痘特异性抗体水平和记忆CD8+ CTL活性将随着时间的推移而下降；c) ifn - γ的产生将是CTL活性的一个很好的替代指标；d)特异性促炎细胞因子或趋化因子的增加将与牛痘接种损伤的强度相关。研究结果将为公众，特别是老年人和其他免疫缺陷人群在发生天花生物攻击时重新接种疫苗的决定提供指导，并为老年人牛痘并发症的机制提供必要的见解。最近接种疫苗者的数据也可作为可能开发的新天花疫苗的参考值，因为目前还没有针对天花免疫的具体相关因素（疫苗“接种”除外）。通过将这些反应与新接种者的牛痘病变大小相关联，可以确定牛痘疫苗接种严重并发症的标志物。

DESCRIPTION (provided by applicant): Discontinuation of smallpox vaccination since 1971 has led to waning of acquired immunity in the U.S. general population, thus, raising the risk of major epidemics from intentional release of smallpox by terrorists or unfriendly governments. The CDC has shown that vaccinia immunization, which led to the eradication of smallpox worldwide, caused excess rates of severe complications and death among older persons. Since higher morbidity and mortality rates from many other viral infections in older persons have been attributed to a deficiency in virus-specific HLA-restricted CTL response, the elderly will be expected to possess reduced vaccinia-specific CTL activity and therefore be more susceptible to smallpox disease as well as complications from vaccinia immunization. The goals of this study are to: 1) profile the sero-prevalence and memory CD8+ CTL activity status against vaccinia in representative young adult and elderly U.S. populations, 2) identify easily detectable and quantifiable surrogates of CTL competence against smallpox infection and 3) explore immunological basis for complications from vaccinia vaccination. Levels of vaccinia-specific memory CD8 CTL activity will be determined in peripheral blood lymphocytes of U.S. born healthy adults (35-49 and 50-64 years old) and elderly adults (~65 years old) who have previously been vaccinated with vaccinia and compared with young adults (18-32 years old) who are recent (<1 year) vaccinia recipients or vaccinia naive. Virus serum neutralization antibody and virus specific IgG antibody titers will be determined and correlated with CTL activity. The frequency of CD8+ T cells expressing interferon (IFN)-gamma (a CTL surrogate) and interleukin (IL)-4 (a CTL antagonist) will be measured using ELISPOT and flow cytometry. The functions of IFN-gamma and IL-4 will be assessed using antisense oligonucleotides and/or antibodies. Selected pro-inflammatory cytokines and chemokines (IL-1¿, TNF-a, IL-6, IL-8 and lL-13) will be analyzed. It is postulated that among previously vaccinated persons: a) the elderly will have significantly reduced CD8+ CTL memory in comparison to younger persons, b) levels of vaccinia-specific antibody and memory CD8+ CTL activity will decline with time, c) IFN-gamma production will be a good surrogate of CTL activity and d) increases in specific pro-inflammatory cytokines or chemokines will be associated with the intensity of vaccinia vaccination lesions. The results should provide guidance in decisions to revaccinate the public, particularly the elderly and other immunodeficient persons, in the event of a biological attack with smallpox and offer needed insight into the mechanism of vaccinia complications in the elderly. Data from recent vaccinees might also serve as reference values for new smallpox vaccines that might be developed since specific correlates of immunity against smallpox (other than a vaccine "take") are currently not available. By correlating these responses with vaccinia lesion sizes in new vaccinees, markers of severe complications from vaccinia vaccination may be identified.