LIFETIME OF EFFECTOR B CELLS

效应 B 细胞的寿命

• 批准号: 8362764
• 负责人: Thomas Dorner
• 金额: $ 2.34万
• 依托单位: UNIVERSITY OF CALIF-LAWRNC LVRMR NAT LAB
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-06-01 至 2012-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8362764
• 关键词: Age; Antibodies; Antigens; B-Lymphocytes; Blood; Cells; DNA; Development; Elements; Ensure; Europe; Funding; Grant; Half-Life; Human; Human body; Immune; Immune system; Immunoglobulin-Secreting Cells; Infection; Isotopes; Longevity; Measures; Memory; Memory B-Lymphocyte; Methodology; Methods; National Center for Research Resources; Play; Population; Principal Investigator; Protein Binding; Protocols documentation; Research; Research Infrastructure; Resource Development; Resources; Role; Sample Size; Sampling; Serum; Source; Staging; Tissues; United States National Institutes of Health; Vaccination; Vaccines; accelerator mass spectrometry; base; cell type; cost; fighting; in vivo; insight; pathogen; prevent

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Human body integrity is largely based on a healthy immune system fighting pathogens. One key feature of the immune system is expression of memory, i.e. maintaining the ability to fight a pathogen upon its re-encounter effectively, even after years of pathogen absence. Based on that, vaccinations can prevent from severe infections. In pathogen defense, proteins binding specifically to the pathogen (antibodies) play a vital role. Antibodies tag the pathogen and thus cause highly efficient degradation of the pathogen by specialized immune cells or neutralize it. The physical half-life of antibodies is less than 4 weeks. By contrast, specific antibodies induced by vaccination or infection can be readily detected in human blood serum after decades of antigen absence (humoral memory). This suggests that serum antibody levels are regulated at the stage of antibody secreting cells, and their specific precursors (memory B cells). For both cell types, longevity vs. continuous renewal is discussed as means to maintain the entire cell population and its composition, but both concepts were neither proven nor disproven physically. In this project we want to measure 14C content in the DNA of antibody-secreting cells and memory B cells obtained from donors exposed to elevated levels of atmospheric 14C in Europe during the 1960s. Elevated levels of the 14C isotope can still be detected in cells being quiescent since then, revealing their age. 14C analysis will require accelerator mass spectrometry (AMS) ensuring a precise quantitation of very low levels of 14C from minimum DNA samples. In analogy to Spalding et al (Cell, Vol. 122, 133143, July 15, 2005), this analysis will determine the in vivo age of candidate subsets of antibody-secreting cells and memory B cells and prove or disprove longevity of these cells, arguing in favor or against B cell longevity as a key element of humoral memory. The expected results will define new requirements for the successful development of effective vaccines. AMS will be used in this project to determine the content of 14C in DNA samples isolated from immune cells, thereby revealing their average age. AMS is indispensable as the cell populations to analyze are very rare, thus yielding very small DNA sample size. According to previous studies, the concentration of the 14C within the sample is again very low. Comparable analysis has already been performed at the LLNL and we believe that close cooperation with the resource is necessary to conduct a successful project. Except for the very small sample size, no additional development of methodology appears to be required. The results from 14C analyses will be combined with different results derived from various immunological methods to provide new insight in functionally different effector B cells and mechanisms of humoral memory. AMS results are a vital part of the whole study. Human blood and tissue will be obtained and specific cells will be extracted. From these cells, DNA will be isolated using established protocols compatible with latter 14C quantitation. DNA will be sent to the resource for 14C analysis.

该子项目是利用资源的众多研究子项目之一 由 NIH/NCRR 资助的中心拨款提供。子项目的主要支持 并且子项目的首席研究员可能是由其他来源提供的， 包括其他 NIH 来源。 子项目可能列出的总成本 代表子项目使用的中心基础设施的估计数量， NCRR 赠款不直接向子项目或子项目工作人员提供资金。 人体的完整性很大程度上取决于对抗病原体的健康免疫系统。免疫系统的一个关键特征是记忆的表达，即即使在病原体消失多年后，仍能在再次遇到病原体时保持有效对抗病原体的能力。基于此，接种疫苗可以预防严重感染。在病原体防御中，与病原体特异性结合的蛋白质（抗体）发挥着至关重要的作用。抗体标记病原体，从而通过专门的免疫细胞高效降解病原体或中和它。抗体的物理半衰期小于4周。相比之下，在抗原缺失数十年后（体液记忆），疫苗接种或感染诱导的特异性抗体可以很容易地在人血清中检测到。这表明血清抗体水平在抗体分泌细胞及其特定前体（记忆B细胞）阶段受到调节。对于这两种细胞类型，长寿与持续更新被讨论为维持整个细胞群及其组成的手段，但这两个概念既没有在物理上得到证实也没有被反驳。 在这个项目中，我们想要测量抗体分泌细胞和记忆 B 细胞 DNA 中的 14C 含量，这些细胞和记忆 B 细胞取自 20 世纪 60 年代欧洲暴露于高浓度大气 14C 的捐赠者。从那时起，在静止的细胞中仍然可以检测到升高的 14C 同位素水平，从而揭示它们的年龄。 14C 分析需要加速器质谱 (AMS)，确保从最少的 DNA 样本中精确定量极低水平的 14C。与 Spalding 等人（Cell，第 122 卷，133 143，2005 年 7 月 15 日）类似，该分析将确定抗体分泌细胞和记忆 B 细胞候选亚群的体内年龄，并证明或反驳这些细胞的寿命细胞，争论赞成或反对 B 细胞寿命作为体液记忆的关键要素。预期结果将为成功开发有效疫苗确定新的要求。 该项目将使用 AMS 来测定从免疫细胞中分离出的 DNA 样本中 14C 的含量，从而揭示它们的平均年龄。 AMS 是不可或缺的，因为要分析的细胞群非常稀有，因此产生的 DNA 样本量非常小。根据之前的研究，样品中 14C 的浓度再次非常低。 LLNL 已经进行了可比分析，我们相信与该资源的密切合作对于开展一个成功的项目是必要的。除了样本量非常小之外，似乎不需要额外开发方法。 14C 分析的结果将与各种免疫学方法得出的不同结果相结合，为功能不同的效应 B 细胞和体液记忆机制提供新的见解。 AMS 结果是整个研究的重要组成部分。 将获得人体血液和组织并提取特定细胞。将使用与后者 14C 定量兼容的既定方案从这些细胞中分离 DNA。 DNA 将被发送至资源进行 14C 分析。