CD4 T cells with specificity to noninherited maternal antigen.

描述（由申请人提供）：妊娠期间母体和胎儿细胞的混合和双向转移创造了一个迷人的和高度相关的免疫耐受生理模型。虽然这种相互作用主要是从母体对胎儿表达的父本抗原的耐受性的角度来解决的，但成熟的胎儿细胞也会遇到同样大量的免疫学上外源的非遗传性母体抗原。因此，调节途径可能在免疫细胞发育的最早阶段就开始存在，以维持对非遗传性母体抗原（NIMA）的耐受性。此外，这些耐受途径也可能长期维持，因为后代中母体细胞的持续微嵌合状态越来越受到重视。在这方面，已经有令人信服的证据表明，从子宫内开始接触NIMA和其他外来抗原，就能产生免疫识别和耐受。然而，仍然未知的是免疫细胞和分子途径，从而实现对NIMA和其他子宫内暴露抗原的耐受性。我们提出了关键的科学瓶颈，包括无法精确识别NIMA应答免疫细胞，这些细胞对于表征其分子表型至关重要，以及以抗原特异性方式区分这些潜在罕见细胞与具有不相关特异性的细胞的工具。我们在概念可行性研究的初步证明中表明，这些限制是通过使用转基因雌性小鼠克服的，所述转基因雌性小鼠被工程化以组成性地和普遍地表达定义的模型抗原，用于与将模型抗原转化为替代NIMA的非转基因雄性小鼠交配。反过来，新开发的四聚体染色和富集技术允许以抗原特异性方式追踪具有替代NIMA特异性的内源性免疫细胞。使用这种方法，我们的初步发现 显示具有NIMA特异性的CD4 T细胞（i）在成年小鼠的外周淋巴组织中发现，（ii）选择性富集Foxp3表达，与幼稚或非遗传性父系抗原暴露对照相比，Foxp3表达水平高>2倍，以及（iii）在用重组单核细胞增生李斯特菌感染后仅适度扩增，同时保留夸大的Foxp3表达，所述重组单核细胞增生李斯特菌经工程改造以表达与重组单核细胞增生李斯特菌相同的NIMA相关肽。病原体相关抗原基于这些初步发现，我们的总体假设是子宫内生理暴露于非遗传性母体抗原通过选择性积累对这些免疫学外源抗原具有特异性的抑制性Foxp3+调节性T细胞触发耐受性。为了研究这一假设，提出了两个相互关联的目标，以建立精确的免疫细胞亚群，决定NIMA诱导的耐受性，并定义母源细胞微嵌合体在具有NIMA特异性的CD4 T细胞的耐受表型中的必要性。这项探索性资助的目标的成功完成将解锁并建立科学框架，以进行更多更深入的功能和机制研究，旨在进一步阐明免疫耐受性的这一未被重视的方面。

DESCRIPTION (provided by applicant): The intermingling and bi-directional transfer of maternal and fetal cells during gestation creates a fascinating and highly relevant physiological model of immune tolerance. Although this interaction has been primarily addressed from the perspective of maternal tolerance to paternal antigens expressed by the fetus, maturing fetal cells also encounter an equally vast array of noninherited maternal antigens that are immunologically foreign. Thus, regulatory pathways are likely in place beginning at the earliest stages of immune cell development to maintain tolerance to noninherited maternal antigen (NIMA). Furthermore, these tolerance pathways are also likely to be maintained long term given the increasingly appreciated sustained microchimerism of maternal cells in offspring. In this regard, there is already compelling evidence illustrating immune recognition and tolerance to NIMA and other foreign antigens with exposure beginning in utero. However, what remains unknown are the immune cell and molecular pathways whereby tolerance to NIMA and other in utero exposed antigens is achieved. We propose key scientific bottlenecks include the inability to precisely identify NIMA responsive immune cells essential for characterizing their molecular phenotype, and the tools for discriminating these potentially rare cells in an antigen specific fashion from those with irrelevant specificity. We show in initial proof of concept feasibility studies these limitations are overcome using transgenic female mice engineered to constitutively and ubiquitously express defined model antigens for mating with non transgenic males that transforms model antigens into surrogate NIMA. In turn, newly developed tetramer staining and enrichment techniques allow endogenous immune cells with surrogate NIMA specificity to be tracked in an antigen specific fashion. Using this approach, our initial findings show CD4 T cells with NIMA specificity are (i) found in the peripheral lymphoid tissue in adult mice, (ii) selectively enriched for Foxp3 expression with >2-fold higher levels compared with naive or noninherited paternal antigen exposed controls, and (iii) expand only modestly while retaining exaggerated Foxp3 expression after infection with recombinant Listeria monocytogenes engineered to express the same NIMA associated peptide as a pathogen associated antigen. Based on these initial findings, our overall hypothesis is physiological exposure to noninherited maternal antigen in utero triggers tolerance through the selective accumulation of suppressive Foxp3+ regulatory T cells with specificity to these immunologically foreign antigens. To investigate this hypothesis, two inter-related aims are proposed to establish the precise immune cell subset that dictates NIMA induced tolerance, and define the necessity for maternal cell microchimerism in tolerance phenotypes for CD4 T cells with NIMA specificity. The successful completion of the aims for this exploratory grant will unlock and establish the scientific framework for additional more in depth functional and mechanistic studies aimed at further unraveling this under appreciated aspect of immunological tolerance.