The role of semen in induction of paternal-specific tolerance during pregnancy

人类妊娠需要母体对胎儿的耐受性。一些流行病学证据表明，在受孕前，伴侣特异性耐受性开始通过暴露于精液而发展，精液携带父亲抗原，胎儿也会表达这些抗原。调节性T细胞（TCFs）在妊娠期间的耐受性中起着关键作用，但目前还不清楚这些细胞如何响应女性粘膜中的父亲抗原。抗原呈递细胞（APC）是最先暴露于父系抗原的细胞之一。它们感知并响应局部微环境，改变成熟状态，并可以在它们遇到的T细胞中诱导活化或调节表型。精液携带高浓度的细胞外囊泡（EV），我们和其他人已经表明与之相关，并诱导APC耐受的标记。我们假设这些囊泡以MHC分子的形式递送父系抗原，并改变机制途径以产生致耐受性APC，其刺激对父系抗原特异性的T细胞的分化。此外，我们预测，在妊娠并发症先兆子痫（PE）的妊娠中，由精液中存在的抗原激活的THBE的频率将低于健康妊娠。在目的1中，我们将研究精液成分如何诱导阴道和宫颈组织APC耐受。我们将利用多种方法：我们最近开发的28色APC表型面板，代谢分析和转录分析，以确定APC亚群中诱导耐受的特定途径。我们还将进行功能研究，以调查APC暴露于精液如何影响共培养T细胞的抑制功能。在目标2中，我们将检查精液EV在阴道暴露后在粘膜中的分布。我们将采用两种创新的EV标记技术（量子点标记和条形码寡核苷酸标记）来跟踪精液EV的组织渗透和体内运输。在目标3中，我们将确定健康妊娠和PE之间分娩后蜕膜和血液中的父系抗原特异性TdR有何不同。为了做到这一点，我们将分离由精液抗原激活的T细胞，并通过T细胞受体测序评估激活群体的克隆性。我们假设，与先兆子痫并发症的妊娠相比，健康妊娠将具有更多数量的抗原反应性Tcl3，以及Tcl3特异性克隆的扩增增强，表明抗原特异性。

Human pregnancy requires maternal tolerance of the fetus. Some epidemiological evidence suggests that before conception, partner-specific tolerance begins to develop through exposure to semen, which carries paternal antigens that will also be expressed by the fetus. Regulatory T cells (Tregs) play key roles in tolerance during pregnancy, but it is unclear how these cells develop in response to paternal antigens in the female mucosa. Antigen-presenting cells (APCs) are among the first cells to be exposed to paternal antigens. They sense and respond to the local microenvironment, shift maturation status, and can induce either activated or regulatory phenotypes in T cells they encounter. Semen carries a high concentration of extracellular vesicles (EV), which we and others have shown associate with, and induce markers of tolerance in APCs. We hypothesize that these vesicles deliver paternal antigens in the form of MHC molecules, and alter mechanistic pathways to generate tolerogenic APCs, which stimulate the differentiation of Tregs specific for paternal antigens. Furthermore, we predict that in pregnancies complicated by the gestational condition preeclampsia (PE), Tregs activated by antigens present in semen will be less frequent than in healthy pregnancies. In Aim 1, we will investigate how components of semen induce tolerance in APCs from vaginal and cervical tissues. We will utilize multiple methods: our recently developed 28 color APC phenotyping panel, metabolic profiling, and transcriptional analysis to define specific pathways of tolerance induction in subsets of APCs. We will also do functional studies to investigate how exposure of APCs to semen affects the suppressive function of co-cultured T cells. In Aim 2, we will examine where semen EV distribute in the mucosa after vaginal exposure. We will employ two innovative new EV tagging technologies (quantum-dot tagging and barcoded oligonucleotide tagging) to follow the penetration into tissue and in vivo trafficking of semen EV. In Aim 3, we will determine how paternal antigen specific Tregs in the decidua and blood following delivery differ between healthy pregnancies and PE. To do this, we will isolate Tregs activated by semen antigens, and assess the clonality of the activated population by T cell receptor sequencing. We hypothesize that healthy pregnancies will have greater numbers of antigen-reactive Tregs, as well as enhanced expansion of specific clones of Tregs, indicating antigen-specificity, as compared to pregnancies complicated by preeclampsia.