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

人类怀孕需要母体对胎儿的耐受性。一些流行病学证据表明，在怀孕之前，伴侣特有的耐受性是通过接触精液而开始形成的，精液携带的父亲抗原也将在胎儿中表达。调节性T细胞(Tregs)在妊娠期间的耐受性中起着关键作用，但这些细胞如何对女性粘膜中的父系抗原做出反应尚不清楚。抗原提呈细胞(APC)是最早接触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，并能诱导它们遇到的T细胞的激活或调节表型。精液携带高浓度的胞外小泡(EV)，我们和其他人已经证明EV与APC相关，并诱导耐受标记。我们假设这些小泡以MHC分子的形式递送父系抗原，并改变机制途径来产生耐受性APC，刺激父系抗原特异性Tregs的分化。此外，我们预测，在妊娠合并先兆子痫(PE)的妊娠中，由精液中存在的抗原激活的Tregs的频率将低于健康妊娠。在目标1中，我们将研究精液成分如何诱导来自阴道和宫颈组织的APC产生耐受。我们将利用多种方法：我们最近开发的28色APC表型面板、代谢图谱和转录分析来确定APC亚群中耐受诱导的特定途径。我们还将进行功能研究，以调查APC与精液接触如何影响共培养的T细胞的抑制功能。在目标2中，我们将检查EV精液在阴道暴露后在粘膜中的分布情况。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EV在组织中的渗透和体内的运输。在目标3中，我们将确定健康妊娠和PE之间蜕膜和血液中父亲抗原特异性Tregs的不同之处。为此，我们将分离被精液抗原激活的Tregs，并通过T细胞受体测序评估激活群体的克隆性。我们假设，与妊娠合并先兆子痫相比，健康妊娠将有更多数量的抗原反应性Tregs，以及特定Tregs克隆的增强扩增，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.