UTERINE NK CELL HOMING FROM THE PERIPHERAL CIRCULATION

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 子宫自然杀伤细胞(Unk)是CD56brightCD16淋巴细胞的一个独特亚群，在排卵后数量增加，并在妊娠早期达到高峰。尽管淋巴细胞通常被认为在宿主防御中发挥作用，但越来越多的证据表明，Unk细胞的主要功能不是免疫学的，而是可能在血管生成、滋养层侵袭和螺旋动脉重建中发挥作用。据报道，反复流产和植入失败的妇女缺乏Unk细胞。因此，Unk细胞的正确定位和功能对于胎儿的正常发育是必要的。 Unk细胞的起源尚不清楚。在怀孕期间，它们的数量大大增加，要么通过增加子宫招募，要么通过扩大常驻人口。大约10%的外周NK细胞(PNK)是Unk表型(CD56brightCD16-)，并被认为在怀孕期间选择性地招募到子宫中。然而，最近有报道称，转化生长因子支持CD56dimCD16+细胞向CD56brightCD16-表型转化，蜕膜基质细胞产生转化生长因子。基于这些发现，我们假设CD56dimCD16+PNK细胞被招募到子宫，在那里它们被转化生长因子诱导为CD56brightCD16-Unk表型。如果这个模型是正确的，那么CD56dimCD16+细胞将被预测优先招募到子宫内皮，而不是CD56brightCD16-细胞。我们将在以下目标中检验这一假设： 具体目的1.测定特定的人NK细胞亚群对小鼠胎盘冰冻切片的相对黏附能力。 特定目的2.利用培养的人子宫微血管内皮细胞(HUtMVEC)，检测Unk募集的整个滚动-停止-迁移级联反应，并检测各亚群的迁移能力。具体地说，我们将(A)检测雌激素、孕激素和黄体生成素对体外培养的HUtMVEC表达黏附分子的影响，以确定它们上调的条件；(B)使用a部分定义的条件，建立人NK细胞与HUtMVEC的细胞-细胞黏附实验；以及(C)建立体外流式细胞术，利用HUtMVEC诱导表达黏附受体，并确定人NK亚群的体外滚动、阻止和跨内皮迁移。 这些研究将有助于更好地了解人类Unk细胞的起源、它们的归巢机制以及它们在维持正常妊娠中的作用。

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. Uterine Natural Killer cells (uNK) are a unique subpopulation of CD56brightCD16- lymphocytes that increase in number after ovulation, and reach their peak numbers during the first trimester of pregnancy. Although lymphocytes are generally thought to play a role in host defense, there is increasing evidence that the primary function of uNK cells is not immunological but rather they may play a role in angiogenesis, trophoblast invasion and spiral artery remodeling. Women who experience recurrent miscarriages and failure of implantation have been reported to be deficient in uNK cells. Thus proper localization and function of uNK cells is necessary for normal fetal development. The origins of uNK cells are unclear. During pregnancy, their number expands greatly, either through increased recruitment to the uterus, or via expansion of resident populations. Approximately 10% of peripheral NK cells (pNK) are of the uNK phenotype (CD56brightCD16-), and have been proposed to be selectively recruited to the uterus during pregnancy. However, recently it has been reported that TGF¿ supports conversion of CD56dimCD16+ cells towards a CD56brightCD16- phenotype, and that decidual stromal cells produce TGF¿. Based on these findings, we hypothesize that CD56dimCD16+ pNK cells are recruited to the uterus, where they are then induced towards the CD56brightCD16- uNK phenotype by TGF¿. If this model is correct, then CD56dimCD16+ cells would be predicted to be preferentially recruited to uterine endothelium as compared to CD56brightCD16- cells. We will test this hypothesis in the following aims: Specific Aim 1. Determine the relative adhesive ability of specific human NK cell subpopulations to frozen sections of mouse placenta. Specific Aim 2. Utilizing cultured human uterine micrrovascular endothelial cells (HUtMVEC), examine the entire rolling-arrest-transmigration cascade of uNK recruitment and test the transmigration capability of each subpopulation. Specifically, we will (a) test the effect of estrogen, progesterone, and LH on expression of adhesion molecules by HUtMVEC in culture, to identify conditions where they are upregulated; (b) using the conditions defined from part a, establish a cell-cell adhesion assay for human NK cells with HUtMVEC; and (c) develop an in vitro flow assay using HUtMVEC induced to express adhesion receptors, and defined human NK subpopulations to recapitulate rolling, arrest and transendothelial migration in vitro. These studies will lead to a better understanding of the origin of human uNK cells, their homing mechanism, and their role in maintenance of normal pregnancy.