Analysis of uterine natural killer cell promotion of endometrial angiogenesis

一个不寻常的免疫细胞亚群(Unk细胞)出现在怀孕早期的子宫中。在小鼠体内，这些细胞分泌信号分子(干扰素-γ和血管内皮细胞生长因子)，导致母体螺旋动脉(SA)的结构变化，SA是滋养胎盘和胎儿的血管。我们认为，当植入部位形成新的血管时，Unk细胞引导血管衬里细胞(内皮)，将它们导向胎儿来源的胎盘细胞。三个具体目标将解决我们的假设。目的1研究妊娠8-12天小鼠唾液酸的主要分子和功能特性。将使用激光捕获显微切割、RNA提取和实时定量RT-PCR来研究血管内皮细胞、血管平滑肌和定位的Unk细胞。这些分子数据将与使用我们的小鼠SA活体显微镜新技术收集的功能数据相关联。子宫动脉节段将与结构稳定的对照肠道血管进行比较。目的2将使用显微镜和分子方法来确定NK细胞和体内移行内皮细胞之间的相互作用。在人子宫微血管内皮细胞和激活的NK细胞的3D悬滴共培养中，将研究淋巴细胞和内皮细胞的相互作用以及其他细胞类型和分子对它们的调节。过度表达白介素15或仅在淋巴细胞中有靶向基因缺失的小鼠的怀孕将用于验证体外结果。目标3将通过比较蜕膜、脾和骨髓前体细胞的基因表达，解决Unk细胞谱系何时与其他组织中的NK细胞分化的问题。这些研究模拟了人类子宫内膜和胎儿健康的重要方面，以及家畜子宫血管生成，其中子宫内膜血管与产仔数相关。这些数据将推动我们在整个动物、细胞和分子水平上了解Unk细胞生物学的长期目标，并可能为深入了解免疫细胞在一般组织愈合或病理性重塑中的血管生成作用提供见解。

An unusual subset of immune cells (uNK cells) appears in the uterus in early pregnancy. In mice, these cells secrete signaling molecules (Interferon-gamma and Vascular Endothelial Cell Growth Factor) which bring about structural changes to maternal spiral arteries (SA), the blood vessels nourishing the placenta and fetus. We believe uNK cells guide blood vessel lining cells (endothelium) as new vessels form in implantation sites, directing them towards fetally-derived placental cells. Three Specific Aims will address our hypothesis. AIM 1 will characterize and correlate key molecular and functional properties of mouse SA during gestation days 8-12. Endothelial, vascular smooth muscle and position-defined uNK cells will be studied using laser capture microdissection, RNA isolation and Realtime quantitative RT-PCR. These molecular data will be correlated with functional data collected using our new technique for intravital microscopy of mouse SA. Uterine artery segments will be compared with control, structurally-stable intestinal vessels. AIM 2 will use microscopic and molecular approaches to define interactions between NK cells and migrating endothelial cells in vitro and in vivo. Lymphocyte-endothelial cell interactions and their regulation by other cell types and molecules will be studied in 3D hanging-drop co-cultures of human uterine microvascular endothelium and activated NK cells. Pregnancies in mice over-expressing interleukin 15 or with targeted gene deletions only in lymphocytes will be used to validate the in vitro results. AIM 3 will address when the uNK cell lineage diverges from NK cells in other tissues by comparing gene expression in decidual, splenic and marrow precursors. These studies model important aspects of human endometrial and fetal health and uterine angiogenesis in livestock where endometrial vascularity correlates with litter size. The data will advance our long-term goal of understanding the biology of uNK cells at the whole animal, cellular and molecular levels, and may provide insights into angiogenic roles of immune cells in general tissue healing or pathogenic remodeling.