CYTOKINES AND NK CELL DIFFERENTIATION IN RODENT DECIDUA

啮齿动物蜕膜中的细胞因子和 NK 细胞分化

• 批准号: 2205693
• 负责人: JUDITH R HEAD
• 金额: $ 18.88万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1995
• 资助国家: 美国
• 起止时间: 1995-09-01 至 2000-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2205693
• 关键词: cell differentiation; cytokine; cytokine receptors; decidua; enzyme linked immunosorbent assay; flow cytometry; fluorescence microscopy; gel electrophoresis; in situ hybridization; interferon gamma; interleukin 2; interleukin 4; laboratory mouse; laboratory rat; lymphocyte proliferation; messenger RNA; mixed tissue /cell culture; natural killer cells; northern blottings; phenotype; polymerase chain reaction; pregnancy; prostaglandin E; receptor expression; tumor necrosis factor alpha

During pregnancy in both rodents and humans, natural killer (NK)-lineage cells accumulate in large numbers in the maternal tissue (decidua) at the implant site, then in a unique structure in rodents, the metrial gland. These cells, termed granulated metrial gland (GMG) cells, have many characteristics of highly lytic IL-2-activated NK cells (e.g., large size and granules with lytic proteins), yet they have poor lytic capacity in vitro towards prototype targets. These cells arise rapidly through proliferation and differentiation from precursors in intimate association with decidual cells, but how the decidual cells influence these events is not known. The long-term goal of our research is to understand how these cells develop and function in pregnancy. Toward this end, the proposed studies are focussed on elucidating the features of the decidual microenvironment in rodents that regulate the development of these cells along a particular phenotypic and functional pathway. In Aim 1, decidual tissue, taken during the time of NK cell differentiation, and metrial gland tissue will be surveyed for the presence of cytokines which can influence NK cells, using ELISA assays and reverse transcription- polymerase chain reaction. Those identified will be localized to determine if they are produced by the decidual cells or by the differentiating NK cells themselves. Aim 2 involves characterization of rat GMG cells with regard to specific markers, lectin binding, and receptor expression, which will further define these cells in relation to classically activated NK populations and provide other markers to monitor during in vitro studies. In Aim 3, two in vitro approaches will be taken to elucidate the factors in decidua that direct proliferation and differentiation of NK cells into the GMG morphological and functional phenotype. First, splenic NK cells and bone marrow progenitor cells will be incubated with the cytokines identified in decidual cells, along with prostaglandins, decidual prolactin-like proteins, and ECM components to determine the combination that induces differentiation of GMG-type cells. Second, NK cells or progenitor cells will be co-cultured with purified decidual cells to establish a model for GMG cell development, then antagonists such as neutralizing antibodies will be used to determine the factors relevant to GMG cell differentiation. In Aim 4, nonpregnant and pregnant animals will be infused with viable fluorescent-labelled precursor cells and endometrium, decidua, and metrial gland tissues subsequently assessed by fluorescence microscopy to determine the time course of GMG precursor cell immigration, using antibodies to identify labelled cells entering these sites. Although their function is unknown, the striking presence of GMG cells in the pregnant uterus suggests that they play some important role in pregnancy. Furthermore, NK-lineage cells are strongly implicated in certain cases of pregnancy failure, suggesting that alterations in the normal regulation of decidual NK cell differentiation may have adverse effects on pregnancy. The proposed studies will provide significant new information towards understanding the development of these cells.

在啮齿动物和人类怀孕期间 细胞在母体组织（Decidua）中大量积累 植入部位，然后在啮齿动物的独特结构中，是公位腺。 这些细胞称为颗粒状的公计腺（GMG）细胞，具有许多 高裂解IL-2激活的NK细胞的特征（例如，大尺寸 和带有裂解蛋白的颗粒），但它们的裂解能力较差 朝向原型目标。 这些细胞迅速通过 亲密关联的增殖和与前体的分化 使用dec骨细胞，但是candual细胞如何影响这些事件 不知道。 我们研究的长期目标是了解如何 这些细胞在怀孕中发育并起作用。 在这一目标中， 提出的研究重点是阐明决法的特征 调节这些细胞发展的啮齿动物的微环境 沿特定的表型和功能途径。 在AIM 1中 组织，在NK细胞分化和公计的时间内服用 将调查腺体组织是否存在细胞因子的存在 使用ELISA分析和逆转录 - 影响NK细胞 - 聚合酶链反应。 确定的那些将本地化 确定它们是由decidual单元或由 区分NK细胞本身。 AIM 2涉及表征 大鼠GMG细胞在特定标记，凝集素结合和 受体表达将进一步定义这些细胞 向经典激活的NK种群，并提供其他标记 在体外研究中监测。 在AIM 3中，两种体外方法将 被采取以阐明直接增殖的Decidua中的因素 NK细胞分化为GMG形态和功能 表型。 首先，脾脏NK细胞和骨髓祖细胞将 与在decidual细胞中鉴定的细胞因子一起孵育 前列腺素，决定性催乳素样蛋白和ECM成分 确定诱导GMG型细胞分化的组合。 其次，NK细胞或祖细胞将与纯化 decidual细胞建立了GMG细胞开发模型，然后 诸如中和抗体之类的拮抗剂将用于确定 与GMG细胞分化有关的因素。 在AIM 4中，未怀孕和 怀孕的动物将注入可行的荧光标记 前体细胞和子宫内膜，DECIDUA和公位组织 随后通过荧光显微镜评估以确定时间 GMG前体细胞移民的过程，使用抗体来识别 标记进入这些位点的细胞。 尽管它们的功能未知，但 GMG细胞在怀孕子宫中的惊人存在表明 他们在怀孕中扮演着重要角色。 此外，NK-Linege 细胞在某些妊娠衰竭的情况下非常含义 表明deciDual NK细胞正常调节的改变 分化可能会对怀孕产生不利影响。 提议 研究将为理解提供重要的新信息 这些细胞的发展。