The Role of Fetal Cell Microchimerism in Maternal Repair

描述(由申请人提供)：我们请求支持一项假设，即通过怀孕从生理上获得并在流产、流产或分娩后保留在成年女性体内的胎儿细胞，包括一组新的细胞，我们将其称为“妊娠相关祖细胞(PAPC)”。到目前为止，围绕成体干细胞可塑性的争论几乎忽略了怀孕对女性的作用。如果PAPC被证明是真正的干细胞，它将具有发育优势，即起源于胎儿，但可以从之前怀孕的成年女性身上提取出来，而不会引发伦理争议。在未输血的成年女性中，我们有大量的初步数据表明，通过妊娠获得的胎儿细胞(基于Y染色体和胎儿特有的DNA多态)可在外周血和临床疾病器官中检测到，并具有多系能力。由于需要从活检和/或尸检标本中获取临床材料，这些研究一直是描述性的，而不是机械性的。我们的假设将在一个动物模型中得到验证，一个表达绿色荧光蛋白(GFP)或荧光素酶的转基因雄性小鼠被培育成野生型雌性小鼠。这将使我们能够控制生育史，并测试关于母体中胎儿细胞的可塑性和活动的多种假说。绿色荧光蛋白和荧光素酶序列是显性转基因。一半的胚胎幼崽将携带转基因，并在它们的部分或全部细胞中表达绿色荧光标记，具体取决于结构。胎儿细胞发出绿色荧光，可以使用各种技术在母体组织中进行识别和跟踪，包括活体整体动物成像、荧光显微镜和实时PCR扩增。在特定目标1中，我们将检验特定因素影响母亲胎儿细胞微嵌合体(FCMC)发育的假设。在具体目标2中，我们将使用化学、外科、遗传和缺血模型，通过分析整体福祉和寿命、目标器官功能、伤口愈合的差异和差异基因表达，确定胎儿细胞是否被招募到特定的组织损伤情景中，并为母体损伤的修复做出贡献。在具体目标3中，我们将检测小鼠微嵌合胎儿细胞的细胞表面特征，并进行微阵列分析，以确定FCMC是由一种还是多种细胞类型引起的。在具体目标4中，我们将检验这样一个假设，即胎儿干细胞比成人干细胞具有优势，并有助于延长生存时间或改善器官功能。长期目标是确定怀孕是否通过获得具有治疗潜力的独特细胞来为女性提供长期优势。

DESCRIPTION (provided by applicant): We are requesting support to test the hypothesis that fetal cells, acquired physiologically through pregnancy, and retained in the adult human female following abortion, miscarriage, or delivery, encompass a novel population of cells that we have termed the "Pregnancy-Associated Progenitor Cell (PAPC)." To date, the controversy surrounding the plasticity of adult stem cells has virtually ignored the role of pregnancy in females. PAPCs, if shown to be true stem cells, would have the developmental advantages of being fetal in origin yet could be retrieved without ethical controversy from an adult female who has previously been pregnant. We have extensive preliminary data in the human adult, non-transfused female that fetal cells (identified on the basis of the Y chromosome as well as fetal-specific DNA polymorphisms), acquired through pregnancy, are detectable in peripheral blood and clinically diseased organs, and have multi-lineage capacity. Due to the necessity of obtaining clinical material from biopsy and/or autopsy specimens, these studies have been descriptive and not mechanistic. Our hypothesis will be tested in an animal model, a transgenic male mouse expressing either green fluorescent protein (GFP) or luciferase bred to wild-type female mice. This will allow us to control reproductive histories, and test multiple hypotheses regarding the plasticity and activity of fetal cells in the maternal body. The GFP and luciferase sequences are dominant transgenes. Half of the fetal pups will carry the transgene, and express the green fluorescent marker in some or all of their cells, depending on the construct. Fetal cells fluoresce green and can be identified and tracked in maternal tissues using a variety of techniques, including in vivo whole animal imaging, fluorescence microscopy, and real-time PCR amplification. In specific aim 1 we will test the hypothesis that specific factors affect the development of fetal cell microchimerism (FCMC) in the mother. In specific aim 2 we will use chemical, surgical, genetic, and ischemic models to determine if fetal cells are recruited in specific tissue injury scenarios and contribute to the repair of maternal injury by analyzing overall well being and longevity, target organ function, differences in wound healing, and differential gene expression. In specific aim 3 we will examine the cell surface characteristics of the murine microchimeric fetal cells and perform microarray analysis to determine whether FCMC is due to 1 or multiple cell types. In specific aim 4 we will test the hypothesis that fetal stem cells have an advantage over adult stem cells and contribute to prolonged survival or improved organ function. The long-term objective is to determine if pregnancy confers a long-term advantage to a female by resulting in the acquisition of unique cells that have therapeutic potential.