Trophoblast MHC-I: Trigger for Immune-Mediated Rejection of Cloned Bovine Fetuses

描述(由申请人提供)：免疫人工流产是妇女妊娠失败的一个重要原因，尽管人们对此知之甚少，而且往往未被认识到。自然流产和先兆子痫与胎盘异常有关，在通过体外受精(IVF)等辅助生殖技术确定的人和牛的怀孕中，发生的频率更高。我们的长期目标是了解免疫介导的流产和胎盘功能不全的免疫学基础，并开发和测试下调对胚胎的免疫反应的策略。将使用一种独特的牛免疫介导的流产和胎盘功能不全的模型，该模型基于牛体细胞核移植(SCNT)妊娠中胎盘发育不足和胎儿丢失的常规发生。几个观察结果支持该模型的有效性：(1)与对照组相比，SCNT胎儿的胎盘滋养层细胞在妊娠早期表达经典的主要组织相容性复合体-I类(MHC-I)抗原；(2)SCNT受体的子宫间质中有T淋巴细胞聚集；(3)SCNT纯合子胎儿的情况好于MHC杂合子胎儿。这些发现表明，SCNT妊娠中的大部分胎儿丢失是由于免疫介导的流产。该项目基于一个普遍假设，即妊娠早期不适当的滋养层细胞MHC-I表达将导致免疫介导的流产或胎盘功能不全。MHC-I蛋白的不适当表达可能包括经典MHC-I蛋白表达增加、非经典MHC-I蛋白表达降低或两者兼而有之。该建议的具体目标是：(1)产生8-10个MHC-I相合的SCNT、8-10个MHC-I不相容的SCNT和8-10个对照妊娠，用于在怀孕32至34天收集子宫和胎盘组织，并比较三种类型妊娠中滋养细胞MHC-I的表达、子宫白细胞数量以及子宫和胎盘基因的表达；以及(2)比较SCNT胚胎移植到MHC-I相合和不相容的受体奶牛中的胚胎死亡率，以确定是否可以通过消除MHC-I触发的抗原来避免免疫介导的流产。该项目将阐明正常和异常妊娠中胎盘和子宫免疫系统之间的相互作用的功能基因组学。功能基因组学和形态数据将被整合，以创建涉及一个重要临床问题的细胞和分子事件的完整图景：免疫介导的堕胎。该项目还将直接测试这一假说，即怀孕早期异常的MHC-I表达会触发免疫中介流产，这是一个长期存在的假说，由于缺乏合适的动物模型，该假说从未得到充分检验。与公共卫生相关：哺乳动物的胚胎必须说服其母亲的免疫系统接受它。当胎儿未能诱导母体耐受时，结果要么是免疫介导的流产，要么是胎盘异常的发展，这可能会导致以后怀孕的问题。随着体外受精等辅助生殖技术的使用，免疫介导的流产和胎盘功能不全在人和牛中的发生率都大大增加。在这项研究中，牛的体细胞核移植怀孕，通常有很高的早期妊娠自然流产率，将被用来研究当胚胎在体外被操纵时，胚胎和母体免疫系统之间的通讯是如何中断的。

DESCRIPTION (provided by applicant): Immune-mediated abortion is an important, although poorly understood and often unrecognized, cause of pregnancy failure in women. Both spontaneous abortion and preeclampsia, which is associated with abnormal placentation, occur at increased frequency in human and bovine pregnancies established by assisted reproductive technologies such as in vitro fertilization (IVF). Our long-term goals are to understand the immunological basis of immune-mediated abortion and placental insufficiency, and to develop and test strategies to down-regulate the immune response to the conceptus. A unique bovine model of immune- mediated abortion and placental insufficiency based on the routine occurrence of inadequate placental development and fetal loss in bovine somatic cell nuclear transfer (SCNT) pregnancies will be used. Several observations support the validity of this model: (1) placental trophoblast cells of SCNT fetuses, in contrast to control fetuses, express classical major histocompatibility complex class I (MHC-I) antigens early in pregnancy, (2) T lymphocytes accumulate in the uterine stroma of SCNT recipients, and (3) MHC homozygous SCNT fetuses fare better than MHC heterozygous fetuses. These findings suggest that much of the fetal loss in SCNT pregnancies is due to immune-mediated abortion. This project is based on the general hypothesis that inappropriate trophoblast MHC-I expression early in pregnancy will induce immune-mediated abortion or placental insufficiency. Inappropriate expression of MHC-I proteins may consist of increased expression of classical MHC-I proteins, decreased expression of non-classical MHC-I proteins or a combination of the two. The proposal's specific aims are: (1) produce 8-10 MHC-I compatible SCNT, 8-10 MHC-I incompatible SCNT and 8-10 control pregnancies for collection of uterine and placental tissues on days 32 to 34 of pregnancy, and compare trophoblast MHC-I expression, uterine leukocyte populations and uterine and placental gene expression in the three types of pregnancies; and (2) compare embryonic mortality rates of SCNT embryos transferred into MHC-I compatible and incompatible recipient cows to determine if immune-mediated abortion can be avoided by eliminating the antigenic, MHC-I trigger. This project will elucidate the functional genomics of the cross-talk between the placenta and uterine immune system in normal and abnormal pregnancies. Functional genomics and morphological data will be integrated to create a complete picture of the cellular and molecular events involved in an important clinical problem: immune-mediated abortion. The project will also directly test the hypothesis that abnormal MHC-I expression early in pregnancy triggers immune-mediated abortion, a long-standing hypothesis that has never been adequately tested because of lack of a suitable animal model. PUBLIC HEALTH RELEVANCE: A mammalian conceptus must convince its mother's immune system to accept it. When the fetus fails to induce maternal tolerance, the outcome is either immune-mediated abortion or the development of abnormal placentation that can result in problems later in pregnancy. The incidence of immune-mediated abortion and placental insufficiency in both humans and cattle are greatly increased with the use of assisted reproductive technologies such as in vitro fertilization. In this study bovine somatic cell nuclear transfer pregnancies, which normally have a high rate of first trimester spontaneous abortions, will be used to study how communications between the conceptus and maternal immune system break down when embryos are manipulated in vitro.