Transgenerational Microchimerism in Pregnancy Loss

描述(申请人提供)：微嵌合体(Mc)指的是拥有少量细胞或来自不同基因个体的DNA。在怀孕期间，Mc是在母亲和胎儿之间的细胞交换中自然获得的。在以前的研究中，我们发现母亲的细胞会在她的后代体内存活到成年。在怀孕期间，已经是母亲Mc宿主的妇女，随着来自胚胎的细胞和无细胞DNA进入她的血液循环，获得了额外的Mc来源。这些观察提出了这样一个问题：人类白细胞抗原世代间的关系是否会影响女性的妊娠结局，以及由于遗传多样性通常被认为是好的，世代间过多的人类白细胞抗原共享是否会导致复发性特发性妊娠丢失(RPL)。人类白细胞抗原(HL A)分子是免疫反应的中心，是区分自身和“他人”的核心，而广泛的多态性是人类白细胞抗原(HL A)基因的特点。由于人类白细胞抗原基因的多态性，大多数家庭成员之间存在着大量的等位基因差异。然而，由于人类白细胞抗原纯合子或杂合的人类白细胞抗原同一性，偶尔在家庭中观察到人类白细胞抗原共享的增加。以前的研究表明，女性和她的伴侣过度共享人类白细胞抗原与特发性RPL有关，尽管研究一直相互矛盾。这项R21提案的第一个具体目标将检验这样一个假设，即当女性先证者患有原发特发性RPL时，三代人之间的HLA关系与有活产和无流产史的先证者相比，具有较小的HLA差异(增加了共享)。DRB1、DRB3、DRB4、DRB5、DQA1、DQB1、A、B和C等位基因将用于特发性RPL患者及其母亲、胎儿和伴侣的基因分型。具体目标2将识别和量化先证者母亲(MP-Mc)在怀孕前、怀孕期间和怀孕后的连续血样中的Mc，这些血样是患有原发特发性RPL并随后流产或活产的先证者和没有流产史的健康妇女。为此，一组人类白细胞抗原特异性和其他遗传多态特异性定量聚合酶链式反应方法已经被开发出来，并将用于检测外周血单核细胞。特定目标3将使用与目标2相同的方法在相同的外周血样中定量来自先证者受孕的Mc(Cp-Mc)，以检测外周血单核细胞和血清。除了目标1的假设外，第二个假设将在目标2和3中检验，即Mc的动态变化，MP-Mc或Cp-Mc或两者都与特发性RPL有关。从多种来源窝藏Mc的后果和长期影响在很大程度上是未知的，因为从怀孕中自然获得的Mc直到最近才被认识到。据我们所知，目前还没有关于特发性RPL中人类白细胞抗原共享或Mc代际的研究。来自不同遗传个体的少量细胞(或DNA)被称为微嵌合体(Mc)。女性从自己的母亲那里获得了Mc，后来又从自己的怀孕中获得了Mc，这一发现引发了一个问题，即不同世代之间的细胞群体之间的相互作用是否有时会影响怀孕成功。因为它们有助于区分一个细胞和另一个特定的基因，称为人类白细胞抗原基因，是特别值得研究的。

DESCRIPTION (provided by applicant): Microchimerism (Mc) refers to harboring a small number of cells or DNA from a genetically distinct individual. During pregnancy Mc is naturally acquired in the exchange of cells between a mother and conceptus. In prior studies we found that a mother's cells persist in her progeny into adult life. During pregnancy a woman who is already host to Mc from her mother acquires an additional source of Mc as cells and cell-free DNA from the conceptus enter her circulation. These observations raise the question of whether HLA-relationships across generations influence the outcome of pregnancy in women and, as genetic diversity is generally thought to be good, whether excessive HLA-sharing across generations contributes to recurrent idiopathic pregnancy loss (RPL). HLA molecules are central to immune responses, to distinguishing self from "other" and extensive polymorphism is the hallmark feature of HLA genes. Due to the polymorphism in HLA genes most of the time there are numerous differences in HLA alleles among family members. However, occasionally an increase of HLA-sharing is observed in families, either due to HLA-homozygosity or heterozygous HLA-identity. Previous studies have suggested that excessive HLA-sharing of a woman and her partner is associated with idiopathic RPL, although studies have been conflicting. The first Specific Aim of this R21 proposal will test the hypothesis that HLA-relationships over three generations have less HLA-disparity (increased sharing) when women probands have primary idiopathic RPL compared to probands with a live birth and no history of miscarriage. HLA-genotyping will be conducted for DRB1, DRB3, DRB4, DRB5, DQA1, DQB1, A, B and C for women with primary idiopathic RPL, their mothers, conceptus, and partners. Specific Aim 2 will identify and quantify Mc from the proband's mother (MP-Mc) in consecutive blood samples from before, during and after pregnancy for probands with primary idiopathic RPL with a subsequent miscarriage or with a live birth and healthy women with no history of miscarriage. For this purpose a panel of HLA-specific and other genetic polymorphism specific quantitative PCR assays has been developed and will be employed to tested peripheral blood mononuclear cells. Specific Aim 3 will quantify Mc from the conceptus of the proband (CP-Mc) in the same peripheral blood samples using the same methods as in Aim 2 to test peripheral blood mononuclear cells as well as serum. In addition to the hypothesis of Aim 1, a second hypothesis will be tested in Aims 2 and 3, that dynamic changes in Mc, either MP-Mc or CP-Mc or both, is associated with idiopathic RPL. The consequences and long-term effects of harboring Mc from multiple sources are largely unknown as naturally acquired Mc from pregnancy has only recently been recognized. To our knowledge there are no prior studies investigating HLA-sharing or Mc across generations in idiopathic RPL. Harboring a small number of cells (or DNA) that originated from a genetically distinct individual is referred to as microchimerism (Mc). The finding that women harbor Mc from their own mothers and later acquire Mc from their own pregnancy(ies) raises questions about whether interactions between cell populations across generations sometimes influences pregnancy success. Because they help distinguish one cell from another particular genes, called HLA genes, are of special interest to investigate.