HCMV infection of human placental trophoblast and hematopoietic progenitors

描述（由申请人提供）：人巨细胞病毒（HCMV）经胎盘传播的机制，HCMV是先天性感染的主要病毒原因，影响美国1-3%的新生儿，是未知的。妊娠早期的原发性母体感染造成40-50%的经胎盘传播风险，15%的患病婴儿有永久性出生缺陷。我们建议研究人胎盘滋养层祖细胞（TB）和造血谱系的HCMV感染，以及发育失调的分子机制的后果。Fisher小组将妊娠早期人类胎盘基质确定为共表达多能性标记物和小鼠TB命运决定因素的细胞的小生境，这表明该区域是人类TB祖细胞（TBPC）的一个来源。他们分离了这些细胞，并开发了连续自我更新的TBPC系。当在触发TB分化的条件下培养时，这些细胞形成成熟的人类TB群体-多核的、运输合胞体滋养层细胞（STB）和侵袭性CTB。在同一区域，Fisher小组鉴定出造血干细胞（HSC）以及髓系和红系定向谱系的中间前体，表明活跃的造血。CD 34 ++ CD 45 low造血干细胞在成分培养基中培养，可分化为红细胞和髓系细胞。总之，这些研究表明，人胎盘是TBPC的来源，TBPC填充绒毛膜绒毛和HSC，可能在胚胎/胎儿造血系统的发育中发挥作用。胎盘HSC可能也有助于胎盘Hofbauer巨噬细胞群体。TBPC对致病性HCMV株的感染高度敏感，该致病性HCMV株诱导控制细胞周期进程的蛋白质的表达以及自我更新和谱系定型之间的平衡。上调的分子被错误定位到细胞质中，并在病毒体组装区室中积累。另外的实验显示HCMV感染胎盘来源的HSC。在这里，我们提出测试HCMV感染改变TBPC自我更新和分化之间的平衡（目的1）的理论。我们还假设HSC的感染损害了它们形成髓系和红系定向谱系的能力（目的2）。这些研究将为病毒对TBPC分化的影响提供新的见解，TBPC执行胎盘的专门功能，HSC有助于胎儿造血和胎盘免疫屏障。然后，我们将使用这些信息来确定是否在子宫内先天性感染的胎盘中观察到相同的表型变化。我们还将测量人中和MoAb保护绒毛膜绒毛免受HCMV感染的能力（目的3）。总之，这些研究将提供有关HCMV对TBPC和HSC自我更新和分化的影响的新信息。我们还希望深入了解预防HCMV传播和伴随的出生缺陷的新方法。

DESCRIPTION (provided by applicant): Mechanisms of transplacental transmission of human cytomegalovirus (HCMV), the leading viral cause of congenital infection affecting 1-3% of births in the U.S., are unknown. Primary maternal infection in first trimester poses a 40-50% risk of transplacental transmission and permanent birth defects in 15% of diseased babies. We propose to study HCMV infection of the human placental progenitors of the trophoblast (TB) and hematopoietic lineages, and the consequences in terms of the molecular mechanisms that dysregulate development. The Fisher group identified the early-gestation human placental stroma as a niche for cells that co-expressed markers of pluripotency and determinants of mouse TB fate, which suggested that this region is one source of human TB progenitor cells (TBPCs). They isolated these cells and developed lines of continuously self-renewing TBPCs. When cultured under conditions that triggered TB differentiation, the cells formed the mature human TB populations-multi-nucleated, transport syncytiotrophoblasts (STBs) and invasive CTBs. In the same region, the Fisher group identified hematopoietic stem cells (HSCs) as well intermediate precursors of the myeloid- and erythroid-committed lineages, suggesting active hematopoiesis. Cultured in defined medium, the CD34++CD45low HSCs contributed to erythrocytes and myeloid cells. Together these studies showed that the human placenta is a source of TBPCs that populate the chorionic villi and HSCs that might play a role in development of the hematopoietic system of the embryo/fetus. It is likely that placental HSCs also contribute to the placental Hofbauer macrophage population. TBPCs are highly susceptible to infection with a pathogenic HCMV strain that induced expression of proteins controlling cell-cycle progression and the balance between self-renewal and lineage-commitment. Upregulated molecules were mislocalized to the cytoplasm and accumulated in the virion assembly compartment. Additional experiments showed that HCMV infected placentally-derived HSCs. Here, we propose testing the theory that HCMV infection alters the balance between TBPC self-renewal and differentiation (Aim 1). We also hypothesize that infection of HSCs impairs their ability to form myeloid- and erythroid-committed lineages (Aim 2). These studies will provide new insights into viral effects on differentiation of TBPCs, which carry out the specialized functions of the placenta, and HSCs, which contribute to fetal hematopoiesis and the placental immunological barrier. Then we will use this information to determine if the same phenotypic changes are observed in placentas that were congenitally infected in utero. We will also measure the ability of human neutralizing MoAbs to protect chorionic villi from HCMV infection (Aim 3). In summary, these studies will provide new information about the effects of HCMV on TBPC and HSC self-renewal and differentiation. We also expect to gain insights into novel approaches for preventing HCMV transmission and the attendant birth defects.