Human Cytomegalovirus-Induced Inhibition of Cytotrophoblast Invasion

描述（由申请人提供）：显著的发病率和死亡率与先天性疱疹病毒人巨细胞病毒（HCMV）感染有关。15%的HCMV原发感染妇女在妊娠早期自发流产；而且显示出感染迹象的是胎盘，而不是胚胎或胎儿。胎盘HCMV感染的病理后果，包括妊娠早期流产、宫内生长受限、先兆子痫和早产，被认为是由妊娠早期卵泡外细胞滋养细胞（EVT）无法充分侵入子宫壁导致母体螺旋动脉重塑受损和浅胎盘介导的。初步研究表明，HCMV感染妊娠早期EVT可抑制EVT侵袭，显著降低促进侵袭的基质金属蛋白酶(MMP)-2和MMP-9的表达和活性。hcmv诱导的EVT抑制伴随着转化生长因子(TGF)-21的表达增加，TGF -21是一种已知的抑制EVT侵袭的因子。由于EVT侵袭发生在妊娠早期，此时胎盘的胎儿侧环境相对缺氧，而在其他系统中，缺氧能够激活疱疹病毒基因表达和裂解复制，因此胎盘过程中的低氧分压实际上可能增强HCMV抑制EVT侵袭的能力。为了确定HCMV抑制EVT侵袭的分子机制，这是本文提出的广泛而长期的研究目标，我们利用生物反应器组织工程技术开发了一种新的、生物学相关的EVT侵袭模型。这些研究提出的假设是，HCMV，更具体地说，病毒包膜糖蛋白B （gB）和/或直接早期基因产物IE1-72和IE2-86，通过激活TGF-21和调节MMP活性来抑制胎盘期间EVT的侵袭；此外，缺氧通过上调HCMV IE表达和裂解复制，增强了HCMV抑制EVT侵袭的能力。验证这些假设的具体目的是：(1)确定HCMV对培养EVT侵袭的抑制是否通过HCMV复制周期的早期事件介导；(2)确定HCMV是否抑制促进侵袭的MMP-2、MMP-3、MMP-9和uPA，并上调抑制侵袭的TIMP-1。TIMP-2和PAI-1在体外培养EVT中的表达，以及HCMV是否通过激活TGF-21抑制体外培养EVT的侵袭性；(3)确定妊娠早期CTB分化所遇到的相对低氧环境是否通过增加溶解性HCMV复制增强HCMV诱导的EVT对间质和血管内侵袭的抑制。阐明HCMV损害胎盘的机制可能是理解与宫内HCMV感染相关的胎儿和母体病理的关键。公共卫生相关性：15%的原发性人巨细胞病毒（HCMV）感染妇女在妊娠早期自发流产，并且是胎盘，而不是胚胎或胎儿，显示感染的证据。此外，HCMV感染可能导致早产、宫内生长受限或先兆子痫，所有这些都与胎盘病理有关。这些并发症被认为至少部分是由于胎盘发育早期子宫壁外细胞滋养细胞侵袭（EVT）不足和母体螺旋动脉重塑受损所致。确定HCMV如何损害EVT侵袭对生殖界具有重要意义，并可能为维持妊娠提供额外的治疗靶点。

DESCRIPTION (provided by applicant): Significant morbidity and mortality is associated with symptomatic congenital infection with the herpes virus human cytomegalovirus (HCMV). Fifteen percent of women with primary HCMV infection spontaneously abort during early pregnancy; and it is the placenta, not the embryo or fetus that shows evidence of infection. Pathological consequences of placental HCMV infection, including first trimester pregnancy loss, intrauterine growth restriction, pre-eclampsia and preterm labor, are believed to be mediated by the inability of extravillous cytotrophoblasts (EVT) to adequately invade the uterine wall during early first trimester pregnancy resulting in impaired remodeling of maternal spiral arteries and shallow placentation. Preliminary studies demonstrate that HCMV infection of first trimester EVT results in inhibition of EVT invasion and in significant reduction of expression and activity of invasion-promoting matrix metalloproteinase (MMP)-2 and MMP-9. HCMV-induced inhibition of EVT occurs along with increased expression of transforming growth factor (TGF)-21, a factor known to inhibit EVT invasion. Since EVT invasion occurs during first trimester pregnancy when the fetal side of the placental environment is relatively hypoxic and since hypoxia, in other systems, is capable of activating herpesvirus gene expression and lytic replication, low oxygen partial pressure during placentation may actually augment the ability of HCMV to inhibit EVT invasion. To determine the molecular mechanism(s) by which HCMV inhibits EVT invasion, which is the broad, long-term objective of the studies proposed herein, a novel, biologically relevant model of EVT invasion has been developed using bioreactor tissue engineering technology. The hypotheses of the studies proposed are that that HCMV, and more specifically, the viral envelope glycoprotein B (gB) and/or immediate- early gene products, IE1-72 and IE2-86, inhibit EVT invasion during placentation through activation of TGF-21 and modulation of MMP activity; and further, that hypoxia increases the ability of HCMV to inhibit EVT invasion through upregulation of HCMV IE expression and lytic replication. The Specific Aims to test these hypotheses are (1) to determine whether the inhibition of invasion of cultured EVT by HCMV is mediated by early events in the HCMV replication cycle (2) to determine whether HCMV represses invasion- promoting MMP-2, MMP-3, MMP-9 and uPA, and upregulates invasion-repressing TIMP-1, TIMP-2 and PAI-1 in cultured EVT and whether HCMV inhibits the invasiveness of cultured EVT through activation of TGF-21 and (3) to determine whether the relatively hypoxic atmosphere encountered by differentiating CTB during first trimester pregnancy enhances HCMV-induced inhibition of interstitial and endovascular invasion by EVT through increased lytic HCMV replication. Elucidating mechanisms by which HCMV impairs placentation may be key to understanding fetal and maternal pathologies associated with intrauterine HCMV infection. PUBLIC HEALTH RELEVANCE: Fifteen percent of women with primary human cytomegalovirus (HCMV) infection spontaneously abort during early pregnancy, and it is the placenta, not the embryo or fetus, that shows evidence of infection. Additionally, HCMV infection may cause premature delivery, intrauterine growth restriction or pre-eclampsia, all of which are associated with placental pathology. These complications are believed to be, at least in part, the result of inadequate extravillous cytotrophoblast invasion (EVT) of the uterine wall and impaired remodeling of the maternal spiral arteries during early stages of placental development. Defining how HCMV impairs EVT invasion is of major importance to the reproductive community and may provide additional therapeutic targets to maintain viable pregnancy.