Human Cytomegalovirus-Induced Inhibition of Cytotrophoblast Invasion

描述（由申请方提供）：显著的发病率和死亡率与疱疹病毒人巨细胞病毒（HCMV）的症状性先天性感染相关。15%的原发性HCMV感染的妇女在怀孕早期自然流产;并且是胎盘，而不是胚胎或胎儿显示感染的证据。胎盘HCMV感染的病理后果，包括早期妊娠丢失、宫内生长受限、先兆子痫和早产，被认为是由绒毛外细胞滋养层细胞（EVT）在早期妊娠期间不能充分侵入子宫壁介导的，从而导致母体螺旋动脉重塑受损和浅胎盘形成。初步研究表明，HCMV感染早期妊娠EVT的结果在EVT的侵袭抑制和显着降低的表达和活性的侵袭促进基质金属蛋白酶（MMP）-2和MMP-9。HCMV诱导的EVT抑制沿着转化生长因子（TGF）-21（一种已知抑制EVT侵袭的因子）的表达增加。由于EVT侵入发生在妊娠早期，此时胎盘环境的胎儿侧相对缺氧，并且由于缺氧在其他系统中能够激活疱疹病毒基因表达和裂解性复制，因此胎盘形成期间的低氧分压实际上可能增强HCMV抑制EVT侵入的能力。为了确定HCMV抑制EVT侵袭的分子机制，这是本文提出的研究的广泛的长期目标，已经使用生物反应器组织工程技术开发了EVT侵袭的新的生物学相关模型。提出的研究假设是HCMV，更具体地说，病毒包膜糖蛋白B（g B）和/或立即早期基因产物IE 1 -72和IE 2 -86，通过激活TGF-21和调节MMP活性抑制胎盘形成期间EVT侵入;此外，缺氧通过上调HCMV IE表达和裂解性复制增加HCMV抑制EVT侵袭的能力。检验这些假设的具体目的是（1）确定HCMV对培养的EVT侵袭的抑制是否由HCMV复制周期中的早期事件介导（2）确定HCMV是否抑制侵袭促进MMP-2、MMP-3、MMP-9和uPA，并上调侵袭抑制TIMP-1，培养的EVT中TIMP-2和派-1的表达，以及HCMV是否通过激活TGF-21抑制培养的EVT的侵袭性，以及（3）确定在妊娠早期通过分化CTB所遇到的相对低氧环境是否增强HCMV-1的表达。EVT通过增加裂解性HCMV复制诱导抑制间质和血管内侵袭。阐明HCMV损害胎盘形成的机制可能是了解宫内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）感染的妇女在妊娠早期自发流产，并且显示感染证据的是胎盘，而不是胚胎或胎儿。此外，HCMV感染可导致早产、宫内生长受限或先兆子痫，所有这些都与胎盘病理学有关。这些并发症被认为是，至少部分是由于子宫壁的绒毛外细胞滋养层浸润（EVT）不足和胎盘发育早期母体螺旋动脉重塑受损。明确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.