Immune Gene Regulation in Human Trophoblast Cells

描述（由申请人提供）：成功怀孕要求抗原性不同的胎儿避免受到母亲免疫系统的攻击。胎盘滋养细胞（tbc）是唯一与母体血液直接接触的胚泡来源细胞，它们在调节母体对妊娠的免疫反应中起重要作用。干扰素-3 （IFN-3）是一种促炎细胞因子，在多种细胞过程中发挥关键作用，包括诱导细胞凋亡、抑制细胞增殖和启动适应性免疫反应。重要的是，在正常怀孕期间，IFN-3存在于子宫中。所有胎龄的人TBCs都对ifn -3介导的细胞凋亡和多态MHC抗原表达的激活具有抗性。这表明人类tbc已经进化出了逃避IFN-3有害影响的机制。对IFN-3的应答是通过JAK-STAT-1途径激活转录介导的。重要的是，多种病原体通过破坏响应IFN-3的JAK-STAT通路来逃避宿主免疫反应。细胞对IFN-3反应的强度和持续时间取决于正作用的jak和STATs活性与负作用的蛋白酪氨酸磷酸酶（PTPs）和细胞因子信号抑制因子（SOCS）活性之间的平衡。调节IFN-3信号传导的PTPs是组成性活性的，但在细胞暴露于IFN-3后会短暂失活。ptp的失活导致jak的最大激活和细胞因子信号的传播。我们之前已经证明，通过ptp介导的jak抑制，IFN-3反应在人原发期tbc和tbc衍生的绒毛膜癌（CC）细胞中被抑制。我们的动力学研究表明，即使在IFN-3治疗的早期时间点，jak在TBCs中也被抑制。这些结果表明，暴露于IFN-3的tbc中，PTPs并未失活。我们最近也证明了SOCS-1和SOCS-3在tbc中组成性表达。因此，在Specific Aim I中，我们将研究1)ptp在IFN-3处理的tbc中是否与其他细胞类型有差异调节，以及2)SOCS-1和SOCS-3在tbc中调节IFN-3反应中的作用。我们的集体研究表明，抑制JAK是tbc抑制IFN-3反应的主要机制，但它们并没有解决JAK或STAT-1异常激活对滋养细胞对IFN-3反应的影响。最近的研究发现，不明原因的妊娠丢失与JAK-2的构成活性（CA）突变体之间存在关联。此外，STAT-1在肿瘤的免疫监视中起着至关重要的作用。因此，在Specific Aim II中，我们将研究JAKs和STAT-1的CA突变体对TBC IFN-3反应的影响。目前尚不清楚TBCs中IFN-3信号的抑制是否发生在整个妊娠期，还是仅限于妊娠的特定三个月。因此，在特异性目标III中，我们将检查IFN-3在妊娠1、2和3个月tbc中的反应。我们的研究将为孕妇如何避开对IFN-3的有害反应，以及tbc中异常的IFN-3信号传导如何导致妊娠并发症提供见解。公共卫生相关性：最近的研究表明，妊娠是一种控制炎症的状态，胎盘中IFN-3水平的改变在胎盘病理与生殖成功之间起着关键作用。我们提出的研究将通过提供tbc调节IFN-3反应的分子机制的全面分析，阐明tbc如何在正常妊娠期间和感染后补偿子宫中促炎IFN-3的存在。因此，我们的研究将为孕妇如何避开IFN-3的潜在有害反应，以及tbc中异常的细胞因子信号如何导致妊娠并发症提供见解。这项工作也对肿瘤免疫具有重要意义，因为肿瘤细胞对IFN-3的反应性丧失在肿瘤免疫逃避中起重要作用。

DESCRIPTION (provided by applicant): Successful pregnancy requires that the antigenically disparate fetus avoid attack by the mother's immune system. Placental trophoblast cells (TBCs) are the only blastocyst-derived cells in direct contact with maternal blood, and they play essential roles in modulating maternal immune responses to the conceptus. Interferon-3 (IFN-3) is a pro-inflammatory cytokine that plays critical roles in diverse cellular processes, including induction of apoptosis, inhibition of cell proliferation and initiation of adaptive immune responses. Importantly, IFN-3 is present in the uterus during normal pregnancy. Human TBCs from all gestational ages examined are resistant to both IFN-3-mediated apoptosis, and activation of polymorphic MHC antigen expression. This suggests that human TBCs have evolved mechanisms to evade the harmful effects of IFN-3. Responses to IFN-3 are mediated by activation of transcription through the JAK-STAT-1 pathway. Importantly, multiple pathogens evade host immune responses by disrupting the JAK-STAT pathway in response to IFN-3. The magnitude and duration of cellular responses to IFN-3 are dictated by the equilibrium between the activities of the positively acting JAKs and STATs, versus negatively acting protein tyrosine phosphatases (PTPs) and suppressors of cytokine signaling (SOCS). The PTPs that regulate IFN-3 signaling are constitutively active, but are transiently inactivated following exposure of cells to IFN-3. Inactivation of the PTPs results in maximal activation of the JAKs and propagation of the cytokine signal. We previously demonstrated that IFN-3 responses are inhibited in human primary term TBCs and TBC-derived choriocarcinoma (CC) cells by PTP-mediated repression of the JAKs. Our kinetic studies demonstrate that JAKs are inhibited in TBCs even at early time points of IFN-3 treatment. These results suggest that the PTPs are not inactivated in TBCs exposed to IFN-3. We also recently demonstrated that SOCS-1 and SOCS-3 are constitutively expressed in TBCs. Thus, in Specific Aim I, we will investigate 1) whether PTPs are differentially regulated in IFN-3-treated TBCs versus other cell types, and 2) the roles of SOCS-1 and SOCS-3 in regulating IFN-3 responses in TBCs. Our collective studies suggest that inhibition of the JAKs is the primary mechanism by which TBCs suppress IFN-3 responses, but they do not address the consequences of aberrant JAK or STAT-1 activation on trophoblastic responses to IFN-3. Recent studies identified an association between unexplained pregnancy loss and a constitutively active (CA) mutant of JAK-2. Moreover, STAT-1 plays an essential role in immunosurveillance of tumors. Thus, in Specific Aim II, we will investigate the effects of CA mutants of the JAKs and STAT-1 on TBC IFN-3 responses. It is currently unclear whether inhibition of IFN-3 signaling in TBCs occurs throughout gestation, or is restricted to specific trimesters of pregnancy. Thus, in Specific Aim III we will examine IFN-3 responses in 1st, 2nd and 3rd trimester TBCs. Our studies will provide insights into how the conceptus evades deleterious responses to IFN-3, and how aberrant IFN-3 signaling in TBCs may lead to complications of pregnancy. PUBLIC HEALTH RELEVANCE: Recent studies suggest that pregnancy is a state of controlled inflammation, and that alterations in placental levels of IFN-3 play a critical role in placental pathology versus reproductive success. Our proposed studies will shed light on how TBCs compensate for the presence of pro-inflammatory IFN-3 in the uterus during normal pregnancy and following infection by providing a comprehensive analysis of the molecular mechanism(s) by which TBCs regulate responses to IFN-3. Thus, our studies will provide insights into how the conceptus evades potentially deleterious responses to IFN-3, and how aberrant cytokine signaling in TBCs may lead to pregnancy complications. This work also has important implications for tumor immunity, for loss of responsiveness to IFN-3 by tumor cells plays an essential role in tumor immunoevasion.