Decidual NK response to infection

蜕膜NK细胞（dNK）是妊娠期母胎界面上最大的母体免疫细胞群， 妊娠早期，直接接触胎儿绒毛外滋养层细胞（EVT），其侵入蜕膜， 重塑血管系统建立胎盘的血液供应dNK和EVT之间的直接联系 挑战母体免疫系统，它必须耐受胎儿细胞，但仍能防止感染。如何 dNK保护胎盘和胎儿免受感染还不清楚。大多数临床显著感染 胎盘和胎儿是由细胞内病原体（细菌、寄生虫和病毒）引起的， 淋巴细胞（NK和细胞毒性T淋巴细胞）是全身保护性免疫的关键。在怀孕的头三个月， 当感染具有最严重的胎儿后果时，蜕膜中几乎没有T细胞。虽然dNK 具有细胞毒性颗粒，表达所有细胞毒性分子，并杀死常规NK细胞靶标，其细胞溶解性 与外周血NK细胞相比，活性降低。此外，尽管dNK与EVT形成联系， 不驱逐或杀害 人巨细胞病毒感染EVT。这些发现强调了 母体免疫细胞清除胎盘感染，防止病原体传播给未出生的孩子。 该方案研究了我们最近发现的一种新的令人兴奋的机制，dNK通过该机制杀死L。 在滋养层细胞内的单核细胞增多症（Lm），而不杀死宿主细胞。dNK表达大量颗粒溶素 （GNLY），一种在细胞毒性颗粒和细胞溶质中发现的抗菌肽， 微生物的，相对于哺乳动物的，膜。我们的初步数据表明，dNK建立纳米管 与EVT的细胞质连接。在不形成常规免疫突触或脱粒的情况下，dNK 通过纳米管将GNLY转移到EVT，而不是其他细胞毒性分子（穿孔素，颗粒酶），这将杀死 宿主细胞这种机制为妊娠防御的免疫困境提供了一个优雅的解决方案 同时保持胎儿和胎盘的耐受性。据我们所知，这是第一个 纳米管免疫功能的证据。GNLY和潜在的其他生物活性物质的纳米管转移 从dNK到EVT的分子有助于控制细胞内感染，并可调节滋养层功能。我们 我们的目标是证实我们的初步数据，即细胞内微生物，而不是胎儿细胞，被杀死， GNLY的dNK转移，独立于穿孔素和颗粒酶;鉴定哪些感染母体和胎儿细胞 胎盘中dNK的保护作用及其机制;探讨nanotube的作用机制 形成，哪些分子被转移，哪些在怀孕中重要的病原体是易感的。的 GNLY和dNK的保护作用也将在人胎盘组织外植体和小鼠中进行评价， 比较GNLY转基因（Tg）和WT小鼠感染后的妊娠结局， 表达GNLY。这些外植体和体内研究将调查三种妊娠病原体- Lm，B组 链球菌（GBS）和刚地弓形虫。

Decidual NK cells (dNK), the largest population of maternal immune cells at the maternal-fetal interface in the first trimester of pregnancy, directly contact fetal extravillous trophoblasts (EVT), which invade the decidua to remodel the vasculature to establish the blood supply to the placenta. The direct contact between dNK and EVT challenges the maternal immune system, which must tolerate fetal cells, but still protect against infection. How dNK protect the placenta and fetus from infection is not well understood. Most clinically significant infections of the placenta and fetus are caused by intracellular pathogens (bacteria, parasites and viruses), for which killer lymphocytes (NK and cytotoxic T lymphocytes) are key to systemic protective immunity. In the first trimester, when infection has the most serious fetal consequences, there are few T cells in the decidua. Although dNK have cytotoxic granules, express all the cytotoxic molecules, and kill conventional NK cell targets, their cytolytic activity is reduced compared to peripheral blood NK cells. Moreover, although dNK form contacts with EVT, they do not degranulate or kill human cytomegalovirus-infected EVT. These findings emphasize the difficulties of maternal immune cells to clear placental infections and prevent transmission of pathogens to the unborn child. This proposal investigates a novel and exciting mechanism we recently discovered by which dNK kill L. monocytogenes (Lm) inside trophoblasts, without killing the host cell. dNK express large amounts of granulysin (GNLY), an antimicrobial peptide found both in cytotoxic granules and the cytosol that preferentially disrupts microbial, relative to mammalian, membranes. Our preliminary data suggest that dNK establish nanotube cytoplasmic connections to EVT. Without forming a conventional immune synapse or degranulating, dNK transfer GNLY via nanotubes to EVT, but not other cytotoxic molecules (perforin, granzymes), which would kill the host cell. This mechanism provides an elegant solution to the immune dilemma of pregnancy – defense against infection while maintaining tolerance of the fetus and placenta. As far as we are aware, this is the first evidence for an immune function of nanotubes. Nanotube transfer of GNLY and potentially other bioactive molecules from dNK to EVT helps control intracellular infection and could regulate trophoblast functions. Our goals are to confirm our preliminary data showing that intracellular microbes, but not fetal cells, are killed by dNK transfer of GNLY, independently of perforin and granzymes; identify which infected maternal and fetal cells in the placenta dNK protect and by what mechanism; explore the mechanism responsible for nanotube formation, which molecules are transferred and which pathogens important in pregnancy are susceptible. The protective role of GNLY and dNK will also be evaluated in human placental tissue explants and in mice by comparing pregnancy outcomes following infection of GNLY-transgenic (Tg) and WT mice, which do not express GNLY. These explant and in vivo studies will investigate three pathogens of pregnancy - Lm, Group B Streptococci (GBS) and Toxoplasma gondii.