Shaun Murphy

Uterine NK Cells Mediate Inflammation-Induced Fetal Demise in IL-10-Null Mice

Specialized NK cells are recruited in high numbers to the mammalian embryo implantation sites, yet remain pregnancy compatible. It is not well understood whether uterine NK (uNK) cells become adversely activated and mediate fetal demise, a common complication of early pregnancy. In this study we show that mating of IL-10−/− mice resulted in fetal resorption or intrauterine growth restriction in response to very low doses of LPS. Pregnancy in congenic wild-type mice was normal even at 10-fold higher LPS doses. Fetal resorption in IL-10−/− mice was associated with a significant increase in uNK cell cytotoxic activation and invasion into the placenta. Depletion of uNK cells, TNF-α neutralization, or IL-10 administration rescued pregnancy in LPS-treated IL-10−/− animals. Our results identify an immune mechanism of fetal demise involving IL-10 deficiency, NK cells, and inflammation. These results may provide insight into adverse pregnancy outcomes in humans.

Evidence for Interleukin-10-Mediated Inhibition of Cyclo- oxygenase-2 Expression and Prostaglandin Production in Preterm Human Placenta

Problem  Interleukin‐10 (IL‐10) is thought to be a key cytokine for the maintenance of pregnancy. Here we examined the expression profiles of IL‐10 and cyclo‐oxygenase‐2 (COX‐2), and the effect of IL‐10 on COX‐2 expression and prostaglandin release in the human placenta from preterm labor deliveries associated with chorioamnionitis.

TLR6 Modulates First Trimester Trophoblast Responses to Peptidoglycan

Intrauterine bacterial infections are a well-established cause of pregnancy complications. One key observation in a number of abnormal pregnancies is that placental apoptosis is significantly elevated. First trimester trophoblast cells are known to express TLR1 and TLR2 and to undergo apoptosis following exposure to Gram-positive bacterial peptidoglycan (PDG). Thus, the objectives of this study were to determine whether PDG-induced pregnancy complications are associated with placental apoptosis and to characterize the cellular mechanisms involved. We have demonstrated, using an animal model, that delivery of PDG to pregnant mice early in gestation resulted in highly elevated placental apoptosis, evidenced by trophoblast M-30 and active caspase 3 immunostaining. Using an in vitro model of human first trimester trophoblasts, apoptosis induced by PDG was found to be mediated by both TLR1 and TLR2 and that this could be blocked by the presence of TLR6. Furthermore, in the presence of TLR6, exposure to PDG resulted in trophoblast NF-κB activation and triggered these cells to secrete IL-8 and IL-6. The findings of this study suggest that a Gram-positive bacterial infection, through TLR2 and TLR1, may directly promote the elevated trophoblast cell death and that this may be the underlying mechanism of pregnancy complications, such as preterm delivery. Furthermore, the expression of TLR6 may be a key factor in determining whether the response to PDG would be apoptosis or inflammation.

Evidence for participation of uterine natural killer cells in the mechanisms responsible for spontaneous preterm labor and delivery

OBJECTIVE The purpose of this study was to determine in a mouse model whether uterine natural killer (uNK) cell cytotoxic activation induces infection/inflammation-associated preterm labor and delivery. STUDY DESIGN Wild type or interleukin (IL)-10(-/-) mice were injected intraperitoneally with lipopolysaccharide on gestational day 14. Mice were either killed for collection of uteroplacental tissue, spleen, and serum or allowed to deliver. Uteroplacental tissue was used for histology and characterization of uNK cells. RESULTS Low-dose lipopolysaccharide treatment triggered preterm labor and delivery in IL-10(-/-), but not wild type mice, in a manner independent of progesterone levels. Preterm labor and delivery in IL-10(-/-) mice was associated with an increased number and placental infiltration of cytotoxic uNK cells and placental cell death. Depletion of NK cells or tumor necrosis factor (TNF)-alpha neutralization in these mice restored term delivery. Furthermore, TNF-alpha neutralization prevented uNK cell infiltration and placental cell apoptosis. CONCLUSION The uNK cell-TNF-alpha-IL-10 axis plays an important role in the genesis of infection/inflammation-induced preterm labor/delivery.

Deficiency of decidual IL-10 in first trimester missed abortion: a lack of correlation with the decidual immune cell profile.

PROBLEM: To determine if first trimester missed abortion decidua is characterized by an altered immune cell profile and/or a modified interleukin (IL)‐10 and interferon (IFN)‐γ production pattern compared with decidua from elective termination.

IFN‐γ‐Mediated Inhibition of COX‐2 Expression in the Placenta from Term and Preterm Labor Pregnancies

Problem:  The inflammatory‐anti‐inflammatory cytokine network is thought to play a critical role in regulated progression and termination of pregnancy. The aim of this study was to evaluate the effects of interferon (IFN)‐γ on the expression of Cyclooxygenase (COX)‐2 and production of prostaglandin E2 (PGE2) in the human placenta from term and preterm labor deliveries.

Inter-Alpha Inhibitor Protein Administration Improves Survival From Neonatal Sepsis in Mice

Inter-alpha inhibitor proteins (IaIp) are serine proteases inhibitors that modulate endogenous protease activity and have been shown to improve survival in adult models of sepsis. We evaluated the effect of IaIp on survival and systemic responses to sepsis in neonatal mice. Sepsis was induced in 2-d-old mice with lipopolysaccharide (LPS), Escherichia coli, and group B Streptococci. Sepsis was associated with 75% mortality. IaIp, given by i.p. administration at doses between 15 and 45 mg/kg from 1 to 6 h after the onset of sepsis, improved survival to ∼90% (p = 0.0159) in both LPS-induced sepsis and with live bacterial infections. The greatest effect was on reversal of hemorrhagic pneumonitis. The effects were dose and time dependent. Systemic cytokine profile and tissue histology were examined. Survival was compared in IL-10 knock out animals. Systemic cytokine levels including TNF-α and IL-10 were increased after induction of sepsis and modulated significantly after IaIp administration. Because the effect of IaIp was still demonstrable in IL-10 deficient mice, we conclude the beneficial effects of IaIp is because of suppression of proinflammatory cytokines such as TNF-α rather than augmentation of IL-10. IaIp may offer significant benefits as a therapeutic adjunct to treatment of sepsis in neonates and adults.

IL-10 and Pregnancy

Soon after the principles of nonself immunological recognition were discovered, it was realized that the state of pregnancy seemingly presents a paradox. In an outbreed population, half of the fetal genes are paternal, thus the fetus may be considered a semi-allograft. Yet, unlike the outcome of a surgical tissue graft, the mother tolerates and nurtures the fetus.

1141635485 Genetic stress (IL‐10 deficiency) and toxicant (PCB)‐induced disruption of pregnancy

Several epidemiologic studies have provided evidence that a significant number of pregnant women living within the World Trade Center disaster area experienced a high rate of low birth weight and premature deliveries. Our hypothesis is that these women were predisposed to pregnancy disruption effects of toxicants detected in the dust of the disaster area. We sought to examine the effects of PCB on pregnancy outcome in an IL‐10‐/‐ mouse model. IL‐10‐/‐ or congenic wild type mice of 6–8 weeks age were given i.p injections of Aroclor 1254 (PCB) at doses of 50 or 500 μg/mouse in 100 μL corn oil on gestational day (gd) 1 and 7 or an equivalent (100 μL) amount of corn oil. Mice were sacrificed on gd 13 and uteri and spleen were excised and collected. Uterine mononuclear cells (UMC) and splenic mononuclear cells (SMC) were isolated. These cell populations were subjected to phenotypic and functional characterization. A portion of uterine tissue was either snap frozen for placental PCB contents or for histology. No doses of Aroclor 1254 affected the pregnancy outcome in wild type mice. However, IL‐10‐/‐ mice experienced fetal resorption when injected with high dose (500 μg/mouse) of Aroclor 1254. Local immune dysregulation and placental anomalies including apoptosis and immunocyte infiltration are currently being investigated. Our results strongly suggest that IL‐10 deficiency and PCB‐mediated immunotoxicity lead to adverse pregnancy outcomes.

1141635158 IL-10 deficiency and uterine NK cell cytotoxic activation link inflammation to preterm parturition

Preterm birth is a major cause of perinatal morbidity and mortality. Intrauterine infection/inflammation is associated with a majority of preterm labor and birth cases. Despite decades of studies recognizing a strong association between infection/inflammation and preterm birth, no effective method of preventing infection‐induced premature labor and delivery is yet available. Importantly, the mechanisms by which intrauterine infection/inflammation may contribute to preterm birth are not known. Based on our observations with human gestational tissue to highlight the role of IL‐10 in normal and comprimised pregnancy outcomes, we have performed experiments with syngeneic and allogeneic pregnant IL‐10‐/‐ mice or congenic wild type mice. Pregnancy outcomes were assessed in response to i.p. administration of low doses of lipopolysacchade (LPS) on gd 14. The mice were allowed to deliver or were sacrificed on gd 16 for isolation of uterine immune cells for functional studies or collected tissue for histological analysis. Attempts were made to prevent preterm parturition. LPS‐treated IL‐10‐/‐, but not wild type mice, displayed a significant acceleration in time of delivery, on gd 16.5 compared to gd 19.6 for wild type controls. The premature delivery observed in LPS‐treated IL‐10‐/‐ mice was associated with an increase in the number of uterine NK (uNK) cells. These cells also displayed a dramatic infiltration of the placenta with a perivascular localization. uNK cells appear to be responsible for the induction of preterm birth in these mice as depletion of NK cells completely restored normal length of gestation. Moreover, neutralization TNF‐α also rescued the premature delivery. Taken together, our results for the first time demonstrate that IL‐10 deficiency and uterine NK cell cytotoxic activation link intrauterine inflammation to preterm parturition.