Nod1 activation by bacterial iE-DAP induces maternal-fetal inflammation and preterm labor.

There is a strong association between infection and prematurity; however, the underlying mechanisms remain largely unknown. Nod1 and Nod2 are intracellular pattern recognition receptors that are activated by bacterial peptides and mediate innate immunity. We previously demonstrated that human first-trimester trophoblasts express Nod1 and Nod2, which trigger inflammation upon stimulation. This study sought to determine the expression and function of Nod1 and Nod2 in third-trimester trophoblasts, and to characterize the in vivo effects of Nod1 activation on pregnancy outcome. Human term placental tissues and isolated term trophoblast expressed Nod1, but not Nod2. Activation of Nod1 by its agonist, bacterial γ-D-glutamyl-meso-diaminopimelic acid (iE-DAP), in term trophoblast cultures induced a proinflammatory cytokine profile, characterized by elevated levels of secreted IL-6, GRO-α, and MCP-1, when compared with the control. However, these cytokines were not upregulated in response to Nod2 stimulation with bacterial MDP. Administration of high-dose bacterial iE-DAP to pregnant C57BL/6J mice on embryonic day 14.5 triggered preterm delivery within 24 h. iE-DAP at a lower dose that did not induce prematurity, reduced fetal weight, altered the cytokine profile at the maternal-fetal interface, and induced fetal inflammation. Thus, functional Nod1 is expressed by trophoblast cells across gestation and may have a role in mediating infection-associated inflammation and prematurity. This study demonstrates that pattern recognition receptors, other than the TLRs, may be implicated or involved in infection-associated preterm labor.

Cutting-edge report: TLR10 plays a role in mediating bacterial peptidoglycan-induced trophoblast apoptosis.

PROBLEM: There is a strong correlation between intrauterine bacterial infection and preterm labor. While inflammation is a common mechanism, certain pathogens may trigger placental apoptosis. TLR2 activation by gram-positive bacterial peptidoglycan (PDG) induces first-trimester trophoblast apoptosis and decreased IL-6 secretion. This is dependent upon the presence of TLR1 and the absence of TLR6 and both TLR2 coreceptors. As TLR10 is also a TLR2 coreceptor, the objective of this study was to determine its expression and function in the trophoblast. METHOD OF STUDY: First-and third-trimester human placental tissue and isolated trophoblast were evaluated for TLR10 expression. A first-trimester human trophoblast cell line stably transfected with a TLR10 dominant negative (TLR10-DN) or vector control was treated with or without PDG and analyzed for apoptosis and IL-6. RESULTS: TLR10 was expressed by trophoblasts during the first and third trimesters of pregnancy. PDG-induced trophoblast caspase-3 activity was inhibited by the presence of the TLR10-DN. The presence of the TLR10-DN had no effect on PDG reduction in trophoblast IL-6 secretion. CONCLUSION: This study demonstrates that trophoblast TLR10 plays a role in promoting apoptosis triggered by gram-positive bacterial components and suggests that TLR10 may regulate the balance between trophoblast survival and cell death.

Uric acid induces trophoblast IL-1ß production via the inflammasome: implications for the pathogenesis of preeclampsia.

PROBLEM: Preeclampsia is associated with hyperuricemia, which correlates with the disease severity. Levels of circulating uric acid increase before the clinical manifestations, suggesting that they may be causally related. Uric acid, or monosodium urate (MSU), activates the Nod-like receptor, Nalp3, leading to inflammasome activation and IL-1ß processing. Because preeclampsia is associated with placental immune/ inflammatory dysregulation, we sought to determine in the trophoblast, the presence of the Nalp3 inflammasome, and the effect of MSU on its activation. METHOD OF STUDY: Isolated first- and third-trimester trophoblasts were assessed for expression of the inflammasome components, Nalp1, Nalp3, and ASC. First-trimester trophoblast cells were incubated with or without MSU, and after which, IL-1ß secretion and processing and caspase-1 activation were determined. RESULTS: Trophoblast cells expressed Nalp1, Nalp3, and ASC under basal conditions. Following incubation with MSU, first-trimester trophoblast IL-1ß secretion was upregulated. This correlated with increased expression levels of active IL-1ß and active caspase-1. ASC knockdown reduced MSU-induced IL-1ß secretion. CONCLUSION: These findings demonstrate that uric acid activates the inflammasome in the trophoblast, leading to IL-1ß production. This may provide a novel mechanism for the induction of inflammation at the maternal­fetal interface leading to placental dysfunction and adverse pregnancy outcome, including preeclampsia.

Antiphospholipid antibodies limit trophoblast migration by reducing IL-6 production and STAT3 activity.

PROBLEM: Women with antiphospholipid antibodies (aPL) are at risk of recurrent miscarriage and pre-eclampsia. aPL target the placenta by binding to beta(2)-glycoprotein I (beta(2) GPI) expressed by the trophoblast. The objective of this study was to evaluate if and how aPL affect first trimester trophoblast migration. METHOD OF STUDY: First trimester trophoblast cells were treated with anti-beta(2) GPI monoclonal antibodies. Migration was determined using a two-chamber assay. Interleukin (IL)-6 production was evaluated by RT-PCR and enzyme-linked immunosorbent assay, and signal transducer and activator of transcription 3 (STAT3) activation was assessed by western blot. RESULTS: Trophoblast cells constitutively secreted IL-6 in a time-dependent manner and this directly correlated with STAT3 phosphorylation. In the presence of anti-beta(2) GPI Abs, trophoblast IL-6 mRNA levels and secretion was downregulated in a Toll-like receptor 4/MyD88-independent manner and this correlated with a reduction in phosphorylated STAT3 levels. In addition, the anti-beta(2) GPI Abs reduced the migratory potential of trophoblast. Heparin was able to reverse aPL-dependent inhibition of trophoblast IL-6 secretion and migration. CONCLUSION: This study demonstrates that aPL limit trophoblast cell migration by downregulating trophoblast IL-6 secretion and STAT3 activity. As heparin was unable to prevent these effects, our findings may explain why women with antiphospholipid syndrome, treated with heparin, remain at risk of developing obstetrical syndromes, associated with impaired deep placentation, such as pre-eclampsia.