Preeclampsia-like Phenotype Research Articles

Periconceptional 1,3-butanediol supplementation suppresses the superimposed preeclampsia-like phenotype in the Dahl salt-sensitive rat.

Preeclampsia is a hypertensive pregnancy disorder with no treatment beyond management of symptoms and delivery of the fetus and placenta. Chronic hypertension increases the risk of developing superimposed preeclampsia. Previous reports showed that 1,3-butanediol attenuates hypertension in rodents; however, the therapeutic potential of 1,3-butanediol for the prevention of preeclampsia has not been investigated. This study tested the hypothesis that attenuating hypertension before pregnancy and through the placentation period via 1,3-butanediol prevents the onset of preeclampsia in female Dahl salt-sensitive (SS/Jr) rats. Female Dahl SS/Jr rats were divided into two groups: 1,3-butanediol treated (20% via drinking water) and control (ad libitum water). Both groups were maintained on low-salt rodent chow (Teklad 7034, 0.3% NaCl; n = 8/group). Animals were treated with 1,3-butanediol for 7 wk (baseline), mated, and treated through day 12 of pregnancy. 1,3-Butanediol treatment increased plasma β-hydroxybutyrate (metabolite of 1,3-butanediol) that negatively correlated with maternal body weight in late pregnancy. Mean arterial pressure was lower in the treated group at baseline, early, and mid pregnancy, but no difference was observed in late pregnancy after treatment ended. Uterine artery resistance index (UARI) was reduced in the treated dams. No adverse fetal effects were observed, and there were no differences in pup weight or length. Placentas from treated dams had decreased vascular endothelial growth factor levels as well as decreased placental basal zone thickness and increased labyrinth zone thickness. These findings support the therapeutic role of physiological ketosis via 1,3-butanediol as a potential therapeutic approach for managing chronic hypertension, thereby preventing and mitigating adverse pregnancy outcomes associated with preeclampsia.NEW & NOTEWORTHY A ketogenic diet or increased β-hydroxybutyrate levels can reduce hypertension, but the potential of 1,3-butanediol, a β-hydroxybutyrate precursor, for treatment of preeclampsia is unknown. We hypothesized that attenuating hypertension before and during pregnancy via 1,3-butanediol prevents preeclampsia in Dahl Salt-sensitive rats. 1,3-Butanediol significantly lowered blood pressure and improved uterine artery resistance with no observable adverse fetal effects. Physiological ketosis via 1,3-butanediol may be a potential therapeutic approach for managing hypertension and mitigating adverse pregnancy outcomes.

Human Leucocyte Antigen G and Murine Qa-2 Are Critical for Myeloid Derived Suppressor Cell Expansion and Activation and for Successful Pregnancy Outcome.

During pregnancy, maternal immune system has to balance tightly between protection against pathogens and tolerance towards a semi-allogeneic organism. Dysfunction of this immune adaptation can lead to severe complications such as pregnancy loss, preeclampsia or fetal growth restriction. In the present study we analyzed the impact of the murine MHC class Ib molecule Qa-2 on pregnancy outcome in vivo. We demonstrate that lack of Qa-2 led to intrauterine growth restriction and increased abortion rates especially in late pregnancy accompanied by a disturbed trophoblast invasion and altered spiral artery remodeling as well as protein aggregation in trophoblast cells indicating a preeclampsia-like phenotype. Furthermore, lack of Qa-2 caused imbalanced immunological adaptation to pregnancy with altered immune cell and especially T-cell homeostasis, reduced Treg numbers and decreased accumulation and functional activation of myeloid-derived suppressor cells. Lastly, we show that application of sHLA-G reduced abortion rates in Qa-2 deficient mice by inducing MDSC. Our results highlight the importance of an interaction between HLA-G and MDSC for pregnancy success and the therapeutic potential of HLA-G for treatment of immunological pregnancy complications.

Characterization of the temporal, cell-specific and interferon-inducible patterns of indoleamine 2,3 dioxygenase 1 (IDO1) expression in the human placenta across gestation.

The human placenta performs multiple functions necessary for successful pregnancy, but the metabolic pathways and molecular mechanisms responsible for regulating placental development and functions remain incompletely understood. Catabolism of the essential amino acid tryptophan has numerous critical roles in normal physiology, including inflammation. The kynurenine pathway, which accounts for ∼90% of tryptophan breakdown, is mediated by indoleamine 2,3 dioxygenase 1 (IDO1) in the placenta. In pregnant mice, alterations of IDO1 activity or expression result in fetal resorption and a preeclampsia-like phenotype. Decreased IDO1 expression at the maternal-fetal interface has also been linked to preeclampsia, in utero growth restriction and recurrent miscarriage in humans. These collective observations suggest essential role(s) for IDO1 in maintaining healthy pregnancy. Despite these important roles, the precise temporal, cell-specific and inflammatory cytokine-mediated patterns of IDO1 expression in the human placenta have not been thoroughly characterized across gestation. Western blot and whole mount immunofluorescence (WMIF) were utilized to characterize and quantify basal and interferon (IFN)-inducible IDO1 expression in 1st trimester (7-13 weeks), 2nd trimester (14-22 weeks) and term (39-41 weeks) placental villi. IDO1 expression is activated in the human placenta between the 13th and 14th weeks of pregnancy, increases through the 2nd trimester and remains elevated at term. Constitutive IDO1 expression is restricted to placental endothelial cells. Interestingly, different types of IFNs have distinct effects on IDO1 expression in the human placenta. Our collective results are consistent with potential role(s) for IDO1 in the regulation of vascular functions in placental villi.

Elevated S100A9 in preeclampsia induces soluble endoglin and IL-1β secretion and hypertension via the NLRP3 inflammasome.

Maternal systemic and placental inflammatory responses participate in the pathogenesis of hypertensive disorders of pregnancy including preeclampsia, a pregnancy-specific syndrome, although the role of inflammation remains unclear. The NLRP3 inflammasome has been implicated in the control of sterile inflammation involved in preeclampsia. In the present study, we hypothesized that S100A9, as major alarmin, are associated with the pathogenesis of preeclampsia and induction of a preeclampsia-like phenotype in pregnant mice. Plasma were taken from normal pregnant women and preeclampsia patients. Human placental tissues, trophoblast cell line Sw.71 cells, and human umbilical vein endothelial cells (HUVEC) were treated with S100A9 with or without inhibitors associated with NLRP3 inflammasome. Pregnant mice were administered S100A9. S100A9 was elevated in plasma and released from placentas of preeclampsia patients. S100A9 activated the NLRP3 inflammasome, resulting in IL-1β secretion, by human placental tissues and trophoblasts. In addition, secretion of soluble endoglin, a main contributor to the pathogenesis of preeclampsia, is regulated via S100A9-stimulated NLRP3 inflammasome activation in the human placenta and HUVECs. S100A9 administration significantly elevated maternal blood pressure and neutrophil accumulation within the placentas of pregnant mice, and both were significantly decreased in Nlrp3-knock out pregnant mice. The results of this study demonstrated that S100A9 acts as a danger signal to activate the NLRP3 inflammasome in the placenta, associating with hypertension during pregnancy.

Investigation of a Paternal-Mediated Preeclampsia-Like Pregnancy Phenotype Mouse Model

Preeclampsia, a pregnancy specific syndrome characterized by new onset hypertension and proteinuria, is a leading cause of maternal and neonatal morbidity and mortality. While no animal model perfectly mimics the human syndrome, breeding C1q-/- (male) to C57 (female) mice results in a preeclampsia-like pregnancy including pregnancy-specific hypertension, vascular dysfunction and altering placental phenotype. As the placental genotype is primarily paternally driven, lack of paternal C1q is likely driving this preeclampsia-like phenotype. However, more work is needed to investigate whether a lack of maternal C1q also contributes to this preeclampsia-like phenotype. The aim of this study was to investigate the pregnancy phenotype of genetic control (C1q-/- female bred to C57 male) mice. Blood pressure was monitored during pregnancy and vascular function assessed during late pregnancy (gestation day 17.5) in genetic control females. These data were compared to similar data obtained from control (C57 male bred to C57 female) and preeclampsia-like (C1q-/- male bred to C57 female) pregnant mice. Genetic control blood pressure and vascular function data were similar to that of the control pregnancy group, indicating no significant effect of maternal C1q deficiency on the “preeclampsia-like” pregnancy phenotype. As understanding preeclampsia and its effect on women’s health is critical, the work presented is important to confirm the C1q-/- x C57 mouse model as a useful model for studying this syndrome further.

Elevated Levels of Soluble Axl (sAxl) Regulates Key Angiogenic Molecules to Induce Placental Endothelial Dysfunction and a Preeclampsia-Like Phenotype.

Preeclampsia (PE), a severe pregnancy-specific syndrome, is characterized by impaired placental angiogenesis. Although the pathogenesis of this condition remains largely unclear, vascular systemic endothelial injury is thought to be the common contributing factor. Soluble Axl (sAxl), a biomarker of endothelial dysfunction, is known to be abnormally increased in a variety of diseases associated with vascular injury. In a previous study, we found that the plasma levels of sAxl were significantly higher in PE with severe features (sPE) than in pregnant women who did not have PE. The current study aimed to further explore the potential role of sAxl in vascular injury in patients with sPE. We found that the upregulation of sAxl in maternal plasma was positively correlated with the plasma levels of sFlt-1 and negatively correlated with placental NO synthase (eNOS) in women with sPE. Furthermore, elevated levels of sAxl suppressed proliferation and endothelial tube formation and promoted cytotoxicity in human umbilical vein endothelial cells (HUVECs) through the downregulation of p-Akt, p-p70S6K, p-mTOR, and Grb2. Subsequently, we established a pregnant rat model with PE-like characteristics by injecting pregnant rats with an adenovirus expressing sAxl. These rats exhibited a typical PE-like phenotype, including increased blood pressure, proteinuria, and fetal growth restriction, along with abnormal placental and fetal renal morphology. In conclusion, our study demonstrated the role of sAxl in systemic vascular injury through the regulation of the expression of key molecules of angiogenesis and described its potential contribution to the development of sPE.

A Novel Peptide Ameliorates TNFα- and LPS-Induced Endothelia Dysfunction in Preeclampsia.

To investigate the protective effects of the novel peptide antiendothelial dysfunction peptide in preeclampsia (AEDPPE) on tumor necrosis factor α (TNFα)- and lipopolysaccharide (LPS)-induced injury in the vascular endothelium in preeclampsia. The effects of AEDPPE on TNFα-induced vascular endothelial injury were assessed by enzyme-linked immunosorbent assay, quantitative real-time PCR, mitochondrial membrane potential assay, Cell Counting Kit-8 assay, THP-1 monocyte-human umbilical vein endothelial cell (HUVEC) adhesion assay, endothelial tube-forming assay, transcriptomic analysis, preeclamptic symptom analysis, and histological analysis in preeclampsia-like rat models induced by LPS. AEDPPE alleviated the upregulation of antiangiogenic factors including soluble fms-like tyrosine kinase-1, endothelin-1, and tissue plasminogen activator and attenuated the reduction in mitochondrial potential induced by TNFα in HUVECs. In addition, AEDPPE treatment counteracted the decrease in tube formation and decreased the numbers of THP-1 monocytes attached to HUVECs caused by TNFα. Mechanistically, cytokine-cytokine receptor interactions enriched many genes and the TNF signaling pathway may be involved in this phenomenon. Moreover, cotreatment with LPS and AEDPPE significantly reversed the preeclampsia-like phenotype including hypertension and proteinuria and improved the functions of the kidney and placenta. AEDPPE effectively ameliorated the vascular endothelial injury induced by TNFα and LPS in preeclampsia. We suggest that AEDPPE may be a novel therapeutic candidate for preeclampsia treatment. These findings demonstrate that AEDPPE may play an effective role in ameliorating vascular endothelial dysfunction and be a potential therapeutic agent for preeclampsia.

30 Viral and bacterial signatures impair trophoblast migration which is reversed by aspirin, aspirin-triggered-lipoxin and hydroxychloroquine

While the etiology of preeclampsia remains elusive, an inflammatory response at the maternal-fetal interface is thought to play a role. Growing evidence suggests a relationship between maternal infection and risk of preeclampsia. Thus, infection may contribute to the pathogenesis by affecting trophoblast function. These cells express Toll-like receptors (TLRs) and Nod-like receptors (NLRs), which mediate infection-induced inflammation. We hypothesize that in addition to inducing inflammation via TLRs and NLRs, infections may limit trophoblast migration leading to a preeclampsia-like phenotype, and this may be reversed by potential therapeutics. A human first trimester trophoblast cell line (Sw.71) was treated with or without bacterial or viral signatures known to activate TLRs and NLRs (Table 1). For some experiments, cells were also treated with the therapeutics aspirin (ASA; 10ug/ml), aspirin-triggered lipoxin (ATL; 100nM), or hydroxychloroquine (HCQ; 1ug/ml). Cell viability was measured using the CellTiter assay. Cell migration was quantified using a two-chamber colorimetric assay. Cell viability was not altered by any infectious triggers with the exception of viral dsRNA which reduced viability by 50.4±4.7% (p<0.05). This was not tested further. Compared to media, viral ssRNA and bacterial CpG ODN significantly reduced trophoblast migration by 48.7±7.2% and 52.5±4.1%, respectively (p<0.05). Bacterial LPS, PGN, flagellin, iE-DAP and MDP did not alter cell migration. Compared to viral ssRNA alone, ASA, ATL and HCQ independently increased trophoblast migration by 1.6±0.3, 2.0±0.3 and 2.0±0.4-fold, respectively (p<0.05). Similarly, compared to CpG ODN alone, ASA, ATL and HCQ independently increased trophoblast migration by 1.6±0.1, 1.5±0.1 and 1.6±0.2-fold, respectively (p<0.05). Our findings suggest that viral ssRNA and bacterial CpG ODN may impair trophoblast migration leading to a preeclampsia-like phenotype. Further research is needed to understand the mechanisms by which ASA, ATL, and HCQ may reverse these effects and improve perinatal outcomes.

Crocin exhibits an antihypertensive effect in a rat model of gestational hypertension and activates the Nrf-2/HO-1 signaling pathway.

Gestational hypertension is a leading cause of both prenatal and maternal mortality and morbidity; however, there have been rather limited advances in the management of gestational hypertension in recent years. There has been evidence supporting the antihypertensive properties of crocin, but the specific mechanism is still unclear. N-Nitro-L-arginine methyl ester (L-NAME) was employed to establish a rat model with a preeclampsia-like phenotype, particularly gestational hypertension. Enzyme-linked immunosorbent assays were conducted to determine the levels of placental growth factor (PlGF) and soluble fms-like tyrosine kinase (sFlt-1); the levels of the circulating cytokines interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α; and oxidative stress factors. Quantitative RT-PCR assays were performed to assess the transcript levels of various cytokines in the placenta, and western blot assays were carried out to evaluate the protein levels of heme oxygenase-1 (HO-1) and nuclear factor-erythroid 2-like 2 (Nrf-2). Treatment with crocin reduced the blood pressure of rats with gestational hypertension, which was accompanied by suppressed circulating levels of PlGF and sFlt-1. Crocin further alleviated the inflammatory signals and oxidative stress in the serum, as well as in placental tissues, in rats with L-NAME-induced hypertension. Crocin treatment also improved pregnancy outcomes in terms of fetal survival, fetal weight, and the fetal/placental weight ratio. Finally, in hypertension elicited by L-NAME, crocin stimulated the placental Nrf-2/HO-1 pathway. Crocin alleviated inflammatory and oxidative stress in placental tissues, thereby protecting against gestational hypertension, one of the major phenotypes of preeclampsia, and activated the Nrf-2/HO-1 pathway.

Adoptive transfer of placental ischemia-stimulated natural killer cells causes a preeclampsia-like phenotype in pregnant rats.

The Reduced Uterine Perfusion Pressure (RUPP) rat model of placental ischemia recapitulates many characteristics of preeclampsia including maternal hypertension, intrauterine growth restriction (IUGR), and increased cytolytic natural killer cells (cNKs). While we have previously shown a 5-fold higher cytotoxicity of RUPP NKs versus normal pregnant NKs, their role in RUPP pathophysiology remains unclear. In this study, we tested the hypotheses that (1) adoptive transfer of RUPP-stimulated NKs will induce maternal hypertension and IUGR in normal pregnant control (Sham) rats and (2) adoptive transfer of Sham NKs will attenuate maternal hypertension and IUGR in RUPP rats. On gestation day (GD)14, vehicle or 5×106 RUPP NKs were infused i.v. into a subset of Sham rats (Sham+RUPP NK), and vehicle or 5×106 Sham NKs were infused i.v. into a subset of RUPP rats (RUPP+Sham NK; n=12/group). On GD18, Uterine Artery Resistance Index (UARI) was measured. On GD19, mean arterial pressure (MAP) was measured, animals were sacrificed, and blood and tissues were collected for analysis. Adoptive transfer of RUPP NKs into Sham rats resulted in elevated NK activation, UARI, placental oxidative stress, and preproendothelin expression as well as reduced circulating nitrate/nitrite. This led to maternal hypertension and IUGR. RUPP recipients of Sham NKs demonstrated normalized NK activation, sFlt-1, circulating and placental VEGF, and UARI, which led to improved maternal blood pressure and normal fetal growth. These data suggest a direct role for cNKs in causing preeclampsia pathophysiology and a role for normal NKs to improve maternal outcomes and IUGR during late gestation.

Mechanisms of vascular dysfunction in the interleukin-10–deficient murine model of preeclampsia indicate nitric oxide dysregulation

Preeclampsia is a pregnancy-specific hypertensive disorder characterized by proteinuria, and vascular injury in the second half of pregnancy. We hypothesized that endothelium-dependent vascular dysfunction is present in a murine model of preeclampsia based on administration of human preeclamptic sera to interleukin-10-/- mice and studied mechanisms that underlie vascular injury. Pregnant wild type and IL-10-/- mice were injected with either normotensive or severe preeclamptic patient sera (sPE) during gestation. A preeclampsia-like phenotype was confirmed by blood pressure measurements; assessment of albuminuria; measurement of angiogenic factors; demonstration of foot process effacement and endotheliosis in kidney sections; and by accumulation of glycogen in placentas from IL-10-/- mice injected with sPE sera (IL-10-/-sPE). Vasomotor function of isolated aortas was assessed. The IL-10-/-sPE murine model demonstrated significantly augmented aortic contractions to phenylephrine and both impaired endothelium-dependent and, to a lesser extent, endothelium-independent relaxation compared to wild type normotensive mice. Treatment of isolated aortas with indomethacin, a cyclooxygenase inhibitor, improved, but failed to normalize contraction to phenylephrine to that of wild type normotensive mice, suggesting the additional contribution from nitric oxide downregulation and effects of indomethacin-resistant vasoconstricting factors. In contrast, indomethacin normalized relaxation of aortas derived from IL-10-/-sPE mice. Thus, our results identify the role of IL-10 deficiency in dysregulation of the cyclooxygenase pathway and vascular dysfunction in the IL-10-/-sPE murine model of preeclampsia and point towards a possible contribution of nitric oxide dysregulation. These compounds and related mechanisms may serve both as diagnostic markers and therapeutic targets for preventive and treatment strategies in preeclampsia.

1123: Diet-induced obesity aggravates preeclampsia-like phenotypes in ASB4-null mice

1123: Diet-induced obesity aggravates preeclampsia-like phenotypes in ASB4-null mice

Reduced mRNA Expression of RGS2 (Regulator of G Protein Signaling-2) in the Placenta Is Associated With Human Preeclampsia and Sufficient to Cause Features of the Disorder in Mice.

Cascade-specific termination of G protein signaling is catalyzed by the RGS (regulator of G protein signaling) family members, including RGS2. Angiotensin, vasopressin, and endothelin are implicated in preeclampsia, and RGS2 is known to inhibit G protein cascades activated by these hormones. Mutations in RGS2 are associated with human hypertension and increased risk of developing preeclampsia and its sequelae. RGS family members are known to influence maternal vascular function, but the role of RGS2 within the placenta has not been explored. Here, we hypothesized that reduced expression of RGS2 within the placenta represents a risk factor for the development of preeclampsia. Although cAMP/CREB signaling was enriched in placentas from human pregnancies affected by preeclampsia compared with clinically matched controls and RGS2 is known to be a CREB-responsive gene, RGS2 mRNA was reduced in placentas from pregnancies affected by preeclampsia. Experimentally reducing Rgs2 expression within the feto-placental unit was sufficient to induce preeclampsia-like phenotypes in pregnant wild-type C57BL/6J mice. Stimulation of RGS2 transcription within immortalized human HTR8/SVneo trophoblasts by cAMP/CREB signaling was discovered to be dependent on the activity of histone deacetylase activity, and more specifically, HDAC9 (histone deacetylase-9), and HDAC9 expression was reduced in placentas from human pregnancies affected by preeclampsia. We conclude that reduced expression of RGS2 within the placenta may mechanistically contribute to preeclampsia. More generally, this work identifies RGS2 as an HDAC9-dependent CREB-responsive gene, which may contribute to reduced RGS2 expression in placenta during preeclampsia.

Evaluation of the effects of Uncaria rhynchophylla alkaloid extract on LPS-induced preeclampsia symptoms and inflammation in a pregnant rat model.

Excessive pro-inflammatory cytokines result in adverse pregnancy outcomes, including preeclampsia-like phenotypes, and fetal growth restriction. Anti-inflammation might be an effective therapy. The aim of this research was to investigate whether Uncaria rhynchophylla alkaloid extract (URE), a highly safe anti-inflammation constituent of the herb, can inhibit inflammation and improve clinical characteristics of preeclampsia in a lipopolysaccharide (LPS)-induced preeclampsia rat model. The rat model was established by daily administration of LPS (1 μg/kg body weight per day) from gestational day (GD) 14 to 19. Different doses of URE (35, 70, and 140 mg/kg body weight per day) were administered from GD 14 to GD 19. The effects of URE on proteinuria, maternal hypertension, pregnancy outcomes, as well as pro-inflammatory cytokines levels in serum and placenta were measured. High-dose URE (HURE) treatment decreased LPS-induced mean 24-h proteinuria and systolic blood pressure, and increased fetal weight, placental weight, and the number of live pups (P<0.05). Moreover, increased serum and placental levels of interleukin (IL)-6, IL-1β, tumor necrosis factor-α, and interferon-γ in the LPS-treated group were obviously inhibited after HURE administration (P<0.01). URE improved preeclampsia symptoms and mitigated inflammatory responses in the LPS-induced preeclampsia rat model, which suggests that the anti-inflammation effect of URE might be an alternative therapy for preeclampsia.

Abstract 128: Interleukin-17 Mediates Hypertension, Intrauterine Growth Restriction, Cytolytic Natural Killer Cells and Vascular Dysfunction in Pregnant Rats

Preeclampsia is a hypertensive disorder of pregnancy characterized by intrauterine growth restriction (IUGR), vascular dysfunction, and chronic immune activation including increased T H 17s and cytolytic NK cells (NK C ). We recently developed a novel model of preeclampsia in which placental ischemia (PI)-induced T H 17s cause a preeclampsia-like phenotype in pregnant rats characterized by hypertension, IUGR, oxidative stress (ROS), and increased cytolytic NK cells (NK C ). In the current study we investigated a novel role for IL-17, the main cytokine secreted from T H 17s, to directly induce IUGR, NK C activation, and vascular dysfunction in pregnancy. IL-17 (150 pg/day) was chronically infused into a subset of normal pregnant (NP) rats from gestation day (GD) 12-19 (NP+IL-17) via i.p. minipump. On GD 18 carotid catheters were implanted and on GD 19 MAP, fetal weight, placental weight, placental NK C , and NK C -associated proteins were measured and vascular reactivity of uterine arteries was assessed. Data are expressed as mean±SEM. MAP significantly increased from 100±3 mmHg in NP (n=9) to 115±1 mmHg in NP+IL-17 (n=12). Fetal weight significantly decreased from 2.5±0.04 g in NP to 2.3±0.03 g in NP+IL-17 (p&lt;0.05). Placental weight significantly decreased from 0.62±0.02 g in NP to 0.55±0.01 g in NP+IL-17 (p&lt;0.05). Placental ROS significantly increased 1353±337 RLU/min/mg in NP to 2210±180 RLU/min/mg in NP+IL-17 (p&lt;0.05). Placental NK C increased from 2.6±1.6% of the total NK population in NP to 11.3±2.2% in NP+IL-17 (p&lt;0.05). Placental granzyme B increased from 22.7±1.6 pg/mg in NP to 30.2±2 pg/mg in NP+IL-17 (p&lt;0.05). Placental granzyme A increased from 3067±225 pg/mg in NP to 3926±210 pg/mg (p&lt;0.05). Additionally, placental levels of VEGF, an important pro-angiogenic factor secreted by non-cytolytic uterine NK cells significantly decreased from 77.6±6.5 pg/mg in NP to 54.2±3.2 pg/mg in NP+IL-17 (p&lt;0.05). We also observed impaired relaxation of uterine arteries in response to acetylcholine. These data suggest a shift from non-cytolytic NK to NK C cells in the placentas of NP+IL17. In addition to hypertension and ROS, this study demonstrates novel roles for IL-17 to directly mediate IUGR, NK C activation, and endothelial vascular dysfunction during pregnancy.

Ameliorative effects of aspirin against lipopolysaccharide-induced preeclampsia-like symptoms in rats by inhibiting the pro-inflammatory pathway.

Preeclampsia is an inflammatory disease and has connection with increased pro-inflammatory cytokines. Aspirin reduces the incidence of preeclampsia complications. However, the effects of aspirin on lipopolysaccharide-induced preeclampsia-like symptoms in rats have not been reported and the underlying molecular mechanism has not been illuminated. Hence, we investigated the anti-inflammatory effects of aspirin on lipopolysaccharide-induced preeclampsia-like phenotypes in pregnant rats and elucidated the potential molecular mechanism. Preeclampsia-like phenotypes were induced by tail vein injection of lipopolysaccharide (1 μg/kg) on gestational day 14. Aspirin (2 mg/kg per day) were administered from gestational day 14 to 19. Clinical phenotypes were recorded. Placenta tissues and serum were obtained to measure inflammatory cytokines levels using ELISA kit on gestational day 20. The mRNA expressions of IL-6, IL-1β, and MCP-1 were measured by real-time PCR. Protein expressions including TLR4, MyD88, NF-κBp65, and TLR2 were determined by Western blot analysis in the rat placentas of each group. Aspirin obviously assuaged lipopolysaccharide-induced preeclampsia-like phenotypes in pregnant rats. Aspirin treatment significantly decreased the levels of pro-inflammatory cytokines in serum and placenta tissues of preeclampsia rats. Aspirin also obviously downregulated the mRNA expressions of IL-6, IL-1β, and MCP-1 and assuaged the activation of TLR4, MyD88, NF-κBp65, and TLR2 in the placental tissue. Our results indicated that aspirin could assuage preeclampsia-like phenotypes, and this improvement effect is possibly the result of the suppression of pro-inflammatory cytokines via the TLR4, MyD88, NF-κBp65, and TLR2 signaling pathway.

ELABELA/APELA Levels Are Not Decreased in the Maternal Circulation or Placenta among Women with Preeclampsia

The genetic deletion of apelin receptor early endogenous ligand (Elabela; official name APELA) produces a preeclampsia-like phenotype in mice. However, evidence linking ELABELA with human disease is lacking. Therefore, we measured placental mRNA and circulating ELABELA in human samples. ELABELA mRNA (measured by RNA sequencing) was unchanged in 82 preeclamptic placentas compared with 82 matched controls (mean difference, 0.53%; 95% CI, −25.9 to 27.0; P = 0.78). We measured circulating ELABELA in 32 women with preterm preeclampsia (delivered at <34 weeks' gestation) and 32 matched controls sampled at the same gestational age. There was no difference in circulating ELABELA concentrations in the preeclamptic cohort compared with controls (median, 28.5 pg/mL; 95% CI, 5.3 to 63.2 versus median, 20.5 pg/mL; 95% CI, 9.2 to 58.0, respectively); the median difference was 8.0 pg/mL (95% CI, −17.7 to 12.1; P = 0.43). In contrast, soluble FLT1 (a protein with an established association with preeclampsia) mRNA was increased in placental tissue (mean difference, 34.9%; 95% CI, 16.6 to 53.1; P = 0.001), and circulating concentrations were 16.8-fold higher among the preeclamptic cohort (P < 0.0001). In conclusion, we were able to recapitulate the association between circulating soluble FLT1 and preeclampsia, but there was no association with ELABELA. The speculated clinical relevance of observations in the murine model linking ELABELA to preeclampsia likely are incorrect.

The intervention effect of aspirin on a lipopolysaccharide-induced preeclampsia-like mouse model by inhibiting the nuclear factor-κB pathway†

Preeclampsia is a severe pregnancy-related disorder, and patients usually present with high circulating inflammatory factor levels and excessive activation of the nuclear factor-κB (NF-κB) pathway. Administration of aspirin (ASP) is effective for preventing preeclampsia, and thus, we propose that ASP might affect placental function by regulating the NF-κB pathway. Systemic lipopolysaccharide (LPS) (20 μg/kg) was used to induce preeclampsia-like pregnant mouse model, and low-dose ASP (15.2 mg/kg) was administrated. Here, we report significantly increased circulatory expression levels of the proinflammatory cytokines tumor necrosis factor-alpha, interleukin-6, and soluble Fms-related tyrosine kinase-1 in LPS-treated pregnant mice, accompanied by kidney and placental dysfunction. Low-dose ASP treatment significantly reversed the preeclampsia-like phenotype, lowering hypertension, decreasing proteinuria, and ameliorating fetal growth retardation. Moreover, the excessive activation of NF-κB signaling in mice placentae induced by LPS was significantly reduced by ASP. In JEG-3 cells, LPS activated the NF-κB signaling pathway by upregulating the expression of cyclooxygenase-2 (COX-2) and related inflammatory factors, whereas the invasion ability of JEG-3 cells was weakened. However, ASP administration impeded NF-κB signaling activation, downregulated COX-2 and inflammatory factor expression, and rescued trophoblast invasion. This study provides new evidence that low-dose ASP is beneficial for preeclampsia prevention by inhibiting NF-κB and its downstream signaling pathways in trophoblast cells.

Causative Effects of Genetically Determined High Maternal/Fetal Endothelin-1 on Preeclampsia-Like Conditions in Mice.

Endothelin-1 (ET-1) is implicated in the pathophysiology of preeclampsia. An association between an EDN1 gene polymorphism with high ET-1 and preeclampsia was reported in humans, but their cause and effect relationships have not been defined. We examined the pregnancy effects in mice with a modified Edn1 allele that increases mRNA stability and thus ET-1 production. Heterozygous Edn1H/+ females showed no obvious abnormalities before pregnancy, but when mated with wild-type (WT) males developed a full spectrum of preeclampsia-like phenotypes, including increased systolic blood pressure, proteinuria, glomerular endotheliosis, and intrauterine fetal growth restriction. At 7.5 days post-coitus, the embryos from Edn1H/+ dams, regardless of their Edn1 genotype, lagged 12 hours in development compared with embryos from WT dams, had disoriented ectoplacental cones, and retained high E-cadherin expression. In contrast, WT females mated with Edn1H/+ males, which also carried half of the fetuses with Edn1H/+ genotype, showed a mild systolic blood pressure increase only. These WT dams had 2× higher plasma soluble fms-like tyrosine kinase-1 than WT dams mated with WT males. In human first trimester trophoblast cells, pharmacological doses of ET-1 increased the cellular sFlt1 transcripts and protein secretion via both type A and B ET-1 receptors. Our data demonstrate that high maternal ET-1 production causes preeclampsia-like phenotypes during pregnancy, affecting both initial stage of trophoblast differentiation/invasion and maternal peripheral vasculature during late gestation. High fetal ET-1 production, however, could cause increased soluble fms-like tyrosine kinase-1 in the maternal circulation and contribute to blood pressure elevation.

Bromine Exposure In Pregnant Mice May Reduce VEGF Signaling Via Increased Circulating VEGF Decoy Receptor sFlt‐1

Inhalation of oxidant gas and vapor like the halogen bromine (Br2) may pose an increased threat to pregnant women and their unborn fetuses. We recently published that pregnant (P) C57Bl/6 mice exposed to 600 ppm Br2 for 30 minutes on gestational day 14.5 (E14.5) exhibited increased mortality, lung injury, and blood pressure at E18.5 when compared to similarly exposed non‐pregnant (NP) mice. Additionally, fetuses of Br2‐exposed pregnant mice displayed severe growth restriction, which cumulatively indicated a preeclampsia‐like phenotype. Administration of the phosphodiesterase 5 (PDE5) inhibitor tadalafil abrogated or significantly mitigated these effects in Br2‐exposed pregnant mice but had no effect on non‐pregnant counterparts. We hypothesize that exposure to Br2 via inhalation in pregnant mice results in placental damage and subsequent release of anti‐angiogenic molecules into maternal circulation resulting in systemic endothelial dysfunction and a vasoconstrictive environment mediated by dysfunctional VEGF signaling. Firstly, we have identified significant increases at 0.5 h post‐Br2 exposure of circulating brominated fatty acids and fatty acid aldehydes (2‐BrPA and 2‐BrPALD) as well as their glutathionylated species at 6h. These may serve as potential biomarkers and/or mediators of Br2‐inhalation induced systemic injury. Furthermore, preliminary results indicate aortas incubated ex vivo in the presence of 2‐BrPA and 2‐BrPALD are more sensitive to the vasoconstricting agent phenylephrine and less sensitive to the vasodilator acetylcholine. Similarly, aortas pregnant mice exposed to Br2 are less sensitive to acetylcholine when compared with similarly exposed non‐pregnant mice. Secondly, utilizing time‐course measurements, we have identified increases in the circulating VEGF decoy receptor sFlt‐1 beginning at 72h post‐exposure in pregnant mice. No increase is observed in similarly exposed non‐pregnant mice (PBr272h 7280.3 ± 1592 vs NPBr272h 1233.5 ± 1063 pg/mL p≤0.001). Additionally, this time point also coincides with a pregnancy specific increase in lung edema/injury as evidenced by lung wet:dry weight ratios (P Br272h 4.68 ± 0.10 vs NP Br272h 4.22 ± 0.04 p≤0.05). Lastly, supplementation of exogenous VEGF‐121 intended to overcome the effects of increased circulating sFlt‐1 improves both survival and weight gain profiles of pregnant mice exposed to Br2 as well as their fetuses. This study implicates diminished VEGF signaling in pregnancy specific Br2 inhalation injury contributing to increased blood pressures, lung injury and subsequent mortality.Support or Funding InformationNIEHS CounterACT ‐5U01ES026458 02, 1 U01 ES027697 01, Ruth L. Kirschstein National Research Service Award Institutional Training Grant ‐ T32HL007918.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.