Details

Autor(en) / Beteiligte

• Travis, Olivia K.D

Titel

Interleukin 17 Signaling and Natural Killer Cell Activation in Preeclampsia

Ort / Verlag

ProQuest Dissertations & Theses

Erscheinungsjahr

2021

Link zum Volltext

• ProQuest

Quelle

ProQuest Dissertations & Theses A&I

Beschreibungen/Notizen

• Preeclampsia (PE) is a multisystem pregnancy disorder that affects around 2-10% of pregnancies and is a leading cause of maternal and fetal morbidity worldwide. It is characterized by new onset maternal hypertension (HTN) after the 20th week of pregnancy, end organ damage, intrauterine growth restriction (IUGR), systemic inflammation, and endothelial dysfunction. Although the etiology is not completely understood, decades of research have determined that PE is initiated by impaired cytotrophoblast invasion and spiral artery remodeling resulting in reduced blood flow to the placenta. The ischemic placenta releases vasoactive substances and inflammatory mediators into the maternal circulation that lead to PE manifestations. Despite this understanding, the only cure remains delivery of the feto-placental unit underscoring an urgent need to identify therapeutic targets for treatment.The aberrations in systemic immune activation observed in PE include an increased TH17:TReg ratio, activation of natural killer cells (NKs), and elevated pro-inflammatory cytokines such as interleukin (IL)-17, tumor necrosis factor alpha (TNF-α) , and interferon gamma (IFNγ). The Reduced Uterine Perfusion Pressure (RUPP) rat model of placental ischemia reflects this immune profile as well as the HTN, IUGR, and endothelial dysfunction seen in preeclamptic women. This model has been used by many investigators to better understand how these immune manifestations contribute to the pathophysiology of PE. Previous studies in the RUPP have shown that total and activated cytolytic NKs (cNKs) are increased in the circulation and placentas of RUPP rats compared to normal pregnant (NP) rats. An additional study showed that adoptive transfer of RUPP TH17s into NP rats induced NK activation; however, the precise mechanism of this activation has not been determined.Other investigations in cancer and anti-fungal immunity have shown that IL-17, the main cytokine secreted by TH17s, plays a role in NK cell activation and cytolytic function, and thus could be a potential mechanism of placental ischemia induced NK activation in PE. Furthermore, experiments depleting NKs in RUPP rats improved maternal blood pressure and fetal weight and lowered levels of TNF-α and IFNγ, suggesting a role for NKs and these associated cytokines in RUPP pathophysiology. However, as these studies depleted the entire NK population, the precise role of cNKs in RUPP and PE pathophysiology has not been elucidated. Moreover, there remains a knowledge gap in the mechanisms by which TNF-α and IFNγ contribute to the HTN, IUGR, and vascular dysfunction seen in PE. Thus the goal of this dissertation was to investigate the overarching hypothesis that placental ischemia-induced IL-17 signaling activates cNKs, and these cNKs induce oxidative stress and secrete inflammatory cytokines which contribute to maternal vascular dysfunction, hypertension and intrauterine growth restriction during pregnancy.The studies designed to test this hypothesis were broken down into three aims. The first aim tested the hypothesis that elevated IL-17 activates cNKs during pregnancy as a mechanism to induce HTN, IUGR, and vascular dysfunction in PE. We first infused 150 pg/day of recombinant IL-17 into a subset of normal pregnant rats from Gestation Days (GD) 12-19 and evaluated changes in mean arterial pressure (MAP), fetal and placental weights, inflammatory cytokines, cNK cell activation, cytotoxic enzymes, in vitro NK cytotoxicity and acetylcholine mediated vasorelaxation in isolated uterine arteries. Our data demonstrated elevations in total and cNK populations in the circulation and placenta, increased NK cytotoxicity, and higher levels of placental cytolytic proteins in NP recipients of IL-17. Furthermore, we also observed increased Mean Arterial Pressure (MAP), decreased fetal and placental weight, endothelial dysfunction, increased placental inflammatory cytokines, and placental oxidative stress in response to chronic IL‐17. These results suggested that cytolytic activation of NKs by IL‐17 may be an important contributor of pathophysiology in PE. We next sought to confirm that IL-17 in response to placental ischemia stimulated NK activation. Thus, we infused 100 pg/day of a soluble IL-17 receptor (IL-17RC) from GD 14-19 into RUPP rats to block IL-17 signaling. We assessed changes in circulating and placental cNK activation, levels of inflammatory and cytolytic proteins, and in vitro placental NK cell cytolytic activity. We also evaluated placental oxidative stress, maternal MAP, placental weight, and fetal outcomes. We found that infusion of IL-17RC into RUPPs prevented circulating and placental NK cell activation, reduced NK cytotoxicity and decreased placental reactive oxygen species (ROS). We also saw that inhibition of IL-17 signaling resulted in a significant decrease in maternal MAP and an increase in both fetal and placental weight. The results of these two studies demonstrate a direct role for increased IL-17 to mediate NK cytotoxic activation during pregnancy.We next wanted to focus on examining the polarity of NKs in RUPP pathophysiology. As the dominant immune cell within the decidua during pregnancy, NKs normally play a supportive role by directing trophoblast invasion, promoting maternal immune tolerance, and secreting growth factors such as vascular endothelial growth factor (VEGF). However the pro-inflammatory type 1 shift seen in PE women and RUPP rats causes increased NK cytotoxicity, reduced VEGF secretion, and increased secretion of pro-inflammatory cytokines TNF-α and IFNγ. Thus, the second aim tested the hypothesis that placental ischemia stimulated cNKs increase placental ROS, inflammatory cytokines (TNF-α and IFNγ), and vascular dysfunction leading to HTN and IUGR in pregnancy. We used adoptive transfer experiments to examine the direct role of cNKs in causing RUPP pathophysiology and assess the role of normal NKs to ameliorate RUPP pathophysiology. On GD14, vehicle or 5×106 RUPP NKs were infused into a subset of Sham rats, and vehicle or 5×106 Sham NKs were infused into a subset of RUPP rats. We evaluated NK populations, maternal MAP, fetal and placental weight, circulating and placental inflammatory cytokines, placental ROS, angiogenic factors, and Uterine Artery Resistance Index (UARI). We found that adoptive transfer of RUPP NKs into Sham rats resulted in NK activation, elevated Uterine Artery Resistance Index (UARI), placental oxidative stress, increased circulating and placental inflammatory cytokines ultimately leading to maternal hypertension, and IUGR in Sham recipients. Furthermore, RUPP recipients of Sham NKs demonstrated reduced NK activation, UARI, and normalized angiogenic balance, which led to improved maternal blood pressure and normal fetal growth. Thus, we have demonstrated that cNKs play a direct role in PE pathophysiology, and that the polarization of NK cell phenotypes can influence pregnancy outcomes.The third aim tested the hypothesis that blockade of cNK associated cytokines (IFNγ and TNF-α) in placental ischemic rats decreases oxidative stress and inflammation resulting in improved vascular function, lower blood pressure, and decreased IUGR during pregnancy. We first treated a subset of NP and RUPP rats with either vehicle or an anti-IFNγ antibody injections on GD 15 and 18. We assessed changes in fetal and placental weight, maternal MAP, and vascular function. We also measured circulating angiogenic factors, placental ROS, circulating and placental inflammatory cytokines, nitric oxide (NO) bioavailability, and endothelial NOS synthase (eNOS) expression and phosphorylation. The data showed a reduction in MAP of RUPP recipients of anti-IFNγ, but fetal and placental weight were not improved compared to RUPPs. UARI, fetal resorptions, placental Reactive Oxygen Species (ROS), and circulating sFlt-1 were also reduced in RUPP+anti-IFNγ compared to vehicle-treated RUPP rats. These results suggest a role for IFNγ in the elevated MAP and UARI in RUPP rats. Finally, we blocked TNF-α signaling via i.p. injections of 0.4 mg/kg of etanercept (a soluble TNF-α receptor) on GD 15 and 18 in a subset of Sham and RUPP rats. We assessed changes in MAP, UARI, fetal and placental growth, placental ROS, NO bioavailability, angiogenic factors and eNOS expression and phosphorylation compared to vehicle treated Shams and RUPPs. UARI and circulating sFlt-1 were reduced and NO bioavailability was elevated in RUPPs treated with ETA compared to vehicle-treated RUPPs, and this was accompanied by a normalization of MAP and fetal growth. These results identified additional mechanisms of TNF-α in mediating HTN, IUGR, and vascular dysfunction in RUPP and PE pathophysiology.