Jessica L. Faulkner

The role of inflammation in the pathology of preeclampsia

Preeclampsia (PE) affects 5-7% of all pregnancies in the United States and is the leading cause of maternal and prenatal morbidity. PE is associated with hypertension after week 20 of gestation, decreased renal function and small-for-gestational-age babies. Women with PE exhibit chronic inflammation and production of autoantibodies. It is hypothesized that during PE, placental ischaemia occurs as a result of shallow trophoblast invasion which is associated with an immune imbalance where pro-inflammatory CD4(+) T-cells are increased and T regulatory cells (Tregs) are decreased. This imbalance leads to chronic inflammation characterized by oxidative stress, pro-inflammatory cytokines and autoantibodies. Studies conducted in our laboratory have demonstrated the importance of this immune imbalance in causing hypertension in response to placental ischaemia in pregnant rats. These studies confirm that increased CD4(+) T-cells and decreased Tregs during pregnancy leads to elevated inflammatory cytokines, endothelin (ET-1), reactive oxygen species (ROS) and agonistic autoantibodies to the angiotensin II (Ang II), typexa01 receptor (AT1-AA). All of these factors taken together play an important role in increasing the blood pressure during pregnancy. Specifically, this review focuses on the decrease in Tregs, and their associated regulatory cytokine interleukin (IL)-10, which is seen in response to placental ischaemia during pregnancy. This study will also examine the effect of regulatory immune cell repopulation on the pathophysiology of PE. These studies show that restoring the balance of the immune system through increasing Tregs, either by adoptive transfer or by infusing IL-10, reduces the blood pressure and pathophysiology associated with placental ischaemia in pregnant rats.

IL-10 supplementation increases Tregs and decreases hypertension in the RUPP rat model of preeclampsia

Objective: The reduced uterine perfusion pressure (RUPP) rat model of preeclampsia was used to determine the effects of added interleukin-10 (IL-10) on Tregs and hypertension in response to placental ischemia and how the decrease in these anti-inflammatory factors mediates the pathophysiology of preeclampsia. Methods: IL-10 (2.5u2009ng/kg/d) was infused via osmotic mini-pump implanted intraperitoneally on day 14 of gestation and, at the same time, the RUPP procedure was performed. Results: IL-10 reduced mean arterial pressure (pu2009<u20090.001), decreased CD4+ T cells (pu2009=u20090.044), while increasing Tregs (pu2009=u20090.043) which led to lower IL-6 and TNF-α (pu2009=u20090.008 and pu2009=u20090.003), reduced AT1-AA production (pu2009<u20090.001), and decreased oxidative stress (pu2009=u20090.029) in RUPP rats. Conclusion: These data indicate that IL-10 supplementation increases Tregs and helps to balance the altered immune system seen during preeclampsia.

Identifying immune mechanisms mediating the hypertension during preeclampsia.

Preeclampsia (PE) is a pregnancy-associated disorder that affects 5-8% of pregnancies and is a major cause of maternal, fetal, and neonatal morbidity and mortality. Hallmark characteristics of PE are new onset hypertension after 20 wk gestation with or without proteinuria, chronic immune activation, fetal growth restriction, and maternal endothelial dysfunction. However, the pathophysiological mechanisms that lead to the development of PE are poorly understood. Recent data from studies of both clinical and animal models demonstrate an imbalance in the subpopulations of CD4+ T cells and a role for these cells as mediators of inflammation and hypertension during pregnancy. Specifically, it has been proposed that the imbalance between two CD4+ T cell subtypes, regulatory T cells (Tregs) and T-helper 17 cells (Th17s), is involved in the pathophysiology of PE. Studies from our laboratory highlighting how this imbalance contributes to vasoactive factors, endothelial dysfunction, and hypertension during pregnancy will be discussed in this review. Therefore, the purpose of this review is to highlight hypertensive mechanisms stimulated by inflammatory factors in response to placental ischemia, thereby elucidating a role.

Vitamin D supplementation improves pathophysiology in a rat model of preeclampsia

Deficiency of vitamin D (VD) is associated with preeclampsia (PE), a hypertensive disorder of pregnancy characterized by proinflammatory immune activation. We sought to determine whether VD supplementation would reduce the pathophysiology and hypertension associated with the reduced uterine perfusion pressure (RUPP) rat model of PE. Normal pregnant (NP) and RUPP rats were supplemented with VD2 or VD3 (270 IU and 15 IU/day, respectively) on gestation days 14-18 and mean arterial pressures (MAPs) measured on day 19. MAP increased in RUPP to 123 ± 2 mmHg compared with 102 ± 3 mmHg in NP and decreased to 113 ± 3 mmHg with VD2 and 115 ± 3 mmHg with VD3 in RUPP rats. Circulating CD4+ T cells increased in RUPP to 7.90 ± 1.36% lymphocytes compared with 2.04 ± 0.67% in NP but was lowered to 0.90 ± 0.19% with VD2 and 4.26 ± 1.55% with VD3 in RUPP rats. AT1-AA, measured by chronotropic assay, decreased from 19.5 ± 0.4 bpm in RUPPs to 8.3 ± 0.5 bpm with VD2 and to 15.4 ± 0.7 bpm with VD3. Renal cortex endothelin-1 (ET-1) expression was increased in RUPP rats (11.6 ± 2.1-fold change from NP) and decreased with both VD2 (3.3 ± 1.1-fold) and VD3 (3.1 ± 0.6-fold) supplementation in RUPP rats. Plasma-soluble FMS-like tyrosine kinase-1 (sFlt-1) was also reduced to 74.2 ± 6.6 pg/ml in VD2-treated and 91.0 ± 16.1 pg/ml in VD3-treated RUPP rats compared with 132.7 ± 19.9 pg/ml in RUPP rats. VD treatment reduced CD4+ T cells, AT1-AA, ET-1, sFlt-1, and blood pressure in the RUPP rat model of PE and could be an avenue to improve treatment of hypertension in response to placental ischemia.

Placental Ischemia and Resultant Phenotype in Animal Models of Preeclampsia.

Preeclampsia is new onset (or worsening of preexisting) hypertension that occurs during pregnancy. It is accompanied by chronic inflammation, intrauterine growth restriction, elevated anti-angiogenic factors, and can occur with or without proteinuria. Although the exact etiology is unknown, it is thought that preeclampsia begins early in gestation with reduced uterine spiral artery remodeling leading to decreased vasculogenesis of the placenta as the pregnancy progresses. Soluble factors, stimulated by the ischemic placenta, shower the maternal vascular endothelium and are thought to cause endothelial dysfunction and to contribute to the development of hypertension during pregnancy. Due to the difficulty in studying such soluble factors in pregnant women, various animal models have been designed. Studies from these models have contributed to a better understanding of how factors released in response to placental ischemia may lead to increased blood pressure and reduced fetal weight during pregnancy. This review will highlight various animal models and the major findings indicating the importance of placental ischemia to lead to the pathophysiology observed in preeclamptic patients.

Serelaxin improves the pathophysiology of placental ischemia in the reduced uterine perfusion pressure rat model of preeclampsia

Preeclampsia is a hypertensive disorder of pregnancy that has limited therapeutic options. In healthy pregnancy, relaxin plays an important vasodilatory role to maintain vascular compliance; however, currently, there is no preclinical evidence to support the use of relaxin during preeclampsia. Therefore, the goal of this study was to test the hypothesis that recombinant human relaxin-2 (Serelaxin, Novartis; RLX) could reduce mean arterial pressure (MAP) and improve uterine artery resistance index (UARI) and nitric oxide bioavailability, and/or decrease prepro-endothelin-1 (PPET-1), soluble fms-like tyrosine kinase-1 (sFlt-1), and TNF-α) in the reduced uterine perfusion pressure (RUPP) model of preeclampsia. On day 14 of gestation (GD14), pregnant rats were assigned to normal pregnant (NP), RUPP, RUPP+RLX, or NP+RLX groups. Treated rats received RLX at 0.4 μg/h or RLX2 4 μg/h RLX via minipump implanted on GD14. On GD18, carotid arterial catheters were inserted, and on GD19, MAP and tissues were collected. MAP was increased in RUPP rats compared with NP but was lowered with either dose of RLX. UARI and sFlt-1 were significantly improved in both treated RUPP groups. Total circulating nitrate-nitrite improved and placental PPET-1 and TNF-α were significantly decreased with the higher dose of RLX. Renal cortex PPET-1 was reduced with both doses of RLX. In conclusion, Serelaxin improved blood pressure, sFlt-1, TNF-α, UARI, and nitric oxide bioavailability and PPET-1 in a rat model of preeclampsia, thereby suggesting a potential therapeutic role for RLX in maintaining maternal health and prolonging pregnancy in the face of placental ischemia.

Proliferation of endogenous regulatory T cells improve the pathophysiology associated with placental ischaemia of pregnancy

Preeclampsia (PE) is associated with inflammation and decreased Treg cells and IL‐10. The reduced uterine perfusion pressure (RUPP) rat model of PE exhibits these characteristics, and we hypothesized that induction of endogenous Tregs by a specific stimulus (CD28 superagonistic monoclonal antibody) would reduce inflammation, vasoactive factors, and hypertension in RUPP rats.

Vitamin D supplementation reduces some AT1-AA induced downstream targets implicated in preeclampsia including hypertension

Autoantibodies to the ANG II type I receptor (AT1-AA) are associated with preeclampsia (PE). We found that vitamin D supplementation reduced AT1-AA and blood pressure (MAP) in the RUPP rat model of PE. However, it was undetermined whether the decrease in AT1-AA was the mechanism whereby vitamin D lowered MAP or if it were through factors downstream of AT1-AA. Uterine artery resistance index, placental ET-1, and soluble FMS-like tyrosine kinase-1 are increased with AT1-AA-induced hypertension and are considered markers of PE in pregnant women. Therefore, we hypothesized that vitamin D would reduce PE factors during AT1-AA-induced hypertension and could lower blood pressure in a model of hypertension during pregnancy without PE features. Either ANG II (50 ng·kg-1·day) or AT1-AA (1:40) was infused from gestational day (GD) 12-19. vitamin D2 (VD2, 270 IU/day) or vitamin D3 (VD3, 15 IU/day) was administered orally from GD14-GD18. MAP (mmHg) increased in AT1-AA (121 ± 4) and ANG II (113 ± 1)-infused pregnant rats compared with normal pregnant rats (NP) (101 ± 2) but was lower in AT1-AA+VD2 (105 ± 2), AT1-AA+VD3 (109 ± 2), ANG II+VD2 (104 ± 4), and ANG II+VD3 (104 ± 3). VD2 and/or VD3 improved PE features associated with AT1-AA during pregnancy, while ANG II did not induce such features, supporting the hypothesis that AT1-AA induces PE features during pregnancy, and these are improved with vitamin D. In this study, we demonstrate that vitamin D improved many factors associated with PE and reduced blood pressure in a hypertensive model without PE features, indicating that vitamin D could be beneficial for various hypertensive disorders of pregnancy.

Arachidonic acid metabolites of CYP4A and CYP4F are altered in women with preeclampsia

Few studies exist on cytochrome P450 (CYP450) metabolites of arachidonic acid (AA) pertaining to the pathophysiological events in pregnancy. We hypothesized that metabolism of AA via the CYP450 pathways is altered within the placenta in women with preeclampsia (PE) and contributes to the pathophysiology of the disease. Thus, placental vascular CYP450 enzyme expression and activity were measured in normal pregnant (NP) and preeclamptic (PE) patients. CYP450 isoform expression (CYP4A11, CYP4A22, CYP4F2, and CYP4F3) was found to be elevated within the placenta of women with PE compared to normal pregnant (NP) women and chronic hypertensive (CHTN) pregnant women. In addition, placental production of 20-HETE was significantly increased in PE women compared to both NP and CHTN women. Moreover, there was an imbalance in circulating 20-HETE:EETs in PE women. To examine whether alterations in CYP450 AA metabolism contribute to the altered placentation in PE, trophoblast function, proliferation and migration were assessed in the presence of exogenous 20-HETE and a 20-HETE specific synthesis inhibitor, HET0016. Trophoblast proliferation was significantly increased in the presence of 20-HETE (1u202fμM) and reduced with 20-HETE blockade by HET0016 (1u202fmM, 5u202fmM, and 10u202fmM). On the contrary, administration of exogenous 20-HETE (1u202fμM) significantly reduced trophoblast migration. In conclusion, metabolism of AA via CYP450 is altered in PE, and increased placental production of 20-HETE may contribute to the pathophysiology of the disease.

Selective inhibition of 20-hydroxyeicosatetraenoic acid lowers blood pressure in a rat model of preeclampsia

Little is currently known of the role(s) of the vasoconstrictor 20-hydroxyeicosatetraenoic acid (20-HETE) in hypertensive pregnancies. We hypothesized that specific inhibition of 20-HETE would attenuate increases in blood pressure in the reduced uterine perfusion pressure (RUPP) rat model of preeclampsia. Specific 20-HETE synthesis inhibitor HET0016 (1mg/kg) was administered daily to RUPP rats from gestational days 14-18. Blood pressure (BP) increased in RUPP rats and was decreased with HET0016 administration. BP was unchanged in NP+HET0016 rats. Fetal death greatly increased in RUPP rats and was reduced in RUPP+HET0016 rats. 20-HETE levels increased modestly in RUPP rats compared to NP and was reduced in both NP+HET0016 and RUPP+HET0016 rats. Furthermore, circulating levels of HETEs, EET, and DHETE were significantly altered between groups. HET0016 shifted CYP metabolism toward EETs, as indicated by a decrease in plasma 20-HETE:EETs in RUPP+HET0016 rats compared to RUPP. In conclusion, 20-HETE inhibition in RUPP rats reduces BP and fetal death, and is associated with an increase in EET/20-HETE ratio.