Preeclampsia (PE) is considered the leading cause of maternal and perinatal morbidity and mortality. The placenta seems to play an essential role in this disease, probably due to factors involved in ...its formation and development. The present study aimed to investigate the association between placental lesions, cytokines and angiogenic factors in pregnant women with preeclampsia (PE). We evaluated 20 normotensive pregnant women, 40 with early-onset PE and 80 with late-onset PE. Placental samples were analyzed for histopathology, immunohistochemistry and determination of granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-10 (IL-10), transforming growth factor-beta 1 (TGF-β1), tumor necrosis factor-alpha (TNF-α), placental growth factor (PlGF), vascular endothelial growth factor (VEGF), fms-like tyrosine-kinase-1 (Flt-1) and endoglin (Eng) levels. Higher percentages of increased syncytial knots and increased perivillous fibrin deposits, and greater levels of TNF-α, TGF-β1and Flt-1 were detected in placentas from early-onset PE. Levels of IL-10, VEGF and PlGF were decreased in PE versus normotensive placentas. Both the TNF-α/IL-10 and sFlt-1/PlGF ratios were higher in placental homogenate of early-onset PE than late-onset PE and control groups. The more severe lesions and the imbalance between TNF-α/IL-10 and PlGF/sFlt-1 in placentas from early-onset PE allows differentiation of early and late-onset PE and suggests higher placental impairment in early-onset PE.
•Hyperactivation of NLRP3 inflammasome was proposed in placentas from pregnant women with severe preeclampsia.•IL-1β, TNF-α and HMGB1may be involved in the exaggerated systemic inflammation of ...preeclampsia.•Placental inflammasome activation may be involved in the pathogenesis of preeclampsia.
Preeclampsia is a pregnancy disorder characterized by imbalance between pro- and anti-inflammatory cytokines associated with high plasma levels of uric acid and Interleukin-1 beta (IL-1β). The inflammasome is a protein complex that mediates innate immune responses via caspase-1 activation promoting secretion of IL-1β and IL-18 in their active forms, and also release of the high-mobility group box 1 protein (HMGB1). As the placenta seems to play an important role in the pathogenesis of PE, the present study investigated the expression of genes and proteins related to the inflammasome in placentas from pregnant women with severe preeclampsia. Placental tissue was collected from 20 normotensive pregnant women and 20 preeclamptic women, and inflammasome components, NLRP3 (NOD-like receptor family, pyrin domain-containing protein 3), caspase-1, IL-1β and IL-18, as well as tumor necrosis factor-alpha (TNF-α) and HMGB1 were evaluated by immunohistochemistry, enzyme-linked immunosorbent assay (ELISA) and also quantified by reverse transcription-qPCR (RT-qPCR). Compared with normotensive pregnant women, placenta from women with PE showed a significant increase in NLRP3, caspase-1, IL-1β, TNF-α and HMGB1 mRNA. Immunohistochemical staining of NLRP3, caspase-1, IL-1β and TNF-α in placental villi, as well as the levels of caspase-1, IL-1β, TNF-α and HMGB1 in placental homogenate were significantly higher in the preeclamptic group than in the normotensive group. However, mRNA expression of IL-18 and its protein concentrations were lower in placentas from preeclamptic women. The results suggest that placentas from pregnant women with preeclampsia show higher expression of NLRP3 inflammasome, which may be involved in the exaggerated inflammatory state in preeclampsia.
Objective
To assess and compare circulating tissue inhibitor of metalloproteinase 3 (TIMP‐3) concentrations between women with pre‐eclampsia and healthy pregnant women. We also aimed to determine the ...relationships between circulating TIMP‐3 and matrix metalloproteinase 2 (MMP‐2), MMP‐9, TIMP‐1, and TIMP‐2 concentrations in pre‐eclampsia.
Methods
A primary case–control study included patients with pre‐eclampsia (n = 219) and gestational hypertension (n = 118), healthy pregnant women (n = 214), and non‐pregnant women (n = 66), and a replication case–control study included patients with pre‐eclampsia (n = 177) and healthy pregnant women (n = 124), all from southeastern Brazil. Plasma TIMP‐3, MMP‐2, MMP‐9, TIMP‐1, and TIMP‐2 concentrations were assessed using commercially available enzyme‐linked immunosorbent assay kits, and the relationships between them were analyzed using Spearman's correlation.
Results
In our primary study, patients with pre‐eclampsia and gestational hypertension exhibited increased TIMP‐3 concentrations compared with healthy pregnant women (both P < 0.0001) and non‐pregnant women (both P < 0.001). These findings were confirmed in the replication study, showing elevated TIMP‐3 concentrations in women with pre‐eclampsia versus healthy pregnant women (P < 0.001). We found no difference in TIMP‐3 concentrations between early‐onset and late‐onset pre‐eclampsia. Moreover, TIMP‐3 concentrations were significantly correlated with plasma concentrations of TIMP‐1 (r = 0.2333; P = 0.0086) and MMP‐2 (r = 0.2159; P = 0.0156) in pre‐eclampsia.
Conclusions
Circulating TIMP‐3 concentration is increased in women with pre‐eclampsia compared with healthy pregnant women, and it is positively correlated with plasma MMP‐2 and TIMP‐1 concentrations in pre‐eclampsia.
Synopsis
Circulating TIMP‐3 concentration is increased in pre‐eclampsia compared with healthy pregnant women, and is positively correlated with plasma MMP‐2 and TIMP‐1 concentrations in pre‐eclampsia.
Summary
Pre‐eclampsia (PE) is an obstetric pathology characterized by abnormal activation of the innate and adaptive immune systems dependent on the imbalance of T helper subsets. The present study ...aimed to evaluate the gene and protein expression of T helper type 1 (Th1)/Th2/Th17/regulatory T (Treg) cell transcription factors in peripheral blood lymphocytes from pregnant women with PE employing quantitative RT‐PCR and flow cytometry techniques, as well as the cytokine profile produced by these CD4+ T‐cell subsets in the plasma of pregnant women with PE, classified as early‐onset PE (n = 20), late‐onset PE (n = 20) and normotensive pregnant women (n = 20). Results showed a higher percentage of CD4+ T cells expressing the RORc transcription factor (Th17) and a lower percentage of cells expressing FoxP3 (Treg) in women with early‐onset PE compared with late‐onset PE and normotensive groups. A lower gene expression of GATA‐3 transcription factor was detected in cells of women with early‐onset PE compared with the late‐onset PE group. Endogenous plasma levels of interleukin‐6 (IL‐6), IL‐17 and tumour necrosis factor‐α were significantly higher in the early‐onset PE group than in the late‐onset PE and normotensive groups, whereas IL‐4 (Th2 profile) and IL‐22 (Th17 profile), were not significantly different between the studied groups. The endogenous levels of transforming growth factor‐β and IL‐10 were significantly lower in the pre‐eclamptic than in the normotensive groups of the same gestational age, with a significant difference between early‐ and late‐onset PE. The results show that in women with PE there is an imbalance between inflammatory and anti‐inflammatory profiles in CD4+ T‐cell subsets, with polarization to Th17 profiles in the early‐onset PE, considered as the severe form of PE.
Endogenous characterization of CD4+ T‐cell subsets by expression of the transcription factors T‐bet, RORc, GATA‐3 and FoxP3 in women with pre‐eclampsia and normotensive pregnant women. A deviation of CD4+ lymphocytes to the T helper type 17 profile was more evident in pregnant women with early‐onset pre‐eclampsia, the more severe form of this obstetric pathology.
•Preeclamptic women showed high plasma levels of sTNFR1 and TNF.•Plasma from preeclamptic women induced high TNFR1 expression by Jurkat cells.•Progesterone played a modulatory role over TNFR1 ...expressed by Jurkat cells.•Progesterone shifted Jurkat cells towards an anti-inflammatory profile.
Pregnant women with preeclampsia (PE) present a shift in the immune response to an inflammatory profile. This deviation could be due to the interaction of tumor necrosis factor (TNF) with TNFR1 and TNFR2 receptors, besides the failure in modulation of inflammation regulatory mechanisms. This study evaluated the effects of progesterone on the expression of TNFR1 and TNFR2 by Jurkat cells after stimulation with plasma from PE and normotensive (NT) pregnant women. Jurkat cells were cultured with or without progesterone in a medium containing 20% (v/v) plasma from PE or NT women. The expression of TNF receptors was evaluated by flow cytometry. The concentration of soluble forms of TNF receptors and cytokines was determined in culture supernatant and plasma by ELISA. The plasma of PE women showed significantly higher concentrations of sTNFR1 and TNF and lower concentrations of sTNFR2 compared to the NT group. TNFR1 receptor expression was increased in Jurkat cells, while TNFR2 was decreased after culture with PE plasma when compared with Jurkat cells cultured with progesterone and plasma from NT women. The concentration of sTNFR1, TNF, and IL-10 in the culture supernatant of Jurkat cells was increased after culture with PE plasma, while the sTNFR2 receptor was decreased when compared to the NT group. Results demonstrate that in preeclamptic women a systemic inflammation occurs with an increase of inflammatory molecules, and progesterone may have a modulating effect on the expression of TNF receptors, shifting Jurkat cells towards an anti-inflammatory profile with greater expression of TNFR2.
•DAMPs involvement in systemic inflammation of pre-eclampsia.•Hyaluronan stimulate inflammasome activation in monocytes from pre-eclamptic women.•Hsp70 induce inflammation in monocytes from ...pre-eclamptic women.
Pre-eclampsia (PE) is a human pregnancy syndrome with abnormal activation of the innate immune response. The study evaluated the involvement of molecular structures called damage-associated molecular patterns (DAMPs), such as hyaluronan (HA) and heat shock proteins (Hsp) on NLRP1 and NLRP3 inflammasomes activation in peripheral blood monocytes. Twenty pre-eclamptic women, 20 normotensive pregnant women (NT) and 20 non-pregnant women (NP) were studied. Enzyme immunoassay was employed for the determination of HA, Hsp70 and High mobility group Box 1 (HMGB1) in plasma, as well as for the detection of Interleukin-1β (IL-1β), IL-18 and tumor necrosis factor alpha (TNF-α) in the supernatant of monocytes cultured with or without HA and Hsp70. The inflammasomes induction was evaluated by the quantification of mRNA for NLRP1, NLRP3, caspase-1, IL-1β, IL-18, HMGB1 and TNF-α by qPCR in monocyte culture. The results showed significantly higher plasma levels of HA, Hsp70 and HMGB1 in pre-eclamptic women than in NT and NP women. Monocytes from women with PE showed endogenous activation of NLRP1 and NLRP3 inflammasomes, and expressed high amounts of IL-1β, IL-18, HMGB1 and TNF-α. The stimulation of monocytes with HA increased the gene expression of NLRP1, NLRP3, caspase-1, TNF-α, IL-1β, HMGB1 and IL-18 and the production of IL-1β in pre-eclamptic women. Monocytes cultured with Hsp70 produced elevated levels of IL-1β and TNF-α through a mechanism independent of inflammasomes activation. These results suggest the participation of these DAMPs in the systemic inflammatory response that is characteristic of PE.
Preeclampsia (PE) is a pregnancy-specific syndrome characterized by a systemic inflammatory response that polarizes peripheral blood monocytes to the M1 phenotype. The classically activated M1 ...monocytes comprise immune effector cells with an acute inflammatory phenotype. CD163 is a scavenger receptor expressed by monocytes/macrophages that may be shed from their cell membrane after proteolytic cleavage, producing the soluble CD163 molecule (sCD163). This study evaluated CD163 expression by monocytes and sCD163 as well as pro- and anti-inflammatory cytokine concentration in the plasma of pregnant women with PE. Fifty-six women with PE and 28 normotensive pregnant women were included. Plasma levels of sCD163, interleukin-1 beta (IL-1β), IL-6, IL-10, transforming growth factor beta (TGF-β1), and tumor necrosis factor-alpha (TNF-α) were determined by ELISA, and CD163 expression by monocytes was assessed by flow cytometry. The expression of CD163 by monocytes was significantly lower in severe and mild PE than in normotensive pregnant. Plasma concentrations of IL-1β, TGF-β1, and TNF-α were higher in severe PE than in mild PE and normotensive pregnant women. Both groups of preeclamptic women showed decreased plasma levels of sCD163 and IL-10. Negative correlations between sCD163 and IL-1β (
r
= − 0.45;
P
= 0.014) and between sCD163 and TNF-α concentrations (
r
= − 0.54;
P
= 0.001) were observed in the severe PE group. The association between the pro-inflammatory cytokine profile and lower concentrations of sCD163 and IL-10 in plasma from women with severe PE suggests an impairment in the modulation of the systemic inflammatory response in this group of pregnant women with preeclampsia.
Preeclampsia (PE) is a human pregnancy-specific syndrome with abnormal activation of cells from the innate immune system. The present study evaluated whether silibinin (SB) treatment of monocytes ...from preeclamptic women could modulate NLRP1 and NLRP3 inflammasomes as well as TLR4/NF-κB pathway activation. Peripheral blood monocytes from 20 preeclamptic and 20 normotensive (NT) pregnant women, as well as the THP-1 cell line, were cultured with or without monosodium urate (MSU) or SB.
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gene expression by monocytes was analysed by quantitative real-time polymerase chain reaction (qPCR), while inflammatory cytokine production and p65NF-κB activity were determined by enzyme-linked immunosorbent assays (ELISAs). TLR4/MyD88/NF-κB and NLRP1/NLRP3 inflammasomes pathways in THP-1 cells were evaluated by flow cytometry and western blot respectively. Compared with NT women, monocytes from preeclamptic women showed The Ethics Committee of the Botucatu Medical School approved the study (protocol number 2.333.216)higher endogenous activation of NLRP1/NLRP3 inflammasomes and the TLR4/NF-κB pathway as well as higher gene and protein expression of IL-1β, IL-18 and TNF-α, and lower expression of IL-10. Monocyte stimulation with MSU increased inflammation-related genes as well as NF-κB activity. In vitro, SB treatment of monocytes from preeclamptic women reduced the basal activation of these cells by decreasing NLRP1/NLRP3 inflammasomes and p65NF-κB activity. THP-1 cells exhibited a similar immunological response profile to monocytes from preeclamptic women when cultured with or without MSU or SB. These results suggest uric acid participates in the systemic inflammatory response characteristic of preeclampsia and that in vitro SB treatment can modulate the sterile inflammation established in monocytes from preeclamptic women.
Hypertensive disorders of pregnancy (HDP), comprising gestational hypertension (GH) and pre-eclampsia (PE), are leading causes of maternal and perinatal morbidity and mortality. Both GH and PE are ...characterized by new-onset hypertension, but PE additionally includes proteinuria and/or end-organ damage. Impaired nitric oxide (NO) bioavailability may lead to endothelial dysfunction in GH and PE, and the primary source of vascular NO is endothelial NO synthase (eNOS). However, no previous study has investigated plasma eNOS concentrations in patients with GH and PE. In this study, we compared plasma eNOS concentrations in healthy pregnancies and HDP in two independent cohorts. The primary study included 417 subjects, with 43 non-pregnant (NP) and 156 healthy pregnant (HP) women and 122 patients with GH and 96 with PE. The replication study included 85 pregnant women (41 healthy and 44 pre-eclamptic). Plasma concentrations of eNOS were measured using a commercial ELISA kit provided by R&D Systems, and plasma nitrite concentrations were assessed using two ozone-based chemiluminescence assays. Correlations between plasma eNOS concentrations and plasma nitrite concentrations, as well as clinical and biochemical parameters, were evaluated by either Spearman’s or Pearson’s tests. In the primary study, NP women and HDP had significantly lower plasma eNOS concentrations compared to HP; concentrations were even lower in PE compared to GH. Plasma eNOS concentrations were reduced but not significant in early-onset PE, PE with severe features, preterm birth, and intrauterine growth restriction. No correlation was observed between plasma eNOS and nitrite levels. In HDP, there was a significant positive correlation between levels of eNOS and hemoglobin (r = 0.1496, p = 0.0336) as well as newborn weight (r = 0.1487, p = 0.0316). Conversely, a negative correlation between eNOS levels and proteinuria was observed (r = −0.2167, p = 0.0179). The replication study confirmed significantly reduced plasma concentrations of eNOS in PE compared to HP. Our findings provide evidence of reduced plasma eNOS concentrations in HDP; they were particularly lower in PE compared to GH and HP in two independent studies.
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