Fetal membrane dysfunction in response to oxidative stress (OS) is associated with adverse pregnancy outcomes. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is one of the regulators of innate OS ...response. This study evaluated changes in Nrf2 expression and its downstream targets heme oxygenase (HO-1) and peroxisome proliferator-activated receptor gamma (PPARγ) in fetal membranes during OS and infection in vitro. Furthermore, we tested the roles of sulforaphane (SFN; an extract from cruciferous vegetables) and trigonelline (TRN; an aromatic compound in coffee) in regulating Nrf2 and its targets. Fetal membranes (
= 6) collected at term were placed in an organ explant system were treated with water-soluble cigarette smoke extract (CSE), an OS inducer (1:10), and lipopolysaccharide (LPS; 100 ng/mL). Nrf2 expression, expression, its enhancement by sulforaphane (SFN, 10 µM/mL) and down regulation by TRN (10uM/mL) was determined by western blots. Expression of Nrf2 response elements PPARγ (western) heme oxygenase (HO-1), and IL-6 were quantified by ELISA. CSE and LPS treatment of fetal membranes increased nrf2, but reduced HO-1 and PPARγ and increased IL-6. Co-treatment of SFN, but not with TRN, with CSE and LPS increased Nrf2 substantially, as well as increased HO-1 and PPARγ and reduced IL-6 expression. Risk factor-induced Nrf2 increase is insufficient to generate an antioxidant response in fetal membranes. Sulforaphane may enhance innate antioxidant and anti-inflammatory capacity by increasing NRF-2 expression.
Environmental exposure to toxicants is a major risk factor for spontaneous preterm birth (PTB, <37 weeks). Toxicants and drugs administered to patients are metabolized primarily by the cytochrome ...P450 (CYP450) system. Along with the adult and fetal liver, the placenta, a critical feto-maternal interface organ, expresses CYP450 enzymes that metabolize these xenobiotics. However, the contribution of the fetal membranes, another tissue of the feto-maternal interface, to the expression of CYP450 enzymes and the detoxification of xenobiotics remains unknown.
This study characterized CYP450 expression and determined the functional activity of CYP450 enzymes in fetal membranes.
RNA sequencing (RNA-Seq) of placental and fetal membrane tissues and cells was done. Differential expressions of CYP450 genes were compared and validated via reverse transcription-quantitative polymerase chain reaction (RT-qPCR) between the two tissues. The functional activity of major CYP450 enzymes was determined using a fluorophore-based enzymatic assay in the presence and absence of their corresponding inhibitors.
With the exception of genes that regulate cholesterol metabolism, the expression profile of CYP450 genes was similar between placental and fetal membranes tissues/cells. RT-qPCR analysis confirmed these findings with significant levels of mRNA for major CYP450 genes being detectable in amnion epithelial cells (AECs) and chorion trophoblasts cells (CTCs). Biochemical analyses revealed significant CYP450 enzymatic activities that were sensitive to specific inhibitors for both AECs and CTCs, suggesting that the genes were expressed as functional enzymes.
This is the first study to determine global expression of CYP450 enzymes in fetal membranes which may play a role in xenobiotic metabolism during pregnancy. Given that many women are exposed to environmental toxins or require medications during pregnancy, a better understanding of their role in metabolism is required to develop safer therapeutics and prevent adverse outcomes.
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•Exposure to drugs and toxicants a major risk factor for preterm birth•Global expression of CYP450 system not yet described in fetal membranes•Similar CYP450 expression profiles were seen in placenta and fetal membranes.•Major CYP450 enzymes were found to be functionally active in fetal membrane cells.•Fetal membranes could play a role in drug and toxicant metabolism during pregnancy.
Polybrominated diphenyl ethers (PBDEs) enhance basal and bacteria-stimulated production of proinflammatory cytokines by the placenta and may also increase the risk of preterm birth and ...neurodevelopmental disorders. Whether or not other brominated flame retardants such as Tetrabromobisphenol A (TBBPA) have similar properties is unclear. Therefore, we evaluated the effects of TBBPA on the production of steroids, as well as biomarkers for inflammation, oxidative stress, and neurodevelopment by the placenta.
Placental explant cultures were established from women undergoing elective Cesarean sections at term and treated with 5–50,000 nM TBBPA in the presence and absence of 107 CFU/ml heat-killed E. coli. Concentrations of P4, E2, testosterone (T), IL-1β, TNF-α, IL-10, HO-1, IL-6, 8-IsoP and BDNF were quantified in the conditioned medium.
In the absence of bacteria, TBBPA tended to increase P4 and T as well as IL-6 and 8-IsoP. For bacteria-treated cultures, TBBPA generally inhibited P4, IL-1β, and HO-1 production but enhanced TNF-α and IL-6 production.
TBBPA alters placental steroidogenesis to favor T production and increases oxidative stress and placental inflammation as suggested by its promotion of 8-IsoP and IL-6 production. TBBPA may also function as a risk modifier where it enhances bacteria-stimulated production TNF-α and IL-6 but reduces HO-1 production, however, this was balanced by reductions in IL-1β. Further studies are warranted to determine if TBBPA increases the risk of adverse pregnancy outcomes such as preterm birth, pPROM and neurodevelopmental disorders such as autism that have been associated with increased production of proinflammatory cytokines, oxidative stress and/or T.
•Tetrabromobisphenol A (TBBPA) had no detectible effect on placental culture viability.•TBBPA increased Testosterone (T), IL-6 and TNF-α production by the placenta.•Elevated IL-6, TNF-α and T levels are linked with preterm birth and/or autism.•Further studies of TBBPA on pregnancy and neurodevelopmental outcomes are warranted.
Exosomes are intercellular signaling vesicles whose cargo reflects the physiological status of the cell of their origin and can regulate gene expression in other tissues. Polybrominated diphenyl ...ethers (PBDEs) and bisphenols (A BPA, Tetrabromobisphenol A TBBPA, and 2,4,6-Tribromophenol TBP) are common environmental pollutants known to increase the risk for spontaneous preterm birth (PTB). We hypothesized that placental exposure to these environmental pollutants causes exosome cargo changes that reflect exposure associated placental response.
Full-term, C-section placenta explants were treated with PBDE congeners (47, 100, 153, 209), TBBPA, TBP or BPA for 24 h. Exosomes were isolated from media by sequential ultracentrifugation and purified by size exclusion chromatography. Exosomes were characterized by electron microscopy, nanoparticle tracking analysis and Western blot. Proteomics identified differentially expressed exosomal proteins and Ingenuity pathway analysis (IPA) determined biological functions and pathways represented by identified proteins.
Regardless of treatment, placental expressed exosomes markers (PLAP, CD9, CD63, 81 and ALIX), had a size distribution between 50 and 175 nm and were present in the conditioned medium at 5–8 x 1011 exosomes/mL. Proteomic analysis identified 2598 proteins which demonstrated that specific pollutants caused differential expression of specific proteins, including alarmin, High Mobility Group Box 1 (HMGB1), MAPK14 (p38 MAPK) and GSK3β. IPA revealed an inhibition of pathways associated with cell survival, tissue repair and proliferation, as well as activation of cell death pathways (e.g. necrosis).
Environmental exposure of placental explants did not change the quantity of exosomes or their characteristics. However, exosome cargo composition exposed to some environment pollutants may be involved in placental nuclear and cellular injury and inflammation.
•Exosomes from environmental pollutants exposed placenta had classic features.•Exosome characteristics were similar in pollutant exposed and control placenta.•Exosomes carry differentially expressed protein cargo reflective of exposure.•PBDEs and BPA/TBBP/246TBP produced exosome cargo reflective of placental injury.
•Dibutyl Phthalate had no effect on viability of placental cultures.•Dibutyl Phthalate enhanced basal Interleukin-6 production.•Bacteria-stimulated IL-1β and TNF-α secretion was enhanced by Dibutyl ...Phthalate.•BDNF production by bacteria-treated placentas was inhibited by Dibutyl Phthalate.
Recent studies suggest that women with high exposures to dibutyl phthalate (DBP) are at increased risk for preterm birth, a condition associated with aberrant inflammation in the placenta often caused by subclinical infections. Placental inflammation is also a risk factor for neurodevelopmental disorders whose risk may also be enhanced by DBP. It is unclear, however, if DBP enhances placental inflammation. Therefore, we studied the effects of DBP on the production of biomarkers of placental inflammation and neurodevelopment under basal conditions and a setting of mild infection. Placental explant cultures established from women undergoing elective caesarean delivery were treated with DBP with and without co-stimulation by 107 CFU/mL heat-killed E. coli for 24 h at 37 °C. Conditioned medium was harvested and concentrations of IL-1β, TNF-α, IL-10, HO-1 and BDNF, a biomarker for neurodevelopment, were quantified. DBP significantly enhanced IL-6 production in basal cultures but had no significant on the other biomarkers quantified. Both TNF-α and IL-1β production was enhanced by DBP for cultures co-stimulated with E. coli. Although marginal enhancement of IL-6, and IL-10 were observed for bacteria co-treated cultures, results were either non-monotonic or only approached statistical significance. HO-1 production tended to be reduced at the highest concentration of DBP tested and BDNF production was reduced by DBP in a dose-dependent manner for bacteria-stimulated cultures. These results suggest that DBP enhances basal IL-6 production but has little or no effect on other biomarkers studied. However, DBP enhances IL-1β and TNF-α production but reduces BDNF production by bacteria-stimulated cultures.
During pregnancy there is an alteration in maternal immunity within the uterus where innate, proinflammatory immune responses are tightly regulated to prevent immunological rejection of the fetal ...allograft. Disruption of the delicate balance of cytokines by bacteria or other factors increases the production of proinflammatory cytokines at the maternal-fetal interface and activates the parturition mechanism prematurely. Despite years of searching, there is still no broadly effective strategy for preventing preterm labor and most therapies are directed at inhibiting myometrial contractions and improving neonatal outcome. Recent studies with progestins and interleukin-10 (IL-10), however, are showing promise in randomized clinical trials and animal studies. Furthermore, the identification of the Toll-like receptors as upstream mediators of inflammation may offer alternative therapeutic targets for preventing this common pregnancy complication.
Globally, ∼50 % of women smoke during pregnancy and the prevalence of vaping is increasing among women of reproductive age. However, the health effects of vaping during pregnancy are largely unknown. ...This study examined the effects of e-cig constituents alone and in combination (propylene glycol PG, vegetable glycerin VG, and nicotine) on human placental tissue viability (MTT assay) and immunoassayed levels of placenta-derived biomarkers, i.e., 8-isoprostane (8-IsoP), heme oxygenase-1 (HO-1), interleukin-6 (IL-6), β-estradiol (E2), progesterone (P4), allopregnanolone (AP), and brain-derived neurotrophic factor (BDNF). Placental explant cultures were exposed ex vivo for 24 h to media-containing either nicotine (0–5000 nM), PG/VG (0–8 % v/v at 50/50 ratio), or a combination of both. No effects on tissue viability were observed at PG/VG concentrations < 8 % (v/v), while viability significantly reduced at PG/VG concentrations ≥ 10 % (v/v); biomarker studies employed only non-cytotoxic doses. Exposure to PG/VG decreased levels of 8-IsoP, IL-6, and E2, and treatment with 2 % or 8 % PG/VG significantly reduced HO-1 levels, compared to non-treated controls. Exposure to nicotine alone at 2,500 nM and 5,000 nM reduced MTT activity by 20 % (P = 0.04) and 70 % (P < 0.001), respectively, and significantly increased (P < 0.001) levels of HO-1 and BDNF, compared to controls. Treatment with nicotine alone and in combination with PG/VG reduced IL-6 and E2 levels. Interestingly, nicotine-induced toxicity was attenuated by PG/VG addition to nicotine-treated groups. These studies demonstrate that e-cig constituents negatively impact the human placenta and alters production of critical placental biomarkers, suggesting that vaping is an unsafe alternative for pregnant women or their unborn fetus.
•E-cigs have become popular among adolescents and young adults of reproductive age.•Potential impacts on pregnancy and fetal development are still unknown.•Analyzed the effects of 2 e-cig components on placenta cytokine production.•Production of biomarkers was altered after various exposure treatments.
Background Bisphenol-A (BPA) is a widespread pollutant whose effects on pregnant women are poorly understood. Therefore, we investigated the effects of BPA on basal and bacteria-stimulated production ...of proinflammatory cytokines interleukin (IL)-1β, tumor necrosis factor-α (TNF-α) and IL-6, anti-inflammatory mediators soluble glycoprotein 130 (sgp) 130, heme oxidase-1 (HO-1) and IL-10 and biomarkers for neurodevelopment brain-derived neurotrophic factor (BDNF), and oxidative stress 8-isoprostane (8-IsoP) by the placenta. Methods Placental explant cultures were treated with BPA (0-10,000 nM) in the presence or absence of 107 colony-forming unit (CFU)/mL heat-killed Escherichia coli for 24 h. Biomarker concentrations in conditioned medium were quantified by the enzyme-linked immunosorbent assay (ELISA). Results Under basal conditions, IL-1β and IL-6 production was enhanced by BPA in a dose-dependent manner. Sgp130, a soluble receptor that reduces IL-6 bioactivity, was suppressed by BPA at 1000-10,000 nM. BPA also enhanced BDNF production at 1000 and 10,000 nM, and 8-IsoP expression at 10 and 100 nM. For bacteria-treated cultures, BPA increased IL-6 production at 100 nM and reduced sgp130 at 1000 nM but had no effect on IL-1β, TNF-α, BDNF, HO-1, 8-IsoP or IL-10 production. Conclusion BPA may increase placental inflammation by promoting IL-1β and IL-6 but inhibiting sgp130. It may also disrupt oxidative balance and neurodevelopment by increasing 8-IsoP and BDNF production.
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