Placental Cell Line Research Articles

The role of Smad4 in the regulation of insulin resistance, inflammation and cell proliferation in HTR8-Svneo cells.

Gestational diabetes mellitus (GDM) is a metabolic disorder whose major pathophysiological basis is demonstrated as placental insulin resistance (IR), while Smad4 always functions in the signal transduction of transforming growth factor beta (TGF-β) pathway. Our study aims to figure out the role of Smad4 in an insulin resistance (IR) cellular model using placental trophoblast cell line. Importantly, HTR8-Svneo cells, in the status of IR, indicated a significant increase in the expression of Smad4. Subsequently, the HTR8-Svneo cell line with up-regulated or depleted Smad4 was respectively achieved by the effective over-expressed plasmid or siRNA of Smad4. We found out that the deficiency of Smad4 could promote the insulin sensitivity and restrict the inflammatory response in IR group of cells with significant augment in glucose uptake, up-regulation of insulin signalling-related molecules and attenuation in inflammatory biomarker expressions. On the contrary, the over-expression of Smad4 showed a reversal effect on these alterations in IR group of cells. Besides, the positive effect of Smad4 on cell viability was also observed in our study. SIGNIFICANCE OF THE STUDY: Gestational diabetes mellitus (GDM) is a metabolic disorder whose major pathophysiological basis is demonstrated as insulin resistance (IR). Importantly, our findings indicate that the deficiency of Smad4 significantly improves the insulin sensitivity and relieves the inflammation in the cellular model of IR. Besides, the positive effect of Smad4 on cell viability was also observed in our study. Our present findings provide novel insights for the investigation on molecular details about the GDM pathogenesis.

Appropriate citation of placenta cell lines 3A(tPA-30-1) and 3A-sub E [post crisis of 3A(tPA-30-1)] in medical literature

IntroductionTo determine how often placenta cell lines 3A (tPA-30-1) and 3A-sub E [post crisis of 3A (tPA-30-1)] are appropriately cited, or identified, as “term”-gestation placental cell lines in medical literature. MethodsWe performed a literature search on two databases, PubMed and One Search, using the terms “3A (tPA-30-1),” “3Asub-E,” “3AsubE,” “tPA-30-1,” “tPA30-1,” and “3A AND (placenta OR placental OR trophoblast OR trophoblastic) AND (cell OR line OR cell line).” Of the 218 citations retrieved, 181 were excluded due to duplication, article content irrelevance or lack of access to a full manuscript. The remaining 37 citations were thoroughly reviewed for 1)the presence of a full citation as designated by the supplier, and 2)the identification of the placental lines as “term.” ResultsOf the 37 eligible citations included in the study, five demonstrated complete identifications of the placental cell lines of interest, while 32 demonstrated partial identifications that failed to match the designations provided by the manufacturer. Furthermore, of the 37 citations, eight accurately identified the cell lines as “term,” while 27 lacked any description of gestational age, and two incorrectly identified them as “first trimester” cell lines. Overall, only three citations contained both a full citation and correct identification as a “term” placenta cell line. DiscussionOnly 5 of the 37 (13.5%) publications demonstrated a complete citation and only 8 publications accurately identified the gestational age of the placenta cell line as “term”. Such findings confirm the need for a representative set of standards for the documentation of cell lines to improve the quality of publications in the scientific community.

Transfer Mechanisms of Compounds between Mother and Fetus/Infant Aimed for Optimized Medication during Pregnancy and Breastfeeding

Potential risks to the fetus or infant should be considered prior to medication during pregnancy and lactation. It is essential to evaluate the exposure levels of drugs and their related factors in addition to toxicological effects. Epilepsy is one of the most common neurological complications in pregnancy; some women continue to use antiepileptic drugs (AEDs) to control seizures. Benzodiazepines (BZDs) are widely prescribed for several women who experience symptoms such as anxiety and insomnia during the postpartum period. In this review, we describe the 1) transport mechanisms of AEDs across the placenta and the effects of these drugs on placental transporters, and 2) the transfer of BZDs into breast milk. Our findings indicated that carrier systems were involved in the uptake of gabapentin (GBP) and lamotrigine (LTG) in placental trophoblast cell lines. SLC7A5 was the main contributor to GBP transport in placental cells. LTG was transported by a carrier that was sensitive to chloroquine, imipramine, quinidine, and verapamil. Short-term exposure to 16 AEDs had no effect on folic acid uptake in placental cells. However, long-term exposure to valproic acid (VPA) affected the expression of folate carriers (FOLR1, SLC46A1). Furthermore, VPA administration changed the expression levels of various transporters in rat placenta, suggesting that sensitivity to VPA differed across gestational stages. Lastly, we developed a method for quantifying eight BZDs in human breast milk and plasma using LC/MS/MS, and successfully applied it to quantify alprazolam in breast milk and plasma donated by a lactating woman.

In vitro function and in situ localization of Multidrug Resistance-associated Protein (MRP)1 (ABCC1) suggest a protective role against methyl mercury-induced oxidative stress in the human placenta

Methyl mercury (MeHg) is an organic highly toxic compound that is transported efficiently via the human placenta. Our previous data suggest that MeHg is taken up into placental cells by amino acid transporters while mercury export from placental cells mainly involves ATP binding cassette (ABC) transporters. We hypothesized that the ABC transporter multidrug resistance-associated protein (MRP)1 (ABCC1) plays an essential role in mercury export from the human placenta. Transwell transport studies with MRP1-overexpressing Madin-Darby Canine Kidney (MDCK)II cells confirmed the function of MRP1 in polarized mercury efflux. Consistent with this, siRNA-mediated MRP1 gene knockdown in the human placental cell line HTR-8/SVneo resulted in intracellular mercury accumulation, which was associated with reduced cell viability, accompanied by increased cytotoxicity, apoptosis, and oxidative stress as determined via the glutathione (GSH) status. In addition, the many sources claiming different localization of MRP1 in the placenta required a re-evaluation of its localization in placental tissue sections by immunofluorescence microscopy using an MRP1-specific antibody that was validated in-house. Taken together, our results show that (1) MRP1 preferentially mediates apical-to-basolateral mercury transport in epithelial cells, (2) MRP1 regulates the GSH status of placental cells, (3) MRP1 function has a decisive influence on the viability of placental cells exposed to low MeHg concentrations, and (4) the in situ localization of MRP1 corresponds to mercury transport from maternal circulation to the placenta and fetus. We conclude that MRP1 protects placental cells from MeHg-induced oxidative stress by exporting the toxic metal and by maintaining the placental cells' GSH status in equilibrium.

Expression and function of lncRNA MALAT1 in gestational diabetes mellitus.

Gestational diabetes mellitus (GDM) severely threatens maternal and fetal health. Long non-coding RNA (lncRNA) participates in the regulation of various cellular processes. Previous studies have identified the role of lncRNA MALAT1 in diabetic retinopathy-related inflammation. However, the role of lncRNA MALAT1 in GDM has not been reported yet. Real-time polymerase chain reaction (RT-PCR) was used to measure the lncRNA MALAT1 expression level in placental tissues from GDM patients and from a normal pregnant group. Placental trophoblastic-derived cell line HTR8 cells were divided into a control group, an siRNA negative control group and a MALAT1 siRNA group. The cells underwent RT-PCR analysis of lncRNA MALAT1 expression, an MTT assay of cell proliferation, and a transwell assay of cell invasion and migration. In addition, enzyme-linked immunosorbent assay (ELISA) was used to analyze the level of tumor necrosis factor α (TNF-α) and interleukin 6 (IL-6). Western blotting was used to measure the changes of the tumor growth factor β (TGF-β)/nuclear factor-kappa B (NF-κB) signaling pathway. Gestational diabetes mellitus placental tissues showed higher lncRNA MALAT1 expression compared to a normal control group (p < 0.05). After siRNA intervention, lncRNA MALAT1 showed decreased expression in the trophoblastic layer; inhibited trophoblastic cell proliferation, migration, or invasion; decreased the secretion of inflammatory factors TNF-α and IL-6; and suppressed the expression of TGF-β and NF-κB compared to that of the control and siRNA-NC groups (p < 0.05). Gestational diabetes mellitus appears to upregulate lncRNA MALAT1. Downregulation of lncRNA MALAT1 inhibits inflammation and suppresses the proliferation, invasion and migration of GDM placental trophoblastic cells, possibly by modulating the TGF-β/NF-κB signaling pathway.

Brown Rice Extract Ameliorates Oxidative Status in Human Placental Cell Line (JEG-3) Induced by Cadmium Chloride

Cadmium (Cd) can induce oxidative stress by producing reactive oxygen species (ROS) and affect the antioxidant defense systems, leading to cellular damage. Supplementation with antioxidants may protect against Cd-induced oxidative stress. This study aimed to investigate the effect of germinated brown rice (GBR), brown rice (BR), and white rice (WR) extracts against Cd-induced toxicity in human placenta (JEG-3) cells. The cytotoxicity of Cd and rice extracts was measured by MTT assay, and pre-treatments with GBR, BR, and WR before Cd exposure were examined. Intracellular ROS levels and the expression of SOD2 and GPx4 were determined by DCFH-DA assay and western blot, respectively. An antioxidant N-acetylcysteine (NAC) was used as a positive control. The antioxidant capacity and total phenolic content of rice extracts were investigated by DPPH and Folin-Ciocalteu methods, respectively. The GBR extract exhibited the highest antioxidant activity and total phenolic content, followed by BR and WR, respectively. Cd decreased cell viability in a dose-dependent manner, whereas incubation with 0.1 - 0.5 mg/mL showed no toxicity in all rice extracts. Moreover, Cd treatments significantly (p &lt; 0.05) increased ROS levels. Conversely, treatments with GBR and BR extracts decreased ROS. Similar results were observed for combinatorial treatments of GBR and BR with Cd, except for WR treatment. Additionally, significant increased SOD2 and decreased GPx4 expression levels were evident in the GBR group and in both GBR and BR pre-treated groups, respectively, compared to Cd exposure alone. These results suggest that brown rice pre-treatments may ameliorate oxidative status induced by Cd in cultured human placental cells.

Unfractionated heparin displaces sFlt-1 from the placental extracellular matrix

Soluble vascular endothelial growth factor receptor-1 (sFlt-1) is an anti-angiogenic protein which is secreted by numerous cell types and acts as a decoy receptor for the angiogenic protein vascular endothelial growth factor (VEGF). Despite its physiologic importance in maintaining angiogenic balance, excess sFlt-1 levels are associated with the pathogenesis of many diseases, especially those with angiogenic imbalance, endothelial dysfunction, and hypertension. Although sFlt-1 is a soluble protein, it contains a binding site for the extracellular matrix component heparan sulfate. This allows cells to retain and localize sFlt-1 in order to prevent excessive VEGF signaling. During pregnancy, placental syncytiotrophoblasts develop a large extracellular matrix which contains significant amounts of heparan sulfate. Consequently, the placenta becomes a potential storage site for large amounts of sFlt-1 bound to extracellular heparan sulfate. Additionally, it should be noted that sFlt-1 can bind to the anticoagulant unfractionated heparin due to its molecular mimicry to heparan sulfate. However, it remains unknown whether unfractionated heparin can compete with heparan sulfate for binding of localized sFlt-1. In this study, we hypothesized that administration of unfractionated heparin would displace and solubilize placental extracellular matrix(ECM)-bound sFlt-1. If unfractionated heparin can displace this large reservoir of sFlt-1 in the placenta and mobilized it into the maternal circulation, we should be able to observe its effects on maternal angiogenic balance and blood pressure. To test this hypothesis, we utilized in vitro, ex vivo, and in vivo methods. Using the BeWo placental trophoblast cell line, we observed increased sFlt-1 in the media of cells treated with unfractionated heparin compared to controls. The increase in media sFlt-1 was found in conjunction with decreased localized cellular Flt (sFlt-1 and Flt-1) as measured by total cell fluorescence. Similar results were observed using ex vivo placental villous explants treated with unfractionated heparin. Real-time quantitative PCR of the explants showed no change in sFlt-1 or heparanase-1 mRNA expression, eliminating increased production and enzymatic cleavage of heparan sulfate as causes for sFlt-1 media increase. Timed-pregnant rats given a continuous infusion of unfractionated heparin exhibited an increased mean arterial pressure as well as decreased bioavailable VEGF compared to vehicle-treated animals. These data demonstrate that chronic unfractionated heparin treatment is able to displace matrix-bound sFlt-1 into the maternal circulation to such a degree that mean arterial pressure is significantly affected. Here we have shown that the placental ECM is a storage site for large quantities of sFlt-1, and that it should be carefully considered in future studies concerning angiogenic balance in pregnancy.

Tert-Butyl Hydroperoxide Stimulated Apoptosis Independent of Prostaglandin E2 and IL-6 in the HTR-8/SVneo Human Placental Cell Line.

Significant gaps exist in our knowledge of how cellular redox status, sometimes referred to as oxidative stress, impacts placental trophoblasts. The present study used tert-butyl hydroperoxide (TBHP) as a known generator of reactive oxygen species (ROS) in the extravillous trophoblast cell line HTR-8/SVneo to examine the role of cellular redox disruption of prostaglandin E2 (PGE2) and the cytokine IL-6 in cell death. Cells were exposed to 0, 12.5, 25, or 50μM TBHP for 4, 8, and 24h to ascertain effects on cell viability, caspase 3/7 activity, PGE2 release, PTGS2 mRNA expression, and IL-6 release. Experiments with inhibitors included the cyclooxygenase inhibitor indomethacin, mitogen-activated protein kinase inhibitors (PD169316, U0126, or SP600125), or treatments to counter expected consequences of TBHP-stimulated generation of ROS (deferoxamine [DFO], butylated hydroxyanisole [BHA], and N,N'-diphenyl-1,4-phenylenediamine [DPPD]) using 24-h exposure to 50μM TBHP. Cell viability, measured by ATP content, decreased 24% relative to controls with a 24-h exposure to 50μM TBHP, but not at lower TBHP concentrations nor at earlier time points. Exposure to 50μM TBHP increased caspase 3/7 activity, an indicator of apoptosis, after 8 and 24h. Antioxidant treatment markedly reduced TBHP-stimulated caspase 3/7 activity, PGE2 release, and IL-6 release. TBHP-stimulated IL-6 release was blocked by PD169316 but unaltered by indomethacin. These data suggest that TBHP-stimulated IL-6 release and caspase 3/7 activation were independent of PGE2 yet were interrupted by treatments with known antioxidant properties, providing new insight into relationships between PGE2, IL-6, and apoptosis under conditions of chemically induced cellular oxidation.

Synthetic PreImplantation Factor (sPIF) reduces inflammation and prevents preterm birth.

Preterm birth (PTB) is the leading cause of neonatal morbidity and mortality and spontaneous PTB is a major contributor. The preceding inflammation/infection contributes not only to spontaneous PTB but is associated with neonatal morbidities including impaired brain development. Therefore, control of exaggerated immune response during pregnancy is an attractive strategy. A potential candidate is synthetic PreImplantation Factor (sPIF) as sPIF prevents inflammatory induced fetal loss and has neuroprotective properties. Here, we tested maternal sPIF prophylaxis in pregnant mice subjected to a lipopolysaccharides (LPS) insult, which results in PTB. Additionally, we evaluated sPIF effects in placental and microglial cell lines. Maternal sPIF application reduced the LPS induced PTB rate significantly. Consequently, sPIF reduced microglial activation (Iba-1 positive cells) and preserved neuronal migration (Cux-2 positive cells) in fetal brains. In fetal brain lysates sPIF decreased IL-6 and INFγ concentrations. In-vitro, sPIF reduced Iba1 and TNFα expression in microglial cells and reduced the expression of pro-apoptotic (Bad and Bax) and inflammatory (IL-6 and NLRP4) genes in placental cell lines. Together, maternal sPIF prophylaxis prevents PTB in part by controlling exaggerated immune response. Given the sPIF`FDA Fast Track approval in non-pregnant subjects, we envision sPIF therapy in pregnancy.

Bavachin suppresses human placental choriocarcinoma cells by targeting electron transport chain complexes and mitochondrial dysfunction

Phytoestrogens are naturally derived estrogen-like therapeutic compounds that have long been studied for their role as anti-cancer agents and supplements during chemotherapy. Bavachin is a therapeutic phytoestrogen used to treat cancer, inflammation, and diabetes mellitus. Though the therapeutic effects of bavachin on various diseases have been explored, its anti-cancer effects and related mechanisms in human placental choriocarcinoma remain unknown. This is the first study to identify the anti-cancer potential of bavachin on human placental choriocarcinoma cell lines JEG3 and JAR. Placental mitochondria support the elevated energy production required for placental development, through oxidative phosphorylation (OXPHOS). Based on this concept, we hypothesized that mitochondrial targeting by bavachin may contribute to anti-cancer activities in high-OXPHOS subtypes of cancer such as placental choriocarcinoma. Apoptosis and caspase activities were increased by bavachin in placental choriocarcinoma cells. Bavachin altered metabolic phenotypes by regulating electron transport chain complex and OXPHOS to suppress choriocarcinoma cell proliferation. It also led to calcium disruption and endoplasmic reticulum stress accompanied by mitochondrial membrane potential depolarization. It showed synergistic anti-cancer effects with paclitaxel on placental choriocarcinoma cells. Taken together, we suggest that bavachin has therapeutic potential against placental choriocarcinoma and may be used to counter paclitaxel-induced toxicity.

1336-P: Trimethylamine N-Oxide Inhibits the Formation of Neutrophil Extracellular Traps to Regulate Placental Development in Gestational Diabetes Mellitus

Objective: Trimethylamine N-oxide (TMAO) is associated with risk of gestational diabetes mellitus (GDM), and TMAO levels in peripheral/cord blood of GDM mothers increased. Diabetes primes neutrophils to produce neutrophil extracellular traps (NETs), but it has not been reported in GDM. We determined the relationship between NETs and GDM, and explored potential role of TMAO. Methods: Maternal peripheral blood was collected during OGTT tests, and cord blood and placenta were collected during delivery. Plasma concentrations of cell-free double-stranded DNA (dsDNA) were compared between GDM and control mothers and association analysis was performed. Expression of myeloperoxidase (MPO) in placental tissues of GDM and control mothers was examined. Neutrophils from the blood of healthy donors were used to observe effects of TMAO on NETs formation. The effect of TMAO on human placental extravillous trophoblast cell line HTR-8/Svneo was examined. Results: Compared with controls, plasma concentrations of dsDNA in peripheral blood and cord blood were significantly increased in GDM mothers. Association analysis found that concentrations of dsDNA in peripheral and cord blood plasma were positively correlated with BMI and blood glucose levels, but not blood pressure. In addition, the placental thickness was positively correlated with dsDNA levels in cord blood plasma. MPO expression in the placental tissues of GDM mothers was also significantly higher than that of control mothers. TMAO-treated neutrophils could resist the formation of NETs stimulated by PMA or LPS, and TMAO promotes the proliferation, migration, invasion and angiogenesis of the cells. Conclusion: The formation of NETs was increased in GDM, and was positively correlated with BMI and blood glucose. TMAO could inhibit NETs formation and promote invasion and angiogenesis of human placental trophoblast cells, which may be involved in the development of the placenta. Disclosure X. Lin: None. X. He: None. X. Xiao: None.

Exposure to wood smoke particles leads to inflammation, disrupted proliferation and damage to cellular structures in a human first trimester trophoblast cell line

The ongoing transition to renewable fuel sources has led to increased use of wood and other biomass fuels. The physiochemical characteristics of biomass combustion derived aerosols depends on appliances, fuel and operation procedures, and particles generated during incomplete combustion are linked to toxicity. Frequent indoor wood burning is related to severe health problems such as negative effects on airways and inflammation, as well as chronic hypoxia and pathological changes in placentas, adverse pregnancy outcome, preterm delivery and increased risk of preeclampsia. The presence of combustion-derived black carbon particles at both the maternal and fetal side of placentas suggests that particles can reach the fetus. Air pollution particles have also been shown to inhibit trophoblast migration and invasion, which are vital functions for the development of the placenta during the first trimester. In this study we exposed a placental first trimester trophoblast cell line to wood smoke particles emitted under Nominal Burn rate (NB) or High Burn rate (HB). The particles were visible inside exposed cells and localized to the mitochondria, causing ultrastructural changes in mitochondria and endoplasmic reticulum. Exposed cells showed decreased secretion of the pregnancy marker human chorionic gonadotropin, increased secretion of IL-6, disrupted membrane integrity, disrupted proliferation and contained specific polycyclic aromatic hydrocarbons (PAHs) from the particles. Taken together, these results suggest that wood smoke particles can enter trophoblasts and have detrimental effects early in pregnancy by disrupting critical trophoblast functions needed for normal placenta development and function. This could contribute to the underlying mechanisms leading to pregnancy complications such as miscarriage, premature birth, preeclampsia and/or fetal growth restriction. This study support the general recommendation that more efficient combustion technologies and burning practices should be adopted to reduce some of the toxicity generated during wood burning.

SAT-734 HSD3B1 Expression in the Human Immune System

3β-hydroxysteroid dehydrogenase-1 (3βHSD1), catalyzing conversion of dehydroepiandrosterone (DHEA) to Δ 4-androstenedione, is an essential enzyme in the pathway toward production of biologically active androgens such as dihydrotestosterone from the adrenally produced precursor DHEA sulfate, the most predominant steroid hormone in circulation. We previously identified, in the gene (HSD3B1) that encodes 3βHSD1, a germline gain-of-function missense-encoding variant that has now been validated in several studies as predicting more rapid progression in prostate cancer patients treated with gonadal testosterone deprivation. Production of androgens from adrenal precursors is important not just in the context of prostate cancer but in other physiologic and pathophysiologic processes, which could include asthma. Androgens are associated with better lung function in both asthma and healthy cohorts, and increasing circulating androgen levels in males help explain the switchover in asthma being more common in boys than girls but then more common in women than men. A main treatment for asthma, as well as other inflammatory processes, is administration of glucocorticoids, yet unresponsiveness to glucocorticoids in a subset of patients remains a major problem. Systemic glucocorticoid administration suppresses adrenally produced DHEA and DHEA-S, suggesting a depleted pool for biologically active androgen production as a mechanism for glucocorticoid resistance. Our surprising preliminary data support a link between glucocorticoid responsiveness and the more active HSD3B1 allele: patients homozygous for the adrenal-permissive HSD3B1(1245C) allele exhibit better response to oral glucocorticoids than those homozygous for the adrenal-restrictive HSD3B1(1245A), with heterozygous patients falling in the middle. This suggests a model in which patients with the more active (adrenal-permissive) form of 3βHSD1 produce sufficient androgens despite the depleted pool of precursor hormones whereas patients with the less active (adrenal-restrictive) form cannot. To further elucidate the link between 3βHSD1 activity and immune response, we assayed HSD3B1 expression in different types of white blood cells. Leukocyte subsets from asthma patients and healthy controls were purified using fluorescence-activated cell sorting, and HSD3B1 expression was analyzed using qPCR. White blood cells of several types expressed HSD3B1 at levels comparable to or greater than both prostate cancer and placental choriocarcinoma cell lines with very robust 3βHSD1 activity. Further determining the cell type specific expression and activity of this key enzyme is an important step in unraveling the link between the HSD3B1 polymorphism and asthma along with potentially many other immune processes.

Schinus terebenthifolius Raddi extracts: From sunscreen activity toward protection of the placenta to Zika virus infection, new uses for a well-known medicinal plant

Schinus terebinthifolius Raddi is a well-known medicinal plant native of South America. This species has demonstrated important biological activities such as antihypertensive and vasodilator, antimicrobial, anti-inflammatory and antioxidant. However, no studies have been, so far, reported with the fruits of S. terebinthifolius as a protector of the placenta against Zika virus infection and as sunscreen agents. The present study aimed to investigate new uses for the ethanolic fruit extracts of S. terebinthifolius, from fruits’peel (STPE) and from the whole fruits (STWFE). Zika virus (ZIKV) has been linked to several fetal malformations, such as microcephaly and other central nervous system abnormalities. Thus, the potential of these natural extracts against ZIKV infection was evaluated, using an in vitro method. The photoprotective potential, determined by spectrometry, along with phenolic content, antioxidant capacity, and chemical composition of both extracts were also evaluated. The chemical composition of the extracts was evaluated by HPLC-UV / vis. The cytotoxicity of peel and whole fruit extracts in vero E6 cell lines, in placental cell lines and placental explant cultures were evaluated by the MTT assay. The infectivity of placental cells and explants was evaluated by qRT-PCR and the effects of extracts on ZIKV infection were investigated using HTR-8/SVneo cells, pre-treated with 100 μg mL−1 of STWFE for 1 h, and infected with MR766 (AD) or PE243 (EH) ZIKV strains. STFE and STWFE were well-tolerated by both placental-derived trophoblast cell line HTR-8/SVneo as well as by term placental chorionic villi explants, which indicate absence of cytotoxicity in all analysed concentrations. Two strains of ZIKV were tested to access if pre-treatment of trophoblast cells with the STWFE would protect them against infection. Flow cytometry analysis revealed that STWFE extract greatly reduced ZIKV infection. The extracts were also photoprotective with SPF values equivalent to the standard, benzophenone-3. The formulations prepared in different concentrations of the extracts (5–10 %) had shown maximum SPF values of 32.21. STWFE represents a potential natural mixture to be used in pregnancy in order to restrain placental infection by ZIKV and might potentially protect fetus against ZIKV-related malformations. The extracts exhibited photoprotective activity and some of the phenolic compounds, mainly resveratrol, catechin and epicatechin, are active ingredients in all assayed activities. The development of biotechnological/medical products, giving extra value to products from family farming, is expected, with strong prospects for success.

FTIR spectroscopy as a novel analytical approach for investigation of glucose transport and glucose transport inhibition studies in transwell in vitro barrier models.

Glucose transport is key for cellular metabolism as well as physiological function and is maintained via passive facilitated and active sodium-glucose linked transport routes. Here, we present for the first time Fourier-transform infrared spectroscopy as a novel approach for quantification of apical-to-basolateral glucose transport of in vitro cell barrier models using liver, lung, intestinal and placental cancer cell lines. Results of our comparative study revealed that distinct differences could be observed upon subjection to transport inhibitors.

Maternal dexamethasone exposure suppresses placental iron transport through GR‐mediated TFR1 pathway in rats

Iron is one of the essential trace elements for almost all living organisms, and of course it is especially important for pregnancy and fetal development. However, inappropriately low or high levels of iron during pregnancy are detrimental and contribute to serious and lasting effects on the fetus, such as fetal growth and development disorders, thereby increasing the risk of adulthood. Here we stress pregnant rats by drinking water with 10−7 mol/L dexamethasone, and then explore its effects on iron metabolism of placenta and fetal rats. The results showed that dexamethasone treatment inhibited placental development, down‐regulated the expression of placental iron transporter TFR1, and decreased iron levels in fetal rats. Furthermore, we use a human placental choriocarcinoma cell line (BeWo) as model to explore the pathway linking inhibit of TFR1 under Dex challenge, our results demonstrate that the decrease in TFR1 caused by dexamethasone is alleviated by the addition of dexamethasone inhibitor RU486. Taken together, our results show that dexamethasone downregulate TFR1 expression adjust iron metabolism via GR‐mediated.Support or Funding InformationThis work was supported by grants from the National Key Research and Development Program of China (2016YFD0500502), the National Natural Science Foundation of China (31872439), the Priority Academic Program Development of Jiangsu Higher Education Institutions and the Jiangsu Collaborative Innovation Center of Meat Production and Processing, Quality and Safety Control.

Molecular analysis of the human placental cysteine dioxygenase type 1 gene

Sulfate is essential for healthy fetal growth and development. Cysteine dioxygenase type 1 (CDO1) plays an important role in the catabolism of cysteine to sulfate. Cdo1 knockout mice exhibit severe and lethal fetal phenotypes but the involvement of CDO1 gene variants in human development is unknown. We searched the NCBI and Ensembl gene databases and identified four alternatively spliced CDO1 coding mRNA transcripts, as well as 148 validated CDO1 gene variants, including 138 missense, 6 nonsense, 1 frameshift, 1 in-frame deletion, and 2 splice site variants. In silico analyses predicted 68 of the missense variants to be deleterious to CDO1 protein structure and function. We examined the relative abundance of the four CDO1 coding mRNA transcripts in human term placentas using qRT-PCR. CDO1 mRNA variant 2 was the most abundant transcript, with intermediate levels of variant 4 and lower levels of variants 1 and 3. Using in situ hybridization, we localised CDO1 mRNA expression to the syncytiotrophoblast layer of human term placenta. To investigate the regulation of CDO1 gene expression, we analysed the transcriptional activity of the human CDO1 5′-flanking region in the JEG-3 placental cell line using luciferase reporter assays. Transcriptional activities were identified in the regions −5 to −269 and − 269 to −1200 nucleotides upstream of the CDO1 transcription initiation site. Mutational analyses of a single nucleotide polymorphism -289C > G that is common in the general population (allele frequency = 0.37) and a putative transcription factor binding motif (CCAAT enhancer binding protein beta) did not alter transcriptional activity of the CDO1 5′-flanking region. Collectively, this study provides an overview and analysis of human CDO1 for future investigations of this gene in human health.

Cellular uptake properties of lamotrigine in human placental cell lines: Investigation of involvement of organic cation transporters (SLC22A1–5)

Lamotrigine (LTG) is an important antiepileptic drug for the treatment of seizures in pregnant women with epilepsy. However, it is not known if the transport of LTG into placental cells occurs via a carrier-mediated pathway. The aim of this study was to investigate the uptake properties of LTG into placental cell lines (BeWo and JEG-3), and to determine the involvement of organic cation transporters (OCTs, SLC22A1–3) and organic cation/carnitine transporter (OCTNs, SLC22A4–5) in the uptake process. The uptake of LTG at 37 °C was higher than that at 4 °C. OCT1 and OCTNs were detected in both cell lines. The uptake of LTG was not greatly affected by the extracellular pH, Na+-free conditions, or the presence of l-carnitine, suggesting that OCTNs were not involved. Although several potent inhibitors of OCTs (chloroquine, imipramine, quinidine, and verapamil) inhibited LTG uptake, other typical inhibitors had no effect. In addition, siRNA targeted to OCT1 had no significant effect on LTG uptake. The mRNA expression in human term placenta followed the order OCTN2 > OCT3 > OCTN1 > OCT1 ≈ OCT2. These observations suggested that LTG uptake into placental cells was carrier-mediated, but that OCTs and OCTNs were not responsible for the placental transport process.

Propylparaben induces apoptotic cell death in human placental BeWo cells via cell cycle arrest and enhanced caspase-3 activity

Propylparaben is one of the most commonly used preservatives in pharmaceuticals, food items, and personal care products. However, growing evidence demonstrates that propylparaben has a harmful effect on various human tissues. In this study, we investigated the cytotoxic effects of propylparaben on the human placental cell line, BeWo. We investigated whether propylparaben caused placental cell death using relevant cytotoxic endpoints, including cell viability, proliferation, cell cycle, and apoptosis, in BeWo cells. However, instead of using conventional methods of protein analysis from pooled samples, we precisely quantified protein expression at the single cell level by flow cytometric analysis. Propylparaben significantly reduced cell viability in a dose-dependent manner. It induced cell cycle arrest at the sub-G1 phase by reducing the expression levels of cyclin D1, whereas the cell population at the G0/G1 and S-phases was decreased. Furthermore, propylparaben induced apoptosis by enhancing the activity of caspase-3. Collectively, our results suggest that propylparaben causes cytotoxic effects in human placental BeWo cells by inducing cell cycle arrest and apoptosis.

Fusarium Mycotoxins Disrupt the Barrier and Induce IL-6 Release in a Human Placental Epithelium Cell Line.

Deoxynivalenol, T-2 toxin, and zearalenone, major Fusarium mycotoxins, contaminate human food on a global level. Exposure to these mycotoxins during pregnancy can lead to abnormalities in neonatal development. Therefore, the aim of this study was to investigate the effects of Fusarium mycotoxins on human placental epithelial cells. As an in vitro model of placental barrier, BeWo cells were exposed to different concentrations of deoxynivalenol, zearalenone or T-2 toxin. Cytotoxicity, effects on barrier integrity, paracellular permeability along with mRNA and protein expression and localization of junctional proteins after exposure were evaluated. Induction of proinflammatory responses was determined by measuring cytokine production. Increasing mycotoxin concentrations affect BeWo cell viability, and T-2 toxin was more toxic compared to other mycotoxins. Deoxynivalenol and T-2 toxin caused significant barrier disruption, altered protein and mRNA expression of junctional proteins, and induced irregular cellular distribution. Although the effects of zearalenone on barrier integrity were less prominent, all tested mycotoxins were able to induce inflammation as measured by IL-6 release. Overall, Fusarium mycotoxins disrupt the barrier of BeWo cells by altering the expression and structure of junctional proteins and trigger proinflammatory responses. These changes in placental barrier may disturb the maternal–fetal interaction and adversely affect fetal development.