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1.
Molecules ; 26(5)2021 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-33652665

RESUMO

The aim of the study was to investigate combined effects of flavonoids (apigenin, baicalein, chrysin, quercetin, and scutellarin) and methyldopa on the expression of selected proinflammatory and vascular factors in vitro for prediction of their action in pregnancy-induced hypertension. The research was conducted on a trophoblast-derived human choriocarcinoma cell line and a primary human umbilical vein endothelial cell line. Cytotoxicity of compounds in selected concentrations (20, 40, and 100 µmol) was measured using the MTT test and the concentration of 40 µmol was selected for further analysis. Subsequently, their effects with methyldopa on the expression of selected markers responsible for inflammation (TNF-α; IL-1ß; IL-6) and vascular effects (hypoxia-inducible factor 1α-HIF-1α; placental growth factor-PIGF; transforming growth factor ß-TGF-ß; vascular endothelial growth factor-VEGF) at the mRNA and protein levels were assessed. It was found that every combined administration of a flavonoid and methyldopa in these cells induced a down-regulating effect on all tested factors, except PIGF, especially at the mRNA expression level. As hypertension generally raises TNF-α, IL-1ß, IL-6, HIF-1α, TGF-ß, and VEGF mRNA expression and/or protein levels, the results obtained in the studied model may provide a positive prognostic factor for such activity in vivo.


Assuntos
Flavonoides/farmacologia , Inflamação/tratamento farmacológico , Metildopa/farmacologia , Doenças Vasculares/tratamento farmacológico , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Técnicas In Vitro , Inflamação/genética , Inflamação/patologia , Placenta/efeitos dos fármacos , Placenta/patologia , Fator de Crescimento Placentário/genética , Gravidez , Trofoblastos/efeitos dos fármacos , Doenças Vasculares/genética , Doenças Vasculares/patologia
2.
Toxicology ; 454: 152741, 2021 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-33662506

RESUMO

Silver bionanoparticles (AgNPs) biosynthesized by Pseudomonas aeruginosa culture supernatant have an important antibacterial activity mediated by ROS increase; however their toxicity in human cells is not known. Due to the high susceptibility of the developing tissues to xenobiotics, the aim of this study was to investigate the AgNPs effect on first trimester human trophoblasts. The HTR8/SVneo cell line was treated with AgNPs (0.3-1.5 pM), for 6 and 24 h. Cell viability, reactive nitrogen and oxygen species (RNS and ROS) production, nitric oxide synthase expression, antioxidant defenses and biomolecule damage were evaluated. The exposure to AgNPs produced changes in HTR8/SVneo cell morphology and decreased cell viability. Alterations in redox balance were observed, with an increase in ROS and RNS levels, and NOS2 protein expression. Superoxide dismutase and catalase augmented their activity accompanied with a decreased in glutathione content and glutathione S-transferase activity. Protein oxidation and genotoxic damage were observed at concentrations greater than 0.6 pM. The pre-incubation with l-NMMA, NAC, mannitol and peroxidase demonstrated that AgNPs-induced cytotoxicity was not mediated by HO and H2O2, but nitric oxide and glutathione pathways were implicated in cell death. Since reported AgNPs microbicidal mechanism is mediated by increasing ROS (mainly HO and H2O2) without an increase in RNS, this work indicates an interesting difference in the reactive species and oxidative pathways involved in AgNPs toxicity in eukaryotic and prokaryotic cells. Highlighting the importance of toxicity evaluation to determine the safety of AgNPs with pharmaceutical potential uses.


Assuntos
Nanopartículas Metálicas/toxicidade , Óxido Nítrico/metabolismo , Prata/toxicidade , Trofoblastos/efeitos dos fármacos , Catalase/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Glutationa/metabolismo , Humanos , Peróxido de Hidrogênio/metabolismo , Pseudomonas aeruginosa/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/metabolismo , Trofoblastos/patologia
3.
Ecotoxicol Environ Saf ; 210: 111872, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33388592

RESUMO

BACKGROUND: Epidemiological studies have revealed that sulfur dioxides (SO2) can increase the risk of pregnancy complications such as missed abortion in the first trimester, stillbirth, preterm birth, small for gestational age, gestational diabetes mellitus and preeclampsia, but the mechanisms underlying these findings remains unknown. What is known, however, is that trophoblasts, a type of fetal cell exerting vital immunologic functions to maintain a successful pregnancy, are usually involved in the pathogenic mechanism of pregnancy complications. OBJECTIVE: To study the effect of SO2 derivatives (bisulfite and sulfite, 1:3 M/M) on the function of trophoblasts. METHODS: Swan.71 trophoblast cells were treated with various concentrations of SO2 derivatives to determine the effect of SO2 derivatives on cellular viability by CKK8. Flow cytometry was performed to analyze the effect of SO2 derivatives on apoptosis, cell cycle and intracellular ROS. Wound healing assay and transwell assay were conducted to examine the migration and invasion of Swan.71 cells. Inflammation-related cytokines in the supernatant (IL-1ß, IL-6, IL-8, IL-10 and TNF-α) were measured by IMMULITE®1000 Systems (SIEMENS). The expression level of NLRP3, Caspase1, MMP9, MMP2, STAT3, and p-STAT3 were evaluated by Western Blotting. RESULTS: Exposure to SO2 derivatives significantly decreased cellular viability, arrested cell cycle at S/G2/M phase and induced cell apoptosis of Swan.71 trophoblasts. In addition, the migration and invasion of Swan.71 cell were significantly inhibited. SO2 derivatives also significantly increased IL-1ß secretion while it is NLRP3/Caspase1 independent. IL-6 secretion was significant inhibited accompanied by decreased STAT3 phosphorylation and expression of MMP2 and MMP9. The intracellular ROS level was significantly suppressed by SO2 derivatives. CONCLUSION: SO2 derivatives exert toxic effects on trophoblasts which results in: suppressing cellular viability and intracellular ROS level, interfering with cell proliferation through arresting cell cycle, inducing cell apoptosis, disturbing inflammation-related cytokines secretion and inhibiting motility. Decreased ROS/IL-6/STAT3 levels play a role in inhibited cell viability, cell cycle arrest, apoptosis and defective motility.


Assuntos
Sulfitos/toxicidade , Trofoblastos/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Citocinas/metabolismo , Feminino , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Gravidez , Espécies Reativas de Oxigênio/metabolismo , Fator de Transcrição STAT3/metabolismo , Trofoblastos/metabolismo
4.
Int J Mol Sci ; 22(2)2021 Jan 12.
Artigo em Inglês | MEDLINE | ID: mdl-33445576

RESUMO

Protease Inhibitors (PI e.g., ritonavir (RTV) and lopinavir (LPV)) used to treat pregnant mothers infected by HIV induce prematurity and endocrine dysfunctions. The maintenance of pregnancy relies on placental hormone production (human Chorionic Gonadotrophin (hCG) and progesterone (P4)). Those functions are ensured by the villous trophoblast and are mainly regulated by the Unfolded Protein Response (UPR) pathway and mitochondria. We investigated, in vitro, if PI impair hCG and P4 production and the potential intracellular mechanisms involved. Term villous cytotrophoblast (VCT) were cultured with or without RTV or LPV from 6 to 48 h. VCT differentiation into syncytiotrophoblast (ST) was followed measuring hCG and P4 secretion. We evaluated the expression of P4 synthesis partners (Metastatic Lymph Node 64 (MLN64), cholesterol side-chain cleavage (P450SCC), Hydroxy-delta-5-Steroid Dehydrogenase and 3 Beta-and steroid delta-isomerase 1 (HSD3B1)), of mitochondrial pro-fusion factors (Mitofusin 2 (Mfn2), Optic Atrophy 1 (OPA1)) and of UPR factors (Glucose-Regulated Protein 78 (GRP78), Activating Transcription Factor 4 (ATF4), Activating Transcription Factor 6 (ATF6), spliced X-box Binding Protein 1 (sXBP1)). RTV had no significant effect on hCG and P4 secretion, whereas lopinavir significantly decreased both secretions. LPV also decreased P450SCC and HSD3B1 expression, whereas it increased Mfn2, GRP78 and sXBP1 expression in ST. RTV has no effect on the endocrine placenta. LPV impairs both villous trophoblast differentiation and P4 production. It is likely to act via mitochondrial fusion and UPR pathway activation. These trophoblastic alterations may end in decreased P4 levels in maternal circulation, inducing prematurity.


Assuntos
Células Endócrinas/efeitos dos fármacos , Células Endócrinas/metabolismo , Inibidores da Protease de HIV/efeitos adversos , Lopinavir/efeitos adversos , Placenta/efeitos dos fármacos , Placenta/metabolismo , Biomarcadores , Células Cultivadas , Vilosidades Coriônicas/efeitos dos fármacos , Vilosidades Coriônicas/metabolismo , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/ultraestrutura , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Feminino , Regulação Enzimológica da Expressão Gênica , Humanos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Gravidez , Progesterona/metabolismo , Trofoblastos/efeitos dos fármacos , Trofoblastos/metabolismo
5.
Ecotoxicol Environ Saf ; 207: 111520, 2021 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-33254395

RESUMO

Methylmercury (MeHg) exposure during pregnancy can lead to adverse outcomes, including miscarriage and intrauterine growth retardation. In this study, MeHg cytotoxicity and its mechanisms in HTR-8/SVneo cells were investigated. MeHg inhibited HTR-8/SVneo cell viability and severely disrupted the cellular submicrostructure, showing a time-dose effect relationship. After MeHg treatment, the reactive oxygen species levels, malondialdehyde content, and superoxide dismutase (SOD) and catalase activities in the HTR-8/SVneo cells increased significantly with increased MeHg concentration (P<0.05). Similarly, MeHg also induced HTR-8/SVneo cell apoptosis in a dose-dependent manner. The proportion of cells in G1 phase decreased with increasing MeHg concentration, while that in the S and G2/M phases gradually increased. Moreover, cell migration and invasion capacities gradually decreased with increasing MeHg concentration, showing a significant difference between the MeHg-treated and control groups. Genes related to oxidative stress (HSPA6, HSPA1A, Nrf2, SOD1, HO-1, NQO1, OSGIN1, and gPX1), cell cycle (P21 and CDC25A), apoptosis (CYCS and AIFM2), and migration and invasion (CXCL8, CXCL3, CLU, IL24, COL3A1, MAPT, and ITGA7) were differentially expressed in the MeHg-treated group, indicating MeHg toxicity and mechanism of action. This study will provide insights into the prevention and treatment of pregnancy-related diseases caused by MeHg.


Assuntos
Poluentes Ambientais/toxicidade , Compostos de Metilmercúrio/toxicidade , Apoptose/efeitos dos fármacos , Morte Celular , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular , Feminino , Humanos , Integrinas , Interleucina-8 , Fator 2 Relacionado a NF-E2 , Estresse Oxidativo/efeitos dos fármacos , Gravidez , Espécies Reativas de Oxigênio/metabolismo , Testes de Toxicidade , Trofoblastos/citologia , Trofoblastos/efeitos dos fármacos , Trofoblastos/metabolismo
6.
PLoS One ; 15(12): e0244684, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33378412

RESUMO

INTRODUCTION: Preeclampsia therapy has not been established, except for the termination of pregnancy. The aim of this study was to identify a potential therapeutic agent from traditional Japanese medicine (Kampo) using the drug repositioning method. MATERIALS AND METHODS: We screened a library of 74 Kampo to identify potential drugs for the treatment of preeclampsia. We investigated the angiogenic effects of these drugs using human umbilical vein endothelial cells (HUVECs). Enzyme-linked immunosorbent assays were performed to measure the levels of placental growth factor (PlGF) in conditioned media treated with 100 µg/mL of each drug. We assessed whether the screened drugs affected cell viability. We performed tube formation assays to evaluate the angiogenic effects of PlGF-inducing drugs. PlGF was measured after administering 10, 50, 100, and 200 µg/mL of the candidate drug in the dose correlation experiment, and at 1, 2, 3, 6, 12, and 24 h in the time course experiment. We also performed tube formation assays with the candidate drug and 100 ng/mL of soluble fms-like tyrosine kinase 1 (sFlt1). PlGF production by the candidate drug was measured in trophoblastic cells (BeWo and HTR-8/SVneo). The Mann-Whitney U test or one-way analyses of variance followed by the Newman-Keuls post-hoc test were performed. P-values < 0.05 were considered significant. RESULTS: Of the 7 drugs that induced PlGF, Tokishakuyakusan (TS), Shoseiryuto, and Shofusan did not reduce cell viability. TS significantly facilitated tube formation (P = 0.017). TS administration increased PlGF expression in a dose- and time-dependent manner. TS significantly improved tube formation, which was inhibited by sFlt1 (P = 0.033). TS also increased PlGF production in BeWo (P = 0.001) but not HTR-8/SVneo cells (P = 0.33). CONCLUSIONS: By using the drug repositioning method in the in vitro screening of the Kampo library, we identified that TS may have a therapeutic potential for preeclampsia. Its newly found mechanisms involve the increase in PlGF production, and improvement of the antiangiogenic state.


Assuntos
Reposicionamento de Medicamentos , Medicamentos de Ervas Chinesas/farmacologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Medicina Kampo , Fator de Crescimento Placentário/metabolismo , Pré-Eclâmpsia/tratamento farmacológico , Adulto , Sobrevivência Celular/efeitos dos fármacos , Medicamentos de Ervas Chinesas/uso terapêutico , Feminino , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Gravidez , Trofoblastos/efeitos dos fármacos , Trofoblastos/metabolismo
7.
Environ Toxicol ; 35(12): 1395-1405, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32790152

RESUMO

Per- and polyfluoroalkyl substances (PFAS), a class of environmental contaminants, have been detected in human placenta and cord blood. The mechanisms driving PFAS-induced effects on the placenta and adverse pregnancy outcomes are not well understood. This study investigated the impact of perfluorooctanesulfonic acid (PFOS), perfluorooctanoic acid (PFOA), and a replacement PFAS known as hexafluoropropylene oxide dimer acid (HFPO-DA, tradename GenX) on placental trophoblasts in vitro. Several key factors were addressed. First, PFAS levels in cell culture reagents at baseline were quantified. Second, the role of supplemental media serum in intracellular accumulation of PFAS in a human trophoblast (JEG3) cell line was established. Finally, the impact of PFAS on the expression of 96 genes involved in proper placental function in JEG3 cells was evaluated. The results revealed that serum-free media (SFM) contained no detectable PFAS. In contrast, fetal bovine serum-supplemented media (SSM) contained PFNA, PFUdA, PFTrDA, and 6:2 FTS, but these PFAS were not detected internally in cells. Intracellular accumulation following 24 hr treatments was significantly higher when cultured in SFM compared to SSM for PFOS and PFOA, but not HFPO-DA. Treatment with PFAS was associated with gene expression changes (n = 32) in pathways vital to placental function, including viability, syncytialization, inflammation, transport, and invasion/mesenchymal transition. Among the most robust PFAS-associated changes were those observed in the known apoptosis-related genes, BAD and BAX. These results suggest a complex relationship between PFAS, in vitro culture conditions, and altered expression of key genes necessary for proper placentation.


Assuntos
Ácidos Alcanossulfônicos/toxicidade , Caprilatos/toxicidade , Fluorcarbonetos/toxicidade , Expressão Gênica/efeitos dos fármacos , Placenta/efeitos dos fármacos , Soro/química , Trofoblastos/efeitos dos fármacos , Ácidos Alcanossulfônicos/sangue , Ácidos Alcanossulfônicos/metabolismo , Apoptose/efeitos dos fármacos , Apoptose/genética , Bioacumulação/efeitos dos fármacos , Bioacumulação/genética , Caprilatos/sangue , Caprilatos/metabolismo , Técnicas de Cultura de Células , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Meios de Cultura Livres de Soro , Feminino , Fluorcarbonetos/sangue , Fluorcarbonetos/metabolismo , Humanos , Placenta/metabolismo , Gravidez , RNA Mensageiro/genética , Trofoblastos/metabolismo
8.
Artigo em Chinês | MEDLINE | ID: mdl-32746565

RESUMO

Objective: To investigate the effect of perfluorooctane sulfonate (PFOS) on inflammatory factors in human placental trophoblast (HTR-8/Svneo) cells. Methods: HTR-8/Svneo cells were exposed to different concentrations of PFOS (0, 0.01, 0.1, 1.0 mg/L) for 24 h, and the cell survival rates were measured by CCK8. Secretion levels of interleukin-6 (IL-6) , tumor necrosis factor-alpha (TNF-α) and interleukin-10 (IL-10) were detected by ELISA. The mRNA expressions of IL-6, TNF-α and IL-10 were detected by fluorescence quantitative PCR. One-way ANOVA was used to analyse the expressions of inflammatory factors. Results: Compared with the control group, the survival rates of 0.1 and 1.0 mg/L PFOS groups were significantly decreased (P<0.05) . Compared with the control group, the secretion levels of IL-6 were decreased in the 0.01, 0.1 and 1.0 mg/L PFOS groups (P<0.05) , the concentrations of TNF-α were increased in the 0.01 and 1.0 mg/L PFOS groups (P<0.05) , and the concentrations of IL-10 were increased in the 0.1 and 1.0 mg/L PFOS groups (P<0.05) . Compared with the control group, the expressions of IL-6 mRNA were increased in the 0.1 and 1.0 mg/L PFOS groups (P<0.05) , and the expressions of IL-10 mRNA were decreased in the 0.01 mg/L, 0.1 mg/L and 1.0 mg/L PFOS groups (P<0.05) . Conclusion: PFOS can induce changes in the secretion levels of inflammatory cytokines in HTR-8/Svneo cells, resulting in decreased activity of placental trophoblast cells and abnomal placental function.


Assuntos
Ácidos Alcanossulfônicos/toxicidade , Fluorcarbonetos/toxicidade , Placenta/efeitos dos fármacos , Trofoblastos/efeitos dos fármacos , Feminino , Humanos , Gravidez , Testes de Toxicidade , Fator de Necrose Tumoral alfa/genética
9.
Life Sci ; 256: 117890, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32497634

RESUMO

OBJECTIVE: We aim to investigate whether there is activation of NLRP1 and autophagy in trophoblast oxidative stress model. Resveratrol was taken to clarify its role in oxidative damage of placental trophoblasts. METHODS: H2O2 was added to HTR-8/SVneo cell for 3 h, then the ROS level and apoptosis panel was performed. The levels of IL-1ß, caspase-1, NLRP1, LC3 and Beclin-1 were detected. Resveratrol was added after 8 h, the ROS level and apoptosis rate were detected, the expression of IL-1ß, caspase-1, NLRP1, LC3 and Beclin-1 were detected. RESULTS: 300 µmol/L H2O2 for 3 h is the optimum combination in establishing the oxidative stress injury model (P < 0.01). LDH, ROS and MDA level was increased, the activity of SOD, CAT were declined (P < 0.01). Apoptosis rate increased (P < 0.01). The expression of IL-1ß, caspase-1, NLRP1, LC3 and Beclin-1 protein was higher (P < .01). Resveratrol (50 µmol/L) treatment for 8 h could improve the changes caused by H2O2, increase the survival rate of cells (P < 0.01), reduce the release of LDH, decrease the level of MDA, increase the level of SOD and CAT (P < 0.01). The expression of IL-1ß, caspase-1, NLRP1, LC3 and Beclin-1 protein decreased (P < 0.01). CONCLUSION: Trophoblast oxidative damage model can be established under 300 µmol/L H2O2 for 3 h, the expression of NLRP1and autophagy after H2O2 treatment were detected. Resveratrol reduces apoptotic cells, thus ensuring the normal biological functions of trophoblasts. CAPSULE: H2O2-induced oxidative stress damage model in HTR-8/SVneo cells can be successfully established under 300 µmol/L H2O2 for 3 h, resveratrol alleviates of H2O2-induced damage by its antioxidant and autophagy regulation function.


Assuntos
Autofagia/efeitos dos fármacos , Inflamassomos/metabolismo , Modelos Biológicos , Estresse Oxidativo , Resveratrol/farmacologia , Trofoblastos/patologia , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Caspase 1/metabolismo , Catalase/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Feminino , Corantes Fluorescentes/química , Humanos , Peróxido de Hidrogênio/toxicidade , Interleucina-1beta/metabolismo , L-Lactato Desidrogenase/metabolismo , Malondialdeído/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Gravidez , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/metabolismo , Trofoblastos/efeitos dos fármacos , Trofoblastos/metabolismo
10.
Sci Rep ; 10(1): 6827, 2020 04 22.
Artigo em Inglês | MEDLINE | ID: mdl-32321940

RESUMO

The placenta forms a maternal-fetal junction that supports many physiological functions such as the supply of nutrition and exchange of gases and wastes. Establishing an in vitro culture model of human and non-human primate trophoblast stem/progenitor cells is important for investigating the process of early placental development and trophoblast differentiation. In this study, we have established five trophoblast stem cell (TSC) lines from cynomolgus monkey blastocysts, named macTSC #1-5. Fibroblast growth factor 4 (FGF4) enhanced proliferation of macTSCs, while other exogenous factors were not required to maintain their undifferentiated state. macTSCs showed a trophoblastic gene expression profile and trophoblast-like DNA methylation status and also exhibited differentiation capacity towards invasive trophoblast cells and multinucleated syncytia. In a xenogeneic chimera assay, these stem cells contributed to trophectoderm (TE) development in the chimeric blastocysts. macTSC are the first primate trophoblast cell lines whose proliferation is promoted by FGF4. These cell lines provide a valuable in vitro culture model to analyze the similarities and differences in placental development between human and non-human primates.


Assuntos
Técnicas de Cultura de Células/métodos , Células-Tronco/citologia , Trofoblastos/citologia , Animais , Bucladesina/farmacologia , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Quimera , Cromossomos de Mamíferos/genética , Metilação de DNA/genética , Ectoderma/citologia , Regulação da Expressão Gênica/efeitos dos fármacos , Células Gigantes/citologia , Macaca fascicularis , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , Especificidade da Espécie , Células-Tronco/efeitos dos fármacos , Trofoblastos/efeitos dos fármacos
11.
Bratisl Lek Listy ; 121(3): 225-229, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32115981

RESUMO

AIM: Nicotine at high concentrations induces apoptosis in trophoblastic cells through induction of cell cytotoxicity and Reactive Oxygen Species (ROS). Methamphetamine in low dose has pharmaceutical properties. It seems that this components in low dose can protect the trophoblastic cells from nicotine-induced cell death. METHOD: Trophoblastic (JEG-3) cells grown in DMEM culture medium. MTT assay test detected the cell viability and Lactate Dehydrogenase test measured the cells cytotoxicity. Griess reaction was used for NO production analysis. Cell migration traced by wounding technique. Human Cytokine Array Focused 13-plex was also used for analysis of IL-1α, IL-1ß, IL-6, INFγ, and TNFα pre-inflammatory cytokines. RESULTS: Methamphetamine, in very low dose (pM), increased the cell viability and NO production, and decreased cell cytotoxicity, IL-1α, IL-1ß, IL-6, INFγ, and TNFα pre-inflammatory cytokines of JEG-3 cell which were exposed to high dose of nicotine, respectively. Cell migration was enhanced by low dose of methamphetamine in JEG-3 cells. CONCLUSION: Methamphetamine in very low dose suppressed the JEG-3 cell death induced by high dose of nicotine (Fig. 5, Ref. 48) Keywords: methamphetamine, nicotine, cell death, NO.


Assuntos
Dopaminérgicos , Inflamação , Metanfetamina , Trofoblastos , Linhagem Celular Tumoral , Sobrevivência Celular , Dopaminérgicos/farmacologia , Humanos , Inflamação/tratamento farmacológico , Metanfetamina/farmacologia , Nicotina/toxicidade , Trofoblastos/efeitos dos fármacos
12.
Sci Adv ; 6(10): eaax6346, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32181339

RESUMO

Placental malaria (PM) is associated with severe inflammation leading to abortion, preterm delivery, and intrauterine growth restriction. Innate immunity responses play critical roles, but the mechanisms underlying placental immunopathology are still unclear. Here, we investigated the role of inflammasome activation in PM by scrutinizing human placenta samples from an endemic area and ablating inflammasome components in a PM mouse model. The reduction in birth weight in babies from infected mothers is paralleled by increased placental expression of AIM2 and NLRP3 inflammasomes. Using genetic dissection, we reveal that inflammasome activation pathways are involved in the production and detrimental action of interleukin-1ß (IL-1ß) in the infected placenta. The IL-1R pharmacological antagonist Anakinra improved pregnancy outcomes by restoring fetal growth and reducing resorption in an experimental model. These findings unveil that IL-1ß-mediated signaling is a determinant of PM pathogenesis, suggesting that IL-1R antagonists can improve clinical outcomes of malaria infection in pregnancy.


Assuntos
Inflamassomos/efeitos dos fármacos , Interleucina-1beta/imunologia , Malária Falciparum/imunologia , Malária/imunologia , Plasmodium falciparum/patogenicidade , Complicações Parasitárias na Gravidez/imunologia , Transdução de Sinais/efeitos dos fármacos , Animais , Caspase 1/genética , Caspase 1/imunologia , Linhagem Celular , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/imunologia , Feminino , Regulação da Expressão Gênica , Humanos , Imunidade Inata , Fatores Imunológicos/farmacologia , Inflamassomos/genética , Inflamassomos/imunologia , Interferon gama/genética , Interferon gama/imunologia , Proteína Antagonista do Receptor de Interleucina 1/farmacologia , Interleucina-1beta/antagonistas & inibidores , Interleucina-1beta/genética , Malária/tratamento farmacológico , Malária/genética , Malária/parasitologia , Malária Falciparum/genética , Malária Falciparum/parasitologia , Malária Falciparum/patologia , Camundongos , Camundongos Knockout , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Plasmodium berghei/imunologia , Plasmodium berghei/patogenicidade , Plasmodium falciparum/imunologia , Gravidez , Complicações Parasitárias na Gravidez/genética , Complicações Parasitárias na Gravidez/parasitologia , Complicações Parasitárias na Gravidez/prevenção & controle , Receptores de Interleucina-1/genética , Receptores de Interleucina-1/imunologia , Transdução de Sinais/imunologia , Células THP-1 , Trofoblastos/efeitos dos fármacos , Trofoblastos/imunologia , Trofoblastos/parasitologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia
13.
Toxicol Appl Pharmacol ; 394: 114960, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32201330

RESUMO

During pregnancy, fetal thyroid hormones (THs) are dependent on maternal placental transport and their physiological level is crucial for normal fetal neurodevelopment. Earlier research has shown that Di-(2-ethylhexyl) phthalate (DEHP) disrupts thyroid function and THs homeostasis in pregnant women and fetuses, and affects placental THs transport. However, the underlying mechanisms are poorly understood. The present study, therefore, aimed to systematically investigate the potential mechanisms of DEHP-induced disruption in the placental THs transport using two human placental trophoblastic cells, HTR-8/SVneo cells and JEG-3 cells. While the exposure of DEHP at the doses of 0-400 µM for 24 h did not affect cell viability, we found reduced consumption of T3 and T4 in the culture medium of HTR-8/Svneo cells treated with DEHP at 400 µM. DEHP treatment did not affect T3 uptake and the expression of monocarboxylate transporters 8 (MCT8) and organic anion transporters 1C1 (OATP1C1). However, DEHP significantly inhibited transthyretin (TTR) internalization, down-regulated TTR, deiodinase 2 (DIO2), and thyroid hormone receptors mRNA expression and protein levels, and up-regulated deiodinase 3 (DIO3) protein levels in a dose-dependent manner. These results indicate that DEHP acts on placental trophoblast cells, inhibits its TTR internalization, down-regulates TTR expression and affects the expression of DIO2, DIO3, and thyroid hormone receptor. These may be the mechanisms by which PAEs affects THs transport through placental.


Assuntos
Dietilexilftalato/toxicidade , Placenta/metabolismo , Pré-Albumina/metabolismo , Trofoblastos/metabolismo , Adulto , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Feminino , Humanos , Iodeto Peroxidase/antagonistas & inibidores , Transportadores de Ácidos Monocarboxílicos/antagonistas & inibidores , Transportadores de Ânions Orgânicos/antagonistas & inibidores , Placenta/citologia , Placenta/efeitos dos fármacos , Pré-Albumina/biossíntese , Gravidez , Receptores dos Hormônios Tireóideos/biossíntese , Receptores dos Hormônios Tireóideos/efeitos dos fármacos , Simportadores/antagonistas & inibidores , Hormônios Tireóideos/metabolismo , Tiroxina/metabolismo , Tri-Iodotironina/biossíntese , Trofoblastos/efeitos dos fármacos
14.
Proc Natl Acad Sci U S A ; 117(9): 4642-4652, 2020 03 03.
Artigo em Inglês | MEDLINE | ID: mdl-32071231

RESUMO

Placental trophoblast cells are potentially at risk from circulating endocrine-disrupting chemicals, such as bisphenol A (BPA). To understand how BPA and the reputedly more inert bisphenol S (BPS) affect the placenta, C57BL6J mouse dams were fed 200 µg/kg body weight BPA or BPS daily for 2 wk and then bred. They continued to receive these chemicals until embryonic day 12.5, whereupon placental samples were collected and compared with unexposed controls. BPA and BPS altered the expression of an identical set of 13 genes. Both exposures led to a decrease in the area occupied by spongiotrophoblast relative to trophoblast giant cells (GCs) within the junctional zone, markedly reduced placental serotonin (5-HT) concentrations, and lowered 5-HT GC immunoreactivity. Concentrations of dopamine and 5-hydroxyindoleacetic acid, the main metabolite of serotonin, were increased. GC dopamine immunoreactivity was increased in BPA- and BPS-exposed placentas. A strong positive correlation between 5-HT+ GCs and reductions in spongiotrophoblast to GC area suggests that this neurotransmitter is essential for maintaining cells within the junctional zone. In contrast, a negative correlation existed between dopamine+ GCs and reductions in spongiotrophoblast to GC area ratio. These outcomes lead to the following conclusions. First, BPS exposure causes almost identical placental effects as BPA. Second, a major target of BPA/BPS is either spongiotrophoblast or GCs within the junctional zone. Third, imbalances in neurotransmitter-positive GCs and an observed decrease in docosahexaenoic acid and estradiol, also occurring in response to BPA/BPS exposure, likely affect the placental-brain axis of the developing mouse fetus.


Assuntos
Compostos Benzidrílicos/toxicidade , Encéfalo/efeitos dos fármacos , Disruptores Endócrinos/toxicidade , Fenóis/toxicidade , Sulfonas/toxicidade , Trofoblastos/efeitos dos fármacos , Animais , Dopamina/metabolismo , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Serotonina/metabolismo , Trofoblastos/metabolismo
15.
PLoS One ; 15(2): e0229332, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32092105

RESUMO

The placenta, a tissue that is metabolically active and rich in mitochondria, forms a critical interface between the mother and developing fetus. Oxidative stress within this tissue, derived from the dysregulation of reactive oxygen species (ROS), has been linked to a number of adverse fetal outcomes. While such outcomes have been associated with mitochondrial dysfunction, the causal role of mitochondrial dysfunction and mitochondrially generated ROS in altering the process of placentation remains unclear. In this study, mitochondrial complex I activity was attenuated using 10 nM rotenone to induce cellular oxidative stress by increasing mitochondrial ROS production in the BeWo choriocarcinoma cell line. Increased mitochondrial ROS resulted in a significant decrease in the transcripts which encode for proteins associated with fusion (GCM1, ERVW-1, and ERVFRD-1) resulting in a 5-fold decrease in the percentage of BeWo fusion. This outcome was associated with increased indicators of mitochondrial fragmentation, as determined by decreased expression of MFN2 and OPA1 along with an increase in a marker of mitochondrial fission (DRP1). Importantly, increased mitochondrial ROS also resulted in a 5.0-fold reduction of human placental lactogen (PL) and a 4.4-fold reduction of insulin like growth factor 2 (IGF2) transcripts; hormones which play an important role in regulating fetal growth. The pre-treatment of rotenone-exposed cells with 5 mM N-acetyl cysteine (NAC) resulted in the prevention of these ROS mediated changes in BeWo function and supports a central role for mitochondrial ROS signaling in the maintenance and function of the materno-fetal interface.


Assuntos
Mitocôndrias/metabolismo , Hormônios Placentários/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Trofoblastos/metabolismo , Fusão Celular , Células Cultivadas , Feminino , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Potencial da Membrana Mitocondrial/fisiologia , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Gravidez , Espécies Reativas de Oxigênio/farmacologia , Rotenona/farmacologia , Transdução de Sinais/efeitos dos fármacos , Trofoblastos/efeitos dos fármacos
16.
Clin Sci (Lond) ; 134(5): 459-472, 2020 03 13.
Artigo em Inglês | MEDLINE | ID: mdl-32068238

RESUMO

Antiphospholipid autoantibodies (aPLs), a major maternal risk factor for preeclampsia, are taken into the syncytiotrophoblast where they bind intracellular vesicles and mitochondria. Subsequently, large quantities of extracellular vesicles (EVs) extruded from syncytiotrophoblast into the maternal circulation are altered such that they cause maternal endothelial cell activation. However, the mechanism driving this change is unknown. First trimester placental explants were treated with aPL for 18 h. The EVs were then collected by different centrifugation. The levels of HSP 70, misfolded proteins, caspase 8 activity, and Mixed Lineage Kinase domain-Like (MLKL) were measured in placental explants and EVs. In addition, the levels of TNF-α and CD95 in conditioned medium were also measured. Treating placental explants with aPL caused an increase in levels of HSP 70, misfolded proteins and MLKL in placental explants and EVs. Increased activity of caspase 8 was also seen in placental explants. Higher levels of TNF-α were seen conditioned medium from aPL-treated placental explant cultures. aPLs appear to induce endoplasmic reticulum stress in the syncytiotrophoblast in a manner that involved caspase 8 and TNF-α. To avoid accumulation of the associated misfolded proteins and MLKL, the syncytiotrophoblast exports these potentially dangerous proteins in EVs. It is likely that the dangerous proteins that are loaded into placental EVs in preeclampsia contribute to dysfunction of the maternal cells.


Assuntos
Anticorpos Antifosfolipídeos/farmacologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Vesículas Extracelulares/metabolismo , Placenta/efeitos dos fármacos , Trofoblastos/efeitos dos fármacos , Caspase 8/metabolismo , Feminino , Proteínas de Choque Térmico HSP70/metabolismo , Humanos , Placenta/metabolismo , Pré-Eclâmpsia/metabolismo , Gravidez , Primeiro Trimestre da Gravidez , Proteínas Quinases/metabolismo , Técnicas de Cultura de Tecidos , Trofoblastos/metabolismo , Fator de Necrose Tumoral alfa/metabolismo
17.
Cell Death Dis ; 11(1): 78, 2020 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-32001671

RESUMO

Preeclampsia (PE) remains a leading cause of maternal and neonatal morbidity and mortality. Numerous studies have shown that women with PE develop autoantibody, termed angiotensin II type 1 receptor autoantibody (AT1-AA), and key features of the disease result from it. Emerging evidence has indicated that inflammatory cell necrosis, such as pyroptosis, could lead to autoantigen exposure and stimulate autoantibody production. Caspase-1, the central enzyme of inflammasome and key target of pyroptosis, may play roles in AT1R exposure and AT1-AA production. Exploring endogenous regulator that could inhibit AT1-AA production by targeting pyroptosis will be essential for treating PE. Lipoxin A4 (LXA4), endogenous dual anti-inflammatory and proresolving lipid mediator, may inhibit AT1-AA production via modulating caspase-1. Thus, we explore whether caspase-1 is essential for AT1-AA production and LXA4 inhibits AT1-AA via modulating caspase-1. PE patients and mice developed AT1-AA associated with caspase-1 activation. Caspase-1 deletion leaded to AT1-AA decrease in PE mice. Consistent with these findings, we confirmed caspase-1 activation, trophoblast pyroptosis and AT1R exposure in PE mice and trophoblast model, while caspase-1 deficiency showed decreased trophoblast pyroptosis and AT1R exposure in vitro and in vivo. Interestingly, LXA4 could suppress AT1-AA production via regulating caspase-1 as well as enhancing phagocytosis of dead trophoblasts by macrophages. These results suggest that caspase-1 promotes AT1-AA production via inducing trophoblast pyroptosis and AT1R exposure, while LXA4 suppresses AT1-AA production via modulating caspase-1, supporting caspase-1 serving as a therapeutic target for attenuating AT1-AA and LXA4 protecting patients from AT1-AA and PE.


Assuntos
Autoanticorpos/metabolismo , Caspase 1/metabolismo , Lipoxinas/farmacologia , Pré-Eclâmpsia/imunologia , Piroptose/efeitos dos fármacos , Receptor Tipo 1 de Angiotensina/imunologia , Trofoblastos/efeitos dos fármacos , Antagonistas de Receptores de Angiotensina , Animais , Autoanticorpos/genética , Autoanticorpos/imunologia , Caspase 1/sangue , Caspase 1/deficiência , Caspase 1/genética , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Células Cultivadas , Citocinas/metabolismo , Feminino , Humanos , Lipopolissacarídeos/farmacologia , Lipoxinas/sangue , Lipoxinas/deficiência , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Pré-Eclâmpsia/sangue , Pré-Eclâmpsia/metabolismo , Gravidez , Piroptose/genética , Piroptose/imunologia , RNA Interferente Pequeno , Receptor Tipo 1 de Angiotensina/sangue , Receptor Tipo 1 de Angiotensina/genética , Receptor Tipo 1 de Angiotensina/metabolismo , Baço/crescimento & desenvolvimento , Baço/imunologia , Baço/patologia , Trofoblastos/metabolismo
18.
FASEB J ; 34(2): 3151-3164, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31908038

RESUMO

Extravillous cytotrophoblasts (EVTs) invade into the uterine wall and remodel spiral arteries for proper placentation. Studies from us and others have demonstrated that the transforming growth factor-ß superfamily member bone morphogenetic protein 2 (BMP2) plays important roles in endometrial decidualization and trophoblast cell invasion. However, BMP2 has also been shown to regulate endothelial cell migration and capillary-like tube formation, as has its downstream signaling molecule inhibitor of DNA binding 1 (ID1). Interestingly, insulin-like growth factor binding protein 3 (IGFBP3) also promotes cell migration and angiogenesis in endothelial precursor cell. Moreover, Id1 has a regulatory effect on Igfbp3 expression in rat prostate epithelial cells. However, whether ID1 and IGFBP3 are integrated in BMP2 signaling and involved in the regulation of trophoblast invasive and endovascular differentiation remains unknown. The objective of our study was to examine the effects of BMP2 on ID1 and IGFBP3 expression and their roles in BMP2-regulated human trophoblast invasion and endothelial-like tube formation. Primary and immortalized (HTR8/SVneo) cultures of human trophoblast cells were employed as study models. BMP2 treatment increased ID1 and IGFBP3 mRNA and protein levels in HTR8/SVneo and primary human EVT cells. Intriguingly, ID1 was essential for BMP2-induced IGFBP3 upregulation in both study models, and BMP2-induced trophoblast invasion was attenuated by knockdown of either ID1 or IGFBP3. In addition, BMP2 significantly increased endothelial-like tube formation and knockdown of ID1 and IGFBP3 reduced basal and BMP2-induced tube formation in HTR8/SVneo cells. Similarly, BMP2 increased placenta growth factor (PlGF) production in HTR8/SVneo cells and these effects were attenuated by knockdown of ID1 or IGFBP3. Our results reveal that BMP2 promotes trophoblast cell invasion and endothelial-like tube formation by ID1-mediated IGFBP3 upregulation.


Assuntos
Proteína Morfogenética Óssea 2/farmacologia , Diferenciação Celular , Movimento Celular , Células Endoteliais/metabolismo , Trofoblastos/metabolismo , Linhagem Celular , Células Cultivadas , Células Endoteliais/citologia , Humanos , Proteína 1 Inibidora de Diferenciação/genética , Proteína 1 Inibidora de Diferenciação/metabolismo , Proteína 3 de Ligação a Fator de Crescimento Semelhante à Insulina/genética , Proteína 3 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Fator de Crescimento Placentário/metabolismo , Trofoblastos/citologia , Trofoblastos/efeitos dos fármacos , Trofoblastos/fisiologia , Regulação para Cima
19.
Toxicology ; 431: 152363, 2020 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-31935421

RESUMO

The neonicotinoid (Neo) insecticide family is a relatively new class of pesticides of growing use. There is an increasing concern that human exposure to environmental pollutants in utero may be associated with diseases in adulthood. A functional placenta and trophoblasts are a requisite for a healthy pregnancy. The aim of this study was to investigate whether the Neo Acetamiprid (Ace) and one of its commercial formulations (Ace CF) display toxic features to a human first trimester trophoblast cell line. HTR-8/SVneo cells were cultured in the presence of Ace or Ace CF (0.1-100 µM) for 4 and 24 h, and changes in cell viability, reactive oxygen species, antioxidant system and macromolecule damage levels were evaluated. Ace and Ace CF are cytotoxic for HTR-8/SVneo trophoblasts. Cell viability loss and oxidative imbalance were triggered by Ace and Ace CF treatments. Impact in the antioxidant enzymes catalase, superoxide dismutase and gluthatione S-transferase activities were observed after 24 h exposure to Ace CF. Moreover, Ace CF caused oxidative damage in proteins, lipids and DNA, whereas Ace only damaged proteins. To test oxidative stress as a toxicity mechanism, cells were pre-incubated with the antioxidant N-acetyl-l-cysteine (NAC), prior Neo treatment. NAC protected trophoblasts from cell death and prevented oxidative damage. Results demonstrate that Ace (as active principle or CF) is cytotoxic for human trophoblasts, and oxidative stress is a toxicity mechanism. Ace CF exhibited a more toxic effect than the active principle, in an identical exposure scenario.


Assuntos
Inseticidas/toxicidade , Neonicotinoides/toxicidade , Trofoblastos/efeitos dos fármacos , Acetilcisteína/metabolismo , Adulto , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Dano ao DNA , Composição de Medicamentos , Feminino , Humanos , Estresse Oxidativo , Gravidez , Espécies Reativas de Oxigênio/metabolismo , Receptores Nicotínicos/efeitos dos fármacos
20.
Am J Pathol ; 190(2): 388-399, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31955792

RESUMO

Preterm premature rupture of membranes (PPROM) and thrombin generation by decidual cell-expressed tissue factor often accompany abruptions. Underlying mechanisms remain unclear. We hypothesized that thrombin-induced colony-stimulating factor-2 (CSF-2) in decidual cells triggers paracrine signaling via its receptor (CSF2R) in trophoblasts, promoting fetal membrane weakening and abruption-associated PPROM. Decidua basalis sections from term (n = 10), idiopathic preterm birth (PTB; n = 8), and abruption-complicated pregnancies (n = 8) were immunostained for CSF-2. Real-time quantitative PCR measured CSF2 and CSF2R mRNA levels. Term decidual cell (TDC) monolayers were treated with 10-8 mol/L estradiol ± 10-7 mol/L medroxyprogesterone acetate (MPA) ± 1 IU/mL thrombin pretreatment for 4 hours, washed, and then incubated in control medium with estradiol ± MPA. TDC-derived conditioned media supernatant effects on fetal membrane weakening were analyzed. Immunostaining localized CSF-2 primarily to decidual cell cytoplasm and cytotrophoblast cell membranes. CSF-2 immunoreactivity was higher in abruption-complicated or idiopathic PTB specimens versus normal term specimens (P < 0.001). CSF2 mRNA was higher in TDCs versus cytotrophoblasts (P < 0.05), whereas CSF2R mRNA was 1.3 × 104-fold higher in cytotrophoblasts versus TDCs (P < 0.001). Thrombin enhanced CSF-2 secretion in TDC cultures fourfold (P < 0.05); MPA reduced this effect. Thrombin-pretreated TDC-derived conditioned media supernatant weakened fetal membranes (P < 0.05), which MPA inhibited. TDC-derived CSF-2, acting via trophoblast-expressed CSFR2, contributes to thrombin-induced fetal membrane weakening, eliciting abruption-related PPROM and PTB.


Assuntos
Descolamento Prematuro da Placenta/fisiopatologia , Decídua/patologia , Membranas Extraembrionárias/patologia , Ruptura Prematura de Membranas Fetais/patologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Nascimento Prematuro/etiologia , Trombina/farmacologia , Decídua/efeitos dos fármacos , Decídua/metabolismo , Membranas Extraembrionárias/metabolismo , Feminino , Ruptura Prematura de Membranas Fetais/etiologia , Ruptura Prematura de Membranas Fetais/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Humanos , Gravidez , Nascimento Prematuro/metabolismo , Nascimento Prematuro/patologia , Transdução de Sinais , Trofoblastos/efeitos dos fármacos , Trofoblastos/metabolismo , Trofoblastos/patologia
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