RESUMO
PROBLEM: Decidual macrophages (dMφ ) play an important role in the formation of maternal-fetal immune tolerance. However, factors that influence the immune status of dMφ and the related potential mechanisms have not been elucidated to date. METHOD OF STUDY: The gene transcription in dMφ , decidual stromal cells (DSCs), extravillous trophoblasts (EVTs), and peripheral monocytes (pMo) from human samples were measured using real-time polymerase chain reaction (PCR). Monocyte-DSC co-culture was established to explore whether DSCs influenced dMφ polarization via C-C motif ligand 2 (CCL2)-C-C chemokine receptor (CCR2) binding using flow cytometry. In vivo, changes in dMφ percentage and M1 and M2 marker expression after treatment with CCR2 or Janus kinase 2 (JAK2) inhibitor were detected with flow cytometry. Embryo resorption percentages in the above groups were also analyzed. RESULTS: We found that dMφ were an M1/M2 mixed status at the maternal-fetal interface during early pregnancy. CCL2 influenced the immune status of dMφ in an autocrine and paracrine manner. As a downstream regulator of CCR2 and triggers the Stat3 pathway, JAK2 was found to be essential for dMφ homeostasis in vivo. JAK2 inhibitor decreased the dMφ proportion and attenuated Ki67, CD36, CD86, CD206, TNF, and IL-10 expression in dMφ at E8.5 d. Moreover, CCR2-JAK2 pathway inhibition decreased the width of the placental labyrinth layer, further influencing the pregnancy outcome. CONCLUSION: The M1/M2 mixed immune status of dMφ was regulated by DSCs via CCR2, and the CCL2/CCR2/JAK2 pathway was essential for the immune status of dMφ and the outcome of early pregnancy.
Assuntos
Quimiocina CCL2/metabolismo , Decídua/enzimologia , Histocompatibilidade Materno-Fetal , Tolerância Imunológica , Janus Quinase 2/metabolismo , Macrófagos/enzimologia , Receptores CCR2/metabolismo , Células Estromais/enzimologia , Adulto , Animais , Células Cultivadas , Técnicas de Cocultura , Decídua/efeitos dos fármacos , Decídua/imunologia , Perda do Embrião/enzimologia , Perda do Embrião/imunologia , Feminino , Humanos , Janus Quinase 2/antagonistas & inibidores , Inibidores de Janus Quinases/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos Endogâmicos C57BL , Fenótipo , Gravidez , Resultado da Gravidez , Receptores CCR2/antagonistas & inibidores , Transdução de Sinais , Células Estromais/efeitos dos fármacos , Células Estromais/imunologia , Adulto JovemRESUMO
Existing evidence suggests that adverse pregnancy outcomes are closely related to dietary factors. Folate plays an important role in neural tube formation and fetal growth, folate deficiency is a major risk factor of birth defects. Our early studies showed that folate deficiency could impair enddecidualization, however, the mechanism is still unclear. Dysfunctional autophagy is associated with many diseases. Here, we aimed to evaluate the adverse effect of folate deficiency on endometrial decidualization, with a particular focus on endometrial cell autophagy. Mice were fed with no folate diet in vivo and the mouse endometrial stromal cell was cultured in a folate-free medium in vitro. The decrease of the number of endometrial autophagosomes and the protein expressions of autophagy in the folate-deficient group indicated that autophagosome formation, autophagosome-lysosome fusion, and lysosomal degradation were inhibited. Autophagic flux examination using mCherry-GFP-LC3 transfection showed that the fusion of autophagosomes with lysosomes was inhibited by folate deficiency. Autophagy inducer rapamycin could reverse the impairment of folate deficiency on endometrial decidualization. Moreover, folate deficiency could reduce autophagy by disrupting AMPK/mTOR signaling, resulting in aberrant endometrial decidualization and adverse pregnancy outcomes. Further co-immunoprecipitation examination showed that decidual marker protein Hoxa10 could interact with autophagic marker protein Cathepsin L, and the interaction was notably reduced by folate deficiency. In conclusion, AMPK/mTOR downregulated autophagy was essential for aberrant endometrial decidualization in early pregnant mice, which could result in adverse pregnancy outcomes. This provided some new clues for understanding the causal mechanisms of birth defects induced by folate deficiency.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Autofagia , Decídua/enzimologia , Deficiência de Ácido Fólico/enzimologia , Ácido Fólico/metabolismo , Células Estromais/enzimologia , Serina-Treonina Quinases TOR/metabolismo , Animais , Autofagossomos/enzimologia , Autofagossomos/ultraestrutura , Células Cultivadas , Decídua/ultraestrutura , Modelos Animais de Doenças , Feminino , Deficiência de Ácido Fólico/genética , Deficiência de Ácido Fólico/patologia , Lisossomos/enzimologia , Lisossomos/ultraestrutura , Camundongos , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Gravidez , Transdução de Sinais , Células Estromais/ultraestruturaRESUMO
Pre-eclampsia (PE) is a pregnancy-related disorder that occurs after 20 weeks of gestation. It seriously affects the health of maternity and the fetus. However, the pathogenesis of PE is still unknown. Decidualization deficiency is considered a contributing factor to the development of PE. CTP synthetase (CTPS) which is the rate-limiting enzyme in the CTP de novo biosynthesis, is essential for nucleic acid synthesis and cellular energy metabolism, and often appears as cytoophidium in many cell types. Here, we found that the expression of CTPS was significantly downregulated in decidual tissues of patients with severe PE compared with healthy pregnant women. During in vitro decidualization, changes in CTPS were accompanied by opposite fluctuation of the AMPK signaling pathway. Moreover, the downregulation of CTPS by glutamine analogs or CTPS small interfering RNA inhibited the decidualization process and the AMPK signaling pathway. Investigating the underlying mechanism of action by co-immunoprecipitation coupled with mass spectrometry showed that CTPS interacted with ATP synthase (ATPS) and maintained the content of ATP on Day 3 of decidualization. However, when combined with mitochondrial stress protein STRESS-70 instead of ATPS, the concentration of ATP on Day 6 of induction was reduced. Corresponding to this, CTPS was mainly distributes in the cytoplasm on Day 3 of induction, while it appeared both in the cytoplasm and the nucleus on Day 6 in decidualized cells, which was similar to that in cells before induction. In summary, we believe that CTPS plays an important role in decidualization by participating in energy metabolism. Abnormal expression of CTPS in decidualization would lead to abnormal decidualization and consequently result in the occurrence of PE.
Assuntos
Carbono-Nitrogênio Ligases/metabolismo , Decídua/enzimologia , Regulação para Baixo , Metabolismo Energético , Pré-Eclâmpsia/enzimologia , Adenilato Quinase/metabolismo , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Diazo-Oxo-Norleucina/farmacologia , Regulação para Baixo/efeitos dos fármacos , Endométrio/patologia , Metabolismo Energético/efeitos dos fármacos , Feminino , Inativação Gênica/efeitos dos fármacos , Células-Tronco Embrionárias Humanas/efeitos dos fármacos , Células-Tronco Embrionárias Humanas/metabolismo , Humanos , ATPases Mitocondriais Próton-Translocadoras/metabolismo , Gravidez , Ligação Proteica/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismoRESUMO
The aim of this study is to investigate the effect of the IDO (indoleamine 2,3-dioxygenase) gene on pregnancy outcome in mice with recurrent pregnancy loss (RPL) and its mechanism of action in the maternal-fetal interface. An RPL model was established via natural mating of female CBA/J mice with male DBA/2 mice; thereafter, the female mice were randomly divided into groups treated with LV-EGFP (enhanced green fluorescent protein)-IDO (lentivirus vector carrying IDO-EGFP gene), LV-EGFP (negative control lentivirus vector), or phosphate-buffered saline (control). The mice were sacrificed at 13.5 days of pregnancy, and the embryo absorption rate was determined. Peripheral blood regulatory T cells (Tregs) from the pregnant mice were detected using flow cytometry. Placental and decidual tissue IDO expression was detected using immunofluorescence and Western blotting. Inflammatory cell infiltration of the placental and decidual tissue was observed using hematoxylin-eosin (HE) staining. The LV-EGFP-IDO group had a significantly lower embryo absorption rate than the LV-EGFP and control groups (P = 0.0006 and P = 0.0049, respectively) and significantly more Tregs than the LV-EGFP and control groups (P = 0.0151 and P = 0.0392, respectively). Placental and decidual IDO protein levels correlated positively with peripheral blood Treg expression levels. The LV-EGFP-IDO group had significantly higher placental and decidual IDO protein levels than the LV-EGFP and control groups (P < 0.005), and it had significantly less inflammatory cell infiltration than the LV-EGFP and control groups. The IDO gene may reduce the embryo absorption rate in an RPL mouse model, possibly improving pregnancy outcome by upregulating Tregs and reducing the inflammatory response.
Assuntos
Aborto Habitual/enzimologia , Decídua/enzimologia , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Placenta/enzimologia , Aborto Habitual/genética , Aborto Habitual/imunologia , Animais , Decídua/imunologia , Modelos Animais de Doenças , Feminino , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Mediadores da Inflamação/metabolismo , Masculino , Camundongos Endogâmicos CBA , Camundongos Endogâmicos DBA , Placenta/imunologia , Gravidez , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismoRESUMO
Indoleamine 2,3-dioxygenase 1 (IDO1) and tryptophan 2,3-dioxygenase (TDO) catalyze the rate-limiting step of tryptophan catabolism along the kynurenine pathway, which has important immuno suppressive properties, particularly in tumor cells and dendritic cells. The prominent expression of IDO1 in the placenta also suggested a role in preventing immune rejection of fetal tissues, and pharmacological inhibition of IDO1 induced abortion of allogeneic fetuses in mice. However, this was later challenged by the lack of rejection of allogeneic fetuses in IDO1-KO mice, suggesting that other mechanisms may compensate for IDO1 deficiency. Here we investigated whether TDO could contribute to feto-maternal tolerance and compensate for IDO1 deficiency in IDO1-KO mice. Expression of TDO mRNA was previously detected in placental tissues. We developed a new chimeric rabbit anti-TDO antibody to confirm TDO expression at the protein level and identify the positive cell type by immunohistochemistry in murine placenta. We observed massive TDO expression in decidual stromal cells, starting at day E3.5, peaking at day E6.5 then declining rapidly while remaining detectable until gestation end. IDO1 was also induced in decidual stromal cells, but only at a later stage of gestation when TDO expression declined. To determine whether TDO contributed to feto-maternal tolerance, we mated TDO-KO and double IDO1-TDO-KO females with allogeneic males. However, we did not observe reduced fertility. These results suggest that, despite its expression in decidual stromal cells, TDO is not a dominant mechanism of feto-maternal tolerance able to compensate for the absence of IDO1. Redundant additional mechanisms of immunosuppression likely take over in these KO mice. The massive expression of TDO during decidualization might suggest a role of TDO in angiogenesis or vessel tonicity, as previously described for IDO1.
Assuntos
Decídua/enzimologia , Tolerância Imunológica , Troca Materno-Fetal/imunologia , Células Estromais/enzimologia , Triptofano Oxigenase/metabolismo , Animais , Decídua/citologia , Decídua/imunologia , Feminino , Fertilidade/imunologia , Idade Gestacional , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Cinurenina/sangue , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Gravidez , Células Estromais/imunologia , Triptofano Oxigenase/genéticaRESUMO
INTRODUCTION: Vitamin D catabolizing enzymes, along with vitamin D receptor (VDR) and vitamin D binding protein (DBP) are expressed in the decidua and placenta during pregnancy and capable of synthesizing active vitamin D. Vitamin D plays roles in immunoregulation and trophoblast invasion, key features of a successful pregnancy. Epidemiological data suggests that vitamin D deficiency is associated with both spontaneous and recurrent miscarriage but few studies have investigated the expression of the key vitamin D catabolizing enzymes in miscarriage. METHODS: Placenta and decidua were collected after termination of apparently normal pregnancies (controls, n = 22) or spontaneous miscarriage (n = 20). Immunohistochemical staining, Western Blot and qRT-PCR were performed for CYP27B1, CYP24A1, CYP2R1, VDR and DBP (not qRT-PCR). HTR-8/SVneo cells were cultured in CoCL2 (hypoxic mimetic) or LPS (bacterial infection mimetic) for 24 h, RNA extracted and qRT-PCR performed for CYP27B1, CYP24A1, CYP2R1 and VDR. RESULTS: In spontaneous miscarriage, placental and decidual expression of CYP27B1 was reduced, while expression of CYP24A1, VDR and DBP was increased. When a trophoblast cell line was treated with CoCL2 expression of CYP27B1 was increased and CYP24A1 was reduced, while LPS induced expression of VDR. DISCUSSION: This is the first report of altered utero-placental vitamin D catabolism in spontaneous miscarriage. It is becoming accepted that women who are undergoing assisted reproductive technologies should ensure they have sufficient vitamin D levels prior to pregnancy, these data support that all women should ensure they are vitamin D replete before planning to get pregnant.
Assuntos
Aborto Espontâneo/etiologia , Sistema Enzimático do Citocromo P-450/metabolismo , Decídua/enzimologia , Receptores de Calcitriol/metabolismo , Proteína de Ligação a Vitamina D/metabolismo , Vitamina D/metabolismo , 25-Hidroxivitamina D3 1-alfa-Hidroxilase/metabolismo , Aborto Espontâneo/sangue , Aborto Espontâneo/enzimologia , Estudos de Casos e Controles , Colestanotriol 26-Mono-Oxigenase/metabolismo , Família 2 do Citocromo P450/metabolismo , Feminino , Humanos , Gravidez , Vitamina D3 24-Hidroxilase/metabolismoRESUMO
Decidualization is required for the successful establishment of pregnancy in rodents and primates. Fatty acid desaturase 3 (Fads3) belongs to the fatty acid desaturase family, which is a crucial enzyme for highly unsaturated fatty acid biosynthesis. However, the expression, regulation and function of Fads3 during early pregnancy in mice are still unknown. In this study, we examined Fads3 expression, regulation and function during mouse decidualization. The expression of Fads3 is detected in the subluminal stromal cells at implantation site on day 5 of pregnancy, but not at inter-implantation site and in day 5 pseudopregnant uteri. Compared to delayed implantation, Fads3 is strongly expressed after delayed implantation is activated by estrogen treatment. From days 6 to 8, Fads3 mRNA signals are significantly detected in the decidua. In ovariectomized mice, estrogen significantly stimulates Fads3 expression. However, estrogen has no effect on Fads3 expression in ovariectomized ERα-deficient mice, suggesting that estrogen regulation on Fads3 expression is ERα dependent. When ovariectomized mice were treated with progesterone, Fads3 expression is significantly increased by progesterone. Progesterone stimulation on Fads3 expression is also detected in cultured stromal cells, which is abrogated by RU486 treatment. These data indicate that progesterone upregulation on Fads3 expression is progesterone receptor-dependent. Fads3 knockdown by siRNA reduces in vitro decidualization of mouse stromal cells. Taken together, Fads3 may play an important role during mouse decidualization.
Assuntos
Decídua/enzimologia , Estrogênios/metabolismo , Ácidos Graxos Dessaturases/metabolismo , Gravidez/metabolismo , Progesterona/metabolismo , Animais , Implantação do Embrião , Feminino , CamundongosRESUMO
Adenosine monophosphate-activated protein kinase (AMPK) is a highly conserved heterotrimeric complex that acts as an intracellular energy sensor. Based on recent observations of AMPK expression in all structures of the female reproductive system, we hypothesized that AMPK is functionally required for maintaining fertility in the female. This hypothesis was tested by conditionally ablating the two catalytic alpha subunits of AMPK, Prkaa1 and Prkaa2, using Pgr-cre mice. After confirming the presence of PRKAA1, PRKAA2 and the active phospho-PRKAA1/2 in the gravid uterus by immunohistochemistry, control (Prkaa1/2 fl/fl ) and double conditional knockout mice (Prkaa1/2 d/d ) were placed into a six-month breeding trial. While the first litter size was comparable between Prkaa1/2 fl/fl and Prkaa1/2 d/d female mice (P = 0.8619), the size of all subsequent litters was dramatically reduced in Prkaa1/2 d/d female mice (P = 0.0015). All Prkaa1/2 d/d female mice experienced premature reproductive senescence or dystocia by the fourth parity. This phenotype manifested despite no difference in estrous cycle length, ovarian histology in young and old nulliparous or multiparous animals, mid-gestation serum progesterone levels or uterine expression of Esr1 or Pgr between Prkaa1/2 fl/fl and Prkaa1/2 d/d female mice suggesting that the hypothalamic-pituitary-ovary axis remained unaffected by PRKAA1/2 deficiency. However, an evaluation of uterine histology from multiparous animals identified extensive endometrial fibrosis and disorganized stromal-glandular architecture indicative of endometritis, a condition that causes subfertility or infertility in most mammals. Interestingly, Prkaa1/2 d/d female mice failed to undergo artificial decidualization. Collectively, these findings suggest that AMPK plays an essential role in endometrial regeneration following parturition and tissue remodeling that accompanies decidualization.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Endometrite/enzimologia , Endométrio/enzimologia , Fertilidade , Regeneração , Reprodução , Proteínas Quinases Ativadas por AMP/deficiência , Proteínas Quinases Ativadas por AMP/genética , Animais , Decídua/enzimologia , Decídua/patologia , Decídua/fisiopatologia , Distocia/enzimologia , Distocia/genética , Distocia/fisiopatologia , Endometrite/genética , Endometrite/patologia , Endometrite/fisiopatologia , Endométrio/patologia , Endométrio/fisiopatologia , Feminino , Fibrose , Tamanho da Ninhada de Vivíparos , Camundongos Knockout , Paridade , GravidezRESUMO
OBJECTIVE: To investigate whether bisphenol A (BPA) exposure is associated with uterine decidualization and embryo implantation failure in mice. DESIGN: Experimental animal study and in vitro study. SETTING: University-based infertility center. ANIMAL(S): ICR mice. INTERVENTION(S): Mice treated with different doses of BPA; Ishikawa cells cultured in medium of different concentrations of BPA. MAIN OUTCOME MEASURE(S): Embryo implantation sites, uterine weight, quantitative real-time reverse transcriptase-polymerase chain reaction, Western blot analysis, hematoxylin and eosin staining, and immunohistochemical, cell proliferation, and statistical analyses. RESULT(S): In the experiment of mouse model, administration of 1-100 µg/kg/day of BPA by gavage led to reduction of the number of embryo implantation sites in a dose-dependent manner; 100 µg/kg/day of BPA statistically significantly reduced the number of implantation sites compared with the control group. The uterine weight change (the wet weight of the decidualized uterine horn divided by the wet weight of the undecidualized uterine horn of the mouse) in groups exposed to BPA (100-10,000 µg/kg/day) were statistically significantly lower compared with the control group. Immunohistochemical analysis demonstrated that administration of 100, 1,000, or 10,000 µg/kg/day of BPA by gavage statistically significantly down-regulated the expression of epithelial Na+ channel α-subunit (ENaCα) in the luminal epithelial cells and desmin in decidual cells of the oil-induced decidualized uterine horns. Administration of 100 µg/kg/day BPA on embryo days 0.5-3.5 by gavage statistically significantly decreased the level of uterine serum and glucocorticoid-regulated kinase 1 (SGK1) protein expression on embryo days 4 and 6. After treatment with 0.001, 0.01, 0.1, or 1.0 µg/mL of BPA for 48 hours, the SGK1, ENaCα, and phospho-SGK1 protein expression of Ishikawa cells was down-regulated, and the effect of BPA on SGK1 could be abrogated by fulvestrant. CONCLUSION(S): Our study provides the first indication that BPA exposure at levels as low as 100 µg/kg/day can impair embryo implantation in mice and BPA can affect decidualization of the uterus in mouse model. Our results suggest that BPA can down-regulate SGK1 and ENaCα protein expression through estrogen receptors in Ishikawa cells.
Assuntos
Compostos Benzidrílicos/toxicidade , Decídua/efeitos dos fármacos , Implantação do Embrião/efeitos dos fármacos , Disruptores Endócrinos/toxicidade , Poluentes Ambientais/toxicidade , Canais Epiteliais de Sódio/metabolismo , Proteínas Imediatamente Precoces/metabolismo , Infertilidade Feminina/induzido quimicamente , Fenóis/toxicidade , Proteínas Serina-Treonina Quinases/metabolismo , Receptores de Estrogênio/efeitos dos fármacos , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Decídua/enzimologia , Decídua/patologia , Decídua/fisiopatologia , Feminino , Infertilidade Feminina/enzimologia , Infertilidade Feminina/patologia , Infertilidade Feminina/fisiopatologia , Camundongos Endogâmicos ICR , Neoplasias Ovarianas/enzimologia , Neoplasias Ovarianas/patologia , Fosforilação , Gravidez , Receptores de Estrogênio/metabolismo , Medição de Risco , Transdução de Sinais/efeitos dos fármacosRESUMO
Matrix metalloproteinase-1 (MMP-1) and neutrophil elastase are proteolytic enzymes involved in tissue remodeling, but a role for them as uterotonic agents has not been considered. However, both these proteases activate protease-activated receptor 1 (PAR-1) that mediates thrombin-induced contractions. Matrix metalloproteinase-1 and elastase are products of neutrophils that infiltrate intrauterine tissues at the time of labor, so we tested the hypothesis that these proteases might be novel uterotonic agents acting via PAR-1. Decidual tissue was collected from fetal membranes of term not-in-labor (TNL), term labor (TL), and preterm labor (PTL) women and analyzed for gene and protein expression of MMP-1 and neutrophil elastase. Contractile effects of MMP-1 and elastase were tested with uterine strips of day 19 and 20 gestation rats. Expression of MMP-1 and neutrophil elastase was increased in TL and PTL as compared to TNL. Expression of both the pro- and active enzymes of MMP-1 increased progressively from TNL to TL to PTL. Tumor necrosis factor-α, a neutrophil product, increased MMP-1 in decidual and myometrial cells. Both MMP-1 and elastase stimulated strong contractions of myometrial strips, which were prevented by inhibition of PAR-1 and inhibition of inositol trisphosphate receptor or calcium channel blockade. Indomethacin did not prevent protease-induced contractions. These data suggest that MMP-1 and neutrophil elastase may be important but heretofore unrecognized players in stimulating uterine contractions at the time of labor, and they may explain why indomethacin delays, but does not prevent, PTL because indomethacin inhibits the prostaglandin component but not the protease component of labor.
Assuntos
Decídua/enzimologia , Metaloproteinase 1 da Matriz/metabolismo , Elastase Pancreática/metabolismo , Receptor PAR-1/metabolismo , Contração Uterina , Adulto , Feminino , Expressão Gênica , Humanos , Trabalho de Parto , Neutrófilos/metabolismo , Trabalho de Parto Prematuro/metabolismo , Gravidez , Adulto JovemRESUMO
The role of placental serotonin has been an active topic of research notably because of its crucial role in brain development. However, which cell types synthesize serotonin in human placenta remains unknown. Moreover, it is not known if the two tryptophan hydroxylase isoforms (TPH1 and TPH2), the rate-limiting enzymes in serotonin biosynthesis, are expressed in placenta. Human placentas were obtained in first trimester or at term, and trophoblast cells were isolated and purified using a magnetic cell sorter and placed in primary culture. The tissue sublocalization of each TPH was determined by immunohistochemistry. TPH expression in primary villous trophoblasts was determined by PCR and immunoblotting, and serotonin secretion by LC-MS/MS. Villous cytotrophoblasts, syncytiotrophoblast, fetal capillaries, extravillous cytotrophoblasts, and decidual cells co-expressed TPH1 and TPH2. Moreover, mRNA and protein levels of both TPHs were detected in human primary trophoblast as well as in mouse placental tissues. Finally, human trophoblast cells were shown to produce serotonin de novo. This study demonstrates that both TPH1 and TPH2 are expressed in human and mouse placenta throughout pregnancy and helps to better understand the placental serotonin system, which is crucial for healthy pregnancy and fetal development. It is therefore important to further understand regulation of the placental serotonin system and how its disruption during pregnancy may impact the developing fetus and subsequent child programming.
Assuntos
Decídua/enzimologia , Regulação Enzimológica da Expressão Gênica/fisiologia , Proteínas da Gravidez/biossíntese , Trofoblastos/enzimologia , Triptofano Hidroxilase/biossíntese , Animais , Decídua/citologia , Feminino , Humanos , Isoenzimas/biossíntese , Camundongos , Gravidez , Serotonina/biossíntese , Trofoblastos/citologiaRESUMO
Aims: The aim of this study was to examine the effect of alcohol on the decidualization of human endometrial stromal cells during early pregnancy. Methods: During in vitro decidualization, human endometrial stromal cells were treated with alcohol, 4-methylpyrazole hydrochloride (FPZ), the inhibitor of alcohol dehydrogenases (ADHs), and tetraethylthiuram disulfide (DSF), the inhibitor of acetaldehyde dehydrogenases (ALDHs), respectively. Cell viability and decidualization were examined. Apoptosis and proliferation were also evaluated. Results: The findings showed that ADHs and ALDHs were up-regulated during decidualization. After alcohol treatment, the cell viability of decidual stromal cells was significantly higher than control, which was abrogated by FPZ or DSF. When cells were treated with alcohol, proliferation-related signal pathways were up-regulated in decidualized cells. Additionally, FOXO1 transcriptionally up-regulates ADH1B. Conclusion: Our study provided an evidence that highly expressed ADHs and ALDHs endow decidual stromal cells an ability to alleviate the harm from alcohol.
Assuntos
Álcool Desidrogenase/biossíntese , Aldeído Oxirredutases/biossíntese , Decídua/efeitos dos fármacos , Decídua/enzimologia , Etanol/toxicidade , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Linhagem Celular Transformada , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Decídua/citologia , Feminino , Humanos , Células Estromais/citologia , Células Estromais/efeitos dos fármacos , Células Estromais/enzimologiaRESUMO
High resistance to oxidative stress is a common feature of mesenchymal stem/stromal cells (MSC) and is associated with higher cell survival and ability to respond to oxidative damage. Aldehyde dehydrogenase (ALDH) activity is a candidate "universal" marker for stem cells. ALDH expression was significantly lower in decidual MSC (DMSC) isolated from preeclamptic (PE) patients. ALDH gene knockdown by siRNA transfection was performed to create a cell culture model of the reduced ALDH expression detected in PE-DMSC. We showed that ALDH activity in DMSC is associated with resistance to hydrogen peroxide (H2O2)-induced toxicity. Our data provide evidence that ALDH expression in DMSC is required for cellular resistance to oxidative stress. Furthermore, candidate ALDH activators were screened and two of the compounds were effective in upregulating ALDH expression. This study provides a proof-of-principle that the restoration of ALDH activity in diseased MSC is a rational basis for a therapeutic strategy to improve MSC resistance to cytotoxic damage.
Assuntos
Aldeído Desidrogenase/metabolismo , Decídua/enzimologia , Células-Tronco Mesenquimais/metabolismo , Pré-Eclâmpsia/enzimologia , Adulto , Aldeído Desidrogenase/genética , Proliferação de Células , Células Cultivadas , Citoproteção , Decídua/patologia , Feminino , Técnicas de Silenciamento de Genes , Humanos , Peróxido de Hidrogênio/metabolismo , Masculino , Estresse Oxidativo , Pré-Eclâmpsia/patologia , Gravidez , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/metabolismo , Reprodutibilidade dos TestesRESUMO
OBJECTIVE: To study the impact of integrin-linked kinase (ILK) in endometrial stromal cells (ESCs) during decidualization. DESIGN: Laboratory study with the use of human endometrium. SETTING: University hospital. PATIENT(S): Fertile reproductive-age women who had not received hormonal treatment for 3 months before tissue collection. INTERVENTION(S): Endometrium tissue collection, in vitro decidualization of isolated ESCs, and small interfering (si) RNA transfection. MAIN OUTCOME MEASURE(S): Immunohistochemistry, ELISA, Western blot analysis, methylthiazolyl tetrazolium assay, and immunofluorescence staining. RESULT(S): In vivo expression of ILK is significantly increased in distended-fusiform stromal cells of late secretory endometrium and in cobblestone-shaped decidual cells of early pregnancy. During in vitro decidualization for up to 8 days, confluent cultures of isolated ESCs consistently displayed increased ILK expression and morphologic transformation from fibroblast-like to polygonal cells. Subsequent ILK knockdown by siRNA transfection reversed this transformation, accompanied by decreased phosphorylation of glycogen synthase kinase (GSK) 3ß and decreased viable cell numbers. Immunofluorescence staining of the decidualized ESCs demonstrated linkage of increased levels of ILK at the tips of the fan-shaped organization of actin stress fibers located in the submembranous area, which expanded the decidual cells into a typical polygonal appearance. Knock-down of ILK abrogated the polymerization and organization of actin fibers, which reverted the cells to their undecidualized morphology. CONCLUSION(S): During human endometrial decidualization, ILK is essential for morphologic transformation of ESCs through organization of the actin cytoskeleton; it may also function through subsequent GSK3ß signaling, which requires further studies.
Assuntos
Movimento Celular , Forma Celular , Decídua/enzimologia , Implantação do Embrião , Proteínas Serina-Treonina Quinases/metabolismo , Células Estromais/enzimologia , Actinas/metabolismo , Sobrevivência Celular , Células Cultivadas , Decídua/patologia , Feminino , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Ciclo Menstrual/metabolismo , Fosforilação , Gravidez , Proteínas Serina-Treonina Quinases/genética , Interferência de RNA , Fibras de Estresse/enzimologia , Células Estromais/patologia , Fatores de Tempo , TransfecçãoRESUMO
Maternal infections with gram-negative bacteria are associated with miscarriage and are one of the most common complications during pregnancy. Previous studies from our group have shown that lipopolysaccharide (LPS)-activated infiltrating peripheral blood mononuclear cells (PBMC) into decidual tissue plays an important role in the establishment of a local inflammatory process that results in embryo cytotoxicity and early embryo resorption. Moreover, we have also shown that an increased endocannabinoid tone mediates LPS-induced deleterious effects during early pregnancy loss. Here, we sought to investigate whether the infiltrating PBMC modulates the decidual endocannabinoid tone and the molecular mechanisms involved. PBMC isolated from 7-day pregnant mice subjected to different treatments were co-cultured in a transwell system with decidual tissue from control 7-day pregnant mice. Decidual fatty acid amide hydrolase (FAAH) activity was measured by radioconvertion, total decidual protein nitration by Western blot (WB), and decidual FAAH nitration by immunoprecipitation followed by WB. We found that co-culture of PBMC obtained from LPS-treated mice increased the level of nitration of decidual FAAH, which resulted in a negative modulation of decidual FAAH activity. Interestingly, co-treatment with progesterone or aminoguanidine prevented this effect. We found that LPS-treated PBMC release high amounts of nitric oxide (NO) which causes tyrosine nitration of decidual FAAH, diminishing its enzymatic activity. Inactivation of FAAH, the main degrading enzyme of anandamide and similar endocannabinoids, could lead to an increased decidual endocannabinoid tone with embryotoxic effects. J. Cell. Physiol. 232: 1441-1447, 2017. © 2016 Wiley Periodicals, Inc.
Assuntos
Amidoidrolases/metabolismo , Decídua/enzimologia , Regulação para Baixo , Perda do Embrião/induzido quimicamente , Perda do Embrião/enzimologia , Leucócitos Mononucleares/metabolismo , Animais , Decídua/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Perda do Embrião/patologia , Feminino , Guanidinas/farmacologia , Leucócitos Mononucleares/efeitos dos fármacos , Lipopolissacarídeos/administração & dosagem , Camundongos Endogâmicos BALB C , Óxido Nítrico/metabolismo , Nitrosação , Progesterona/farmacologia , Quercetina/farmacologiaRESUMO
Glucocorticoids are implicated in successful blastocyst implantation, whereas alterations in glucocorticoid levels are associated with various pregnancy disorders including preeclampsia. Tissue concentration of active glucocorticoids depends on the expression of 11ß-hydroxysteroid dehydrogenase (11ß-HSD). This study investigated the contribution of first trimester decidua to glucocorticoid availability at the fetal-maternal interface by assessing the expression and regulation of 11ß-HSD in human first trimester decidual tissues and cells and by evaluating 11ß-HSD levels in preeclamptic vs. gestational age-matched decidua. 11ß-HSD1 was the predominant isoform in first trimester decidua. In vitro, decidual cell 11ß-HSD1 levels and enzymatic activity were up-regulated by ovarian steroids and inflammatory cytokines. Higher levels of 11ß-HSD1 were found in preeclamptic decidua compared to controls. The present study indicates the predominance of 11ß-HSD oxoreductase isoform in early decidua. Observations that ovarian hormones and inflammatory cytokines up-regulate 11ß-HSD1, together with increased 11ß-HSD1 expression in preeclampsia, highlight a role for decidual cells in controlling biologically active glucocorticoids in early pregnancy.
Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 1/genética , Decídua/enzimologia , Decídua/patologia , Regulação Enzimológica da Expressão Gênica , Pré-Eclâmpsia/enzimologia , Pré-Eclâmpsia/genética , Primeiro Trimestre da Gravidez/genética , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/metabolismo , Citocinas/farmacologia , Decídua/efeitos dos fármacos , Feminino , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Humanos , Imuno-Histoquímica , Placenta/efeitos dos fármacos , Placenta/enzimologia , Placenta/patologia , Gravidez , Primeiro Trimestre da Gravidez/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Esteroides/metabolismoRESUMO
INTRODUCTION: In this study we have tracked glycogen and glycoprotein flux associated with nutrient uptake into trophoblast in early deciduochorial and later haemochorial placenta. METHODS: α-amylase, glycogen synthase and glycogen phosphorylase were immunohistochemically localised in 6-14 week and term placenta and first trimester decidua. Placentae of 4-18 weeks' gestation and term were also stained with 22 biotinylated lectins. RESULTS: Histochemical data were consistent with glycogenolysis in decidual gland epithelium and placental cyto- and syncytiotrophoblast; α-amylase was present in decidual secretions but absent in placenta. Glycogen and glycogen synthase were both apparent in villous cytotrophoblast cells and columns. Profound changes were observed in placental glycosylation during gestation. Syncytial microvilli were richly glycosylated as were first trimester vacuoles but, by term, syncytiotrophoblast showed little lectin binding except in microvillous and basal membranes. Cytotrophoblast Golgi bodies were active in the first trimester; at term the cells were generally more glycosylated than syncytiotrophoblast. DISCUSSION: We deduce that decidual cell glycogen is broken down for transport into the placenta where the products may be reassembled into glycogen or used for metabolic processes. First trimester histiotrophe is internalised by syncytiotrophoblast, then broken down in apical vacuoles containing lysosomal markers. This process declines after haemotrophic nutrition commences. Transition from histiotrophic to haemotrophic nutrition involves reduced amounts of uterine secretory derivatives reaching the placenta, and reduction in internalisation of glycoprotein by syncytiotrophoblast, presumably reflecting the shift to low molecular weight nutrients. Glycogen accumulates in cytotrophoblast from early pregnancy and is mobilised for utilisation by fetoplacental tissues.
Assuntos
Decídua/metabolismo , Glicogênio/metabolismo , Glicoproteínas/metabolismo , Fenômenos Fisiológicos da Nutrição Materna , Troca Materno-Fetal , Placenta/metabolismo , Placentação , Adulto , Decídua/citologia , Decídua/enzimologia , Feminino , Glicogênio/biossíntese , Glicogênio Fosforilase/metabolismo , Glicogênio Sintase/metabolismo , Glicogenólise , Glicosilação , Humanos , Especificidade de Órgãos , Placenta/citologia , Placenta/enzimologia , Gravidez , Primeiro Trimestre da Gravidez , Terceiro Trimestre da Gravidez , Processamento de Proteína Pós-Traducional , alfa-Amilases/metabolismoRESUMO
The three isoforms of AKT: AKT1, AKT2, and AKT3, are crucial regulators of both normal and pathological cellular processes. Each of these isoforms exhibits a high level of homology and functional redundancy with each other. However, while being highly similar and structurally homologous, a rising amount of evidence is showing that each isoform possesses specific targets as well as preferential subcellular localization. The role of AKT has been studied extensively in reproductive processes, but isoform-specific roles are yet to be fully understood. This review will focus on the role of AKT in the uterus and its function in processes related to cell death and proliferation such as embryo implantation, decidualization, endometriosis, and endometrial cancer in an isoform-centric manner. In this review, we will cover the activation of AKT in various settings, localization of isoforms in subcellular compartments, and the effect of isoform expression on cellular processes. To fully understand the dynamic molecular processes taking place in the uterus, it is crucial that we better understand the physiological role of AKT isoforms as well as their function in the emergence of diseases.
Assuntos
Proteínas Proto-Oncogênicas c-akt/metabolismo , Reprodução , Útero/enzimologia , Animais , Decídua/enzimologia , Implantação do Embrião , Endometriose/enzimologia , Endometriose/patologia , Endometriose/fisiopatologia , Ativação Enzimática , Feminino , Regulação Enzimológica da Expressão Gênica , Humanos , Isoenzimas , Proteínas Proto-Oncogênicas c-akt/genética , Transdução de Sinais , Neoplasias Uterinas/enzimologia , Neoplasias Uterinas/patologia , Neoplasias Uterinas/fisiopatologia , Útero/patologia , Útero/fisiopatologiaRESUMO
OBJECTIVE: To clarify the role and mechanism of interleukin (IL)-25 in regulating the biological functions of decidual stromal cells (DSCs) in human early pregnancy. DESIGN: Laboratory study of the effect of IL-25 induced by hCG on the proliferation of DSCs. SETTING: Research laboratories. PATIENT(S): Women aged 23-47 years with normal pregnancy and abortion. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Signal transduction from IL-25. RESULT(S): Here we show that DSCs coexpress IL-25/IL-17RB. Human chorionic gonadotropin promotes the expression of IL-25/IL-17RB. In contrast to anti-human IL-25 neutralizing antibody, recombinant human IL-25 (rhIL-25) significantly stimulates the proliferation of DSCs in dosage- and time-dependent manners. RhIL-25 promotes the phosphorylation of AKT, extracellular-signal-regulated kinases 1/2 (ERK1/2), c-Jun N-terminal kinases (JNK), and nuclear factor κB in DSCs. Interestingly, blocking JNK or AKT signal with inhibitors down-regulates the stimulatory effect on DSC proliferation induced by rhIL-25. In addition, the results of Western blot show that the expression of IL-25/IL-17RB in DSCs from normal pregnancy was higher than that from abortion. CONCLUSION(S): Our results have revealed that hCG derived of trophoblasts up-regulates the expression of IL-25/IL-17RB in DSCs and that IL-25 further stimulates the proliferation of DSCs through activating JNK and AKT signals, which finally contributes to the establishment and maintenance of physiological pregnancy.
Assuntos
Proliferação de Células , Gonadotropina Coriônica/metabolismo , Decídua/enzimologia , Interleucina-17/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Células Estromais/enzimologia , Adulto , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Decídua/efeitos dos fármacos , Ativação Enzimática , Feminino , Humanos , Interleucina-17/farmacologia , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Pessoa de Meia-Idade , Fosforilação , Gravidez , Primeiro Trimestre da Gravidez , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Receptores de Interleucina/metabolismo , Receptores de Interleucina-17 , Proteínas Recombinantes/farmacologia , Transdução de Sinais/efeitos dos fármacos , Células Estromais/efeitos dos fármacos , Fatores de Tempo , Regulação para Cima , Adulto JovemRESUMO
Arachidonic acid (ARA) metabolites maintain pregnancy and control parturition. We generated a network of 77 proteins involved in placental ARA metabolism to identify novel proteins in this pathway. We identified a long pathway within this network which showed that secretory and cytosolic phospholipase A2 proteins act in concert. The functions of all network proteins expressed in the placental decidua were determined by database searches. Thus ARA metabolism was linked to carbohydrate metabolism. One protein, transmembrane protein 62 (TMEM62), expressed in decidua was previously uncharacterized, and was identified as a putative lipoxygenase. TMEM62 may play a role in pregnancy and/or parturition.