RESUMO
Objective: To evaluate the usefulness of the serum concentration of nine matrix metalloproteinases (MMPs) as biomarkers of spontaneous abortion.Methods: A retrospective nested cohort case-control study was carried out in Zacatecas, Mexico. MMP-1-3, MMP-7-10, and MMP-12-13 were analyzed in serum from women who had spontaneous abortion of unknown causes (n = 7), who suffered abortions attributed to urinary tract infection (n = 7) and from those with healthy pregnancies without complications (controls; n = 20). Protein profiles were determined between 11 and 13 weeks of gestation (GW) using the Bio-Plex Pro Human MMP Panel. Differences in serum MMP concentrations between the study groups and their correlation with clinical findings were evaluated statistically.Results: There were differences in serum concentrations of MMP-9 between groups of spontaneous abortion of unknown cause (13.2 ± 7.5 ng/µL), abortion attributed to urinary tract infection (11.6 ± 5.8 ng/µL) and the controls (11.8 ± 16.5 ng/µL) (p = .022). Compared with controls, higher serum concentrations of MMP-8, MMP-9, and MMP-10 were observed in the group of spontaneous abortions of unknown causes (p value < .05). A negative correlation between MMP-8 and MMP-9 and urine density was also identified (r = -0.949, p value = .0167; and r = -0.947, p = .0167).Conclusions: Elevated serum concentrations of MMP-8, MMP-9, and MMP-10 were associated and preceded by the appearance of spontaneous interruption of pregnancy of unknown causes. Our results support the hypothesis that altered MMP modulation may be related with the pathogenesis of spontaneous abortion.
Assuntos
Aborto Espontâneo , Metaloproteinase 9 da Matriz , Aborto Espontâneo/sangue , Aborto Espontâneo/metabolismo , Estudos de Casos e Controles , Feminino , Humanos , Metaloproteinase 9 da Matriz/sangue , Metaloproteinase 9 da Matriz/metabolismo , México , Gravidez , Estudos RetrospectivosRESUMO
During decidualization, endometrial stromal cells undergo reticular stress (RS) and unfolded protein response (UPR), allowing the endoplasmic reticulum-expansion and immunomodulators production. Physiological RS generates the activation of sensing proteins, inflammasome activation and mature-IL-1ß secretion, associated with pro-implantatory effects. We focus on the impact of RS and UPR on decidualized cells and whether they induce a physiological sterile inflammatory response through IL-1ß production. Human endometrial stromal cell line (HESC) after decidualization treatment with MPA + dibutyryl-cAMP (Dec) increased the expression of RS-sensors (ATF6, PERK and IRE1α) and UPR markers (sXBP1 and CHOP) in comparison with Non-dec cells. Then we found increased NLRP3 expression in Dec cells compared with Non-dec cells. In fact STF-083010 (an IRE1α inhibitor) prevented this increase. Downstream, increased levels of active caspase-1 on Dec cells were detected by FAM-Flica Caspase-1 associated with an increase in IL-1ß production. Moreover, the treatment with STF-083010 decreased the invasion index observed in Dec cells, evaluated by an in vitro model of implantation. In endometrial biopsies from recurrent spontaneous abortion patients an increased expression of IRE1α was found in comparison with fertile women; while recurrent implantation failure samples showed a lower expression of sXBP1, TXNIP and NLRP3 than fertile women, suggesting that RS/UPR tenors might condition endometrial receptivity.
Assuntos
Endométrio/patologia , Estresse do Retículo Endoplasmático , Resposta a Proteínas não Dobradas , Aborto Espontâneo/metabolismo , Aborto Espontâneo/patologia , Aborto Espontâneo/fisiopatologia , Adulto , Caspase 1/metabolismo , Linhagem Celular , Decídua/patologia , Implantação do Embrião , Feminino , Humanos , Inflamação/patologia , Interleucina-1beta/biossíntese , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Recidiva , Células Estromais/metabolismo , Células Estromais/patologia , Trofoblastos/patologiaRESUMO
STUDY QUESTION: What is the role of the endocannabinoid system (eCS) on the lipopolysaccharide (LPS) effects on uterine explants from 7-day pregnant mice in a murine model of endotoxin-induced miscarriage? SUMMARY ANSWER: We found evidence for cannabinoid receptor type2 (CB2) involvement in LPS-induced increased prostaglandin-F2α (PGF2α) synthesis and diminished cyclic adenosine monophosphate (cAMP) intracellular content in uterine explants from early pregnant mice. WHAT IS KNOWN ALREADY: Genital tract infections by Gram-negative bacteria are a common complication of human pregnancy that results in an increased risk of pregnancy loss. LPS, the main component of the Gram-negative bacterial wall, elicits a strong maternal inflammatory response that results in embryotoxicity and embryo resorption in a murine model endotoxin-induced early pregnancy loss. We have previously shown that the eCS mediates the embryotoxic effects of LPS, mainly via CB1 receptor activation. STUDY DESIGN, SIZE, DURATION: An in vitro study of mice uterine explants was performed to investigate the eCS in mediating the effects of LPS on PGF2α production and cAMP intracellular content. PARTICIPANTS/MATERIALS, SETTING, METHODS: Eight to 12-week-old virgin female BALB/c or CD1 (wild-type [WT] or CB1-knockout [CB1-KO]) mice were paired with 8- to 12-week-old BALB/c or CD1 (WT or CB1-KO) males, respectively. On day 7 of pregnancy, BALB/c, CD1 WT or CD1 CB1-KO mice were euthanized, the uteri were excised, implantation sites were removed and the uterine tissues were separated from decidual and embryo tissues. Uterine explants were cultured and exposed for an appropriate amount of time to different pharmacological treatments. The tissues were then collected for cAMP assay and PGF2α content determination by radioimmunoassay. MAIN RESULTS AND THE ROLE OF CHANCE: In vitro treatment of uteri explants from 7-day pregnant BALB/c or CD1 (WT or CB1-KO) mice with LPS induced an increased production of PGF2α (P < 0.05) and a reduction of the tissue content of cAMP (P < 0.05). These effects were mediated by CB2 receptors since exposure to AM630 (a specific CB2 receptor antagonist) prevented these LPS-induced effects (P < 0.05). Collectively, our results suggest a role for the eCS mediating LPS-induced deleterious effects on reproductive tissues. LIMITATIONS, REASONS FOR CAUTION: Since our experimental design involves in vitro experiments of uterine explants, the extrapolation of the results presented here to humans is limited. WIDER IMPLICATIONS OF THE FINDINGS: Our findings provide evidence for the role of CB2 receptors in reproductive events as well as their participation as a mediator of LPS deleterious effects on reproductive tissues. LARGE SCALE DATA: None. STUDY FUNDING AND COMPETING INTEREST(S): Dr Ana María Franchi was funded by Agencia Nacional para la Promoción Científica y Tecnológica (PICT 2010/0813 and PICT 2013/0097) and by Consejo Nacional de Investigaciones Científicas y Técnicas (PIP 2012/0061). Dr Carlos Davio was funded by Agencia Nacional para la Promoción Científica y Tecnológica (PICT 2013/2050). The authors have no competing interests.
Assuntos
Aborto Espontâneo/metabolismo , AMP Cíclico/metabolismo , Lipopolissacarídeos/farmacologia , Receptor CB1 de Canabinoide/genética , Receptor CB2 de Canabinoide/genética , Útero/efeitos dos fármacos , Aborto Espontâneo/induzido quimicamente , Aborto Espontâneo/genética , Aborto Espontâneo/patologia , Animais , Agonistas de Receptores de Canabinoides/farmacologia , AMP Cíclico/antagonistas & inibidores , Dinoprosta/biossíntese , Modelos Animais de Doenças , Feminino , Deleção de Genes , Expressão Gênica , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Técnicas de Cultura de Órgãos , Gravidez , Receptor CB1 de Canabinoide/deficiência , Receptor CB2 de Canabinoide/metabolismo , Útero/metabolismo , Útero/patologiaRESUMO
STUDY QUESTION: What is the role of the endocannabinoid system (eCS) in the alterations of the endocrine system in a murine model of lipopolysaccharide (LPS)-induced miscarriage? SUMMARY ANSWER: In 7-days pregnant wild type, but not cannabinoid receptor type 1 knockout (CB1-KO) mice, LPS increased COX-2 expression and prostaglandin F2α (PGF2α) production in the uterus leading to lower expression of prolactin receptor in the ovary and a marked regression of corpora lutea (CL), suggesting that the eCS mediates the deleterious effects of LPS on reproductive events. WHAT IS KNOWN ALREADY: Appropriate systemic progesterone levels are critical for a successful pregnancy outcome. Precocious loss of luteal progesterone (P4) secretion leads to miscarriage in rodents. We have previously shown that LPS administration to pregnant mice induces embryonic resorption accompanied by a dramatic decrease in systemic progesterone levels in a murine model of inflammatory miscarriage, with the eCS mediating these LPS-induced deleterious effects. STUDY DESIGN SAMPLES/MATERIALS, METHODS: CD1 wild-type (WT) and CB1-KO mice were randomly allocated to Vehicle (saline; i.p.) or LPS (0.5 µg/g body weight; i.p.) treated groups: (WT-Vehicle; WT-LPS; CB1-KO-Vehicle and CB1-KO-LPS). A single injection was given on day 7 of pregnancy and tissues (blood, ovary, uterus) were collected 6, 12, 24 and 48 h later. P4 and PGF2α plasma levels were determined by radioimmunoassay. Cyclooxygenase-2 (COX-2) mRNA (RT-PCR) and protein (Western blot) content in uterus was assayed. COX-2 and prolactin receptor (PrlR) mRNA levels in the ovary were assayed by RT-PCR. Tissue morphology of the CL was assessed by haematoxylin-eosin staining. MAIN RESULTS AND THE ROLE OF CHANCE: Treatment of 7-day pregnant WT mice with LPS induced a P4 withdrawal (p < 0.05), increased in uterine COX-2 mRNA and protein expression (p < 0.05) as well as an increase in uterine PGF2α production (p < 0.05). These changes were absent in LPS-treated 7-day pregnant CB1-KO mice. In ovarian tissues, LPS treatment to 7-day pregnant WT mice induced a downregulation of PrlR mRNA expression (p < 0.05) together with an increase in COX-2 mRNA expression (p < 0.05) and PGF2α content (p < 0.05). These effects were absent in the CB1-KO mice. Collectively, our results suggest a role for the eCS mediating LPS-induced deleterious effects on reproductive tissues. LIMITATIONS, REASONS FOR CAUTION: An important caveat of this study is the endocrine differences between mice and humans during pregnancy (e.g. P4 is produced by the CL throughout pregnancy in mice, whereas this is not the case in humans), which limits the extrapolation of the results presented here. WIDER IMPLICATIONS OF THE FINDINGS: Our findings provide new insights in the role of the endocannabinoid system in the physiopathology of reproduction as well as the role of this endogenous system as a mediator of LPS deleterious effects on reproductive tissues. LARGE SCALE DATA: None. STUDY FUNDING AND COMPETING INTERESTS: Dr Ana María Franchi was funded by Agencia Nacional para la Promoción Científica y Tecnológica (PICT 2010/0813 and PICT 2013/0097) and by Consejo Nacional de Investigaciones Científicas y Técnicas (PIP 2012/0061). The authors have no competing interests.
Assuntos
Aborto Espontâneo/tratamento farmacológico , Aborto Espontâneo/metabolismo , Endocanabinoides/metabolismo , Lipopolissacarídeos/toxicidade , Fase Luteal/metabolismo , Progesterona/metabolismo , Animais , Corpo Lúteo/metabolismo , Feminino , Luteólise/metabolismo , Camundongos , Camundongos Knockout , Gravidez , RadioimunoensaioRESUMO
Miscarriage caused by Gram-negative bacteria infecting the female genital tract is one of the most common complications of human pregnancy. Intraperitoneal administration of LPS to 7-days pregnant mice induces embryo resorption after 24 h. Here, we show that LPS induced apoptosis on uterine explants from 7-days pregnant mice and that CB1 receptor was involved in this effect. On the other hand, heparin has been widely used for the prevention of pregnancy loss in women with frequent miscarriage with or without thrombophilia. Besides its anticoagulant properties, heparin exerts anti-inflammatory, immunomodulatory and anti-apoptotic effects. Here, we sought to investigate whether the administration of heparin prevented LPS-induced apoptosis in uterine explants from 7-days pregnant mice. We found that heparin enhanced cell survival in LPS-treated uterine explants and that this effect was mediated by increasing uterine FAAH activity. Taken together, our results point towards a novel mechanism involved in the protective effects of heparin.
Assuntos
Aborto Espontâneo/metabolismo , Aborto Espontâneo/fisiopatologia , Apoptose/efeitos dos fármacos , Endocanabinoides/metabolismo , Heparina/farmacologia , Útero/metabolismo , Aborto Espontâneo/genética , Animais , Sobrevivência Celular/efeitos dos fármacos , Implantação do Embrião , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Gravidez , Receptor CB1 de Canabinoide/genética , Receptor CB1 de Canabinoide/metabolismoRESUMO
INTRODUCTION: Symptomatic or asymptomatic Shiga toxin producing Escherichia coli (STEC) infections during early pregnancy may cause maternal or fetal damage mediated by Shiga toxin type 2 (Stx2). The aim of this study is to elucidate the mechanisms responsible for early pregnancy loss in rats treated with Stx2. METHODS: Sprague Dawley pregnant rats were intraperitoneally injected at day 8 of gestation with a sublethal dose (0.5 ng of Stx2/g of total body weight, 250 µl) of purified Stx2. Control rats were injected with the same volume of PBS. The expression of globotriaosylceramide (Gb3) glycosphingolipid receptor for Stx2 was evaluated by thin-layer chromatography (TLC). Regions of hypoxia in decidual tissue were determined by pimonidazole immunohistochemistry and vascular endothelial growth factor (VEGF) expression by Western blot and immunohistochemistry. Tumor necrosis factor-alpha (TNF-α) levels in serum and decidual tissue were evaluated by ELISA. Serum progesterone levels were determined by RIA. RESULTS: Decidual tissue from both, control and Stx2-treated rats showed similar expression of Gb3 receptor. Intrauterine growth restriction was observed in Stx2-treated rats, associated with hypoxia and an increase of decidual TNF-α levels. Decrease of serum progesterone levels and decidual VEGF expression were also demonstrated. DISCUSSION: Our findings indicate that Stx2 reaches the uteroplacental unit, binds Gb3 and triggers damage in decidual tissue. Poor oxygen supply accompanied with damage in the uteroplacental unit and inflammation could be responsible for the early pregnancy loss. Decrease in the pregnancy protective factors, serum progesterone and local VEGF, may contribute to the pregnancy loss.
Assuntos
Aborto Espontâneo/patologia , Hipóxia/patologia , Inflamação/patologia , Toxina Shiga II , Aborto Espontâneo/induzido quimicamente , Aborto Espontâneo/metabolismo , Animais , Feminino , Hipóxia/metabolismo , Inflamação/metabolismo , Gravidez , Ratos , Ratos Sprague-Dawley , Triexosilceramidas/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Sound stress exposure increases fetal loss via inflammatory pathways. Inflammation is known to up-regulate cell adhesion molecules, such as vascular cell adhesion molecule-1 (VCAM-1), which mediates the adhesion of leukocytes to the vascular endothelium. In this work, we studied the frequency of VCAM-1(+) vessels at the fetomaternal interface in stressed and non-stressed pregnant CBA/J female mice mated with DBA/2J (high fetal loss model) or BALB/c (low fetal loss model) males. The high fetal loss model had fewer large vessels on gestation day 6.5, and stress reduced the frequency of large vessels to a similar number in both high and low fetal loss models. In the high fetal loss model, however, the frequency of VCAM-1+ vessels was dramatically increased. This study shows that VCAM-1 expression is modulated by stress at the fetomaternal interface in abortion-prone cross-breeding.
Assuntos
Aborto Espontâneo/metabolismo , Regulação da Expressão Gênica , Placenta/metabolismo , Estresse Fisiológico , Molécula 1 de Adesão de Célula Vascular/biossíntese , Aborto Espontâneo/patologia , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos CBA , Placenta/patologia , GravidezRESUMO
To analyze immunomodulating effects related to parity status, we studied trophoblast invasion grade, placental expression and systemic concentration of VEGF and its receptor Flt-1 in normal fertile (CBA/JxBALB/c) mice and abortion-prone (CBA/JxDBA/2) H-2(d)xH-2(k) mice. BALB/c or DBA/2 mated CBA/J females were, respectively, divided into the following groups: primiparous young (3.0+/-0.5 months old); primiparous old (8.5+/-0.5 months old) and multiparous old (8.5+/-0.5 months old, with 4 pregnancies). Immunohistochemical analysis of term placentae from both multiparous groups revealed various layers of invasive trophoblast tissue, identified as cytokeratin+/vimentin- cells, in contrast to the single layer detected in the placentae of primiparous animals, indicating that multiparity increases trophoblast invasion regardless of the success of the pregnancy outcome. Invasive trophoblast tissue from primiparous CBA/JxDBA/2 placentae showed diminished VEGF expression in comparison with the normal fertile group, while both multiparous groups demonstrated high expression of VEGF in the invasive trophoblast tissue. Placental expression of Flt-1 was similar in all groups. However, the primiparous CBA/JxBALB/c group showed the highest plasma concentration of sFlt-1 at term, while both multiparous groups demonstrated low circulating levels. No differences in circulating VEGF levels were observed among the groups. These results demonstrate an increase in trophoblast invasion tissue and expression of VEGF in the maternal-fetal interface in multiparous mice compared to primiparous mice. Moreover, the placenta appears to be able to regulate the circulating levels of VEGF by releasing sFlt-1.
Assuntos
Aborto Espontâneo/fisiopatologia , Troca Materno-Fetal , Paridade , Trofoblastos/metabolismo , Fator A de Crescimento do Endotélio Vascular/biossíntese , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/biossíntese , Aborto Espontâneo/sangue , Aborto Espontâneo/genética , Aborto Espontâneo/metabolismo , Animais , Proliferação de Células , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Troca Materno-Fetal/fisiologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos CBA , Camundongos Endogâmicos DBA , Paridade/fisiologia , Gravidez , Trofoblastos/patologia , Fator A de Crescimento do Endotélio Vascular/genética , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/sangue , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genéticaRESUMO
OBJECTIVE: To explore the utility of histological, immunohistochemical and chromosome in situ hybridization (CISH) test in the differential diagnosis of Complete Hydatidiform Mole (CHM), Partial Hydatidiform Mole (PHM) and Hydropic Abortion (HA). STUDY DESIGN: We analyzed the histological characteristics, p57kip2 and Factor VIII expression and CISH test in 38 cases with some diagnostic concerns, comprising 13 CHM, 14 PHM and 11 HA. RESULTS: Our results indicate that p57kip2 expression and the ploidy assessed by CISH were essential for the reclassification of 2 cases, one from CHM to PHM and another from PHM to HA, as well as for confirming the previous diagnosis in cases where there were conflicting features. p57kip2 expression is diagnostic if no cells at all present it (CHM) or when there are over 10% of cells expressing it (PHM and HA). CONCLUSIONS: We concluded that there is no single criterion for the distinction of CHM, PHM and HA. So p57kip2 expression and CISH test can be used in association with the histological findings for the differential diagnosis of the three conditions in cases presenting some concern for definitive diagnosis.
Assuntos
Aborto Espontâneo/diagnóstico , Mola Hidatiforme/diagnóstico , Neoplasias Uterinas/diagnóstico , Aborto Espontâneo/genética , Aborto Espontâneo/metabolismo , Adolescente , Adulto , Inibidor de Quinase Dependente de Ciclina p57/metabolismo , Diagnóstico Diferencial , Fator VIII/metabolismo , Feminino , Humanos , Mola Hidatiforme/genética , Mola Hidatiforme/metabolismo , Imuno-Histoquímica , Hibridização In Situ , Gravidez , Estudos Retrospectivos , Neoplasias Uterinas/genética , Neoplasias Uterinas/metabolismoRESUMO
PROBLEM: Several studies indicate that RANTES (regulated on activation, normal T cell expressed and secreted) is able to downregulate T-cell responses which suggest it might be relevant for fetal tolerance induction. However, the role of RANTES in pregnancy had not been established. Here we investigate RANTES regulation during early pregnancy and potential failures leading to losses of pregnancies. METHOD OF STUDY: RANTES and progesterone levels were determined in sera and feto-placental units from high resorption rate CBA/JxDBA/2 pregnant females and compared with CBA/JxBALB/c normal pregnant mice. RANTES in vitro modulation was also studied in nulliparous, primiparous and multiparous CBA/J and BALB/c cells in response to paternal alloantigen and progesterone stimulation. RESULTS: Nulliparous CBA/J females were quantitatively deficient in RANTES sera levels, whereas pregnancies with male BALB/c or DBA/2 increased its production. However, feto-placental units from CBA/J females are high producers of progesterone and RANTES. CONCLUSION: These data suggest that the beneficial effect of RANTES on feto-maternal interface requires an optimal concentration range and might be modulated by progesterone, hence exacerbated placental expression could be associated with high resorption rate.