RESUMO
Ovarian endometrioma (OMA) and deep infiltrating endometriosis (DIE) are the most severe forms of endometriosis, but different pathogenetic mechanisms and clinical symptoms distinguish these two forms. Corticotrophin-releasing hormone (CRH) and urocortin (Ucn) are endometrial neuropeptides involved in tissue differentiation and inflammation. The expression of CRH, Ucn, Ucn2, CRH-receptors (type-1 and type-2) and inflammatory enzymes phospholipase-A2 group IIA (PLA2G2A) and cycloxygenase-2 (COX2) were evaluated in OMA (n = 22) and DIE (n = 26). The effect of CRH or Ucn on COX2 mRNA expression was evaluated in cultured human endometrial stromal cells. In DIE lesions, CRH, Ucn and CRH-R2 mRNA levels were significantly higher than in OMA (P < 0.01, P < 0.001 and P < 0.05, respectively); DIE lesions showed a higher expression of COX2 (P < 0.01) and PLA2G2A (P < 0.05) mRNA than OMA, which was positively correlated with CRH-R2 mRNA expression (P < 0.05). Intense immunostaining for CRH and Ucn was shown in DIE. Treatment of cultured endometrial stromal cells with Ucn significantly increased COX2 mRNA expression (P < 0.01); this effect was reversed by the CRH-R2 antagonist astressin-2B. In DIE, DIE lesions highly express neuropeptide and enzyme mRNAs, supporting a strong activation of inflammatory pathways.
Assuntos
Endometriose/metabolismo , Doenças Ovarianas/metabolismo , Receptores de Hormônio Liberador da Corticotropina/metabolismo , Urocortinas/metabolismo , Adulto , Células Cultivadas , Hormônio Liberador da Corticotropina/genética , Hormônio Liberador da Corticotropina/metabolismo , Hormônio Liberador da Corticotropina/farmacologia , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Relação Dose-Resposta a Droga , Endometriose/genética , Endometriose/patologia , Endométrio/efeitos dos fármacos , Endométrio/metabolismo , Endométrio/patologia , Feminino , Fosfolipases A2 do Grupo II/genética , Fosfolipases A2 do Grupo II/metabolismo , Humanos , Doenças Ovarianas/genética , Doenças Ovarianas/patologia , Fragmentos de Peptídeos/farmacologia , Peptídeos Cíclicos/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Hormônio Liberador da Corticotropina/genética , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Urocortinas/genética , Urocortinas/farmacologia , Adulto JovemRESUMO
In chorioamnionitis (CAM), a major cause of preterm birth (PTB), maternal-fetal inflammation of the decidua and amniochorion cause the release of cytokines that elicit cervical ripening, fetal membrane rupture and myometrial activation. We posit that this inflammatory milieu triggers PTB by inhibiting progesterone receptor (PR) expression and increasing decidual prostaglandin (PG) production. Immunohistochemical staining of decidua detected significantly lower PR levels in decidual cells (DCs) from CAM-complicated PTB. Incubation of DCs with IL-1ß decreased PR expression and significantly increased PGE2 and PGF2α production and COX-2 expression. The addition of PGF2α to DC cultures also suppressed PR expression. However, the COX inhibitor, indomethacin, did not reverse IL-1ß suppression of PR expression in DC cultures. Although IL-1ß treatment activated the NF-KB, ERK1/2 and p38 MAPK signalling cascades in DCs, inhibition of ERK1/2 MAPK signalling alone was sufficient to completely reverse the suppression of PR levels by IL-1ß. These findings suggest that CAM-associated PTB is induced at least in part by IL-1ß-mediated functional progesterone withdrawal.
Assuntos
Corioamnionite/metabolismo , Decídua/metabolismo , Interleucina-1beta/metabolismo , Nascimento Prematuro/etiologia , Receptores de Progesterona/biossíntese , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Immunoblotting , Imuno-Histoquímica , Análise de Sequência com Séries de Oligonucleotídeos , Gravidez , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Dysmenorrhea, defined as painful cramps occurring immediately before or during the menstrual period, is a common symptom of different gynecological diseases. An acute uterine inflammatory response driven by prostaglandins (PGs) is responsible for painful symptoms. Progesterone withdrawal is responsible for activation of cyclooxygenase (COX-2) enzyme and decrease of hydroxyprostaglandin dehydrogenase (HPDG) with consequent increased secretion of PGs secretion, inducing uterine contractility and pain. The most widely used drugs for the treatment of pelvic pain associated with menstrual cycle are non steroidal anti-inflammatory drugs (NSAIDs). The uterine site of action of these drugs is still not defined and the present study evaluated the effect of naproxen sodium in cultured human endometrial stromal cells (HESC) collected from healthy women. PGE2 release was measured by ELISA; COX-2 and HPDG mRNA expression were assessed by qRT-PCR. Naproxen sodium did not affect HESC vitality. Naproxen sodium significantly decreased PGE2 secretion (p < 0.01) and COX-2 mRNA expression (p < 0.01). TNF-α induced PGE2 release was reduced in presence of naproxen sodium (p < 0.05), in association with decreased COX-2 and increased HPDG mRNAs expression. Naproxen sodium decreases endometrial PGE2 release induced by inflammatory stimulus acting on endometrial COX-2 and HPDG expression, suggesting endometrial synthesis of prostaglandins as a possible target for reduction of uterine inflammatory mechanism in dysmenorrhea.
Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , Dinoprostona/metabolismo , Endométrio/efeitos dos fármacos , Naproxeno/farmacologia , Adulto , Células Cultivadas , Ciclo-Oxigenase 2/metabolismo , Endométrio/citologia , Endométrio/enzimologia , Endométrio/metabolismo , Feminino , Humanos , Células Estromais/efeitos dos fármacos , Células Estromais/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Adulto JovemRESUMO
Burkitt lymphoma (BL) is characterized by tumor microenvironment (TME) in which macrophages represent the main component, determining a distinct histological appearance known as "starry sky" pattern. However, in some instances, BL may exhibit a granulomatous reaction that has been previously linked to a favorable prognosis and spontaneous regression. The aim of our study was to deeply characterize the immune landscape of 7 cases of EBV + BL with granulomatous reaction compared to 8 cases of EBV + BL and 8 EBV- BL, both with typical "starry sky" pattern, by Gene expression profiling performed on the NanoString nCounter platform. Subsequently, the data were validated by multiplex and combined immunostaining. Based on unsupervised clustering of differentially expressed genes, BL samples formed 3 distinct clusters differentially enriched in BL with a diffuse granulomatous reaction (cluster 1), EBV+ BL with typical starry sky pattern (Cluster 2), EBV - BL with typical "starry sky" (cluster 3). We observed variations in the immune response signature among BL with granulomatous reaction and BL with typical "starry sky", both EBV + and EBV -. The TME signature in BL with diffuse granulomatous reaction showed a proinflammatory response, while BLs with "starry sky" were characterized by up-regulation of M2- polarization and pro-tumor response. Moreover, the analysis of additional signatures revealed an up-regulation of Dark zone-signature and epigenetic-signature in BL with typical "starry sky". Tumor associated macrophages (TAM) and epigenetic regulators may be promising targets for additional therapies in BL lymphoma opening novel immunotherapeutic strategies.
RESUMO
Iron overload is associated with pregnancy complications. Ferroportin (FPN) is the only known iron exporter in mammalian cells. We hypothesize that FPN is functionally important in ferrotopsis, a process of iron-dependent non-apoptotic programmed cell death, and may have a critical role to play in pregnancy success. We investigated the expression of FPN in placenta/fetal membranes by immunohistochemistry in tissues collected from pregnancies with/without preeclampsia (PE) and spontaneous preterm birth (SPTB). FPN was highly expressed in both trophoblasts and decidual cells found in placenta/fetal membranes. Staining was significantly reduced in fetal membranes from SPTB versus healthy pregnancies (P = 0.046). FPN expression in immortalized human endometrial stromal cells (HESC) increased with in vitro decidualization induction using 1 µM of medroxyprogesterone acetate and 0.5 mM of dibutyryl-cAMP. In addition, both HESC cells and immortalized extravillous trophoblast SW71 cells with FPN knockdown showed significant sensitivity to ferroptosis inducer, erastin (P < 0.001 and P = 0.009, respectively). The survival of both HESC and SW71 cells was not negatively affected by iron supplementation with ferric ammonium citrate in the medium. However, SW71 cells were more sensitive than HESC cells to physiologic iron in the presence of a non-lethal dose of erastin (P < 0.001). Taken together, our data demonstrating increased sensitivity of FPN knockdown HESC and SW71 cells to erastin and increased sensitivity of trophoblasts to iron overload under ferroptotic stress support the hypothesis that FPN protects against ferroptosis during pregnancy.
Assuntos
Ferroptose , Sobrecarga de Ferro , Nascimento Prematuro , Recém-Nascido , Gravidez , Feminino , Animais , Humanos , Resultado da Gravidez , Nascimento Prematuro/metabolismo , Placenta/metabolismo , Ferro , Sobrecarga de Ferro/metabolismo , Mamíferos/metabolismoRESUMO
Preeclampsia is characterized by an exaggerated systemic inflammatory state as well as shallow placentation. In the decidual implantation site, preeclampsia is accompanied by an excessive number of both macrophages and dendritic cells as well as their recruiting chemokines, which have been implicated in the impairment of endovascular trophoblast invasion. Granulocyte-macrophage colony-stimulating factor is known to regulate the differentiation of both macrophages and dendritic cells, prompting both in vivo and in vitro evaluation of granulocyte-macrophage colony-stimulating factor expression in human decidua as well as in a mouse model of preeclampsia. This study revealed increased granulocyte-macrophage colony-stimulating factor expression levels in preeclamptic decidua. Moreover, both tumor necrosis factor-α and interleukin-1 ß, cytokines that are implicated in the genesis of preeclampsia, markedly up-regulated granulocyte-macrophage colony-stimulating factor production in cultured first-trimester human decidual cells. The conditioned media of these cultures promoted the differentiation of both macrophages and dendritic cells from a monocyte precursor. Evaluation of a murine model of preeclampsia revealed that the decidua of affected animals displayed higher levels of immunoreactive granulocyte-macrophage colony-stimulating factor as well as increased numbers of both macrophages and dendritic cells when compared to control animals. Because granulocyte-macrophage colony-stimulating factor is a potent inducer of differentiation and activation of both macrophages and dendritic cells, these findings suggest that this factor plays a crucial role in the pathogenesis of preeclampsia.
Assuntos
Decídua/citologia , Decídua/metabolismo , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Pré-Eclâmpsia/imunologia , Pré-Eclâmpsia/fisiopatologia , Animais , Diferenciação Celular/fisiologia , Células Cultivadas , Células Dendríticas/citologia , Células Dendríticas/fisiologia , Modelos Animais de Doenças , Feminino , Humanos , Interleucina-1beta/imunologia , Macrófagos/citologia , Macrófagos/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Monócitos/citologia , Monócitos/fisiologia , Placenta/citologia , Placenta/metabolismo , Placenta/patologia , Pré-Eclâmpsia/patologia , Gravidez , Fator de Necrose Tumoral alfa/imunologiaRESUMO
Endometriosis is a major cause of chronic pain, infertility, medical and surgical interventions, and health care expenditures. Tissue factor (TF), the primary initiator of coagulation and a modulator of angiogenesis, is not normally expressed by the endothelium; however, prior studies have demonstrated that both blood vessels in solid tumors and choroidal tissue in macular degeneration express endothelial TF. The present study describes the anomalous expression of TF by endothelial cells in endometriotic lesions. The immunoconjugate molecule (Icon), which binds with high affinity and specificity to this aberrant endothelial TF, has been shown to induce a cytolytic immune response that eradicates tumor and choroidal blood vessels. Using an athymic mouse model of endometriosis, we now report that Icon largely destroys endometriotic implants by vascular disruption without apparent toxicity, reduced fertility, or subsequent teratogenic effects. Unlike antiangiogenic treatments that can only target developing angiogenesis, Icon eliminates pre-existing pathological vessels. Thus, Icon could serve as a novel, nontoxic, fertility-preserving, and effective treatment for endometriosis.
Assuntos
Endometriose/terapia , Imunoconjugados/farmacologia , Neovascularização Patológica/terapia , Doenças Peritoneais/terapia , Tromboplastina/antagonistas & inibidores , Tromboplastina/imunologia , Adulto , Animais , Células CHO , Cricetinae , Cricetulus , Sistemas de Liberação de Medicamentos , Endometriose/metabolismo , Endometriose/patologia , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Feminino , Humanos , Imunoconjugados/administração & dosagem , Fragmentos Fc das Imunoglobulinas/administração & dosagem , Fragmentos Fc das Imunoglobulinas/farmacologia , Imunoterapia/métodos , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Neovascularização Patológica/metabolismo , Doenças Peritoneais/metabolismo , Doenças Peritoneais/patologia , Tromboplastina/metabolismo , Transplante HeterólogoRESUMO
Chorioamnionitis frequently precedes both genital tract and placental inflammation and is both a primary cause of maternal morbidity and a major antecedent of preterm premature rupture of the membranes (PPROM) as well as preterm delivery (PTD). In most cases of chorioamnionitis, neutrophils dominate the decidua. In a subset of these cases, a predominance of monocytes is uniquely associated with both neonatal intraventricular hemorrhage and death. The multifunctional cytokine, interleukin-6, promotes local monocyte dominance via several mechanisms. In this study, immunostaining of placental sections revealed significantly higher interleukin-6 HSCOREs in decidual cells (DCs) but not in interstitial trophoblasts, in chorioamnionitis versus gestational age-matched control placentas (P < 0.05). In confluent leukocyte-free term DCs, secreted interleukin-6 levels in incubations with estradiol-17ß were increased 2500-fold by IL-1ß (P < 0.05). This up-regulation was inhibited by more than 50% in parallel incubations that included medroxyprogesterone acetate (n = 12, P < 0.05). Western blotting data confirmed these enzyme-linked immunosorbent assay results; quantitative RT-PCR findings demonstrated corresponding changes in interleukin-6 mRNA levels. Specific inhibitors of signaling for both nuclear factor-κB activation and p38-mitogen-activated protein kinase, but not for protein kinase C, significantly decreased IL-1ß-enhanced interleukin-6 expression levels in cultured DCs. In conclusion, in situ and in vitro results indicate that significantly enhanced interleukin-6 expression levels in DCs during chorioamnionitis could be pivotal in skewing decidual monocyte differentiation to macrophages.
Assuntos
Corioamnionite/metabolismo , Decídua/metabolismo , Regulação da Expressão Gênica , Interleucina-6/metabolismo , Placenta/metabolismo , Adulto , Antineoplásicos Hormonais/farmacologia , Western Blotting , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Corioamnionite/tratamento farmacológico , Corioamnionite/patologia , Decídua/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Estradiol/farmacologia , Estrogênios/farmacologia , Feminino , Humanos , Técnicas Imunoenzimáticas , Interleucina-1beta/farmacologia , Acetato de Medroxiprogesterona/farmacologia , Monócitos/citologia , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Placenta/efeitos dos fármacos , Gravidez , Nascimento Prematuro , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Macrophage Migration Inhibitory Factor (MIF) is a pivotal regulator of innate and acquired immunity affecting the response and behavior of macrophages and lymphocytes. However, a number of studies indicated wider physiological functions for this cytokine to include key-roles in reproductive biology. The present study was designed to clone the coding sequence of sheep MIF, to examine the characteristics of the protein in vitro, and to evaluate its expression in sheep tissues and in the ewe reproductive tract in vivo. Ovine MIF cDNA consisted of 348 nucleotides encoding a 115 amino acids protein with an estimated molecular mass of 12,343 Da and an isoelectric point of 7.68. Sheep MIF shared high amino acid identity with the other mammalian MIF family members and showed parallel functions to human MIF, displaying enzymatic oxoreductase activity and inducing monocyte transmigration. Expression studies detected a MIF transcript in all the sheep tissues examined. Among reproductive tissues, MIF mRNA and protein were detected in the ovary, oviduct, uterus and placenta. These results indicate that sheep MIF shares crucial features with other MIF family members and delineate its potential involvement in several aspects of ovine physiology.
Assuntos
Regulação da Expressão Gênica , Fatores Inibidores da Migração de Macrófagos/metabolismo , Placenta/metabolismo , Sequência de Aminoácidos , Animais , Clonagem Molecular , Biologia Computacional/métodos , Feminino , Linfócitos/citologia , Macrófagos/citologia , Dados de Sequência Molecular , Ovário/metabolismo , Oviductos/metabolismo , Gravidez , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Ovinos , Distribuição Tecidual , Útero/metabolismoRESUMO
BACKGROUND: Uterine extracellular matrix (ECM) remodeling occurs throughout pregnancy and at parturition. Imbalanced availability of key mediators in ECM degradation, namely, matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs), is implicated in the pathogenesis of preterm labor (PTL). OBJECTIVES: Examine the expression of MMPs and their inhibitors TIMPs in (a) the mouse uterus throughout normal gestation, at labor, and during inflammation-induced PTL and (b) the human term and preterm myometrium. METHODS: The expression of Mmp-2/9/3/10 and Timp-1/2 was determined in the uterus of C57BL/6 mice (n = 6/group) during pregnancy (on days (d) 5, 8, 12, 15, 17, and 18), at normal labor, and during lipopolysaccharide-induced PTL (n = 6/group). The expression of MMP-10 and TIMP-1 was determined in human term and preterm myometrium before the onset of labor (TNL, n = 7; PTNL, n = 7) and during active labor (TL, n = 8; PTL, n = 8). Gene expression and tissue localization were assessed by quantitative polymerase chain reaction and immunohistochemistry, respectively. RESULTS: Mmp-10 was higher during murine labor (53-fold vs early pregnancy) in contrast to Mmp-2/3/9 and Timp-1, the expression of which reached a nadir at labor ( P < .001 vs d5 [ Mmp-2/ 9] or P < .05 vs d8 [ Mmp-3 and Timp-1]). The Mmp-3/10 and Timp-1 were localized to the uterine epithelium and stroma/myometrium. In the human myometrium, TIMP-1 messenger RNA was higher and MMP-10 was lower in TL versus TNL ( P < .05), PTL ( P < .001), and PTNL ( P < .001). MMP-10 and TIMP-1 were localized to the myometrial smooth muscle cells, interstitial fibroblasts, and inflammatory cells. CONCLUSIONS: These data implicate MMP-3, TIMP-1, and MMP-10 in the uterine ECM remodeling during physiological and pathological parturition.
Assuntos
Trabalho de Parto , Metaloproteinases da Matriz/metabolismo , Miométrio/metabolismo , Trabalho de Parto Prematuro , Gravidez , Útero/metabolismo , Animais , Feminino , Humanos , Inflamação/induzido quimicamente , Inflamação/metabolismo , Lipopolissacarídeos/administração & dosagem , Metaloproteinase 10 da Matriz/metabolismo , Metaloproteinase 3 da Matriz/metabolismo , Camundongos Endogâmicos C57BL , RNA Mensageiro/metabolismo , Inibidor Tecidual de Metaloproteinase-1/metabolismoRESUMO
Preeclampsia (PE) (gestational proteinuric hypertension) is the leading cause of maternal and perinatal mortality worldwide. Although placental endothelial dysfunction and oxidative stress are known to contribute to PE, the exact pathological basis for this disorder remains unclear. Previously, we demonstrated that DNA damage at the maternal-fetal interface is more common in the placentas of women with PE than normotensive controls. In this study, we utilized an in vivo comparative study, including 20 preeclamptic women and 8 healthy control subjects, and an in vitro hypoxia/reperfusion model to mimic the effects of oxidative stress at the maternal-fetal interface. We tracked the spatial pattern of expression of 2 base excision repair proteins, 8-oxoguanine glycosylase (OGG1) and apurinic/apyrimidinic endonuclease-1 (APE1), at the maternal-fetal interface in response to oxidative stress. In vivo, we found a significant increase in OGG1 and APE1 concentrations in PE placental tissues as compared to normotensive controls ( P < .0001). Further, our in vitro study revealed that OGG1 and APE1 expression is much greater in maternal cells (decidua) than in fetal cells (cytotrophoblasts) of placental tissue subjected to oxidative stress ( P < .0001). Our results suggest that OGG1 and APE1 likely protect decidual cells from oxidative base damage.
Assuntos
Reparo do DNA/genética , Estresse Oxidativo/genética , Placenta/metabolismo , Pré-Eclâmpsia/genética , DNA Glicosilases/genética , DNA Glicosilases/metabolismo , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Feminino , Humanos , Pré-Eclâmpsia/metabolismo , Gravidez , Trofoblastos/metabolismoRESUMO
OBJECTIVE: Adenomyosis is a uterine disorder characterized by dysmenorrhea, dyspareunia, abnormal uterine bleeding, and infertility. Pathogenesis indicates that endometrial cells invade and proliferate within myometrium, and inflammatory mediators participate to the intense painful symptoms. The aim of the present study was to investigate the messenger RNA (mRNA) and protein expression of inflammatory (interleukin 1ß [IL-1ß], corticotropin-releasing hormone [CRH], urocortin [Ucn]) and neurogenic (nerve growth factors [NGFs], synaptophysin [SYN], microtubule-associated protein 2 [MAP2]) factors in adenomyotic nodules. MATERIALS AND METHODS: This prospective study enrolled 16 women, 8 women with nodular adenomyosis and 8 control women undergoing to hysterectomy. Specimens from adenomyotic nodules and eutopic endometrium were collected after surgery. Endometrial tissue was also obtained from the control group and also used for preparing primary culture of human endometrial stromal cells (HESCs). Messenger RNA expression of inflammatory mediators (IL-1ß, CRH, and Ucn) and neurogenic factors (NGF, SYN, and MAP2) was analyzed by real-time polymerase chain reaction. The in vitro effects of CRH/Ucn on NGF or SYN mRNA expression were also investigated. RESULTS: Adenomyotic nodules highly expressed IL-1ß, CRH, and Ucn mRNAs, as well as NGF, SYN, and MAP2 mRNAs ( P < .001 vs eutopic endometrium and control). Endometrium of women with adenomyosis showed high expression of IL-1ß and CRH ( P < .001 vs control). Protein expression of CRH, NGF, and SYN in adenomyotic nodules was confirmed by immunohistochemical and immunofluorescence analyses. Urocortin increased NGF mRNA expression in cultured HESCs. CONCLUSION: The present study showed that adenomyotic nodules are novel site of expression of inflammatory and neurogenic factors, probably involved in the pathogenesis of adenomyosis.
Assuntos
Adenomiose/metabolismo , Hormônio Liberador da Corticotropina/metabolismo , Interleucina-1beta/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Fatores de Crescimento Neural/metabolismo , Sinaptofisina/metabolismo , Urocortinas/metabolismo , Útero/metabolismo , Adenomiose/genética , Hormônio Liberador da Corticotropina/genética , Endométrio/metabolismo , Feminino , Humanos , Interleucina-1beta/genética , Proteínas Associadas aos Microtúbulos/genética , Miométrio/metabolismo , Fatores de Crescimento Neural/genética , Neurogênese/fisiologia , Sinaptofisina/genética , Urocortinas/genéticaRESUMO
BACKGROUND: Adenomyosis was found to have negative impacts on embryo implantation. Leukemia inhibitory factor (LIF), proposed to be a molecular marker for endometrial receptivity, works through the LIF receptor (LIFR) on both the embryo and the endometrium. We aimed to evaluate the endometrial expression of LIF and LIFR and its subsequent signaling in patients with adenomyosis during the window of implantation (WOI). METHODS: Endometrium was obtained during the WOI from patients with adenomyosis (age <45 years) who underwent hysterectomy and from age-matched controls who had no endometriosis or adenomyosis. The LIF and LIFR expressions were measured by polymerase chain reaction for messenger RNA expression, immunohistochemistry for protein intensity and localization, and immunofluorescent staining for colocalization. The ratio of signal transducer and activator of transcription 3 (STAT3) to extracellular signal-regulated kinase (ERK) phosphorylation was measured by Western blot of both the endometrium and the isolated human endometrial stromal cells (ESCs). RESULTS: Patients with adenomyosis showed significantly and parallelly reduced LIF and LIFR expressions in the eutopic endometrium during WOI as compared with the control women and subsequently with remarkably reduced activation of STAT3 and ERK signaling. The significantly increased STAT3 and ERK phosphorylation induced by the LIF treatment in the cultured ESCs supported the linkage between the LIF-LIFR reaction and the signaling cascade. CONCLUSION: Significant reduction in LIFR expression and the reduced activation of subsequent signaling strongly suggest a working model of how the implantation markers, LIF, may affect the endometrium of patients with adenomyosis. These molecular changes supported the declined implantation rates reported in patients with adenomyosis.
Assuntos
Adenomiose/metabolismo , Implantação do Embrião/fisiologia , Endométrio/metabolismo , Receptores de OSM-LIF/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/fisiologia , Fosfatos de Dinucleosídeos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Humanos , Fosforilação/fisiologia , Células Estromais/metabolismoRESUMO
Macrophage migration inhibitory factor (MIF) is a widely expressed cytokine involved in various biological processes. Although MIF's functions in cancer have not been completely elucidated, its expression has usually been correlated with tumour progression and aggressiveness, and it is currently discussed as a new promising target for novel therapies. Recent studies seem to confirm its active role in melanoma pathobiology; however, its expression has not yet been extensively studied in melanocytic tumours. We evaluated MIF protein expression in 126 skin lesions, including benign and atypical nevi, melanoma and melanoma metastases. In 55 cases, we also assessed MIF mRNA expression by real-time RT-PCR. Benign nevi were subdivided into nevocytic and Spitz/blue types; and melanomas into the radial, and vertical growth phase. A strong cytoplasmic MIF positivity was found in most samples, although it was more heterogeneous in malignant tumours; MIF nuclear expression characterized Spitz/blue nevi, atypical nevi, melanomas and metastases. All samples expressed MIF mRNA but it was significantly lower in benign nevi vs atypical nevi, melanomas and metastases (p=0.001; p<0.0001; p=0.002, respectively). Our study shows a widespread distribution of MIF among melanocytic tumours. Whereas we observed a trend towards higher expression levels of mRNA in atypical and malignant tumours, MIF protein was highly expressed in all lesions, although limited to the cytoplasm in most benign nevi. These observations suggest differences in MIF protein storage, subcellular location and properties in most benign nevi vs atypical and malignant tumours that should be confirmed by further investigation and correlation with clinical outcome.
Assuntos
Fatores Inibidores da Migração de Macrófagos/metabolismo , Melanoma/metabolismo , Nevo Pigmentado/metabolismo , RNA Mensageiro/metabolismo , Neoplasias Cutâneas/metabolismo , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Humanos , Imuno-Histoquímica , Fatores Inibidores da Migração de Macrófagos/genética , Melanoma/patologia , Metástase Neoplásica , Nevo Pigmentado/patologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Neoplasias Cutâneas/patologiaRESUMO
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
OBJECTIVE: To evaluate the expression pattern of activins and related growth factor messenger RNA (mRNA) levels in adenomyotic nodules and in their endometrium. DESIGN: Prospective study. SETTING: University hospital. PATIENT(S): Symptomatic premenopausal women scheduled to undergo hysterectomy for adenomyosis. INTERVENTION(S): Samples from adenomyotic nodules and homologous endometria were collected. Endometrial tissue was also obtained from a control group. MAIN OUTCOME MEASURE(S): Quantitative real-time polymerase chain reaction (PCR) analysis and immunohistochemical localization of activin-related growth factors (activin A, activin B, and myostatin), binding protein (follistatin), antagonists (inhibin-α, cripto), and receptors (ActRIIa, ActRIIb) were performed. RESULT(S): Myostatin mRNA levels in adenomyotic nodule were higher than in eutopic endometrium and myostatin, activin A, and follistatin concentrations were higher than in control endometrium. No difference was observed for inhibin-α, activin B, and cripto mRNA levels. Increased mRNA levels of ActRIIa and ActRIIb were observed in adenomyotic nodules compared with eutopic endometrium and control endometrium. Immunofluorescent staining for myostatin and follistatin confirmed higher protein expression in both glands and stroma of patients with adenomyosis than in controls. CONCLUSION(S): The present study showed for the first time that adenomyotic tissues express high levels of myostatin, follistatin, and activin A (growth factors involved in proliferation, apoptosis, and angiogenesis). Increased expression of their receptors supports the hypothesis of a possible local effect of these growth factors in adenomyosis. The augmented expression of ActRIIa, ActRIIb, and follistatin in the endometrium of these patients may play a role in adenomyosis-related infertility.
Assuntos
Receptores de Activinas Tipo II/análise , Adenomiose/metabolismo , Endométrio/química , Folistatina/análise , Miostatina/análise , Receptores de Activinas Tipo II/genética , Ativinas/análise , Adenomiose/genética , Adenomiose/cirurgia , Adulto , Estudos de Casos e Controles , Endométrio/cirurgia , Feminino , Folistatina/genética , Humanos , Imuno-Histoquímica , Miostatina/genética , Estudos Prospectivos , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Aberrations of genes/proteins regulating cell cycle and growth, increased proliferation and telomerase activity (TA) are documentable in glioblastoma multiforme. TA is more frequently detectable in secondary glioblastoma, which is also characterized by p53 mutation/overexpression. Discordant telomere (Te) length values have been reported in glioblastomas with and without TA. In 31 glioblastomas, in which pre-existing astrocytoma was not documented, we compared cases with and without TA for the expression of p53, EGFR, c-Myc, MIB-1 and Topoisomerase IIalpha; p53 mutations were also investigated by SSCP-PCR. Correlations were made with Te parameters [TePs: number (TeNo), length and area] as evaluated by image analysis in interphase nuclei of fluorescence in situ hybridization (FISH)-processed sections. We found no differences in the expression of the proteins evaluated and in TePs, except Te/nuclear area %, which was significantly lower in TA+ cases (p=0.02). TePs were, instead, inversely correlated with TA (p=0.0001). TA was positively correlated with MIB1 staining index in the TA+ cases (p=0.033), which also showed a positive correlation between TeNo and EGFR expression (p=0.042), and a trend towards a negative correlation between TeNo and p53 expression (p=0.05). Tumors overexpressing EGFR had a significantly shorter lifetime (p=0.0001). TeNo seems to be inversely correlated to tumor proliferation and lifetime in glioblastoma multiforme.
Assuntos
Glioblastoma/enzimologia , Hibridização in Situ Fluorescente/métodos , Telomerase/metabolismo , Telômero/ultraestrutura , Fosfatase Ácida/metabolismo , Adolescente , Adulto , Antígenos de Neoplasias , Neoplasias Encefálicas/enzimologia , Divisão Celular , Criança , DNA Topoisomerases Tipo II/metabolismo , Proteínas de Ligação a DNA , Receptores ErbB/metabolismo , Glioblastoma/metabolismo , Glioblastoma/patologia , Humanos , Processamento de Imagem Assistida por Computador , Isoenzimas/metabolismo , Antígeno Ki-67/biossíntese , Antígeno Ki-67/metabolismo , Pessoa de Meia-Idade , Polimorfismo Conformacional de Fita Simples , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fosfatase Ácida Resistente a Tartarato , Proteína Supressora de Tumor p53/metabolismoRESUMO
OBJECTIVE: To evaluate antimüllerian hormone (AMH) and AMH receptor II (AMHRII) mRNA and protein expression in endometrium and in ovarian or deep lesions of women with endometriosis. DESIGN: Prospective study. SETTING: University hospitals in Italy and Brazil. PATIENTS: Patients with endometriosis (n = 55) and healthy women (n = 45). INTERVENTIONS: Specimens of endometrium obtained by hysteroscopy from patients with endometriosis and from healthy control subjects; specimens of ovarian endometriosis (n = 29) or of deep endometriosis (n = 26) were collected by laparoscopy. Serum samples were collected in some endometriotic patients (n = 23) and healthy control subjects (n = 20). MAIN OUTCOME MEASURE(S): AMH and AMHRII mRNA levels were evaluated by quantitative reverse-transcription polymerase chain reaction and protein localization by immunohistochemistry. AMH levels in tissue homogenates and in serum were assessed by ELISA. RESULT(S): Endometrium from women with endometriosis showed higher AMH and AMHRII mRNA levels than control women, with no significant differences between proliferative and secretory phases. Specimens collected from ovarian or deep endometriosis showed the highest AMH and AMHRII mRNA expression. Immunolocalization study confirmed the high AMH and AMHRII protein expression in endometriotic lesions. No difference of serum AMH levels between the groups was found. CONCLUSION(S): The increased AMH and AMHRII mRNA and protein expression in endometrium and in endometriotic lesions suggests a possible involvement of AMH in endometriosis.
Assuntos
Hormônio Antimülleriano/biossíntese , Endometriose/diagnóstico , Endometriose/metabolismo , Regulação da Expressão Gênica , Receptores de Peptídeos/biossíntese , Receptores de Fatores de Crescimento Transformadores beta/biossíntese , Adulto , Hormônio Antimülleriano/sangue , Biomarcadores/sangue , Biomarcadores/metabolismo , Endometriose/cirurgia , Endométrio/metabolismo , Endométrio/patologia , Feminino , Humanos , Estudos Prospectivos , Receptores de Peptídeos/sangue , Receptores de Fatores de Crescimento Transformadores beta/sangue , Adulto JovemRESUMO
The present study investigated expression and protein localization of FOXL2 messenger RNA (mRNA) in endometrium of healthy women and in patients with endometriosis during endometrial cycle. In endometriotic lesions, FOXL2 mRNA and protein were evaluated and a possible correlation with activin A mRNA expression changes was also studied. Endometrium was collected from healthy women (n = 52) and from women with endometriosis (n = 31) by hysteroscopy; endometriotic tissues were collected by laparoscopy (n = 38). FOXL2 gene expression analysis in endometrium of healthy women showed a significant expression and no significant changes in mRNA levels between proliferative and secretory phases; a similar pattern was observed in endometrium of patients with endometriosis. Immunohistochemical evaluation showed that FOXL2 protein localized in stromal and glandular cells and colocalized with SUMO-1. FOXL2 mRNA expression was 3-fold higher in endometriosis than in healthy endometrium (P < .01) and a positive correlation between FOXL2 and activin A mRNA was found (P < .05) in endometriosis. In conclusion, FOXL2 mRNA expression and its protein localization do not change during endometrial cycle in eutopic endometrium from healthy individuals or patients with endometriosis; the hyperexpression of FOXL2 in endometriotic lesions suggests an involvement of this transcriptional regulator, probably associated with activin A expression and related to the pathogenesis of endometriosis.