RESUMO
Ectopic pregnancy is the major cause of maternal morbidity and mortality in the first trimester of pregnancy. Tubal ectopic pregnancy (TEP) accounts for nearly 98% of all ectopic pregnancies. TEP is usually associated with salpingitis but the underlying mechanism in salpingitis leading to TEP remains unclear. Adrenomedullin (ADM) is a peptide hormone abundantly expressed in the fallopian tube with potent anti-inflammatory activities. Its expression peaks at the early luteal phase when the developing embryo is being transported through the fallopian tube. In the present study, we demonstrated reduced expression of ADM in fallopian tubes of patients with salpingitis and TEP. Using macrophages isolated from the fallopian tubes of these women, our data revealed that the salpingistis-associated ADM reduction contributed to aggravated pro-inflammatory responses of the tubal macrophages resulting in production of pro-inflammatory and pro-implantation cytokines IL-6 and IL-8. These cytokines activated the expression of implantation-associated molecules and Wnt signaling pathway predisposing the tubal epithelium to an adhesive and receptive state for embryo implantation. In conclusion, this study provided evidence for the role of ADM in the pathogenesis of TEP through regulating the functions of tubal macrophages.
Assuntos
Adrenomedulina/metabolismo , Tubas Uterinas/imunologia , Tubas Uterinas/metabolismo , Ativação de Macrófagos/imunologia , Macrófagos/imunologia , Macrófagos/metabolismo , Gravidez Ectópica/etiologia , Adrenomedulina/sangue , Adrenomedulina/deficiência , Adrenomedulina/genética , Adulto , Biomarcadores , Linhagem Celular , Plasticidade Celular/genética , Plasticidade Celular/imunologia , Citocinas/metabolismo , Suscetibilidade a Doenças , Implantação do Embrião/genética , Implantação do Embrião/imunologia , Epitélio/metabolismo , Tubas Uterinas/patologia , Feminino , Expressão Gênica , Humanos , Imuno-Histoquímica , Imunofenotipagem , Pessoa de Meia-Idade , NF-kappa B/metabolismo , Gravidez , Gravidez Ectópica/metabolismo , Gravidez Ectópica/patologia , Receptores de Adrenomedulina/genética , Receptores de Adrenomedulina/metabolismo , Salpingite/complicações , Salpingite/etiologia , Salpingite/metabolismo , Salpingite/patologia , Transdução de SinaisRESUMO
Tumor metastasis is a primary source of morbidity and mortality in cancer. Adrenomedullin (AM) is a multifunctional peptide regulated by receptor activity-modifying proteins (RAMPs). We previously reported that the AM-RAMP2 system is involved in tumor angiogenesis, but the function of the AM-RAMP3 system remains largely unknown. Here, we investigated the actions of the AM-RAMP2 and 3 systems in the tumor microenvironment and their impact on metastasis. PAN02 pancreatic cancer cells were injected into the spleens of mice, leading to spontaneous liver metastasis. Tumor metastasis was enhanced in vascular endothelial cell-specific RAMP2 knockout mice (DI-E-RAMP2-/-). By contrast, metastasis was suppressed in RAMP3-/- mice, where the number of podoplanin (PDPN)-positive cancer-associated fibroblasts (CAFs) was reduced in the periphery of tumors at metastatic sites. Because PDPN-positive CAFs are a hallmark of tumor malignancy, we assessed the regulation of PDPN and found that Src/Cas/PDPN signaling is mediated by RAMP3. In fact, RAMP3 deficiency CAFs suppressed migration, proliferation, and metastasis in co-cultures with tumor cells in vitro and in vivo. Moreover, the activation of RAMP2 in RAMP3-/- mice suppressed both tumor growth and metastasis. Based on these results, we suggest that the upregulation of PDPN in DI-E-RAMP2-/- mice increases malignancy, while the downregulation of PDPN in RAMP3-/- mice reduces it. Selective activation of RAMP2 and inhibition of RAMP3 would therefore be expected to suppress tumor metastasis. This study provides the first evidence that understanding and targeting to AM-RAMP systems could contribute to the development of novel therapeutics against metastasis.
Assuntos
Adrenomedulina/deficiência , Fibroblastos Associados a Câncer/química , Neoplasias Hepáticas Experimentais/prevenção & controle , Glicoproteínas de Membrana/metabolismo , Neoplasias Pancreáticas/prevenção & controle , Proteína 3 Modificadora da Atividade de Receptores/fisiologia , Animais , Células Cultivadas , Neoplasias Hepáticas Experimentais/metabolismo , Neoplasias Hepáticas Experimentais/secundário , Masculino , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Metástase Neoplásica , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologiaRESUMO
Adrenomedullin (AM) is a vasodilating peptide involved in the regulation of circulatory homeostasis and in the pathophysiology of certain cardiovascular diseases. AM plays critical roles in blood vessels, including regulation of vascular stability and permeability. To elucidate the autocrine/paracrine function of AM in endothelial cells (EC) in vivo, a conditional knockout of AM in EC (AM(EC-KO)) was used. The amount of vascularization of the matrigel implants was lower in AM(EC-KO) mice indicating a defective angiogenesis. Moreover, ablation of AM in EC revealed increased vascular permeability in comparison with wild type (WT) littermates. In addition, AM(EC-KO) lungs exhibited significantly less tumor growth than littermate WT mice using a syngeneic model of metastasis. Furthermore, following middle cerebral artery permanent occlusion, there was a significant infarct size decrease in animals lacking endothelial AM when compared to their WT counterparts. AM is an important regulator of EC function, angiogenesis, tumorigenesis, and brain response to ischemia. Studies of AM should bring novel approaches to the treatment of vascular diseases.
Assuntos
Adrenomedulina/deficiência , Encéfalo/patologia , Permeabilidade Capilar , Células Endoteliais/metabolismo , Neoplasias Pulmonares/secundário , Neovascularização Fisiológica , Adrenomedulina/metabolismo , Animais , Pressão Sanguínea , Isquemia Encefálica/complicações , Isquemia Encefálica/patologia , Movimento Celular , Proliferação de Células , Regulação da Expressão Gênica , Infarto da Artéria Cerebral Média/complicações , Infarto da Artéria Cerebral Média/patologia , Camundongos KnockoutRESUMO
Bronchopulmonary dysplasia (BPD) is a chronic lung disease of premature infants that is characterized by alveolar simplification and decreased lung angiogenesis. Hyperoxia-induced oxidative stress and inflammation contributes to the development of BPD in premature infants. Adrenomedullin (AM) is an endogenous peptide with potent angiogenic, anti-oxidant, and anti-inflammatory properties. Whether AM regulates hyperoxic injury in fetal primary human lung cells is unknown. Therefore, we tested the hypothesis that AM-deficient fetal primary human pulmonary microvascular endothelial cells (HPMEC) will have increased oxidative stress, inflammation, and cytotoxicity compared to AM-sufficient HPMEC upon exposure to hyperoxia. Adrenomedullin gene (Adm) was knocked down in HPMEC by siRNA-mediated transfection and the resultant AM-sufficient and -deficient cells were evaluated for hyperoxia-induced oxidative stress, inflammation, cytotoxicity, and Akt activation. AM-deficient HPMEC had significantly increased hyperoxia-induced reactive oxygen species (ROS) generation and cytotoxicity compared to AM-sufficient HPMEC. Additionally, AM-deficient cell culture supernatants had increased macrophage inflammatory protein 1α and 1ß, indicating a heightened inflammatory state. Interestingly, AM deficiency was associated with an abrogated Akt activation upon exposure to hyperoxia. These findings support the hypothesis that AM deficiency potentiates hyperoxic injury in primary human fetal HPMEC via mechanisms entailing Akt activation.
Assuntos
Adrenomedulina/deficiência , Células Endoteliais/metabolismo , Hiperóxia/etiologia , Pulmão/irrigação sanguínea , Adrenomedulina/genética , Adrenomedulina/metabolismo , Displasia Broncopulmonar/etiologia , Displasia Broncopulmonar/metabolismo , Displasia Broncopulmonar/patologia , Morte Celular , Células Cultivadas , Células Endoteliais/patologia , Feto/irrigação sanguínea , Feto/metabolismo , Técnicas de Silenciamento de Genes , Humanos , Hiperóxia/metabolismo , Hiperóxia/patologia , Recém-Nascido , Pulmão/metabolismo , Lesão Pulmonar/genética , Lesão Pulmonar/metabolismo , Lesão Pulmonar/patologia , Estresse Oxidativo , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
Adrenomedullin was originally isolated from pheochromocytoma cells and reduces insulin resistance by decreasing oxidative stress. White matter lesions induced by aging and hyperglycemia play a crucial role in cognitive impairment in poststroke patients. Here, we examine whether adrenomedullin deficiency and aging exacerbate ischemic white matter injury after prolonged cerebral hypoperfusion. Adrenomedullin heterozygous, wild-type young/aged mice were subjected to prolonged hypoperfusion. Prolonged cerebral hypoperfusion followed by immunohistochemical analysis was used to evaluate white matter injury. After prolonged hypoperfusion, white matter damage progressed in a time-dependent manner in AM(+/-) group compared with the wild-type group. The number of oligodendrocyte progenitor cells gradually increased after prolonged hypoperfusion, whereas oligodendrocytes decreased following a transient increase, but the ratio of increase was mild in the AM(+/-) group (P < 0.05). Oxidative stress was detected in oligodendrocytes, with a larger increase in the AM(+/-) group (P < 0.05). Aged mice showed the same tendency, but white matter damage was worse, especially in the aged AM(+/-) group. Our results demonstrated that white matter injury was increased in adrenomedullin deficiency, which induced oxidative stress. White matter injury was more exacerbated because of hyperglycemia in aged AM(+/-) group. Adrenomedullin may be an important target in the control of ischemic white matter injury.
Assuntos
Adrenomedulina/deficiência , Envelhecimento/metabolismo , Hipóxia-Isquemia Encefálica/metabolismo , Células-Tronco Neurais/metabolismo , Oligodendroglia/metabolismo , Substância Branca/lesões , Adrenomedulina/genética , Envelhecimento/genética , Animais , Doença Crônica , Hipóxia-Isquemia Encefálica/patologia , Masculino , Camundongos , Camundongos Mutantes , Células-Tronco Neurais/patologia , Oligodendroglia/patologia , Substância Branca/metabolismo , Substância Branca/patologiaRESUMO
Intracellular cargo transport relies on microtubules and motor proteins such as kinesins and dyneins. Currently we have ample knowledge of the mechanisms by which motor proteins propel themselves along the microtubules, but little is known about intracellular factors that regulate motor speed. Here we show that proadrenomedullin N-terminal 20 peptide (PAMP) increases kinesin velocity and ATP consumption in a dose-dependent manner, using a variety of human kinesins. Structure-activity studies found that the terminal amide of PAMP is required for modulating kinesin activity and that the smallest peptide fragment retaining this role is PAMP12â20. On the other hand, peptide fragments as small as PAMP18â20 maintained the ability of delaying tubulin polymerization, another function previously described for PAMP, indicating that these two activities depend on different regions of the molecule. To demonstrate that these observations are also relevant in vivo, hippocampal neurons were isolated from mice lacking the gene coding for PAMP and from wild type littermates. Intravital stains followed by time-lapse microscopy analysis revealed that mitochondrial speed inside neurons lacking PAMP was significantly slower than in cells expressing the peptide. External addition of synthetic PAMP reversed this phenotype in PAMP-null neurons. Besides the obvious implications for better understanding cell biology, these results may be also relevant for the rapidly evolving discipline of nanotechnology because PAMP may be used as an accelerator of nanodevices based on microtubules and motor proteins.
Assuntos
Adrenomedulina/farmacologia , Adrenomedulina/fisiologia , Cinesinas/efeitos dos fármacos , Cinesinas/fisiologia , Adenosina Trifosfatases/efeitos dos fármacos , Adenosina Trifosfatases/metabolismo , Adrenomedulina/deficiência , Animais , Relação Dose-Resposta a Droga , Hipocampo/citologia , Humanos , Técnicas In Vitro , Camundongos , Camundongos Knockout , Microtúbulos/efeitos dos fármacos , Microtúbulos/fisiologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/fisiologia , Modelos Animais , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , PolimerizaçãoRESUMO
This study was designed to examine the effect of adrenomedullin deficiency on cerebral infarction and the relationship between adrenomedullin and cyclic AMP-protein kinase A pathway in regulating reactive oxygen species (ROS). Adrenomedullin heterozygous and wild-type mice were subjected to 60-mins focal ischemia. We used adrenomedullin heterozygous mice because adrenomedullin homozygotes die in utero. Infarct volume, neurologic deficit scores, and immunohistochemical analyses were evaluated at several time points after ischemia. The infarct volume and neurologic deficit scores were significantly worse in adrenomedullin heterozygous mice. Significant accumulation of inducible nitric oxide, oxidative DNA damage, and lipid peroxidation was noted after reperfusion in adrenomedullin heterozygous mice. Treatment of wild-type mice with H89, a protein kinase A inhibitor, resulted in increased infarct size, and worsening of neurologic deficit score and other parameters to levels comparable to those of adrenomedullin heterozygous mice. In contrast, cilostazol, which increases cyclic AMP, rescued neurologic deficit and ROS accumulation in adrenomedullin heterozygous mice. This study showed that adrenomedullin downregulation results in increase in ROS after transient focal ischemia in mice. The results also indicated that adrenomedullin has an important function against ischemic injury through the cyclic AMP-protein kinase A pathway.