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Chronic inflammation with progressive age, called inflammaging, contributes to the pathogenesis of cardiovascular diseases. Previously, we have shown increased vascular expression of the Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) in aged mice and humans, presumably via mutual upregulation with the pro-inflammatory cytokine TNF-α. CEACAM1 is critical for aging-associated vascular alterations like endothelial dysfunction, fibrosis, oxidative stress, and sustained inflammation and can be regarded as a main contributor to vascular inflammaging. This study was conducted to elucidate the mechanisms underlying endothelial CEACAM1 upregulation by TNF-α in detail. Using wildtype (WT) and TNF-α knockout (Tnf-/-) mice, we confirmed that the aging-related upregulation of endothelial CEACAM1 critically depends on TNF-α. The underlying mechanisms were analyzed in an endothelial cell culture model. TNF-α time-dependently upregulated CEACAM1 in vitro. In pharmacological experiments, we identified an early NF-κB- and a delayed ß-catenin-mediated response. Involvement of ß-catenin was further substantiated by siRNA-mediated knockdown of the ß-catenin-targeted transcription factor TCF4. Both signaling pathways acted independent from each other. Elucidating the delayed response, co-immunoprecipitation analysis revealed release of ß-catenin from adherens junctions by TNF-α. Finally, TNF-α activated Akt kinase by increasing its Ser473 phosphorylation. Consequently, Akt kinase facilitated ß-catenin signaling by inhibiting its degradation via phosphorylation of GSK3ß at Ser9 and by increased phosphorylation of ß-catenin at Ser552 that augments its transcriptional activity. Taken together, our study provides novel mechanistic insights into the aging-related, inflammation-mediated endothelial upregulation of CEACAM1. Beyond the pathogenesis of cardiovascular diseases, these findings may be significant to all fields of inflammaging.
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Cancer is a disease characterized by the acquisition of a multitude of unique traits. It has long been understood that cancer cells divert significantly from normal cell metabolism. The most obvious of metabolic changes is that cancer cells strongly rely on glucose conversion by aerobic glycolysis. In addition, they also regularly develop mechanisms to use lipids and fatty acids for their energy needs. Peroxisomes lie central to these adaptive changes of lipid metabolism. Peroxisomes are metabolic organelles that take part in over 50 enzymatic reactions crucial for cellular functioning. Thus, they are essential for an effective and comprehensive use of lipids' energy supplied to cells. Cancer cells display a substantial increase in the biogenesis of peroxisomes and an increased expression of proteins necessary for the enzymatic functions provided by peroxisomes. Moreover, the enzymatic conversion of FAs in peroxisomes is a significant source of reactive oxygen and nitrogen species (ROS/RNS) that strongly impact cancer malignancy. Important regulators in peroxisomal FA oxidation and ROS/RNS generation are the transcription factors of the peroxisome proliferator-activated receptor (PPAR) family. This review describes the metabolic changes in tumorigenesis and cancer progression influenced by peroxisomes. We will highlight the ambivalent role that peroxisomes and PPARs play in the different stages of tumor development and summarize our current understanding of how to capitalize on the comprehension of peroxisomal biology for cancer treatment.
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Pathological angiogenesis promotes tumor growth, metastasis, and atherosclerotic plaque rupture. Macrophages are key players in these processes. However, whether these macrophages differentiate from bone marrow-derived monocytes or from local vascular wall-resident stem and progenitor cells (VW-SCs) is an unresolved issue of angiogenesis. To answer this question, we analyzed vascular sprouting and alterations in aortic cell populations in mouse aortic ring assays (ARA). ARA culture leads to the generation of large numbers of macrophages, especially within the aortic adventitia. Using immunohistochemical fate-mapping and genetic in vivo-labeling approaches we show that 60% of these macrophages differentiate from bone marrow-independent Ly6c+/Sca-1+ adventitial progenitor cells. Analysis of the NCX-/- mouse model that genetically lacks embryonic circulation and yolk sac perfusion indicates that at least some of those progenitor cells arise yolk sac-independent. Macrophages represent the main source of VEGF in ARA that vice versa promotes the generation of additional macrophages thereby creating a pro-angiogenetic feedforward loop. Additionally, macrophage-derived VEGF activates CD34+ progenitor cells within the adventitial vasculogenic zone to differentiate into CD31+ endothelial cells. Consequently, depletion of macrophages and VEGFR2 antagonism drastically reduce vascular sprouting activity in ARA. In summary, we show that angiogenic activation induces differentiation of macrophages from bone marrow-derived as well as from bone marrow-independent VW-SCs. The latter ones are at least partially yolk sac-independent, too. Those VW-SC-derived macrophages critically contribute to angiogenesis, making them an attractive target to interfere with pathological angiogenesis in cancer and atherosclerosis as well as with regenerative angiogenesis in ischemic cardiovascular disorders.
Assuntos
Túnica Adventícia , Células Endoteliais , Túnica Adventícia/patologia , Animais , Medula Óssea/patologia , Células Endoteliais/patologia , Macrófagos/patologia , Camundongos , Neovascularização Patológica/patologia , Células-Tronco/patologia , Fator A de Crescimento do Endotélio VascularRESUMO
Aging is an independent risk factor for cardiovascular diseases and therefore of particular interest for the prevention of cardiovascular events. However, the mechanisms underlying vascular aging are not well understood. Since carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is crucially involved in vascular homeostasis, we sought to identify the role of CEACAM1 in vascular aging. Using human internal thoracic artery and murine aorta, we show that CEACAM1 is upregulated in the course of vascular aging. Further analyses demonstrated that TNF-α is CEACAM1-dependently upregulated in the aging vasculature. Vice versa, TNF-α induces CEACAM1 expression. This results in a feed-forward loop in the aging vasculature that maintains a chronic pro-inflammatory milieu. Furthermore, we demonstrate that age-associated vascular alterations, that is, increased oxidative stress and vascular fibrosis, due to increased medial collagen deposition crucially depend on the presence of CEACAM1. Additionally, age-dependent upregulation of vascular CEACAM1 expression contributes to endothelial barrier impairment, putatively via increased VEGF/VEGFR-2 signaling. Consequently, aging-related upregulation of vascular CEACAM1 expression results in endothelial dysfunction that may promote atherosclerotic plaque formation in the presence of additional risk factors. Our data suggest that CEACAM1 might represent an attractive target in order to delay physiological aging and therefore the transition to vascular disorders such as atherosclerosis.
Assuntos
Envelhecimento/metabolismo , Antígenos CD/metabolismo , Moléculas de Adesão Celular/metabolismo , Endotélio Vascular/metabolismo , Transdução de Sinais , Idoso , Animais , Antígenos CD/genética , Moléculas de Adesão Celular/genética , Células Cultivadas , Endotélio Vascular/patologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-IdadeRESUMO
Carcinoembryonic antigen-related cell adhesion molecule-1 (CEACAM1) is known to be crucial to vasculogenesis and angiogenesis. Recently, CEACAM1 deficiency was shown to result in the formation of aortic plaque-like lesions, indicating a role for CEACAM1 in adult vessels as well. The underlying mechanisms remained largely elusive. Therefore, we aimed to elucidate the role of CEACAM1 in endothelial homeostasis. Here, we show that CEACAM1 deficiency causes subcellular eNOS redistribution in endothelial cells ( i.e., by eNOS depalmitoylation) and alters endothelial glycocalyx that confers antiadhesive properties to the endothelium ( i.e., by repression of glycocalyx-degrading enzymes). Accordingly, our analysis revealed an increased leukocyte-endothelial interaction in CEACAM1-deficient endothelium. In addition, CEACAM1 age dependently modulated basal and TNF-α-mediated endothelial barrier (EB) leakiness. In younger mice, CEACAM1 was protective for EB, whereas in aged mice it promoted EB leakiness. EB function depends on interendothelial adherence junctions formed by ß-catenin/vascular endothelial-cadherin complexes. We show here that CEACAM1 influenced basal and TNF-α-mediated phosphorylation of ß-catenin and caveolin-1, which are essential players in EB modulation. Both increased adhesiveness to leukocytes and EB modulation due to CEACAM1 deficiency may facilitate inflammatory cell transmigration into the vascular wall and subsequent plaque formation. Collectively, these results identify a crucial role for CEACAM1 in endothelial homeostasis of adult blood vessels.-Ghavampour, S., Kleefeldt, F., Bömmel, H., Volland, J., Paus, A., Horst, A., Pfeiffer, V., Hübner, S., Wagner, N., Rueckschloss, U., Ergün, S. Endothelial barrier function is differentially regulated by CEACAM1-mediated signaling.
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Antígeno Carcinoembrionário/metabolismo , Células Endoteliais/metabolismo , Endotélio Vascular/metabolismo , Transdução de Sinais , Migração Transendotelial e Transepitelial , Animais , Caderinas/metabolismo , Caveolina 1/metabolismo , Células Endoteliais/patologia , Endotélio Vascular/patologia , Camundongos , Camundongos Knockout , Óxido Nítrico Sintase Tipo III/genética , Óxido Nítrico Sintase Tipo III/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
En-face fat staining is frequently used to visualize atherosclerotic lesions. This method, however, is not suitable to visualize endothelial barrier damage prior to microscopically detectable morphological alterations of the arterial wall such as sub-endothelial lipid deposition. To enable the investigation of early endothelial barrier damage and in particular the initial steps of atherosclerosis, a new method has to fulfill three requirements: (i) easy and fast to perform, (ii) low cost of applicability without requirement for highly sophisticated technical equipment, and (iii) reliable reproducibility of valid results. To this end, we used intracardial Evans blue dye injection after washout of blood and measured dye deposition within the aortic wall as a parameter of endothelial barrier leakiness, which is recognized as one of the earliest signs of atherosclerotic plaque formation. These analyses were performed in ApoE -/-, LDL receptor -/- and Cc1 -/- mouse models which have been reported to develop aortic plaques with or without high cholesterol diet. Our data show that sub-endothelial dye deposition is a reliable and reproducible readout parameter to assess endothelial barrier damage. Along these lines, measurements of aortic intima areas with Evans blue deposition in relation to total intima circumference enabled quantitative assessments of the results. Our technique enables the imaging of endothelial barrier damage prior to detectable aortic lipid deposition and plaque development. Thus, it will facilitate the detection of the initial vascular pathogenetic processes that lead to cardiovascular diseases. It will also enable the testing of new drugs and therapeutic procedures to prevent these disorders.
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Barreira Hematoencefálica/patologia , Células Endoteliais/patologia , Placa Aterosclerótica/patologia , Animais , Células Cultivadas , Camundongos , Camundongos KnockoutRESUMO
Ciliary neurotrophic factor (Cntf) plays an essential role in postnatal maintenance of spinal motoneurons. Whereas the expression of this neurotrophic factor is low during embryonic development, it is highly up-regulated after birth in myelinating Schwann cells of rodents. To characterize the underlying transcriptional mechanisms, we have analyzed and compared the effects of various glial transcription factors. In contrast to Pit-1, Oct-1, Unc-86 homology region (POU) domain class 3, transcription factor 1 (Oct6/SCIP/Tst-1) and paired box gene 3 (Pax3), SRY-box-containing gene 10 (Sox10) induces Cntf expression in Schwann cells. Subsequent promoter analysis using luciferase reporter gene and EMSA identified the corresponding response elements within the Cntf promoter. Overexpression of Sox10 in primary sciatic nerve Schwann cells leads to a >100-fold up-regulation of Cntf protein, and suppression of Sox10 by RNA interference in the spontaneously immortalized Schwann cell line 32 reduces Cntf expression by >80%. Mice with heterozygous inactivation of the Sox10 gene show significantly reduced Cntf protein levels in sciatic nerves, indicating that Sox10 is necessary and sufficient for regulating Cntf expression in the peripheral nervous system.
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Fator Neurotrófico Ciliar , Proteínas de Ligação a DNA/metabolismo , Regulação da Expressão Gênica , Proteínas de Grupo de Alta Mobilidade/metabolismo , Células de Schwann/fisiologia , Fatores de Transcrição/metabolismo , Animais , Sítios de Ligação , Linhagem Celular , Fator Neurotrófico Ciliar/genética , Fator Neurotrófico Ciliar/metabolismo , Proteínas de Ligação a DNA/genética , Genes Reporter , Proteínas de Grupo de Alta Mobilidade/genética , Camundongos , Regiões Promotoras Genéticas , Ratos , Fatores de Transcrição SOXE , Células de Schwann/citologia , Nervo Isquiático/citologia , Nervo Isquiático/metabolismo , Fatores de Transcrição/genética , Sítio de Iniciação de TranscriçãoRESUMO
Environmental pollutants can influence the expression of immunoregulatory molecules and, in this way, promote allergies. The local synthesis of proinflammatory chemokines is an important aspect in the development of allergic airway inflammation. We have characterized the influence of pyrene, a polycyclic aromatic hydrocarbon (PAH) contained, for example, in diesel exhaust particles (DEP), on transcription and secretion of the chemokines interleukin-8 (IL-8) and eotaxin. Reporter genes under control of the respective promoters were tested in the human cell lines A549 and HeLa, mRNA production was assayed in A549 cells and protein production was measured by ELISA in cell supernatants from primary human fibroblasts. Pyrene content of cell supernatants was measured by analytical HPLC. Promoter activity, mRNA production and protein expression of IL-8 were increased by pyrene. The activating effect in reporter gene studies was abolished by mutating either an NF-kappaB or an AP-1 binding site in the IL-8 promoter. In contrast, pyrene showed no effect on transcription from the eotaxin promoter, despite the important role of this chemokine in asthma. Our data show that pyrene has specific effects on chemokine synthesis, which are not restricted to mediators primarily associated with atopic diseases. Pyrene also affected cells not derived from lung tissue, which suggests a broader immunoregulatory influence for this pollutant.
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Poluentes Atmosféricos/toxicidade , Quimiocinas CC/biossíntese , Células Epiteliais/efeitos dos fármacos , Interleucina-8/biossíntese , Pirenos/toxicidade , Emissões de Veículos , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Quimiocina CCL11 , Quimiocinas CC/genética , Relação Dose-Resposta a Droga , Ensaio de Imunoadsorção Enzimática , Células Epiteliais/metabolismo , Expressão Gênica/efeitos dos fármacos , Genes Reporter/genética , Humanos , Interleucina-8/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Progressive motor neuronopathy (pmn) mutant mice have been widely used as a model for human motoneuron disease. Mice that are homozygous for the pmn gene defect appear healthy at birth but develop progressive motoneuron disease, resulting in severe skeletal muscle weakness and respiratory failure by postnatal week 3. The disease starts at the motor endplates, and then leads to axonal loss and finally to apoptosis of the corresponding cell bodies. We localized the genetic defect in pmn mice to a missense mutation in the tubulin-specific chaperone E (Tbce) gene on mouse chromosome 13. The human orthologue maps to chromosome 1q42.3. The Tbce gene encodes a protein (cofactor E) that is essential for the formation of primary alpha-tubulin and beta-tubulin heterodimeric complexes. Isolated motoneurons from pmn mutant mice exhibit shorter axons and axonal swelling with irregularly structured beta-tubulin and tau immunoreactivity. Thus, the pmn gene mutation provides the first genetic evidence that alterations in tubulin assembly lead to retrograde degeneration of motor axons, ultimately resulting in motoneuron cell death.
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Chaperonas Moleculares/genética , Doença dos Neurônios Motores/genética , Mutação de Sentido Incorreto , Sequência de Aminoácidos , Animais , Tamanho Celular , Sobrevivência Celular , Cromossomos Humanos Par 1 , Modelos Animais de Doenças , Embrião de Mamíferos/anatomia & histologia , Feminino , Genótipo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos , Chaperonas Moleculares/química , Chaperonas Moleculares/metabolismo , Dados de Sequência Molecular , Doença dos Neurônios Motores/metabolismo , Neurônios Motores/citologia , Neurônios Motores/fisiologia , Linhagem , Alinhamento de SequênciaRESUMO
Cellular functions induced by cytokine interleukin (IL)-4 and IL-4 signaling through signal transducer and activator of transcription (Stat)6 typify a Th2-type immune response. We investigated the inhibitor effect of the NFkappaB blocker parthenolide in the late-phase, Th2-type immune response. Parthenolide blocked by 90.6 +/- 7.4% the IL-4-induced expression of the endothelial vascular cell adhesion molecule (VCAM)-1, a hallmark of extravasation of very late antigen-4-positive leukocytes. The noncytotoxic concentrations of 10 microM parthenolide left a section of the IL-4 receptor signal transduction intact. Parthenolide did not interfere with the immediate IL-4-induced phosphorylation of endothelial Stat6 on its tyrosine residue Y641 and with tyrosine phosphorylation of the adapter molecule, Jak2-both processes are obligatory for dimerization and nuclear translocation of Stat6. But parthenolide inhibited the Stat6 DNA-binding activity in IL-4-stimulated endothelial cells and inhibited the IL-4-driven activation of a luciferase reporter gene under the control of Stat6-responsive elements (IC(50) 5.11 +/- 0.67 microM). Together, these data suggest an anti-chronic disease profile for the sesquiterpene lactone parthenolide.