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Idiopathic pulmonary fibrosis (IPF) is a progressive, irreversible respiratory disease with limited therapeutic options. A hallmark of IPF is excessive fibroblast activation and extracellular matrix (ECM) deposition. The resulting increase in tissue stiffness amplifies fibroblast activation and drives disease progression. Dampening stiffness-dependent activation of fibroblasts could slow disease progression. We performed an unbiased, next generation sequencing (NGS) screen to identify signaling pathways involved in stiffness-dependent lung fibroblast activation. Adipocytokine signaling was downregulated in primary lung fibroblasts (PFs) cultured on stiff matrices. Re-activating adipocytokine signaling with adiponectin suppressed stiffness-dependent activation of human PFs. Adiponectin signaling depended on CDH13 expression and p38 mitogen-activated protein kinase gamma (p38MAPKγ) activation. CDH13 expression and p38MAPKγ activation were strongly reduced in lungs from IPF donors. Our data suggest that adiponectin-signaling via CDH13 and p38MAPKγ activation suppresses pro-fibrotic activation of fibroblasts in the lung. Targeting of the adiponectin signaling cascade may provide therapeutic benefits in IPF.
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Idiopathic pulmonary fibrosis (IPF) is a fatal disease of the lower respiratory tract with restricted therapeutic options. Repetitive injury of the bronchoalveolar epithelium leads to activation of pulmonary fibroblasts, differentiation into myofibroblasts and excessive extracellular matrix (ECM) deposition resulting in aberrant wound repair. However, detailed molecular and cellular mechanisms underlying initiation and progression of fibrotic changes are still elusive. Here, we report the generation of a representative fibroblast reporter cell line (10-4A BFP ) to study pathophysiological mechanisms of IPF in high throughput or high resolution in vitro live cell assays. To this end, we immortalized primary fibroblasts isolated from the distal lung of Sprague-Dawley rats. Molecular and transcriptomic characterization identified clone 10-4A as a matrix fibroblast subpopulation. Mechanical or chemical stimulation induced a reversible fibrotic state comparable to effects observed in primary isolated fibroblasts. Finally, we generated a reporter cell line (10-4A BFP ) to express nuclear blue fluorescent protein (BFP) under the promotor of the myofibroblast marker alpha smooth muscle actin (Acta2) using CRISPR/Cas9 technology. We evaluated the suitability of 10-4A BFP as reporter tool in plate reader assays. In summary, the 10-4A BFP cell line provides a novel tool to study fibrotic processes in vitro to gain new insights into the cellular and molecular processes involved in fibrosis formation and propagation.
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Diphtheria toxin (DT) efficiently inhibits protein synthesis in human cells, resulting in severe disease diphtheria. The sensitivity towards DT varies between mammalian species. Mice and rats are resistant to DT. However, the reason underlying this insensitivity is controversially discussed and not well understood. Therefore, we investigated the steps of DT uptake, i.e. receptor binding and internalization into mouse J774A.1 macrophages and primary rat fibroblasts. We exploited the non-toxic DT-mutant cross-reacting material 197 (CRM197) and three additional receptor binding-deficient mutants (250 nM each) to investigate binding to cell surface and internalization into murine cells via flow cytometry and stimulated emission depletion (STED) super-resolution optical microscopy. Dual-color STED imaging unveiled CRM197 interacting with the murine precursor of the heparin-binding epidermal growth factor-like growth factor (HB-EGF). Moreover, we identified CRM197's transmembrane domain as an additional HB-EGF binding site, which is also involved in the receptor-mediated internalization into murine cells. However, we do not find evidence for translocation of the catalytically active subunit (DTA) into the cytosol when 250 nM DT were applied. In conclusion, we provide evidence that the resistance of murine cells to DT is caused by an insufficiency of DTA to escape from endosomes and reach the cytosol. Possibly, a higher affinity interaction of DT and the HB-EGF is required for translocation, which highlights the role of the receptor in the endosomes during the translocation step. We extend the current knowledge about cellular uptake of the medically relevant DT and CRM197.
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Proteínas de Bactérias , Toxina Diftérica/toxicidade , Sequência de Aminoácidos , Animais , Sítios de Ligação , Fator de Crescimento Epidérmico , Fibroblastos , Fator de Crescimento Semelhante a EGF de Ligação à Heparina , Humanos , Camundongos , Microscopia , Ligação Proteica , Ratos , Receptores de Superfície CelularRESUMO
Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal lung disease with limited therapeutic options. The current model suggests that chronic or repetitive "micro-injuries" of the alveolar epithelium lead to activation and proliferation of fibroblasts and excessive extracellular matrix (ECM) deposition. Disruption of alveolar type II (ATII) epithelial cell homeostasis and the characteristics of mesenchymal cell populations in IPF have received particular attention in recent years. Emerging data from single cell RNA sequencing (scRNAseq) analysis shed novel light on alterations in ATII cell progenitor dysfunction and the diversity of mesenchymal cells within the fibrotic lung. Within this minireview, we summarize the data from most recent human scRNAseq studies. We aim to collate the current knowledge on cellular plasticity and heterogeneity in the development and progression of IPF, effects of drug treatment on transcriptional changes. Finally, we provide a brief outlook on future challenges and promises for large scale sequencing studies in the development of novel therapeutics for IPF.
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Degeneration of dopaminergic neurons in the substantia nigra causes the motor symptoms of Parkinson's disease. The mechanisms underlying this age-dependent and region-selective neurodegeneration remain unclear. Here we identify Cav2.3 channels as regulators of nigral neuronal viability. Cav2.3 transcripts were more abundant than other voltage-gated Ca2+ channels in mouse nigral neurons and upregulated during aging. Plasmalemmal Cav2.3 protein was higher than in dopaminergic neurons of the ventral tegmental area, which do not degenerate in Parkinson's disease. Cav2.3 knockout reduced activity-associated nigral somatic Ca2+ signals and Ca2+-dependent after-hyperpolarizations, and afforded full protection from degeneration in vivo in a neurotoxin Parkinson's mouse model. Cav2.3 deficiency upregulated transcripts for NCS-1, a Ca2+-binding protein implicated in neuroprotection. Conversely, NCS-1 knockout exacerbated nigral neurodegeneration and downregulated Cav2.3. Moreover, NCS-1 levels were reduced in a human iPSC-model of familial Parkinson's. Thus, Cav2.3 and NCS-1 may constitute potential therapeutic targets for combatting Ca2+-dependent neurodegeneration in Parkinson's disease.
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Envelhecimento/genética , Canais de Cálcio Tipo R/genética , Proteínas de Transporte de Cátions/genética , Sobrevivência Celular/genética , Neurônios Dopaminérgicos/metabolismo , Proteínas Sensoras de Cálcio Neuronal/genética , Neuropeptídeos/genética , Doença de Parkinson/genética , Envelhecimento/metabolismo , Animais , Canais de Cálcio Tipo R/metabolismo , Sinalização do Cálcio , Proteínas de Transporte de Cátions/metabolismo , Neurônios Dopaminérgicos/patologia , Humanos , Células-Tronco Pluripotentes Induzidas , Camundongos , Camundongos Knockout , Proteínas Sensoras de Cálcio Neuronal/metabolismo , Neuropeptídeos/metabolismo , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Substância Negra/metabolismo , Substância Negra/patologia , Regulação para Cima , Área Tegmentar Ventral/metabolismo , Área Tegmentar Ventral/patologiaRESUMO
UNLABELLED: Death rates from hepatocellular carcinoma (HCC) are steadily increasing, yet therapeutic options for advanced HCC are limited. We identify a subset of mouse and human HCCs harboring VEGFA genomic amplification, displaying distinct biologic characteristics. Unlike common tumor amplifications, this one seems to work via heterotypic paracrine interactions; stromal VEGF receptors (VEGFR), responding to tumor VEGF-A, produce hepatocyte growth factor (HGF) that reciprocally affects tumor cells. VEGF-A inhibition results in HGF downregulation and reduced proliferation, specifically in amplicon-positive mouse HCCs. Sorafenib-the first-line drug in advanced HCC-targets multiple kinases, including VEGFRs, but has only an overall mild beneficial effect. We found that VEGFA amplification specifies mouse and human HCCs that are distinctly sensitive to sorafenib. FISH analysis of a retrospective patient cohort showed markedly improved survival of sorafenib-treated patients with VEGFA-amplified HCCs, suggesting that VEGFA amplification is a potential biomarker for HCC response to VEGF-A-blocking drugs. SIGNIFICANCE: Using a mouse model of inflammation-driven cancer, we identified a subclass of HCC carrying VEGFA amplification, which is particularly sensitive to VEGF-A inhibition. We found that a similar amplification in human HCC identifies patients who favorably responded to sorafenib-the first-line treatment of advanced HCC-which has an overall moderate therapeutic efficacy.
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Antineoplásicos/uso terapêutico , Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Niacinamida/análogos & derivados , Compostos de Fenilureia/uso terapêutico , Fator A de Crescimento do Endotélio Vascular/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/deficiência , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Animais , Antineoplásicos/farmacologia , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Hepatócitos/metabolismo , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Macrófagos/metabolismo , Masculino , Camundongos Knockout , Niacinamida/farmacologia , Niacinamida/uso terapêutico , Compostos de Fenilureia/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Sorafenibe , Carga Tumoral/efeitos dos fármacos , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Membro 4 da Subfamília B de Transportadores de Cassetes de Ligação de ATPRESUMO
UNLABELLED: The receptor for advanced glycation endproducts (RAGE) is a multiligand receptor and member of the immunoglobulin superfamily. RAGE is mainly involved in tissue damage and chronic inflammatory disorders, sustaining the inflammatory response upon engagement with damage-associated molecular pattern molecules (DAMPs) such as S100 proteins and high-mobility group box 1 (HMGB1). Enhanced expression of RAGE and its ligands has been demonstrated in distinct tumors and several studies support its crucial role in tumor progression and metastasis by still unknown mechanisms. Here we show that RAGE supports hepatocellular carcinoma (HCC) formation in the Mdr2(-/-) mouse model, a prototype model of inflammation-driven HCC formation, which mimics the human pathology. Mdr2(-/-) Rage(-/-) (dKO) mice developed smaller and fewer HCCs than Mdr2(-/-) mice. Interestingly, although in preneoplastic Mdr2(-/-) livers RAGE ablation did not affect the onset of inflammation, premalignant dKO livers showed reduced liver damage and fibrosis, in association with decreased oval cell activation. Oval cells expressed high RAGE levels and displayed reduced proliferation upon RAGE silencing. Moreover, stimulation of oval cells with HMGB1 promoted an ERK1/2-Cyclin D1-dependent oval cell proliferation in vitro. Finally, genetic and pharmacologic blockade of RAGE signaling impaired oval cell activation in an independent mouse model of oval cell activation, the choline deficient ethionine-supplemented dietary regime. CONCLUSION: Our data identified a novel function of RAGE in regulating oval cell activation and tumor development in inflammation-associated liver carcinogenesis.
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Carcinoma Hepatocelular/etiologia , Neoplasias Hepáticas/etiologia , Receptores Imunológicos/fisiologia , Células-Tronco/fisiologia , Subfamília B de Transportador de Cassetes de Ligação de ATP/deficiência , Animais , Carcinoma Hepatocelular/patologia , Transformação Celular Neoplásica , Proteína HMGB1/metabolismo , Inflamação/metabolismo , Cirrose Hepática/patologia , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Knockout , Receptor para Produtos Finais de Glicação Avançada , Receptores Imunológicos/biossíntese , Células-Tronco/patologia , Membro 4 da Subfamília B de Transportadores de Cassetes de Ligação de ATPRESUMO
BACKGROUND: Calprotectin consists of the Ca2+-binding proteins S100a8 and S100a9 that are induced in epithelial cells in response to tissue damage and infection. Both proteins are also secreted by activated innate immune cells and numerous studies demonstrate their crucial role in pathological conditions of acute and chronic inflammation. RESULTS: Here, we established a conditional mouse model with simultaneous S100a8 and S100a9 transgene expression in hepatocytes (TgS100a8a9hep) under the control of doxycycline to unravel the role of epithelial-derived Calprotectin on tissue homeostasis and inflammation. TgS100a8a9hep mice displayed a significant enrichment of neutrophils in peripheral blood and tissues with high blood content. Interestingly, Cxcl1 transcription was significantly induced in the liver of TgS100a8a9hep mice and primary hepatocytes derived thereof as compared to Control mice, accompanied by an increase of Cxcl1 serum levels. However, expression of other chemokines with a known function in neutrophil mobilization from the bone marrow, e.g. Csf3 and Cxcl2, was not altered. Doxycycline treatment of TgS100a8a9hep mice reduced Cxcl1 expression in the liver and resulted in normal numbers of neutrophils. CONCLUSION: In summary, our data demonstrate for the first time that hepatocyte-specific S100a8 and S100a9 expression induces a systemic mobilization of neutrophils by a specific activation of Cxcl1 transcription in the liver.
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OBJECTIVE: From data in the literature, we hypothesized that high vascular resistance values in the uterine arteries at the end of the first trimester would increase adverse pregnancy outcomes and therefore might be accompanied by changes in VEGF/VEGFR1 immunoreactivities. STUDY DESIGN: In our university hospital 82 women (Study I n=62 and Study II n=20) were divided into two groups according to their uterine vascular resistance values. Uterine vascular resistance values were measured in the 10-13th weeks of gestation by color-Doppler ultrasonography. Women were divided into low and high vascular resistance groups. In the prospective follow-up study (Study I) the data of the pregnancy outcome were recorded. In cross-sectional study (Study II), VEGF and VEGFR1 immunoreactivities were measured on the tissue samples from women who underwent termination of pregnancy. RESULTS: In the high vascular resistance group (PI>2.3), the probability of adverse pregnancy outcome was significantly higher (40.0% vs. 12.8%). No differences in VEGF and VEGFR1 immunoreactivities were observed between groups. In both groups, intense VEGF immunoreactivity was observed in the maternal glandular epithelium and in the decidual cells. Weak reactivity was observed in the villous trophoblast. VEGFR1 immunoreactivity was intense in all regions. CONCLUSIONS: Our data suggest that high vascular resistance values in the first trimester are independent from VEGF/VEGFR1 immunoreactivities and markedly increase the probability of adverse pregnancy outcomes. This may be used for early screening of pregnant women in the first trimester.
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Placenta/metabolismo , Complicações na Gravidez/etiologia , Artéria Uterina/fisiologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Resistência Vascular , Adulto , Estudos Transversais , Feminino , Seguimentos , Humanos , Gravidez , Primeiro Trimestre da Gravidez/fisiologia , Estudos ProspectivosRESUMO
BACKGROUND: In the past, molecular mechanisms that drive the initiation of an inflammatory response have been studied intensively. However, corresponding mechanisms that sustain the expression of inflammatory response genes and hence contribute to the establishment of chronic disorders remain poorly understood. Recently, we provided genetic evidence that signaling via the receptor for advanced glycation end products (Rage) drives the strength and maintenance of an inflammatory reaction. In order to decipher the mode of Rage function on gene transcription levels during inflammation, we applied global gene expression profiling on time-resolved samples of mouse back skin, which had been treated with the phorbol ester TPA, a potent inducer of skin inflammation. RESULTS: Ranking of TPA-regulated genes according to their time average mean and peak expression and superimposition of data sets from wild-type (wt) and Rage-deficient mice revealed that Rage signaling is not essential for initial changes in TPA-induced transcription, but absolutely required for sustained alterations in transcript levels. Next, we used a data set of differentially expressed genes between TPA-treated wt and Rage-deficient skin and performed computational analysis of their proximal promoter regions. We found a highly significant enrichment for several transcription factor binding sites (TFBS) leading to the prediction that corresponding transcription factors, such as Sp1, Tcfap2, E2f, Myc and Egr, are regulated by Rage signaling. Accordingly, we could confirm aberrant expression and regulation of members of the E2f protein family in epidermal keratinocytes of Rage-deficient mice. CONCLUSIONS: In summary, our data support the model that engagement of Rage converts a transient cellular stimulation into sustained cellular dysfunction and highlight a novel role of the Rb-E2f pathway in Rage-dependent inflammation during pathological conditions.
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Redes Reguladoras de Genes/genética , Inflamação/genética , Receptores Imunológicos/metabolismo , Pele/metabolismo , Pele/patologia , Animais , Análise por Conglomerados , Biologia Computacional , Modelos Animais de Doenças , Fatores de Transcrição E2F/genética , Fatores de Transcrição E2F/metabolismo , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Inflamação/patologia , Camundongos , Modelos Biológicos , Regiões Promotoras Genéticas/genética , Receptor para Produtos Finais de Glicação Avançada , Receptores Imunológicos/deficiência , Receptores Imunológicos/genética , Proteína do Retinoblastoma/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/efeitos dos fármacos , Acetato de Tetradecanoilforbol/farmacologia , Fatores de TempoRESUMO
OBJECTIVE: The current study was performed in order to determine the prevalence of different myositis-specific and myositis-associated antibodies, as well as their association with clinical characteristics, disease course and response to therapy in 169 Hungarian patients with idiopathic inflammatory myopathy. METHODS: Sera of 130 primary and 39 overlap myositis including systemic sclerosis (13), rheumatoid arthritis (12), systemic lupus erythematosus (5) and Sjögren's syndrome (9) cases were analyzed. Antinuclear antibody, scleroderma-associated antibodies (anti-centromere, anti-topoisomerase I), anti-Jo-1, anti-PL-7, anti-PL-12, anti-Mi-2, anti-SRP and anti-PM-Scl, anti-Ku, anti-SS-A, anti-SS-B, anti-U1snRNP were tested. Autoantibody results were compared with clinical characteristics, disease course of overlap versus primary myositis patients, as well as with response to therapy. RESULTS: Associated connective tissue disease occurred in 23.1% of the patients. Myositis-associated antibodies were found in 8.5% of primary myositis patients, indicating that 11 additional primary myositis patients (23% vs. 29.6%) can be classified as overlap in all cohort according to the newly proposed diagnostic criteria. Polymyositis was found to be the most common myositis form in overlap myositis (87.2%), while scleroderma was the most common disease associated (33.3%). ANA was positive in 25.4% of primary and in 61.5% of overlap myositis cases. Altogether 39.6% of myositis patients (n=67) had autoantibodies, most commonly anti Jo-1 (18.3%) correlating with a polycyclic disease course. CONCLUSION: Inclusion of myositis-specific and associated antibodies into the newly proposed diagnostic criteria for inflammatory myopathies is of great importance in order to determine subclasses and to introduce adequate therapy in time.
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Autoanticorpos/imunologia , Dermatomiosite/classificação , Dermatomiosite/imunologia , Polimiosite/classificação , Polimiosite/imunologia , Adolescente , Adulto , Idoso , Autoanticorpos/sangue , Estudos Transversais , Dermatomiosite/tratamento farmacológico , Dermatomiosite/epidemiologia , Feminino , Seguimentos , Humanos , Imunossupressores/uso terapêutico , Masculino , Pessoa de Meia-Idade , Polimiosite/tratamento farmacológico , Polimiosite/epidemiologia , Estudos Retrospectivos , Estudos Soroepidemiológicos , Resultado do Tratamento , Adulto JovemRESUMO
Claudins, known as major contributors in the formation of the tight junction, are differentially expressed in malignant tumors as compared to the corresponding healthy tissues. Therefore, they are thought to play a role in carcinogenesis and tumor progression. Altered expression of claudin-1 has been reported in several tumor types including endometrial, papillary renal cell and colonic carcinoma, and increased claudin-1 mRNA levels have been observed in papillary thyroid carcinoma (PTC). In this study, we aimed at determining the pattern of claudin-1 expression in various types of thyroid lesions at the protein level and investigating the immunolocalization of beta-catenin reported to regulate claudin-1 expression. Samples included 19 PTCs, ten cases of corresponding regional lymph node metastasis, eight papillary microcarcinomas (PMC), 17 follicular thyroid carcinomas (FTC) and 19 follicular adenomas (FA). All cases were evaluated by quantitative immunohistochemistry. Conspicuous claudin-1 immunostaining was detected in the majority of PTC/PMC primary tumors and lymph node metastases (19/27 and 9/10, respectively). On the other hand, we found weak or no claudin-1 expression in any of the FA and FTC cases or peritumoral non-malignant thyroid tissues. Our data prove that high claudin-1 protein expression is specific for PTC and its regional lymph node metastases, while we failed to verify that claudin-1 is regulated by beta-catenin in thyroid tumors. Based on these results, claudin-1 may be a useful tumor marker for PTC.
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Adenocarcinoma Papilar/metabolismo , Adenocarcinoma Papilar/patologia , Proteínas de Membrana/biossíntese , Neoplasias da Glândula Tireoide/metabolismo , Neoplasias da Glândula Tireoide/patologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/análise , Criança , Claudina-1 , Feminino , Humanos , Imuno-Histoquímica , Metástase Linfática/patologia , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
UNLABELLED: The nuclear factor-kappaB (NF-kappaB) signaling pathway has been recently shown to participate in inflammation-induced cancer progression. Here, we describe a detailed analysis of the NF-kappaB-dependent gene regulatory network in the well-established Mdr2 knockout mouse model of inflammation-associated liver carcinogenesis. Expression profiling of NF-kappaB-deficient and NF-kappaB-proficient hepatocellular carcinoma (HCC) revealed a comprehensive list of known and novel putative NF-kappaB target genes, including S100a8 and S100a9. We detected increased co-expression of S100A8 and S100A9 proteins in mouse HCC cells, in human HCC tissue, and in the HCC cell line Hep3B on ectopic RelA expression. Finally, we found a synergistic function for S100A8 and S100A9 in Hep3B cells resulting in a significant induction of reactive oxygen species (ROS), accompanied by enhanced cell survival. CONCLUSION: We identified S100A8 and S100A9 as novel NF-kappaB target genes in HCC cells during inflammation-associated liver carcinogenesis and provide experimental evidence that increased co-expression of both proteins supports malignant progression by activation of ROS-dependent signaling pathways and protection from cell death.
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Calgranulina A/metabolismo , Calgranulina B/metabolismo , Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , NF-kappa B/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Animais , Apoptose/fisiologia , Calgranulina A/genética , Calgranulina B/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Modelos Animais de Doenças , Humanos , Neoplasias Hepáticas/patologia , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/fisiologia , Membro 4 da Subfamília B de Transportadores de Cassetes de Ligação de ATPRESUMO
The receptor for advanced glycation end products (RAGE) is a single transmembrane receptor of the immunoglobulin superfamily that is mainly expressed on immune cells, neurons, activated endothelial and vascular smooth muscle cells, bone forming cells, and a variety of cancer cells. RAGE is a multifunctional receptor that binds a broad repertoire of ligands and mediates responses to cell damage and stress conditions. It activates programs responsible for acute and chronic inflammation, and is implicated in a number of pathological diseases, including diabetic complications, stroke, atheriosclerosis, arthritis, and neurodegenerative disorders. The availability of Rage knockout mice has not only advanced our knowledge on signalling pathways within these pathophysiological conditions, but also on the functional importance of the receptor in processes of cancer. Here, we will summarize molecular mechanisms through which RAGE signalling contributes to the establishment of a pro-tumourigenic microenvironment. Moreover, we will review recent findings that provide genetic evidence for an important role of RAGE in bridging inflammation and cancer.
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BACKGROUND: Serum beta2-microglobulin (beta2m) has been established as a marker of disease activity in malignancies, autoimmune conditions and infections. Despite its important role in prognosis assessment and disease monitoring, relatively few studies are available on its expression in healthy individuals. Furthermore, interpretation of results is hampered by the variety in reference limits due to differences in methodology, sample population and statistics. METHODS: Serum beta2m concentrations were measured using a microparticle-enhanced immunonephelometric method in 183 healthy blood donors aged 29-75 years. RESULTS: The median beta2m concentration was 1.67 (0.88-2.75) mg/L with no difference between men and women (1.71 mg/L vs. 1.62 mg/L, p<0.07). A linear correlation was found between beta2m and age (p<0.0001), serum concentrations significantly higher in older subjects (1.55, 1.59, 1.70, and 1.87 mg/L in age groups of 29-40, 40-50, 50-60 and 60-75 years, respectively, p<0.0001). Reference intervals obtained by non-parametric estimation after partitioning by age were 1.02-2.46 mg/L vs. 1.29-2.70 mg/L in younger (29-49 years) vs. older (50-75 years) individuals. CONCLUSIONS: These data can help standardise beta2m reference limits and support age-adjusted comparisons in clinical studies.
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Envelhecimento/sangue , Imunoensaio/métodos , Nefelometria e Turbidimetria/métodos , Microglobulina beta-2/sangue , Adulto , Idoso , Biomarcadores/sangue , Estudos de Coortes , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Valores de Referência , Fatores Sexuais , Estatísticas não ParamétricasRESUMO
The incidence of cholangiocarcinomas originating from intrahepatic and extrahepatic bile ducts, as well as of gallbladder carcinoma is increasing worldwide. The malignant transformation of biliary epithelia involves profound alterations of proteins in the intercellular junctions, among others zonula occludens-1 (ZO-1), occludin, and E-cadherin. Each plays important role in the maintenance of epithelial cell polarity and regulation of cell growth and differentiation. Our aim was to investigate ZO-1, occludin, and E-cadherin immunohistochemical reactions in tissue microarray blocks containing 57 normal and 62 neoplastic biliary tract samples. We demonstrated that the tight junction components ZO-1, occludin, and E-cadherin are downregulated in carcinomas arising from various compartments of the biliary tract (normal intrahepatic and extrahepatic bile ducts, gallbladder) as compared with their normal sites of origin. These results were confirmed by discriminant analysis yielding clear separation of the three normal sample groups from carcinomas in the corresponding locations.
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Neoplasias dos Ductos Biliares/metabolismo , Ductos Biliares Intra-Hepáticos/metabolismo , Caderinas/biossíntese , Colangiocarcinoma/metabolismo , Proteínas de Membrana/biossíntese , Fosfoproteínas/biossíntese , Idoso , Neoplasias dos Ductos Biliares/patologia , Ductos Biliares Intra-Hepáticos/patologia , Biomarcadores Tumorais/análise , Colangiocarcinoma/patologia , Feminino , Humanos , Imuno-Histoquímica , Masculino , Ocludina , Análise Serial de Tecidos , Proteína da Zônula de Oclusão-1RESUMO
Biliary tract cancers are relatively common malignant gastrointestinal tumors in the elderly. Claudins are integral components of tight junctions that play important roles in maintaining epithelial cell polarity, controlling paracellular diffusion, and regulating cell growth and differentiation. The expression profile of claudins has been extensively characterized, but few reports exist on their expression in the normal and neoplastic biliary tract. Our aim was therefore to study claudins by IHC reactions in normal and neoplastic biliary tract samples. We detected that claudin expressions differ in the normal sample groups: the normal gallbladder strongly expressed claudin-2, -3, -4, and -10, but only weak reactions were seen in normal intrahepatic bile ducts. Although each cancer type expressed several claudins with various intensities, only claudin-4 presented especially strong immunoreactions in extrahepatic bile duct cancers and gallbladder carcinomas, whereas claudin-1 and -10 presented in intrahepatic bile duct cancers. Comparing the normal and carcinoma groups, the most significant decrease was detected in the expression of claudin-10. In conclusion, the expression pattern of claudins is different in the various parts of the normal and neoplastic biliary tract; moreover, an unequivocal decrease was detected in the carcinomas compared with their corresponding normal samples. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.
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Neoplasias do Sistema Biliar/metabolismo , Proteínas de Membrana/biossíntese , Sistema Biliar/metabolismo , Biomarcadores Tumorais/biossíntese , Claudina-1 , Claudina-3 , Claudina-4 , Claudinas , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Junções Íntimas/metabolismoRESUMO
In this study we analyzed the recovery of the immune system in children after completion of the therapy. We analysed 88 children (51 boys, 37 girls, mean age at diagnosis: 7.8 years) receiving chemotherapy for malignant diseases (43 acute lymphoblastic leukemia, 15 lymphoma, 20 bone tumor, ten other solid tumors). Serum immunoglobulin levels (Ig), natural killer activity (NK), antibody-dependent cellular cytotoxicity (ADCC) and T and B cell proliferation were determined 1 year after cessation of therapy. The mean levels of Ig were in the normal range at a mean of 13 months after chemotherapy (IgG: 11.2 +/- 3.3, IgA: 1.6 +/- 0.9, IgM: 1.0 +/- 0.5 g/l), however in the leukemic patients serum IgG was below the lower limit of the normal range in 3/43 (7.0%) cases, serum IgA was low in 5/43 (11.6%) and serum IgM was decreased in 4/43 (9.3%) cases. In the solid tumor patients IgG values were within the normal range and only 2-2/45 children had lower values for IgA and IgM (4.4%). NK activity decreased in 7/43 (16.3%) leukemic patients, and in 3/45 (6.7%) solid tumor patients, ADCC decreased in 8/43 (18.6%) and 3/45 (6.7%), respectively (p < 0.001). B-cell blastic transformation was decreased in 3/43 (7%) leukemic patients and in 4/45 (8.9%) solid tumor patients. At the same time T-cell blastic transformation was altered in 5/43 (11.6%) and in 4/45 (8.9%) cases, respectively. Leukemic patients had significantly more infections during the first year after chemotherapy than solid tumor patients (1.60 +/- 1.18 vs 0.96 +/- 1.14; p = 0.011). No significant correlations could be found between the investigated immune parameters and the number and severity of infections. It is concluded, that cytotoxic therapy can lead to long-term depression of the immune system, first of all in leukemic patients.
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Formação de Anticorpos/efeitos dos fármacos , Antineoplásicos/efeitos adversos , Imunidade Celular/efeitos dos fármacos , Neoplasias/imunologia , Formação de Anticorpos/fisiologia , Linfócitos B/efeitos dos fármacos , Linfócitos B/imunologia , Proliferação de Células/efeitos dos fármacos , Criança , Feminino , Humanos , Imunidade Celular/fisiologia , Imunoglobulinas/sangue , Imunoglobulinas/efeitos dos fármacos , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/imunologia , Contagem de Leucócitos , Masculino , Neoplasias/tratamento farmacológico , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologiaRESUMO
A broad range of experimental and clinical evidence has highlighted the central role of chronic inflammation in promoting tumor development. However, the molecular mechanisms converting a transient inflammatory tissue reaction into a tumor-promoting microenvironment remain largely elusive. We show that mice deficient for the receptor for advanced glycation end-products (RAGE) are resistant to DMBA/TPA-induced skin carcinogenesis and exhibit a severe defect in sustaining inflammation during the promotion phase. Accordingly, RAGE is required for TPA-induced up-regulation of proinflammatory mediators, maintenance of immune cell infiltration, and epidermal hyperplasia. RAGE-dependent up-regulation of its potential ligands S100a8 and S100a9 supports the existence of an S100/RAGE-driven feed-forward loop in chronic inflammation and tumor promotion. Finally, bone marrow chimera experiments revealed that RAGE expression on immune cells, but not keratinocytes or endothelial cells, is essential for TPA-induced dermal infiltration and epidermal hyperplasia. We show that RAGE signaling drives the strength and maintenance of an inflammatory reaction during tumor promotion and provide direct genetic evidence for a novel role for RAGE in linking chronic inflammation and cancer.
Assuntos
Inflamação/imunologia , Receptores Imunológicos/imunologia , Proteínas S100/imunologia , Neoplasias Cutâneas/imunologia , 9,10-Dimetil-1,2-benzantraceno , Animais , Feminino , Regulação Neoplásica da Expressão Gênica , Inflamação/induzido quimicamente , Inflamação/patologia , Proteínas Inflamatórias de Macrófagos/genética , Proteínas Inflamatórias de Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor para Produtos Finais de Glicação Avançada , Receptores Imunológicos/deficiência , Receptores Imunológicos/genética , Proteínas S100/genética , Neoplasias Cutâneas/induzido quimicamente , Neoplasias Cutâneas/patologia , Acetato de TetradecanoilforbolRESUMO
INTRODUCTION: Currently, malignancies in childhood can be cured in 70 percent of the cases. However, the intensive cytostatic therapy may lead to late side effects influencing quality of life. AIM OF THE STUDY: Analysis of the reconvalescence of the immune functions after completion of therapy for malignancies in children. PATIENTS: 88 long-term survivors (51 boys, 37 girls) were investigated (43 acute lymphoid leukemia, 15 lymphoma, 20 bone tumors, 10 other solid tumors). Mean age at the time of diagnosis was 7.8 years (1 mo-17.7 years). METHODS: The following parameters were investigated: serum immunoglobulin levels after completion of the chemotherapy and in the next 4 years thereafter, lymphocyte subpopulations in the peripheral blood by flow-cytometry and cellular immunity by in vitro tests (natural killer activity, antibody-dependent cellular cytotoxicity, mitogen-induced T- and B-cell blastic transformations). RESULTS: Lower serum immunoglobulin (IgG) levels could be detected in patients with leukemia after completion of the chemotherapy (8.8 +/- 3.2 g/l). One year thereafter serum IgG levels increased significantly (10.1 +/- 2.9 g/l) (p<0.05). In patients with solid tumors the serum IgG levels were in the normal range at the end of the chemotherapy (12.1 +/- 4.3 g/l). At a mean of 1.3 years after the end of chemotherapy NK activity decreased in 7/43 (16.3%) leukemia patients, and in 3/45 (6.7%) solid tumor patients, ADCC decreased in 8/43 (18.6%) and 3/45 (6.7%), respectively (p<0.05 leukemia vs. solid tumor). At a mean of 15 months after the end of the therapy B-cell blastic transformation was decreased in 3/43 (7%) leukemia patients and in 4/45 (8.9%) solid tumor patients. At the same time point T-cell blastic transformation was altered in 5/43 (11.6%) and in 4/45 (8.9%) cases, respectively. CONCLUSION: Cytotoxic therapies lead to severe, long-term depression of the immune system. At the end of the chemotherapy this effect is more pronounced in leukemia patients. Years (1.5-3) after completion of the therapy in a significant proportion of the patients some in vitro parameters of the immune system are yet altered, so careful monitoring of this patient population is mandatory.