RESUMO
To study the estrogen regulated transcription of the uteroglobin (UG) gene, the founding member of the secretoglobin family widely expressed in many different mammalian species, we re-created functional estrogen response elements (EREs) in the UG gene promoter from a species where UG expression is not regulated by estrogens: the hamster Mesocricetus auratus (Ma), to ascertain if the lack of functional EREs is the real cause of its estrogen insensitivity. Functional EREs in the hamster promoter, including the consensus ERE (cERE), failed to respond to an appropriate estrogen stimulus compared with its estrogen regulated ortholog from the brown hare Lepus capensis (Lc). As the nucleotide sequence is the only difference between genetic constructs from these two species, we suspected that the UG promoter from the hamster probably contains cis-acting genetic elements that negatively impairs the estrogen-regulated transcription mediated by the functional ERE. Accordingly, we prepared chimeric DNA constructs which eventually allowed to identify a region located 29 base pairs (bp) downstream of the ERE as responsible for the lack of estrogen-responsiveness of the Ma-UG gene in the breast cancer cell line MCF-7. This region contains the sequence ACACCCC which has been identified as the core sequence of the Sp/ Krüppel-like factor (KLF) family of transcription factors. This finding is relevant, not only due to the observation on a novel mechanism that control estrogen-induced transcription, but also because it may encourage further investigation for better defining specific genes with an ERE that do not respond to estrogen signaling in MCF-7 cells, a cell line widely employed as an in vitro model in breast cancer research.
Assuntos
Neoplasias da Mama , Lebres , Cricetinae , Animais , Humanos , Feminino , Células MCF-7 , Uteroglobina/genética , Sequência de Bases , Estrogênios/farmacologia , Estrogênios/metabolismo , Neoplasias da Mama/genética , Lebres/metabolismo , Transcrição Gênica , Estradiol/farmacologiaRESUMO
OBJECTIVE: To evaluate serum uteroglobin-related protein 1 expression early after smoke inhalation injuries and its association with the severity of inhalation injury in burned patients. METHODS: Smoke or chemical inhalation injury is associated with morbidity and mortality. The consequences of inhalation result from an inflammatory response. Uteroglobin-related protein 1 is an anti-inflammatory protein and may improve lung inflammation. We hypothesized that uteroglobin-related protein 1 levels could reflect disease severity and predict outcome in patients with inhalation injury. Sixteen patients diagnosed with acute respiratory distress syndrome secondary to smoke inhalation injury were prospectively included in the study. Plasma was collected upon intensive care unit admission and within 24 hours of the inhalation injury. Bronchoscopies were carried out in all patients to assess the severity of inhalation injury within 72 hours. Uteroglobin-related protein 1 plasma levels were determined in duplicate with enzyme-linked immunosorbent assay. RESULTS: The mean age was 23 ± 5 years, and the inhalation injury distribution was as follows: three of grade 1, four of grade 2, and nine of grade 3. The level of uteroglobin-related protein 1 was related to inhalation severity (grade 1: 0.389 ± 0.053 arbitrary units versus grade 2: 0.474 ± 0.0423 arbitrary units versus grade 3: 0.580 ± 0.094 arbitrary units; p = 0.007). CONCLUSION: Plasma levels of uteroglobin-related protein 1 are associated with the degree of lung inhalation injury.
OBJETIVO: Avaliar a expressão sérica da proteína 1 relacionada à uteroglobulina na fase inicial após lesões por inalação de fumaça e sua associação com a gravidade da lesão por inalação em pacientes queimados. MÉTODOS: A lesão por inalação de fumaça ou produtos químicos se associa com morbidade e mortalidade. As consequências da inalação resultam de uma resposta inflamatória. A proteína 1 relacionada à uteroglobulina é anti-inflamatória e pode melhorar a inflamação pulmonar. Nossa hipótese é que os níveis de proteína 1 relacionada à uteroglobulina podem refletir a gravidade da doença e predizer o desfecho em pacientes com lesão por inalação. Incluíram-se prospectivamente neste estudo 16 pacientes com diagnóstico de síndrome do desconforto respiratório agudo decorrente de lesão por inalação de fumaça. Em todos os pacientes, colheu-se amostra de plasma quando da admissão à unidade de terapia intensiva, para avaliar a gravidade da lesão por inalação dentro de 72 horas. Os níveis plasmáticos de proteína 1 relacionada à uteroglobulina foram determinados em duplicata por meio de ensaio de imunoabsorção ligado à enzima. RESULTADOS: A média de idade foi de 23 ± 5 anos, e a distribuição da lesão por inalação foi: três em grau 1, quatro em grau 2 e nove em grau 3. O nível de proteína 1 relacionada à uteroglobulina foi relacionado ao grau de severidade (grau 1: 0,389 ± 0,053 unidade arbitrária versus grau 2: 0,474 ± 0,0423 unidade arbitrária versus grau 3: 0,580 ± 0,094 unidade arbitrária; p = 0,007). CONCLUSÃO: Os níveis plasmáticos de proteína 1 relacionada à uteroglobulina se associam com o grau da lesão pulmonar por inalação.
Assuntos
Queimaduras , Síndrome do Desconforto Respiratório , Lesão por Inalação de Fumaça , Adolescente , Adulto , Humanos , Unidades de Terapia Intensiva , Uteroglobina , Adulto JovemRESUMO
RESUMO Objetivo: Avaliar a expressão sérica da proteína 1 relacionada à uteroglobulina na fase inicial após lesões por inalação de fumaça e sua associação com a gravidade da lesão por inalação em pacientes queimados. Métodos: A lesão por inalação de fumaça ou produtos químicos se associa com morbidade e mortalidade. As consequências da inalação resultam de uma resposta inflamatória. A proteína 1 relacionada à uteroglobulina é anti-inflamatória e pode melhorar a inflamação pulmonar. Nossa hipótese é que os níveis de proteína 1 relacionada à uteroglobulina podem refletir a gravidade da doença e predizer o desfecho em pacientes com lesão por inalação. Incluíram-se prospectivamente neste estudo 16 pacientes com diagnóstico de síndrome do desconforto respiratório agudo decorrente de lesão por inalação de fumaça. Em todos os pacientes, colheu-se amostra de plasma quando da admissão à unidade de terapia intensiva, para avaliar a gravidade da lesão por inalação dentro de 72 horas. Os níveis plasmáticos de proteína 1 relacionada à uteroglobulina foram determinados em duplicata por meio de ensaio de imunoabsorção ligado à enzima. Resultados: A média de idade foi de 23 ± 5 anos, e a distribuição da lesão por inalação foi: três em grau 1, quatro em grau 2 e nove em grau 3. O nível de proteína 1 relacionada à uteroglobulina foi relacionado ao grau de severidade (grau 1: 0,389 ± 0,053 unidade arbitrária versus grau 2: 0,474 ± 0,0423 unidade arbitrária versus grau 3: 0,580 ± 0,094 unidade arbitrária; p = 0,007). Conclusão: Os níveis plasmáticos de proteína 1 relacionada à uteroglobulina se associam com o grau da lesão pulmonar por inalação.
ABSTRACT Objective: To evaluate serum uteroglobin-related protein 1 expression early after smoke inhalation injuries and its association with the severity of inhalation injury in burned patients. Methods: Smoke or chemical inhalation injury is associated with morbidity and mortality. The consequences of inhalation result from an inflammatory response. Uteroglobin-related protein 1 is an anti-inflammatory protein and may improve lung inflammation. We hypothesized that uteroglobin-related protein 1 levels could reflect disease severity and predict outcome in patients with inhalation injury. Sixteen patients diagnosed with acute respiratory distress syndrome secondary to smoke inhalation injury were prospectively included in the study. Plasma was collected upon intensive care unit admission and within 24 hours of the inhalation injury. Bronchoscopies were carried out in all patients to assess the severity of inhalation injury within 72 hours. Uteroglobin-related protein 1 plasma levels were determined in duplicate with enzyme-linked immunosorbent assay. Results: The mean age was 23 ± 5 years, and the inhalation injury distribution was as follows: three of grade 1, four of grade 2, and nine of grade 3. The level of uteroglobin-related protein 1 was related to inhalation severity (grade 1: 0.389 ± 0.053 arbitrary units versus grade 2: 0.474 ± 0.0423 arbitrary units versus grade 3: 0.580 ± 0.094 arbitrary units; p = 0.007). Conclusion: Plasma levels of uteroglobin-related protein 1 are associated with the degree of lung inhalation injury.
Assuntos
Humanos , Adolescente , Adulto , Adulto Jovem , Síndrome do Desconforto Respiratório do Recém-Nascido , Queimaduras , Lesão por Inalação de Fumaça , Uteroglobina , Unidades de Terapia IntensivaRESUMO
The Non-Ciliated Bronchiolar Cell (NCBC) is responsible for the defense and maintenance of the bronchiolar epithelium. Several cellular defense mechanisms have been associated with an increase in the secretion of CC16 and changes in the phenotype of the cell; these mechanisms could be linked to tolerance to the damage due to exposure to inhaled Particulate Matter (PM) of the epithelium. These defense mechanisms have not been sufficiently explored. In this article, we studied the response of the NCBC to inhaled vanadium, an element which adheres to PM. This response was measured by the changes in the phenotype of the NCBC and the secretion of CC16 in a mouse model. Mice were exposed in two phases to different vanadium concentrations; 1.27 mg/m³ in the first phase and 2.56 mg/m³ in the second phase. Mice were sacrificed on the 2nd, 4th, 5th, 6th and 8th weeks. In the second phase, we observed the following: sloughing of the NCBC, hyperplasia and small inflammatory foci remained without changes and that the expression of CC16 was higher in this phase than in phase I. We also observed a change in the phenotype with a slow decrease in both phases. The increase in the secretion of CC16 and the phenotype reversion could be due to the anti-inflammatory activity of CC16. The changes observed in the second phase could be attributed to the tolerance to inhaled vanadium.
Assuntos
Bronquíolos , Células Epiteliais , Uteroglobina/metabolismo , Vanádio/toxicidade , Poluentes Atmosféricos/toxicidade , Animais , Anti-Inflamatórios/metabolismo , Bronquíolos/citologia , Bronquíolos/metabolismo , Bronquíolos/patologia , Tolerância a Medicamentos/fisiologia , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Epitélio/metabolismo , Epitélio/patologia , Inflamação , Inalação , Pulmão/metabolismo , Camundongos , Material Particulado/toxicidadeRESUMO
Clara cells are the main airway secretory cells able to regenerate epithelium in the distal airways through transdifferentiating into goblet cells, a process under negative regulation of the Notch pathway. Pneumocystis is a highly prevalent fungus in humans occurring between 2 and 5 months of age, a period when airways are still developing and respiratory morbidity typically increases. Pneumocystis induces mucus hyperproduction in immunocompetent host airways and whether it can stimulate Clara cells is unknown. Markers of Clara cell secretion and Notch1 activation were investigated in lungs of immunocompetent rats at 40, 60, and 80 days of age during Pneumocystis primary infection with and without Valproic acid (VPA), a Notch inducer. The proportion of rats expressing mucin increased in Pneumocystis-infected rats respect to controls at 60 and 80 days of age. Frequency of distal airways Clara cells was maintained while mRNA levels for the mucin-encoding genes Muc5B and Muc5ac in lung homogenates increased 1.9 and 3.9 times at 60 days of infection (P. = 0.1609 and P. = 0.0001, respectively) and protein levels of the Clara cell marker CC10 decreased in the Pneumocystis-infected rats at 60 and 80 days of age (P. = 0.0118 & P. = 0.0388). CC10 and Muc5b co-localized in distal airway epithelium of Pneumocystis-infected rats at day 60. Co-localization of Muc5b and Ki67 as marker of mitosis in distal airways was not observed suggesting that Muc5b production by Clara cells was independent of mitosis. Notch levels remained similar and no transnucleation of activated Notch associated to Pneumocystis infection was detected. Unexpectedly, mucus was greatly increased at day 80 in Pneumocystis-infected rats receiving VPA suggesting that a Notch-independent mechanism was triggered. Overall, data suggests a Clara to goblet cell transdifferentiation mechanism induced by Pneumocystis and independent of Notch.
Assuntos
Pulmão/metabolismo , Pulmão/microbiologia , Mucina-5AC/biossíntese , Mucina-5B/biossíntese , Infecções por Pneumocystis/metabolismo , Infecções por Pneumocystis/microbiologia , Pneumocystis/patogenicidade , Receptores Notch/metabolismo , Animais , Transdiferenciação Celular/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Feminino , Antígeno Ki-67/metabolismo , Mitose/efeitos dos fármacos , Mucina-5AC/genética , Mucina-5AC/metabolismo , Mucina-5B/genética , Mucina-5B/metabolismo , Pneumocystis/efeitos dos fármacos , Infecções por Pneumocystis/patologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , Transdução de Sinais , Uteroglobina/metabolismo , Ácido Valproico/farmacologiaRESUMO
Objective: To evaluate the effects of burnt sugarcane harvesting on the plasmatic and urinary concentrations of the club cell secretory protein (CC16) and inflammatory systemic biomarkers in a group of sugarcane cutters. Methods: Seventy-eight sugar cane workers were evaluated. The plasmatic and urinary concentrations of CC16, a pulmonary damage marker and inflammatory systemic biomarkers were collected at three time points: before, three months after and six months after the onset of the burnt sugarcane harvesting period. All evaluations were performed at â¼7 am, before the daily work shift. In the three-month evaluation, a post-work shift assessment (acute effect) was also performed. Results: The age of the workers was 37.9 ± 11.0 years. The PM2.5 concentrations were 27.0 (23.0-33.0) and 101.0 (31.0-139.5) µg/m3 in the pre harvest and harvest periods, respectively (p < .001). Burnt sugarcane harvesting was associated with a reduction, throughout the work during burnt sugarcane harvesting (subchronic effect), in plasmatic and urinary CC16 concentrations. Acutely, there was a decrease in plasmatic concentrations. There were acute and subchronic increases in inflammatory markers (neutrophils, monocytes) and muscle damage markers (CK and LDH) and a decrease in red blood cells. Conclusions: Harvesting of burnt sugarcane was associated with acute and subchronic reductions in the plasmatic and urinary concentrations of CC16 protein and changes in systemic inflammatory markers.
Assuntos
Poluentes Atmosféricos/efeitos adversos , Produção Agrícola , Exposição por Inalação/efeitos adversos , Exposição Ocupacional/efeitos adversos , Material Particulado/efeitos adversos , Saccharum , Adulto , Biomarcadores/sangue , Biomarcadores/urina , Humanos , Inflamação/sangue , Inflamação/induzido quimicamente , Inflamação/imunologia , Inflamação/urina , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Masculino , Pessoa de Meia-Idade , Monócitos/imunologia , Neutrófilos/imunologia , Uteroglobina/sangue , Uteroglobina/urinaRESUMO
The aim of this pilot study was to determine Clara cell protein (CC16) concentration in bronchoalveolar lavages (BAL) fluid from full-term and preterm (<37 weeks' gestational age) neonates requiring respiratory support, having symptoms of neonatal respiratory distress syndrome, and at risk of bronchopulmonary dysplasia (BPD). We hypothesized that CC16 may be predictive of BPD diagnosis regardless of gestational age. BAL fluid CC16 was measured by ELISA at birth and at day 7 of life. Both groups that developed BPD showed significantly decreased BAL fluid CC16 levels compared to those infants that did not develop the disease. CC16 positively correlated with diagnosis of BPD and negatively with the severity of the disease. These results suggest that BAL fluid CC16 levels may have a diagnostic value at day 7 for BPD in both term and preterm infants. This study demonstrates the potential utility of BAL fluid CC16 levels as a biomarker for BPD in term infants.
Assuntos
Displasia Broncopulmonar/metabolismo , Respiração Artificial/efeitos adversos , Síndrome do Desconforto Respiratório do Recém-Nascido/metabolismo , Uteroglobina/metabolismo , Líquido da Lavagem Broncoalveolar/química , Displasia Broncopulmonar/etiologia , Feminino , Humanos , Recém-Nascido , Recém-Nascido Prematuro , Masculino , Projetos Piloto , Síndrome do Desconforto Respiratório do Recém-Nascido/complicações , Síndrome do Desconforto Respiratório do Recém-Nascido/terapia , Uteroglobina/análiseRESUMO
Atopic asthma is a chronic allergic disease that involves T-helper type 2 (Th2)-inflammation and airway remodeling. Bronchiolar club cells (CC) and alveolar macrophages (AM) are sentinel cells of airway barrier against inhaled injuries, where allergy induces mucous metaplasia of CC and the alternative activation of AM, which compromise host defense mechanisms and amplify Th2-inflammation. As there is evidence that high levels of environmental endotoxin modulates asthma, the goal of this study was to evaluate if the activation of local host defenses by Lipopolysaccharide (LPS) previous to allergy development can contribute to preserving CC and AM protective phenotypes. Endotoxin stimulus before allergen exposition reduced hallmarks of allergic inflammation including eosinophil influx, Interleukin-4 and airway hyperreactivity, while the T-helper type 1 related cytokines IL-12 and Interferon-γ were enhanced. This response was accompanied by the preservation of the normal CC phenotype and the anti-allergic proteins Club Cell Secretory Protein (CCSP) and Surfactant-D, thereby leading to lower levels of CC metaplasia and preventing the increase of the pro-Th2 cytokine Thymic stromal lymphopoietin. In addition, classically activated alveolar macrophages expressing nitric oxide were promoted over the alternatively activated ones that expressed arginase-1. We verified that LPS induced a long-term overexpression of CCSP and the innate immune markers Toll-like receptor 4, and Tumor Necrosis Factor-α, changes that were preserved in spite of the allergen challenge. These results demonstrate that LPS pre-exposition modifies the local bronchioalveolar microenvironment by inducing natural anti-allergic mechanisms while reducing local factors that drive Th2 type responses, thus modulating allergic inflammation.
Assuntos
Asma/imunologia , Macrófagos Alveolares/imunologia , Mucosa Respiratória/citologia , Mucosa Respiratória/imunologia , Animais , Western Blotting , Modelos Animais de Doenças , Endotoxinas/imunologia , Endotoxinas/toxicidade , Ensaio de Imunoadsorção Enzimática , Feminino , Imunofluorescência , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos BALB C , Fenótipo , Uteroglobina/metabolismoRESUMO
To get further insights on the estrogen regulation of the uteroglobin (UG) gene, the 5'-flanking region of the UG gene from the brown hare (Lepus capensis) (Lc) was cloned and compared with those from two phylogenetically related species: the rabbit (Orictolagus cuniculus) (Oc) and the volcano rabbit (Romerolagus diazi) (Rd). The Lc-UG gene is very similar to those from rabbits (94%) and volcano rabbits (95%), and shares a number of genetic elements, including an estrogen response element (ERE). The estrogen-regulated transcription of a series of progressive 5'-deletion mutants of the Lc-UG gene, identified a functional ERE in the promoter region exhibiting the same orientation and relative position than that previously described in rabbits. The Lc-ERE is identical to the Oc-ERE, but different from both the Rd-ERE and the consensus ERE (c-ERE) by one nucleotide. We also detected important species-specific differences in the estrogen-regulated transcription of the UG gene. A luciferase reporter driven by 333 base pairs (bp) of the Lc-UG promoter elicited a higher response to estradiol than its related counterparts when expressed in estrogen-sensitive MCF-7 cells. Several ERE-like motifs which failed to act as functional EREs were also identified; one of them exhibited two mismatches in its palindromic sequence, a characteristic exhibited in many other natural occurring EREs, including the Rd-ERE.
Assuntos
Estrogênios/farmacologia , Lebres/genética , Transcrição Gênica/efeitos dos fármacos , Uteroglobina/genética , Região 5'-Flanqueadora/genética , Animais , Sequência de Bases , Estradiol/farmacologia , Receptor alfa de Estrogênio/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Células MCF-7 , Dados de Sequência Molecular , Motivos de Nucleotídeos/genética , Coelhos , Elementos de Resposta/genética , Homologia de Sequência do Ácido Nucleico , Especificidade da Espécie , Uteroglobina/metabolismoRESUMO
Inherent recipient factors, including pretransplant diagnosis, obesity and elevated pulmonary pressures, are established primary graft dysfunction (PGD) risks. We evaluated the relationship between preoperative lung injury biomarkers and PGD to gain further mechanistic insight in recipients. We performed a prospective cohort study of recipients in the Lung Transplant Outcomes Group enrolled between 2002 and 2010. Our primary outcome was Grade 3 PGD on Day 2 or 3. We measured preoperative plasma levels of five biomarkers (CC-16, sRAGE, ICAM-1, IL-8 and Protein C) that were previously associated with PGD when measured at the postoperative time point. We used multivariable logistic regression to adjust for potential confounders. Of 714 subjects, 130 (18%) developed PGD. Median CC-16 levels were elevated in subjects with PGD (10.1 vs. 6.0, p<0.001). CC-16 was associated with PGD in nonidiopathic pulmonary fibrosis (non-IPF) subjects (OR for highest quartile of CC-16: 2.87, 95% CI: 1.37, 6.00, p=0.005) but not in subjects with IPF (OR 1.38, 95% CI: 0.43, 4.45, p=0.59). After adjustment, preoperative CC-16 levels remained associated with PGD (OR: 3.03, 95% CI: 1.26, 7.30, p=0.013) in non-IPF subjects. Our study suggests the importance of preexisting airway epithelial injury in PGD. Markers of airway epithelial injury may be helpful in pretransplant risk stratification in specific recipients.