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1.
Am J Respir Cell Mol Biol ; 68(3): 267-278, 2023 03.
Artigo em Inglês | MEDLINE | ID: mdl-36287630

RESUMO

Bronchopulmonary dysplasia (BPD) is a common lung disease of premature infants. Hyperoxia exposure and microbial dysbiosis are contributors to BPD development. However, the mechanisms linking pulmonary microbial dysbiosis to worsening lung injury are unknown. Nrf2 (nuclear factor erythroid 2-related factor 2) is a transcription factor that regulates oxidative stress responses and modulates hyperoxia-induced lung injury. We hypothesized that airway dysbiosis would attenuate Nrf2-dependent antioxidant function, resulting in a more severe phenotype of BPD. Here, we show that preterm infants with a Gammaproteobacteria-predominant dysbiosis have increased endotoxin in tracheal aspirates, and mice monocolonized with the representative Gammaproteobacteria Escherichia coli show increased tissue damage compared with germ-free (GF) control mice. Furthermore, we show Nrf2-deficient mice have worse lung structure and function after exposure to hyperoxia when the airway microbiome is augmented with E. coli. To confirm the disease-initiating potential of airway dysbiosis, we developed a novel humanized mouse model by colonizing GF mice with tracheal aspirates from human infants with or without severe BPD, producing gnotobiotic mice with BPD-associated and non-BPD-associated lung microbiomes. After hyperoxia exposure, BPD-associated mice demonstrated a more severe BPD phenotype and increased expression of Nrf2-regulated genes, compared with GF and non-BPD-associated mice. Furthermore, augmenting Nrf2-mediated antioxidant activity by supporting colonization with Lactobacillus species improved dysbiotic-augmented lung injury. Our results demonstrate that a lack of protective pulmonary microbiome signature attenuates an Nrf2-mediated antioxidant response, which is augmented by a respiratory probiotic blend. We anticipate antioxidant pathways will be major targets of future microbiome-based therapeutics for respiratory disease.


Assuntos
Displasia Broncopulmonar , Hiperóxia , Lesão Pulmonar , Pneumonia , Animais , Recém-Nascido , Humanos , Camundongos , Hiperóxia/metabolismo , Lesão Pulmonar/metabolismo , Animais Recém-Nascidos , Antioxidantes , Fator 2 Relacionado a NF-E2/genética , Disbiose , Escherichia coli , Recém-Nascido Prematuro , Pulmão/metabolismo , Displasia Broncopulmonar/metabolismo , Pneumonia/metabolismo , Oxirredução , Modelos Animais de Doenças
2.
Respir Res ; 22(1): 57, 2021 Feb 17.
Artigo em Inglês | MEDLINE | ID: mdl-33596914

RESUMO

BACKGROUND: MicroRNA (miR) are small conserved RNA that regulate gene expression post-transcription. Previous genome-wide analysis studies in preterm infants indicate that pathways of miR 219-5p are important in infants with Bronchopulmonary Dysplasia (BPD). METHODS: Here we report a prospective cohort study of extremely preterm neonates wherein infants diagnosed with severe BPD expressed increased airway miR-219-5p and decreased platelet derived growth factor receptor alpha (PDGFR-α), a target of mir-219-5p and a key regulator of alveolarization, compared to post-conception age-matched term infants. RESULTS: miR-219-5p was highly expressed in the pulmonary epithelial lining in lungs of infants with BPD by in situ hybridization of human infant lungs. In both in vitro and in vivo (mouse) models of BPD, miR-219-5p was increased on exposure to hyperoxia compared with the normoxia control, with a complementary decrease of PDGFR-α. To further confirm the target relationship between miR-219 and PDGFR-α, pulmonary epithelial cells (MLE12) and lung primary fibroblasts were treated with a mimic of miR-219-5p and a locked nucleic acid (LNA) based inhibitor of miR-219-5p. In comparison with the control group, the level of miR-219 increased significantly after miR-219 mimic treatment, while the level of PDGFR-α declined markedly. LNA exposure increased PDGFR-α. Moreover, in BPD mouse model, over-expression of miR-219-5p inhibited alveolar development, indicated by larger alveolar spaces accompanied by reduced septation. CONCLUSIONS: Taken together, our results demonstrate that increased miR-219-5p contributes to the pathogenesis of BPD by targeting and reducing PDGFR-α. The use of specific miRNA antagonists may be a therapeutic strategy for preventing the development of BPD.


Assuntos
Displasia Broncopulmonar/metabolismo , MicroRNAs/biossíntese , Alvéolos Pulmonares/metabolismo , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/antagonistas & inibidores , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/biossíntese , Animais , Animais Recém-Nascidos , Displasia Broncopulmonar/patologia , Displasia Broncopulmonar/terapia , Estudos de Coortes , Pressão Positiva Contínua nas Vias Aéreas/métodos , Humanos , Recém-Nascido , Recém-Nascido Prematuro/metabolismo , Pulmão/metabolismo , Pulmão/patologia , Camundongos , Camundongos Endogâmicos C57BL , Estudos Prospectivos , Alvéolos Pulmonares/patologia
3.
Am J Physiol Renal Physiol ; 320(3): F442-F453, 2021 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33459165

RESUMO

Diabetic nephropathy (DN) is the leading cause of end-stage kidney disease. Besides glycemic and blood pressure control, environmental factors such as cigarette smoking (CS) adversely affect the progression of DN. The effects of CS on DN progression have been attributed to combustion-generated molecules without consideration to the role of nicotine (NIC), responsible for the addictive properties of both CS and electronic cigarettes (ECs). Podocytes are essential to preserve the structure and function of the glomerular filtration barrier, and strong evidence indicates that early podocyte loss promotes DN progression. We performed experiments in human podocytes and in a mouse model of diabetes that develops nephropathy resembling human DN. We determined that NIC binding to podocytes in concentrations achieved with CS and ECs activated NADPH oxidase, which sets in motion a dysfunctional molecular network integrated by cyclooxygenase 2, known to induce podocyte injury; downregulation of AMP-activated protein kinase, important for maintaining cellular energy stores and antioxidation; and upregulation of CD36, which increased lipid uptake and promoted apoptosis. In diabetic mice, NIC increased proteinuria, a recognized marker of chronic kidney disease progression, accompanied by reduced glomerular podocyte synaptopodin, a crucial stabilizer of the podocyte cytoskeleton, and increased fibronectin expression. This novel study critically implicates NIC itself as a contributor to DN progression in CS and EC users.NEW & NOTEWORTHY In this study, we demonstrate that nicotine increases the production of reactive oxygen species, increases cyclooxygenase-2 expression, and upregulates Cd36 while inducing downregulation of AMP-activated protein kinase. In vivo nicotine increases proteinuria and fibronectin expression in diabetic mice. This study demonstrates that effects of nicotine on podocytes are responsible, at least in part, for the deleterious effects of smoking in the progression of chronic kidney disease, including diabetic nephropathy.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Nefropatias Diabéticas/metabolismo , Nicotina/farmacologia , Podócitos/metabolismo , Fumar/efeitos adversos , Animais , Apoptose/efeitos dos fármacos , Diabetes Mellitus Experimental/metabolismo , Nefropatias Diabéticas/tratamento farmacológico , Humanos , Camundongos , Podócitos/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo
4.
Sci Rep ; 10(1): 12035, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32694705

RESUMO

Dry eye affects millions of individuals. In experimental models, dry eye disease is associated with T helper cell 17-mediated inflammation of the ocular surface that may cause persistent damage to the corneal epithelium. However, the initiating and perpetuating factors associated with chronic inflammation of the ocular surface remain unclear. The ocular microbiota alters ocular surface inflammation and may influence dry eye disease development and progression. Here, we collected serial samples of tears on awakening from sleep, closed eye tears, during a randomized clinical trial of a non-pharmaceutical dry eye therapy and used 16S rRNA metabarcoding to characterize the microbiome. We show the closed dry eye microbiome is distinct from the healthy closed eye microbiome, and that the microbiome remains distinct despite daily saline eye wash upon awakening. The ocular microbiome was described only recently, and this report implicates a distinct microbiome in ocular disease development. Our findings suggest an interplay between microbial commensals and inflammation on the ocular surface. This information may inform future studies of the pathophysiological mechanisms of dry eye disease.


Assuntos
Síndromes do Olho Seco/etiologia , Microbiota , Adulto , Estudos de Casos e Controles , Síndromes do Olho Seco/diagnóstico , Feminino , Humanos , Aprendizado de Máquina , Masculino , Metagenômica/métodos , Pessoa de Meia-Idade , RNA Ribossômico 16S/genética , Lágrimas/microbiologia , Índices de Gravidade do Trauma
5.
Am J Physiol Lung Cell Mol Physiol ; 318(2): L421-L428, 2020 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-31644312

RESUMO

Airway microbial dysbiosis is associated with subsequent bronchopulmonary dysplasia (BPD) development in very preterm infants. However, the relationship of airway microbiome in normal pulmonary development has not been defined. To better understand the role of the airway microbiome, we compared normal and abnormal alveolar and pulmonary vascular development in mice with or without a microbiome. We hypothesized that the lungs of germ-free (GF) mice would have an exaggerated phenotypic response to hyperoxia compared with non-germ-free (NGF) mice. With the use of a novel gnotobiotic hyperoxia chamber, GF and NGF mice were exposed to either normoxia or hyperoxia. Alveolar morphometry, pulmonary mechanics, echocardiograms, inflammatory markers, and measures of pulmonary hypertension were studied. GF and NGF mice in normoxia showed no difference, whereas GF mice in hyperoxia showed protected lung structure and mechanics and decreased markers of inflammation compared with NGF mice. We speculate that an increase in abundance of pathogenic bacteria in NGF mice may play a role in BPD pathogenesis by regulating the proinflammatory signaling and neutrophilic inflammation in lungs. Manipulation of the airway microbiome may be a potential therapeutic intervention in BPD and other lung diseases.


Assuntos
Vida Livre de Germes , Hiperóxia/patologia , Alvéolos Pulmonares/irrigação sanguínea , Alvéolos Pulmonares/crescimento & desenvolvimento , Animais , Animais Recém-Nascidos , Fenômenos Biomecânicos , Pressão Sanguínea , Modelos Animais de Doenças , Ventrículos do Coração/patologia , Ventrículos do Coração/fisiopatologia , Hiperóxia/fisiopatologia , Inflamação/complicações , Inflamação/patologia , Camundongos , Camundongos Endogâmicos C57BL , Microvasos/patologia , Alvéolos Pulmonares/fisiopatologia , Sístole
6.
Am J Physiol Lung Cell Mol Physiol ; 317(6): L740-L747, 2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31432715

RESUMO

Hyperoxia-induced oxidant stress contributes to the pathogenesis of bronchopulmonary dysplasia (BPD) in preterm infants. Mitochondrial functional differences due to mitochondrial DNA (mtDNA) variations are important modifiers of oxidant stress responses. The objective of this study was to determine whether mtDNA variation independently modifies lung development and mechanical dysfunction in newborn mice exposed to hyperoxia. Newborn C57BL6 wild type (C57n/C57mt, C57WT) and C3H/HeN wild type (C3Hn/C3Hmt, C3HWT) mice and novel Mitochondrial-nuclear eXchange (MNX) strains with nuclear DNA (nDNA) from their parent strain and mtDNA from the other-C57MNX (C57n/C3Hmt) and C3HMNX (C3Hn/C57mt)-were exposed to 21% or 85% O2 from birth to postnatal day 14 (P14). Lung mechanics and histopathology were examined on P15. Neonatal mouse lung fibroblast (NMLF) bioenergetics and mitochondrial superoxide (O2-) generation were measured. Pulmonary resistance and mitochondrial O2- generation were increased while alveolarization, compliance, and NMLF basal and maximal oxygen consumption rate were decreased in hyperoxia-exposed C57WT mice (C57n/C57mt) versus C57MNX mice (C57n/C3Hmt) and in hyperoxia-exposed C3HMNX mice (C3Hn/C57mt) versus C3HWT (C3Hn/C3Hmt) mice. Our study suggests that neonatal C57 mtDNA-carrying strains have increased hyperoxia-induced hypoalveolarization, pulmonary mechanical dysfunction, and mitochondrial bioenergetic and redox dysfunction versus C3H mtDNA strains. Therefore, mtDNA haplogroup variation-induced differences in mitochondrial function could modify neonatal alveolar development and BPD susceptibility.


Assuntos
DNA Mitocondrial/genética , Modelos Animais de Doenças , Variação Genética , Hiperóxia/fisiopatologia , Pulmão/patologia , Mitocôndrias/patologia , Alvéolos Pulmonares/patologia , Animais , Animais Recém-Nascidos , Metabolismo Energético , Feminino , Hiperóxia/complicações , Pulmão/metabolismo , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , Alvéolos Pulmonares/metabolismo , Superóxidos/metabolismo
7.
Cell ; 176(1-2): 113-126.e15, 2019 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-30633902

RESUMO

Here, we describe a novel pathogenic entity, the activated PMN (polymorphonuclear leukocyte, i.e., neutrophil)-derived exosome. These CD63+/CD66b+ nanovesicles acquire surface-bound neutrophil elastase (NE) during PMN degranulation, NE being oriented in a configuration resistant to α1-antitrypsin (α1AT). These exosomes bind and degrade extracellular matrix (ECM) via the integrin Mac-1 and NE, respectively, causing the hallmarks of chronic obstructive pulmonary disease (COPD). Due to both ECM targeting and α1AT resistance, exosomal NE is far more potent than free NE. Importantly, such PMN-derived exosomes exist in clinical specimens from subjects with COPD but not healthy controls and are capable of transferring a COPD-like phenotype from humans to mice in an NE-driven manner. Similar findings were observed for another neutrophil-driven disease of ECM remodeling (bronchopulmonary dysplasia [BPD]). These findings reveal an unappreciated role for exosomes in the pathogenesis of disorders of ECM homeostasis such as COPD and BPD, providing a critical mechanism for proteolytic damage.


Assuntos
Exossomos/fisiologia , Neutrófilos/metabolismo , Animais , Líquido da Lavagem Broncoalveolar/citologia , Células Cultivadas , Matriz Extracelular/metabolismo , Feminino , Humanos , Inflamação , Integrinas , Elastase de Leucócito/metabolismo , Pulmão/metabolismo , Pulmão/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/fisiologia , Doença Pulmonar Obstrutiva Crônica/fisiopatologia , alfa 1-Antitripsina/metabolismo
8.
Am J Physiol Renal Physiol ; 316(3): F463-F472, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30600684

RESUMO

Heterozygosity for human polycystic kidney and hepatic disease 1 ( PKHD1) mutations was recently associated with cystic liver disease and radiographic findings resembling medullary sponge kidney (MSK). However, the relevance of these associations has been tempered by a lack of cystic liver or renal disease in heterozygous mice carrying Pkhd1 gene trap or exon deletions. To determine whether heterozygosity for a smaller Pkhd1 defect can trigger cystic renal disease in mice, we generated and characterized mice with the predicted truncating Pkhd1C642* mutation in a region corresponding to the middle of exon 20 cluster of five truncating human mutations (between PKHD1G617fs and PKHD1G644*). Mouse heterozygotes or homozygotes for the Pkhd1C642* mutation did not have noticeable liver or renal abnormalities on magnetic resonance images during their first weeks of life. However, when aged to ~1.5 yr, the Pkhd1C642* heterozygotes developed prominent cystic liver changes; tissue analyses revealed biliary cysts and increased number of bile ducts without signs of congenital hepatic fibrosis-like portal field inflammation and fibrosis that was seen in Pkhd1C642* homozygotes. Interestingly, aged female Pkhd1C642* heterozygotes, as well as homozygotes, developed radiographic changes resembling MSK. However, these changes correspond to proximal tubule ectasia, not an MSK-associated collecting duct ectasia. In summary, by demonstrating that cystic liver and kidney abnormalities are triggered by heterozygosity for the Pkhd1C642* mutation, we provide important validation for relevant human association studies. Together, these investigations indicate that PKHD1 mutation heterozygosity (predicted frequency 1 in 70 individuals) is an important underlying cause of cystic liver disorders and MSK-like manifestations in a human population.


Assuntos
Cistos/diagnóstico por imagem , Nefropatias/diagnóstico por imagem , Túbulos Renais Proximais/diagnóstico por imagem , Hepatopatias/diagnóstico por imagem , Rim em Esponja Medular/diagnóstico por imagem , Receptores de Superfície Celular/metabolismo , Animais , Cistos/genética , Cistos/metabolismo , Diagnóstico Diferencial , Dilatação Patológica/diagnóstico por imagem , Dilatação Patológica/genética , Dilatação Patológica/metabolismo , Modelos Animais de Doenças , Nefropatias/genética , Nefropatias/metabolismo , Túbulos Renais Proximais/metabolismo , Hepatopatias/genética , Hepatopatias/metabolismo , Imageamento por Ressonância Magnética , Rim em Esponja Medular/genética , Rim em Esponja Medular/metabolismo , Camundongos , Camundongos Knockout , Receptores de Superfície Celular/genética
9.
JCI Insight ; 3(5)2018 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-29515035

RESUMO

Premature infants are at high risk for developing bronchopulmonary dysplasia (BPD), characterized by chronic inflammation and inhibition of lung development, which we have recently identified as being modulated by microRNAs (miRNAs) and alterations in the airway microbiome. Exosomes and exosomal miRNAs may regulate cell differentiation and tissue and organ development. We discovered that tracheal aspirates from infants with severe BPD had increased numbers of, but smaller, exosomes compared with term controls. Similarly, bronchoalveolar lavage fluid from hyperoxia-exposed mice (an animal model of BPD) and supernatants from hyperoxia-exposed human bronchial epithelial cells (in vitro model of BPD) had increased exosomes compared with air controls. Next, in a prospective cohort study of tracheal aspirates obtained at birth from extremely preterm infants, utilizing independent discovery and validation cohorts, we identified unbiased exosomal miRNA signatures predictive of severe BPD. The strongest signal of reduced miR-876-3p in BPD-susceptible compared with BPD-resistant infants was confirmed in the animal model and in vitro models of BPD. In addition, based on our recent discovery of increased Proteobacteria in the airway microbiome being associated with BPD, we developed potentially novel in vivo and in vitro models for BPD combining Proteobacterial LPS and hyperoxia exposure. Addition of LPS led to a larger reduction in exosomal miR 876-3p in both hyperoxia and normoxia compared with hyperoxia alone, thus indicating a potential mechanism by which alterations in microbiota can suppress miR 876-3p. Gain of function of miR 876-3p improved the alveolar architecture in the in vivo BPD model, demonstrating a causal link between miR 876-3p and BPD. In summary, we provide evidence for the strong predictive biomarker potential of miR 876-3p in severe BPD. We also provide insights on the pathogenesis of neonatal lung disease, as modulated by hyperoxia and microbial product-induced changes in exosomal miRNA 876-3p, which could be targeted for future therapeutic development.


Assuntos
Células Epiteliais Alveolares/imunologia , Displasia Broncopulmonar/diagnóstico , Exossomos/metabolismo , Lactente Extremamente Prematuro/imunologia , MicroRNAs/metabolismo , Células Epiteliais Alveolares/citologia , Células Epiteliais Alveolares/microbiologia , Animais , Animais Recém-Nascidos , Biomarcadores/metabolismo , Líquido da Lavagem Broncoalveolar/citologia , Displasia Broncopulmonar/imunologia , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Linhagem Celular , Modelos Animais de Doenças , Exossomos/genética , Exossomos/imunologia , Feminino , Humanos , Hiperóxia/imunologia , Recém-Nascido de Peso Extremamente Baixo ao Nascer/imunologia , Recém-Nascido , Lipopolissacarídeos/imunologia , Masculino , Camundongos , MicroRNAs/genética , MicroRNAs/imunologia , Microbiota/imunologia , Prognóstico , Estudos Prospectivos , Proteobactérias/imunologia , Índice de Gravidade de Doença
10.
Sci Rep ; 6: 31023, 2016 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-27488092

RESUMO

Alterations of pulmonary microbiome have been recognized in multiple respiratory disorders. It is critically important to ascertain if an airway microbiome exists at birth and if so, whether it is associated with subsequent lung disease. We found an established diverse and similar airway microbiome at birth in both preterm and term infants, which was more diverse and different from that of older preterm infants with established chronic lung disease (bronchopulmonary dysplasia). Consistent temporal dysbiotic changes in the airway microbiome were seen from birth to the development of bronchopulmonary dysplasia in extremely preterm infants. Genus Lactobacillus was decreased at birth in infants with chorioamnionitis and in preterm infants who subsequently went on to develop lung disease. Our results, taken together with previous literature indicating a placental and amniotic fluid microbiome, suggest fetal acquisition of an airway microbiome. We speculate that the early airway microbiome may prime the developing pulmonary immune system, and dysbiosis in its development may set the stage for subsequent lung disease.


Assuntos
Líquido Amniótico/microbiologia , Displasia Broncopulmonar/imunologia , Corioamnionite/microbiologia , Disbiose/microbiologia , Recém-Nascido Prematuro/imunologia , Lactobacillus/fisiologia , Pulmão/microbiologia , Microbiota/imunologia , Placenta/microbiologia , Sistema Respiratório/microbiologia , Feminino , Idade Gestacional , Humanos , Lactente , Recém-Nascido , Parto , Gravidez
11.
Hypertension ; 65(4): 813-20, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25624342

RESUMO

Transcription factor E26 transformation-specific sequence-1 (ETS-1) is a transcription factor that regulates the expression of a variety of genes, including growth factors, chemokines, and adhesion molecules. We recently demonstrated that angiotensin II increases the glomerular expression of ETS-1 and that blockade of ETS-1 ameliorates the profibrotic and proinflammatory effects of angiotensin II. The Dahl salt-sensitive rat is a paradigm of salt-sensitive hypertension associated with local activation of the renin-angiotensin system. In these studies, we determined whether: (1) salt-sensitive hypertension is associated with renal expression of ETS-1 and (2) ETS-1 participates in the development of end-organ injury in salt-sensitive hypertension. Dahl salt-sensitive rats were fed a normal-salt diet (0.5% NaCl diet) or a high-salt diet (4% NaCl) for 4 weeks. Separate groups on high-salt diet received an ETS-1 dominant-negative peptide (10 mg/kg/d), an inactive ETS-1 mutant peptide (10 mg/kg/d), the angiotensin II type 1 receptor blocker candesartan (10 mg/kg/d), or the combination high-salt diet/dominant-negative peptide/angiotensin II type 1 receptor blocker for 4 weeks. High-salt diet rats had a significant increase in the glomerular expression of the phosphorylated ETS-1 that was prevented by angiotensin II type 1 receptor blocker. ETS-1 blockade reduced proteinuria, glomerular injury score, fibronectin expression, urinary transforming growth factor-ß excretion, and macrophage infiltration. Angiotensin II type 1 receptor blocker reduced proteinuria, glomerular injury score, and macrophage infiltration, whereas concomitant ETS-1 blockade and angiotensin II type 1 receptor blocker had additive effects and reduced interstitial fibrosis. Our studies demonstrated that salt-sensitive hypertension results in increased glomerular expression of phosphorylated ETS-1 and suggested that ETS-1 plays an important role in the pathogenesis of end-organ injury in salt-sensitive hypertension.


Assuntos
Injúria Renal Aguda/genética , Angiotensinas/metabolismo , DNA/genética , Regulação da Expressão Gênica , Hipertensão/genética , Glomérulos Renais/metabolismo , Proteína Proto-Oncogênica c-ets-1/genética , Injúria Renal Aguda/etiologia , Injúria Renal Aguda/metabolismo , Animais , Western Blotting , Modelos Animais de Doenças , Hipertensão/complicações , Hipertensão/metabolismo , Imuno-Histoquímica , Glomérulos Renais/patologia , Masculino , Proteína Proto-Oncogênica c-ets-1/biossíntese , Ratos , Ratos Endogâmicos Dahl
12.
Clin Exp Metastasis ; 31(6): 625-38, 2014 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-24832758

RESUMO

A significant percentage of breast cancer victims will suffer from metastases indicating that new approaches to preventing breast cancer metastasis are thus needed. Dietary stearate (ST) and chemotherapy have been shown to reduce breast cancer metastasis. We tested the complementary use of dietary ST with a taxol-based chemotherapy which work through separate mechanisms to reduce breast cancer metastasis. We therefore carried out a prevention study in which diets were initiated prior to human MDA-MB-435 cancer cells being injected into the host and a treatment study in which diets were combined with paclitaxel (PTX). Using an orthotopic athymic nude mouse model and three diets [corn oil (CO) control diet, low fat (LF) or ST] the prevention study demonstrated that the ST diet decreased the incidence of lung metastasis by 50 % compared to both the LF and CO diets. The ST diet also reduced the number and size of metastatic lung nodules compared to the LF diet. Results of the treatment study indicated that both the CO and ST diets decreased the number of mice with lung metastasis compared to the LF diet. Both CO and ST also decreased the number of lung metastases per mouse compared to the LF diet however only the ST diet cohort was significant. Histomorphometric analysis of the lung tumor tissue indicated that the ST diet plus PTX decreased angiogenesis compared to the LF diet plus PTX. In conclusion these results support combining diet with chemotherapy in both treatment and prevention settings.


Assuntos
Neoplasias da Mama/patologia , Dieta , Neoplasias Pulmonares/secundário , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Neoplasias da Mama/dietoterapia , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Comportamento Alimentar , Feminino , Humanos , Neoplasias Pulmonares/dietoterapia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Nus , Paclitaxel/farmacologia , Paclitaxel/uso terapêutico , Ácidos Esteáricos/administração & dosagem , Aumento de Peso
13.
Hypertension ; 60(5): 1226-33, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22966006

RESUMO

Angiotensin II (Ang II) plays a major role in the pathogenesis of end-organ injury in hypertension via its diverse hemodynamic and nonhemodynamic effects. Erythroblastosis virus E26 oncogen homolog-1 (ETS-1) is an important transcription factor recently recognized as an important mediator of cell proliferation, inflammation, and fibrosis. In the present studies, we tested the hypothesis that ETS-1 is a common mediator of the renal proinflammatory and profibrotic effects of Ang II. C57BL6 mice (n=6 per group) were infused with vehicle (control), Ang II (1.4 mg/kg per day), Ang II and an ETS-1 dominant-negative peptide (10 mg/kg per day), or Ang II and an ETS-1 mutant peptide (10 mg/kg per day) via osmotic minipump for 2 or 4 weeks. The infusion of Ang II resulted in significant increases in blood pressure and left ventricular hypertrophy, which were not modified by ETS-1 blockade. The administration of ETS-1 dominant-negative peptide significantly attenuated Ang II-induced renal injury as assessed by urinary protein excretion, mesangial matrix expansion, and cell proliferation. Furthermore, ETS-1 dominant-negative peptide but not ETS-1 mutant peptide significantly reduced Ang II-mediated upregulation of transforming growth factor-ß, connective tissue growth factor, and α-smooth muscle actin. In addition, ETS-1 blockade reduced several proinflammatory effects of Ang II, including macrophage infiltration, nitrotyrosine expression, and NOX4 mRNA expression. Our studies suggest that ETS-1 is a common mediator of the proinflammatory and profibrotic effects of Ang II-induced hypertensive renal damage and may result in the development of novel strategies in the treatment and prevention of end-organ injury in hypertension.


Assuntos
Hipertensão/prevenção & controle , Hipertrofia Ventricular Esquerda/prevenção & controle , Rim/efeitos dos fármacos , Peptídeos/farmacologia , Proteína Proto-Oncogênica c-ets-1/fisiologia , Sequência de Aminoácidos , Angiotensina II , Animais , Pressão Sanguínea/efeitos dos fármacos , Western Blotting , Fibrose/prevenção & controle , Expressão Gênica/efeitos dos fármacos , Hipertensão/induzido quimicamente , Hipertensão/fisiopatologia , Hipertrofia Ventricular Esquerda/induzido quimicamente , Hipertrofia Ventricular Esquerda/fisiopatologia , Imuno-Histoquímica , Inflamação/prevenção & controle , Antígeno Ki-67/metabolismo , Rim/metabolismo , Rim/patologia , Córtex Renal/efeitos dos fármacos , Córtex Renal/metabolismo , Córtex Renal/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Mutação , NADPH Oxidase 4 , NADPH Oxidases/genética , NADPH Oxidases/metabolismo , Proteína Proto-Oncogênica c-ets-1/genética , Proteína Proto-Oncogênica c-ets-1/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa
14.
Am J Physiol Renal Physiol ; 303(2): F304-12, 2012 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-22552933

RESUMO

Clinical studies have established the role of cigarette smoking as a risk factor in the progression of chronic kidney disease (CKD). We have shown that nicotine promotes mesangial cell proliferation and hypertrophy via nonneuronal nicotinic acetylcholine receptors (nAChRs). The α7-nAChR is one of the most important subunits of the nAChRs. These studies were designed to test the hypothesis that nicotine worsens renal injury in rats with 5/6 nephrectomy (5/6Nx) and that the α7-nAChR subunit is required for these effects. We studied five different groups: Sham, 5/6Nx, 5/6Nx + nicotine (Nic; 100 µg/ml dry wt), 5/6Nx + Nic + α7-nAChR blocker methyllicaconitine (MLA; 3 mg·kg(-1)·day(-1) sq), and Sham + Nic. Blood pressure was measured by the tail-cuff method, and urine was collected for proteinuria. After 12 wk, the rats were euthanized and kidneys were collected. We observed expression of the α7-nAChR in the proximal and distal tubules. The administration of nicotine induced a small increase in blood pressure and resulted in cotinine levels similar to those found in the plasma of smokers. In 5/6Nx rats, the administration of nicotine significantly increased urinary protein excretion (onefold), worsened the glomerular injury score and increased fibronectin (∼ 50%), NADPH oxidase 4 (NOX4; ∼100%), and transforming growth factor-ß expression (∼200%). The administration of nicotine to sham rats increased total proteinuria but not albuminuria, suggesting direct effects on tubular protein reabsorption. These effects were prevented by MLA, demonstrating a critical role for the α7-nAChR as a mediator of the effects of nicotine in the progression of CKD.


Assuntos
Progressão da Doença , Nefropatias/fisiopatologia , Nicotina/efeitos adversos , Receptores Nicotínicos/fisiologia , Aconitina/análogos & derivados , Aconitina/farmacologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Pressão Sanguínea/fisiologia , Doença Crônica , Modelos Animais de Doenças , Nefropatias/etiologia , Nefropatias/metabolismo , Masculino , NADPH Oxidase 4 , NADPH Oxidases/metabolismo , Nefrectomia/efeitos adversos , Nicotina/farmacologia , Antagonistas Nicotínicos/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Ratos , Ratos Sprague-Dawley , Receptores Nicotínicos/efeitos dos fármacos , Fator de Crescimento Transformador beta/metabolismo , Receptor Nicotínico de Acetilcolina alfa7
15.
Am J Physiol Renal Physiol ; 302(11): F1418-29, 2012 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-22357921

RESUMO

Angiotensin II (ANG II) produced as result of activation of the renin-angiotensin system (RAS) plays a critical role in the pathogenesis of chronic kidney disease via its hemodynamic effects on the renal microcirculation as well as by its nonhemodynamic actions including the production of extracellular matrix proteins such as fibronectin, a multifunctional extracellular matrix protein that plays a major role in cell adhesion and migration as well as in the development of glomerulosclerosis. ETS-1 is an important transcription factor essential for normal kidney development and glomerular integrity. We previously showed that ANG II increases ETS-1 expression and is required for fibronectin production in mesangial cells. In these studies, we determined that ANG II induces phosphorylation of ETS-1 via activation of the type 1 ANG II receptor and that Erk1/2 and Akt/PKB phosphorylation are required for these effects. In addition, we characterized the role of ETS-1 on the transcriptional activation of fibronectin production in mesangial cells. We determined that ETS-1 directly activates the fibronectin promoter and by utilizing gel shift assays and chromatin immunoprecipitation assays identified two different ETS-1 binding sites that promote the transcriptional activation of fibronectin in response to ANG II. In addition, we identified the essential role of CREB and its coactivator p300 on the transcriptional activation of fibronectin by ETS-1. These studies unveil novel mechanisms involved in RAS-induced production of the extracellular matrix protein fibronectin in mesangial cells and establish the role of the transcription factor ETS-1 as a direct mediator of these effects.


Assuntos
Angiotensina II/farmacologia , Fibronectinas/biossíntese , Mesângio Glomerular/metabolismo , Proteína Proto-Oncogênica c-ets-1/fisiologia , Animais , Western Blotting , Linhagem Celular , Núcleo Celular/metabolismo , Imunoprecipitação da Cromatina , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/fisiologia , Proteína p300 Associada a E1A/fisiologia , Ensaio de Desvio de Mobilidade Eletroforética , Matriz Extracelular/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/fisiologia , Genes Reporter/genética , Mesângio Glomerular/citologia , Mesângio Glomerular/efeitos dos fármacos , Imunoprecipitação , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Plasmídeos/genética , Proteína Proto-Oncogênica c-ets-1/genética , Ratos , Estimulação Química , Transcrição Gênica , Transfecção
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