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1.
PLoS One ; 15(1): e0220019, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31945053

RESUMO

The migration of cancer cells is highly regulated by the biomechanical properties of their local microenvironment. Using 3D scaffolds of simple composition, several aspects of cancer cell mechanosensing (signal transduction, EMC remodeling, traction forces) have been separately analyzed in the context of cell migration. However, a combined study of these factors in 3D scaffolds that more closely resemble the complex microenvironment of the cancer ECM is still missing. Here, we present a comprehensive, quantitative analysis of the role of cell-ECM interactions in cancer cell migration within a highly physiological environment consisting of mixed Matrigel-collagen hydrogel scaffolds of increasing complexity that mimic the tumor microenvironment at the leading edge of cancer invasion. We quantitatively show that the presence of Matrigel increases hydrogel stiffness, which promotes ß1 integrin expression and metalloproteinase activity in H1299 lung cancer cells. Then, we show that ECM remodeling activity causes matrix alignment and compaction that favors higher tractions exerted by the cells. However, these traction forces do not linearly translate into increased motility due to a biphasic role of cell adhesions in cell migration: at low concentration Matrigel promotes migration-effective tractions exerted through a high number of small sized focal adhesions. However, at high Matrigel concentration, traction forces are exerted through fewer, but larger focal adhesions that favor attachment yielding lower cell motility.


Assuntos
Colágeno/farmacologia , Células Epiteliais/efeitos dos fármacos , Matriz Extracelular/efeitos dos fármacos , Adesões Focais/efeitos dos fármacos , Laminina/farmacologia , Mecanotransdução Celular , Proteoglicanas/farmacologia , Adesão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Colágeno/química , Combinação de Medicamentos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Matriz Extracelular/química , Matriz Extracelular/metabolismo , Adesões Focais/ultraestrutura , Expressão Gênica , Humanos , Integrina beta1/genética , Integrina beta1/metabolismo , Laminina/química , Modelos Biológicos , Proteoglicanas/química , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Propriedades de Superfície , Microambiente Tumoral/efeitos dos fármacos
2.
Life Sci ; 242: 117213, 2020 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-31881228

RESUMO

Acute respiratory distress syndrome (ARDS) is a multifactorial, inflammatory lung injury disease with high morbidity and mortality. However, the underlying pathogenic mechanism remains unknown. In this study, lipopolysaccharide (LPS)-stimulated alveolar epithelial cells were used to mimic the inflammatory pathogenesis of ARDS in vitro. We here investigated the role of miR-424 in LPS-stimulated alveolar epithelial cells and found it to be substantially downregulated. Overexpression of miR-424 inhibited apoptosis and inflammation in LPS-stimulated alveolar epithelial cells, and the miR-424 inhibitor exhibited the opposite effect. A bioinformatic analysis revealed a potential binding site of miR-424 in the 3'-UTR of fibroblast growth factor 2 (FGF2). A luciferase reporter assay suggested that miR-424 targeted FGF2 in alveolar epithelial cells. The level of FGF2 protein was inhibited by miR-424 mimic, whereas was significantly upregulated after miR-424 suppression in LPS-stimulated alveolar epithelial cells. MiR-424 also exhibited the protective role in LPS-induced apoptosis and inflammation by directly targeting FGF2 via the NF-κB pathway. In conclusion, our results demonstrate that miR-424 had a protective role in LPS-induced apoptosis and inflammation of alveolar epithelial cells by targeting FGF2 via regulating NF-κB pathway. This might contribute novel evidence to help identify a therapeutic target for treating ARDS.


Assuntos
Células A549/metabolismo , Apoptose/efeitos dos fármacos , Fator 2 de Crescimento de Fibroblastos/fisiologia , Inflamação/fisiopatologia , Lipopolissacarídeos/farmacologia , MicroRNAs/metabolismo , NF-kappa B/metabolismo , Alvéolos Pulmonares/metabolismo , Mucosa Respiratória/metabolismo , Transdução de Sinais , Células A549/fisiologia , Apoptose/fisiologia , Western Blotting , Fator 2 de Crescimento de Fibroblastos/metabolismo , Imunofluorescência , Humanos , Inflamação/metabolismo , MicroRNAs/fisiologia , Alvéolos Pulmonares/citologia , Alvéolos Pulmonares/fisiologia , Reação em Cadeia da Polimerase em Tempo Real , Mucosa Respiratória/citologia , Mucosa Respiratória/fisiologia , Transdução de Sinais/fisiologia
3.
Medicine (Baltimore) ; 98(38): e17267, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31568004

RESUMO

Smoking is a substantial risk factor for many respiratory diseases. This study aimed to identify the gene and microRNA changes related to smoking in human airway epithelium by bioinformatics analysis.From the Gene Expression Omnibus (GEO) database, the mRNA datasets GSE11906, GSE22047, GSE63127, and microRNA dataset GSE14634 were downloaded, and were analyzed using GEO2R. Functional enrichment analysis of the differentially expressed genes (DEGs) was enforced using DAVID. The protein-protein interaction (PPI) network and differentially expressed miRNAs (DEMs)- DEGs network were executed by Cytoscape.In total, 107 DEGs and 10 DEMs were determined. Gene Ontology (GO) analysis revealed that DEGs principally enriched in oxidation-reduction process, extracellular space and oxidoreductase activity. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway demonstrated that DEGs were principally enriched in metabolism of xenobiotics by cytochrome P450 and chemical carcinogenesis. The PPI network revealed 15 hub genes, including NQO1, CYP1B1, AKR1C1, CYP1A1, AKR1C3, CEACAM5, MUCL1, B3GNT6, MUC5AC, MUC12, PTGER4, CALCA, CBR1, TXNRD1, and CBR3. Cluster analysis showed that these hub genes were associated with adenocarcinoma in situ, squamous cell carcinoma, cell differentiation, inflammatory response, oxidative DNA damage, oxidative stress response and tumor necrosis factor. Hsa-miR-627-5p might have the most target genes, including ITLN1, TIMP3, PPP4R4, SLC1A2, NOVA1, RNFT2, CLDN10, TMCC3, EPHA7, SRPX2, PPP1R16B, GRM1, HS3ST3A1, SFRP2, SLC7A11, and KLHDC8A.We identified several molecular changes induced by smoking in human airway epithelium. This study may provide some candidate genes and microRNAs for assessing the risk of lung diseases caused by smoking.


Assuntos
Expressão Gênica/efeitos dos fármacos , MicroRNAs/metabolismo , Mucosa Respiratória/metabolismo , Fumar/efeitos adversos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Mapas de Interação de Proteínas/efeitos dos fármacos , Mucosa Respiratória/efeitos dos fármacos , Fumar/metabolismo
4.
Int J Mol Sci ; 20(19)2019 Oct 05.
Artigo em Inglês | MEDLINE | ID: mdl-31590401

RESUMO

Mutations in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene lead to cystic fibrosis (CF). The most common mutation F508del inhibits folding and processing of CFTR protein. FDA-approved correctors rescue the biosynthetic processing of F508del-CFTR protein, while potentiators improve the rescued CFTR channel function. Transforming growth factor (TGF-ß1), overexpressed in many CF patients, blocks corrector/potentiator rescue by inhibiting CFTR mRNA in vitro. Increased TGF-ß1 signaling and acquired CFTR dysfunction are present in other lung diseases. To study the mechanism of TGF-ß1 repression of CFTR, we used molecular, biochemical, and functional approaches in primary human bronchial epithelial cells from over 50 donors. TGF-ß1 destabilized CFTR mRNA in cells from lungs with chronic disease, including CF, and impaired F508del-CFTR rescue by new-generation correctors. TGF-ß1 increased the active pool of selected micro(mi)RNAs validated as CFTR inhibitors, recruiting them to the RNA-induced silencing complex (RISC). Expression of F508del-CFTR globally modulated TGF-ß1-induced changes in the miRNA landscape, creating a permissive environment required for degradation of F508del-CFTR mRNA. In conclusion, TGF-ß1 may impede the full benefit of corrector/potentiator therapy in CF patients. Studying miRNA recruitment to RISC under disease-specific conditions may help to better characterize the miRNAs utilized by TGF-ß1 to destabilize CFTR mRNA.


Assuntos
Brônquios/metabolismo , Regulador de Condutância Transmembrana em Fibrose Cística/genética , MicroRNAs/metabolismo , Estabilidade de RNA , Mucosa Respiratória/metabolismo , Fator de Crescimento Transformador beta/farmacologia , Brônquios/citologia , Células Cultivadas , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Inativação Gênica , Humanos , MicroRNAs/genética , Mucosa Respiratória/efeitos dos fármacos
5.
Toxicol Lett ; 317: 1-12, 2019 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-31562913

RESUMO

During extrusion of some polymers, fused filament fabrication (FFF) 3-D printers emit billions of particles per minute and numerous organic compounds. The scope of this study was to evaluate FFF 3-D printer emission-induced toxicity in human small airway epithelial cells (SAEC). Emissions were generated from a commercially available 3-D printer inside a chamber, while operating for 1.5 h with acrylonitrile butadiene styrene (ABS) or polycarbonate (PC) filaments, and collected in cell culture medium. Characterization of the culture medium revealed that repeat print runs with an identical filament yield various amounts of particles and organic compounds. Mean particle sizes in cell culture medium were 201 ±â€¯18 nm and 202 ±â€¯8 nm for PC and ABS, respectively. At 24 h post-exposure, both PC and ABS emissions induced a dose dependent significant cytotoxicity, oxidative stress, apoptosis, necrosis, and production of pro-inflammatory cytokines and chemokines in SAEC. Though the emissions may not completely represent all possible exposure scenarios, this study indicate that the FFF could induce toxicological effects. Further studies are needed to quantify the detected chemicals in the emissions and their corresponding toxicological effects.


Assuntos
Resinas Acrílicas/toxicidade , Butadienos/toxicidade , Células Epiteliais/efeitos dos fármacos , Nanopartículas/toxicidade , Cimento de Policarboxilato/toxicidade , Poliestirenos/toxicidade , Impressão Tridimensional , Mucosa Respiratória/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Células Cultivadas , Citocinas/metabolismo , Relação Dose-Resposta a Droga , Células Epiteliais/metabolismo , Células Epiteliais/ultraestrutura , Humanos , Mediadores da Inflamação/metabolismo , Necrose , Estresse Oxidativo/efeitos dos fármacos , Tamanho da Partícula , Mucosa Respiratória/metabolismo , Mucosa Respiratória/ultraestrutura , Medição de Risco , Fatores de Tempo
6.
Respir Res ; 20(1): 172, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31370853

RESUMO

Genome wide association (GWA) studies have reproducibly identified signals on chromosome 4q24 associated with lung function and COPD. GSTCD (Glutathione S-transferase C-terminal domain containing) represents a candidate causal gene in this locus, however little is currently known about the function of this protein. We set out to further our understanding of the role of GSTCD in cell functions and homeostasis using multiple molecular and cellular approaches in airway relevant cells. Recombinant expression of human GSTCD in conjunction with a GST activity assay did not identify any enzymatic activity for two GSTCD isoforms questioning the assignment of this protein to this family of enzymes. Protein structure analyses identified a potential methyltransferase domain contained within GSTCD, with these enzymes linked to cell viability and apoptosis. Targeted knockdown (siRNA) of GSTCD in bronchial epithelial cells identified a role for GSTCD in cell viability as proliferation rates were not altered. To provide greater insight we completed transcriptomic analyses on cells with GSTCD expression knocked down and identified several differentially expressed genes including those implicated in airway biology; fibrosis e.g. TGFBR1 and inflammation e.g. IL6R. Pathway based transcriptomic analyses identified an over-representation of genes related to adipogenesis which may suggest additional functions for GSTCD. These findings identify potential additional functions for GSTCD in the context of airway biology beyond the hypothesised GST activity and warrant further investigation.


Assuntos
Estudo de Associação Genômica Ampla/métodos , Homeostase/fisiologia , Pulmão/fisiologia , Miócitos de Músculo Liso/fisiologia , Proteínas/genética , Mucosa Respiratória/metabolismo , Animais , Células CHO , Cricetinae , Cricetulus , Humanos , Pulmão/citologia , Proteínas/metabolismo , Mucosa Respiratória/citologia
7.
Respir Res ; 20(1): 181, 2019 Aug 09.
Artigo em Inglês | MEDLINE | ID: mdl-31399087

RESUMO

BACKGROUND: KRAS is a GTPase that activates pathways involved in cell growth, differentiation and survival. In normal cells, KRAS-activity is tightly controlled, but with specific mutations, the KRAS protein is persistently activated, giving cells a growth advantage resulting in cancer. While a great deal of attention has been focused on the role of mutated KRAS as a common driver mutation for lung adenocarcinoma, little is known about the role of KRAS in regulating normal human airway differentiation. METHODS: To assess the role of KRAS signaling in regulating differentiation of the human airway epithelium, primary human airway basal stem/progenitor cells (BC) from nonsmokers were cultured on air-liquid interface (ALI) cultures to mimic the airway epithelium in vitro. Modulation of KRAS signaling was achieved using siRNA-mediated knockdown of KRAS or lentivirus-mediated over-expression of wild-type KRAS or the constitutively active G12 V mutant. The impact on differentiation was quantified using TaqMan quantitative PCR, immunofluorescent and immunohistochemical staining analysis for cell type specific markers. Finally, the impact of cigarette smoke exposure on KRAS and RAS protein family activity in the airway epithelium was assessed in vitro and in vivo. RESULTS: siRNA-mediated knockdown of KRAS decreased differentiation of BC into secretory and ciliated cells with a corresponding shift toward squamous cell differentiation. Conversely, activation of KRAS signaling via lentivirus mediated over-expression of the constitutively active G12 V KRAS mutant had the opposite effect, resulting in increased secretory and ciliated cell differentiation and decreased squamous cell differentiation. Exposure of BC to cigarette smoke extract increased KRAS and RAS protein family activation in vitro. Consistent with these observations, airway epithelium brushed from healthy smokers had elevated RAS activation compared to nonsmokers. CONCLUSIONS: Together, these data suggest that KRAS-dependent signaling plays an important role in regulating the balance of secretory, ciliated and squamous cell differentiation of the human airway epithelium and that cigarette smoking-induced airway epithelial remodeling is mediated in part by abnormal activation of KRAS-dependent signaling mechanisms.


Assuntos
Diferenciação Celular/fisiologia , Fumar Cigarros/efeitos adversos , Fumar Cigarros/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/fisiologia , Mucosa Respiratória/metabolismo , Poluição por Fumaça de Tabaco/efeitos adversos , Adulto , Remodelação das Vias Aéreas/efeitos dos fármacos , Remodelação das Vias Aéreas/fisiologia , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Fumar Cigarros/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/patologia , Adulto Jovem
8.
Acta Histochem ; 121(7): 852-865, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31445760

RESUMO

Anatolian ground squirrel (Spermophilus xanthoprymnus) is a true hibernator. This animal transiently reduces pulmonary function during hibernation. Continuance of pulmonary function is very important to survive ground squirrels during the hibernation. Natriuretic peptides may be key players in the modulation of pulmonary hemostasis. However, NPs' role in pulmonary function during hibernation remains unclear. We aimed to investigate the localization and distribution of atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP) and C-type natriuretic peptide (CNP) in squirrel lungs during pre-hibernation and hibernation periods using immunohistochemistry. Our immunohistochemical data indicate that ANP, BNP, and CNP were produced by the mucosal epithelium of terminal and respiratory bronchioles, smooth muscle cells in the lamina propria of terminal bronchioles and vascular smooth muscle cells, alveolar type II cells, and macrophages. ANP immunoreactivity was weaker than BNP and CNP immunoreactivities in these cells. The results also demonstrate that the number of ANP, BNP and CNP positive alveolar type II cells tended to increase, although statistically non-significant, during the hibernation period, but the expression of NPs in other pulmonary cells is unaffected by hibernation. This study firstly investigates ANP, BNP and CNP distribution in the Anatolian ground squirrel lung. However, further studies are required to dissect their functional roles during the hibernation.


Assuntos
Regulação da Expressão Gênica/fisiologia , Hibernação/fisiologia , Pulmão , Peptídeos Natriuréticos/biossíntese , Mucosa Respiratória , Sciuridae/metabolismo , Animais , Imuno-Histoquímica , Pulmão/citologia , Pulmão/metabolismo , Masculino , Mucosa Respiratória/citologia , Mucosa Respiratória/metabolismo
9.
Nat Commun ; 10(1): 3841, 2019 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-31451696

RESUMO

Human lung tissue-resident NK cells (trNK cells) are likely to play an important role in host responses towards viral infections, inflammatory conditions and cancer. However, detailed insights into these cells are still largely lacking. Here we show, using RNA sequencing and flow cytometry-based analyses, that subsets of human lung CD69+CD16- NK cells display hallmarks of tissue-residency, including high expression of CD49a, CD103, and ZNF683, and reduced expression of SELL, S1PR5, and KLF2/3. CD49a+CD16- NK cells are functionally competent, and produce IFN-γ, TNF, MIP-1ß, and GM-CSF. After stimulation with IL-15, they upregulate perforin, granzyme B, and Ki67 to a similar degree as CD49a-CD16- NK cells. Comparing datasets from trNK cells in human lung and bone marrow with tissue-resident memory CD8+ T cells identifies core genes co-regulated either by tissue-residency, cell-type or location. Together, our data indicate that human lung trNK cells have distinct features, likely regulating their function in barrier immunity.


Assuntos
Imunidade nas Mucosas , Células Matadoras Naturais/metabolismo , Pneumopatias/imunologia , Pulmão/citologia , Transcriptoma/imunologia , Idoso , Idoso de 80 Anos ou mais , Células da Medula Óssea/imunologia , Células da Medula Óssea/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Conjuntos de Dados como Assunto , Feminino , Humanos , Células Matadoras Naturais/imunologia , Pulmão/imunologia , Pulmão/cirurgia , Pneumopatias/patologia , Pneumopatias/cirurgia , Masculino , Pessoa de Meia-Idade , Pneumonectomia , Mucosa Respiratória/citologia , Mucosa Respiratória/imunologia , Mucosa Respiratória/metabolismo
10.
Nutrients ; 11(9)2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31466230

RESUMO

Taste receptors, first identified on the tongue, are best known for their role in guiding our dietary preferences. The expression of taste receptors for umami, sweet, and bitter have been demonstrated in tissues outside of the oral cavity, including in the airway, brain, gastrointestinal tract, and reproductive organs. The extra-oral taste receptor chemosensory pathways and the endogenous taste receptor ligands are generally unknown, but there is increasing data suggesting that taste receptors are involved in regulating some aspects of innate immunity, and may potentially control the composition of the nasal microbiome in healthy individuals or patients with upper respiratory diseases like chronic rhinosinusitis (CRS). For this reason, taste receptors may serve as potential therapeutic targets, providing alternatives to conventional antibiotics. This review focuses on the physiology of sweet (T1R) and bitter (T2R) taste receptors in the airway and their activation by secreted bacterial products. There is particular focus on T2R38 in sinonasal ciliated cells, as well as the sweet and bitter receptors found on specialized sinonasal solitary chemosensory cells. Additionally, this review explores the impact of genetic variations in these receptors on the differential susceptibility of patients to upper airway infections, such as CRS.


Assuntos
Imunidade Inata , Imunidade nas Mucosas , Receptores Acoplados a Proteínas-G/metabolismo , Mucosa Respiratória/metabolismo , Sistema Respiratório/metabolismo , Infecções Respiratórias/metabolismo , Paladar , Animais , Antibacterianos/uso terapêutico , Bactérias/imunologia , Bactérias/metabolismo , Cílios/imunologia , Cílios/metabolismo , Interações Hospedeiro-Patógeno , Humanos , Imunidade Inata/efeitos dos fármacos , Imunidade nas Mucosas/efeitos dos fármacos , Receptores Acoplados a Proteínas-G/efeitos dos fármacos , Receptores Acoplados a Proteínas-G/imunologia , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/imunologia , Mucosa Respiratória/microbiologia , Sistema Respiratório/efeitos dos fármacos , Sistema Respiratório/imunologia , Sistema Respiratório/microbiologia , Infecções Respiratórias/tratamento farmacológico , Infecções Respiratórias/imunologia , Infecções Respiratórias/microbiologia , Transdução de Sinais , Paladar/efeitos dos fármacos
11.
Mol Med Rep ; 20(2): 1569-1574, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31257531

RESUMO

Short­chain fatty acids (SCFAs; butyrate, propionate and acetate) are metabolites derived from the gut microbiota via dietary fiber fermentation. In colon cancer, treatment with SCFAs, mainly butyrate and propionate, suppresses cell proliferation, migration and invasion. Furthermore, although sodium butyrate is known to induce cell apoptosis in lung cancer, the anticancer effects of sodium propionate (SP) on lung cancer are not well understood. In the present study, SP treatment induced cell cycle arrest, especially in the G2/M phase, and cell apoptosis in the H1299 and H1703 lung cancer cell lines. As determined by reverse transcription­quantitative PCR and western blotting, Survivin and p21 expression levels were significantly affected by SP treatment, suggesting that SP treatment suppressed cell proliferation in these lung cancer cell lines. Thus, it was proposed that the SP­mediated regulation of Survivin and p21 in lung cancer may be applicable to lung cancer therapy.


Assuntos
Antineoplásicos/farmacologia , Ácido Butírico/farmacologia , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica , Propionatos/farmacologia , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p21/agonistas , Inibidor de Quinase Dependente de Ciclina p21/genética , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Pontos de Checagem da Fase G2 do Ciclo Celular/genética , Microbioma Gastrointestinal/fisiologia , Humanos , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Transdução de Sinais , Survivina/antagonistas & inibidores , Survivina/genética , Survivina/metabolismo
12.
Immunol Allergy Clin North Am ; 39(3): 309-319, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31284922

RESUMO

The infant's developing immune response is central to establishing a balanced system that reacts appropriately to infectious stimuli, but does not induce altered disease states with potential long-term sequelae. Respiratory syncytial virus may alter the immune system, affecting future responses. Early infection may have direct effects on the lung itself. Other early life processes contribute to the development of immune responses including assembly of the microbiome, which seems to have a particularly important role for establishing the immune environment. This review covers studies that have set up important paradigms and discusses recent data that direct research toward informative hypotheses.


Assuntos
Asma/etiologia , Infecções por Vírus Respiratório Sincicial/complicações , Infecções por Vírus Respiratório Sincicial/virologia , Vírus Sinciciais Respiratórios , Imunidade Adaptativa/genética , Fatores Etários , Animais , Asma/epidemiologia , Suscetibilidade a Doenças , Metabolismo Energético , Epigênese Genética , Microbioma Gastrointestinal/imunologia , Humanos , Imunidade Inata/genética , Mucosa Respiratória/imunologia , Mucosa Respiratória/metabolismo , Mucosa Respiratória/virologia , Infecções por Vírus Respiratório Sincicial/imunologia , Vírus Sinciciais Respiratórios/imunologia
13.
Part Fibre Toxicol ; 16(1): 29, 2019 07 09.
Artigo em Inglês | MEDLINE | ID: mdl-31288843

RESUMO

BACKGROUND: Industrially produced quantities of TiO2 nanoparticles are steadily rising, leading to an increasing risk of inhalation exposure for both professionals and consumers. Particle inhalation can result in inflammatory and allergic responses, and there are concerns about other negative health effects from either acute or chronic low-dose exposure. RESULTS: To study the fate of inhaled TiO2-NP, adult rats were exposed to 2-h intra-tracheal inhalations of 48V-radiolabeled, 20 nm TiO2-NP aerosols (deposited NP-mass 1.4 ± 0.5 µg). At five time points (1 h, 4 h, 24 h, 7d, 28d) post-exposure, a complete balance of the [48V]TiO2-NP fate was quantified in organs, tissues, carcass, lavage and body fluids, including excretions. After fast mucociliary airway clearance (fractional range 0.16-0.31), long-term macrophage-mediated clearance (LT-MC) from the alveolar region is 2.6-fold higher after 28d (integral fraction 0.40 ± 0.04) than translocation across the air-blood-barrier (integral fraction 0.15 ± 0.01). A high NP fraction remains in the alveoli (0.44 ± 0.05 after 28d), half of these on the alveolar epithelium and half in interstitial spaces. There is clearance from both retention sites at fractional rates (0.02-0.03 d- 1) by LT-MC. Prior to LT-MC, [48V]TiO2-NP are re-entrained to the epithelium as reported earlier for 20 nm inhaled gold-NP (AuNP) and iridium-NP (IrNP). CONCLUSION: Comparing the 28-day biokinetics patterns of three different inhaled NP materials TiO2-NP, AuNP and IrNP, the long-term kinetics of interstitial relocation and subsequent re-entrainment onto the lung-epithelium is similar for AuNP and Ir-NP but slower than for TiO2-NP. We discuss mechanisms and pathways of NP relocation and re-entrainment versus translocation. Additionally, after 28 days the integral translocated fractions of TiO2-NP and IrNP across the air-blood-barrier (ABB) are similar and become 0.15 while the translocated AuNP fraction is only 0.04. While NP dissolution proved negligible, translocated TiO2-NP and IrNP are predominantly excreted in urine (~ 0.1) while the urinary AuNP excretion amounts to a fraction of only 0.01. Urinary AuNP excretion is below 0.0001 during the first week but rises tenfold thereafter suggesting delayed disagglomeration. Of note, all three NP dissolve minimally, since no ionic radio-label release was detectable. These biokinetics data of inhaled, same-sized NP suggest significant time-dependent differences of the ABB translocation and subsequent fate in the organism.


Assuntos
Exposição por Inalação/análise , Pulmão/metabolismo , Nanopartículas/química , Titânio/farmacocinética , Aerossóis , Animais , Líquido da Lavagem Broncoalveolar , Feminino , Taxa de Depuração Metabólica , Especificidade de Órgãos , Tamanho da Partícula , Ratos , Ratos Endogâmicos WKY , Mucosa Respiratória/metabolismo , Fatores de Tempo , Distribuição Tecidual , Titânio/química
14.
Cells ; 8(7)2019 07 18.
Artigo em Inglês | MEDLINE | ID: mdl-31323757

RESUMO

Mucociliary clearance is an important innate host defense of the mammalian respiratory system, as it traps foreign substances, including pollutants, pathogens, and allergens, and transports them out of the airway. The underlying mechanism of the actuation and coordination of cilia, the interplay between the cilia and mucus, and the formation of the metachronal wave have been explored extensively both experimentally and mathematically. In this mini-review, we provide a survey of the mathematical models of mucociliary clearance, from the motion of one single cilium to the emergence of the metachronal wave in a group of them, from the fundamental theoretical study to the state-of-the-art three-dimensional simulations. The mechanism of cilium actuation is discussed, together with the mathematical simplification and the implications or caveats of the results.


Assuntos
Modelos Teóricos , Depuração Mucociliar , Animais , Cílios/metabolismo , Cílios/ultraestrutura , Humanos , Mucosa Respiratória/metabolismo
15.
Int J Mol Sci ; 20(14)2019 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-31340547

RESUMO

BACKGROUND: Lung cancer cells are known to change proliferation and migration under simulated microgravity. In this study, we sought to evaluate cell adherence, apoptosis, cytoskeleton arrangement, and gene expression under simulated microgravity. METHODS: Human lung cancer cells were exposed to simulated microgravity in a random-positioning machine (RPM). Cell morphology and adherence were observed under phase-contrast microscopy, cytoskeleton staining was performed, apoptosis rate was determined, and changes in gene and protein expression were detected by real-time PCR with western blot confirmation. RESULTS: Three-dimensional (3D)-spheroid formation was observed under simulated microgravity. Cell viability was not impaired. Actin filaments showed a shift in alignment from longitudinal to spherical. Apoptosis rate was significantly increased in the spheroids compared to the control. TP53, CDKN2A, PTEN, and RB1 gene expression was significantly upregulated in the adherent cells under simulated microgravity with an increase in corresponding protein production for p14 and RB1. SOX2 expression was significantly upregulated in the adherent cells, but protein was not. Gene expressions of AKT3, PIK3CA, and NFE2L2 remained unaltered. CONCLUSION: Simulated microgravity induces alteration in cell adherence, increases apoptosis rate, and leads to upregulation of tumor suppressor genes in human lung cancer cells.


Assuntos
Apoptose/genética , Adesão Celular/genética , Células Epiteliais/metabolismo , Regulação Neoplásica da Expressão Gênica , Ausência de Peso , Citoesqueleto de Actina/metabolismo , Citoesqueleto de Actina/ultraestrutura , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Classe I de Fosfatidilinositol 3-Quinases/genética , Classe I de Fosfatidilinositol 3-Quinases/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Células Epiteliais/ultraestrutura , Humanos , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Proteínas de Ligação a Retinoblastoma/genética , Proteínas de Ligação a Retinoblastoma/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Transdução de Sinais , Esferoides Celulares/metabolismo , Esferoides Celulares/patologia , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Simulação de Ausência de Peso/instrumentação , Simulação de Ausência de Peso/métodos
17.
Respir Res ; 20(1): 129, 2019 Jun 24.
Artigo em Inglês | MEDLINE | ID: mdl-31234850

RESUMO

BACKGROUND: Azithromycin (Azm) is a macrolide recognized for its disease-modifying effects and reduction in exacerbation of chronic airway diseases. It is not clear whether the beneficial effects of Azm are due to its anti-microbial activity or other pharmacological actions. We have shown that Azm affects the integrity of the bronchial epithelial barrier measured by increased transepithelial electrical resistance. To better understand these effects of Azm on bronchial epithelia we have investigated global changes in gene expression. METHODS: VA10 bronchial epithelial cells were treated with Azm and cultivated in air-liquid interface conditions for up to 22 days. RNA was isolated at days 4, 10 and 22 and analyzed using high-throughput RNA sequencing. qPCR and immunostaining were used to confirm key findings from bioinformatic analyses. Detailed assessment of cellular changes was done using microscopy, followed by characterization of the lipidomic profiles of the multivesicular bodies present. RESULTS: Bioinformatic analysis revealed that after 10 days of treatment genes encoding effectors of sterol and cholesterol metabolism were prominent. Interestingly, expression of genes associated with epidermal barrier differentiation, KRT1, CRNN, SPINK5 and DSG1, increased significantly at day 22. Together with immunostaining, these results suggest an epidermal differentiation pattern. We also found that Azm induced the formation of multivesicular and lamellar bodies in two different airway epithelial cell lines. Lipidomic analysis revealed that Azm was entrapped in multivesicular bodies linked to different types of lipids, most notably palmitate and stearate. Furthermore, targeted analysis of lipid species showed accumulation of phosphatidylcholines, as well as ceramide derivatives. CONCLUSIONS: Taken together, we demonstrate how Azm might confer its barrier enhancing effects, via activation of epidermal characteristics and changes to intracellular lipid dynamics. These effects of Azm could explain the unexpected clinical benefit observed during Azm-treatment of patients with various lung diseases affecting barrier function.


Assuntos
Antibacterianos/farmacologia , Azitromicina/farmacologia , Diferenciação Celular/efeitos dos fármacos , Epiderme/efeitos dos fármacos , Corpos Multivesiculares/efeitos dos fármacos , Mucosa Respiratória/efeitos dos fármacos , Diferenciação Celular/fisiologia , Linhagem Celular , Epiderme/metabolismo , Humanos , Corpos Multivesiculares/metabolismo , Mucosa Respiratória/citologia , Mucosa Respiratória/metabolismo
18.
Int Immunopharmacol ; 74: 105706, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31254955

RESUMO

Linalool is a natural product present in fruits and aromatic plants with biological activities. Researchers have reported that the inhalation of linalool exerts anti-inflammatory activities. In this study, we examined the therapeutic effects of linalool on airway inflammation and mucus overproduction in mice with allergic asthma. Oral administration of linalool significantly inhibited the levels of eosinophil numbers, Th2 cytokines and immunoglobulin E (IgE) caused by ovalbumin (OVA) exposure. Linalool exerted preventive effects against the influx of inflammatory cells and mucus hypersecretion in the lung tissues. Linalool also dose-dependently decreased the levels of inducible nitric oxide synthase (iNOS) expression and protein kinase B (AKT) activation in the lung tissues. Linalool effectively downregulated the activation of mitogen-activated protein kinases (MAPKs) and nuclear factor-κB (NF-κB) caused by OVA exposure. Furthermore, linalool exerted inhibitory effect on OVA-induced airway hyperresponsiveness (AHR). In the in vitro study, the increased secretion of MCP-1 was attenuated with linalool treatment in lipopolysaccharide (LPS)-stimulated H292 airway epithelial cells. In conclusion, linalool effectively exerts a protective role in OVA-induced airway inflammation and mucus hypersecretion, and its protective effects are closely related to the downregulation of inflammatory mediators and MAPKs/NF-κB signaling.


Assuntos
/uso terapêutico , Anti-Inflamatórios/uso terapêutico , Asma/tratamento farmacológico , Hipersensibilidade/tratamento farmacológico , Pulmão/metabolismo , Hipersensibilidade Respiratória/tratamento farmacológico , Células Th2/imunologia , Administração Oral , Alérgenos/imunologia , Animais , Células Cultivadas , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Imunoglobulina E/metabolismo , Pulmão/patologia , Camundongos , Camundongos Endogâmicos BALB C , NF-kappa B/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Ovalbumina/imunologia , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia
19.
Physiol Res ; 68(4): 675-679, 2019 08 29.
Artigo em Inglês | MEDLINE | ID: mdl-31177801

RESUMO

Chemokine (C-X-C motif) receptor 4 (CXCR4) agonists have been shown to protect lung endothelial barrier function in vitro. In vivo effects of CXCR4 modulation on lung endothelial permeability are unknown. Here we tested the effects of the CXCR4 agonist ubiquitin and the antagonist AMD3100 on lung vascular permeability and cytokine concentrations in a rat hemorrhage model. Animals were hemorrhaged (mean arterial blood pressure 30 mmHg for 30 min), treated with vehicle, ubiquitin (0.7 and 3.5 µmol/kg) or AMD3100 (3.5 µmol/kg), and resuscitated with crystalloids. Evans blue extravasation was employed to quantify lung vascular permeability. Ubiquitin dose-dependently reduced Evans blue extravasation into the lung. AMD3100 increased Evans blue extravasation. With AMD3100, TNFalpha levels in lung homogenates were increased; while TNFalpha levels were lower with ubiquitin, these differences did not reach statistical significance. Our findings suggest that CXCR4 regulates lung vascular permeability and further point towards CXCR4 as a drug target to confer lung protection during resuscitation from traumatic-hemorrhagic shock.


Assuntos
Permeabilidade Capilar/fisiologia , Receptores CXCR4/fisiologia , Mucosa Respiratória/metabolismo , Ressuscitação , Choque Hemorrágico/metabolismo , Choque Hemorrágico/terapia , Animais , Permeabilidade Capilar/efeitos dos fármacos , Relação Dose-Resposta a Droga , Compostos Heterocíclicos/farmacologia , Mediadores da Inflamação/antagonistas & inibidores , Mediadores da Inflamação/metabolismo , Masculino , Ratos , Ratos Sprague-Dawley , Receptores CXCR4/agonistas , Receptores CXCR4/antagonistas & inibidores , Mucosa Respiratória/efeitos dos fármacos , Ressuscitação/tendências , Ubiquitina/farmacologia
20.
Dokl Biochem Biophys ; 485(1): 150-152, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31201638

RESUMO

A hybrid 6XRE-hTERT promoter consisting of the hTERT tumor-specific promoter and six copies of the XRE element from the CYP1A1 human gene promoter was created. Using a human lung cancer cells as a model, we showed that XRE elements in the hybrid promoter greatly increase the activity of the hTERT promoter and ensure the reporter gene transcriptional activation in response to the treatment of the cells with the AhR ligand benzo(a)pyrene. However, similar effects were also observed in normal human bronchial epithelial cells HBEpC, which indicates the loss of the tumor-specific activity by the 6XRE-hTERT hybrid promoter. XRE elements can be used for nonspecific transcription enhancement but are unsuitable for the creation of tumor-specific promoters with enhanced activity.


Assuntos
Citocromo P-450 CYP1A1 , Elementos de Resposta , Telomerase , Ativação Transcricional/efeitos dos fármacos , Células A549 , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Benzo(a)pireno/toxicidade , Brônquios/metabolismo , Brônquios/patologia , Citocromo P-450 CYP1A1/biossíntese , Citocromo P-450 CYP1A1/genética , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Humanos , Receptores de Hidrocarboneto Arílico/genética , Receptores de Hidrocarboneto Arílico/metabolismo , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Telomerase/biossíntese , Telomerase/genética
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