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Extracellular vesicles (EVs) are membrane-enclosed bio-nanoparticles secreted by cells and naturally evolved to transport various bioactive molecules between cells and even organisms. These cellular objects are considered one of the most promising bio-nanovehicles for the delivery of native and exogenous molecular cargo. However, many challenges with state-of-the-art EV-based candidates as drug carriers still exist, including issues with scalability, batch-to-batch reproducibility, and cost-sustainability of the final therapeutic formulation. Microalgal extracellular vesicles, which we named nanoalgosomes, are naturally released by various microalgal species. Here, we evaluate the innate biological properties of nanoalgosomes derived from cultures of the marine microalgae Tetraselmis chuii, using an optimized manufacturing protocol. Our investigation of nanoalgosome biocompatibility in preclinical models includes toxicological analyses, using the invertebrate model organism Caenorhabditis elegans, hematological and immunological evaluations ex vivo and in mice. We evaluate nanoalgosome cellular uptake mechanisms in C. elegans at cellular and subcellular levels, and study their biodistribution in mice with accurate space-time resolution. Further examination highlights the antioxidant and anti-inflammatory bioactivities of nanoalgosomes. This holistic approach to nanoalgosome functional characterization demonstrates that they are biocompatible and innate bioactive effectors with unique bone tropism. These findings suggest that nanoalgosomes have significant potential for future therapeutic applications.
Assuntos
Anti-Inflamatórios , Antioxidantes , Vesículas Extracelulares , Microalgas , Vesículas Extracelulares/metabolismo , Animais , Microalgas/metabolismo , Camundongos , Anti-Inflamatórios/farmacologia , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Caenorhabditis elegans/metabolismo , Materiais Biocompatíveis/química , Clorófitas/metabolismo , Osso e Ossos/metabolismo , TropismoRESUMO
IL-17A drives inflammation and oxidative stress, affecting the progression of chronic lung diseases (asthma, chronic obstructive pulmonary disease (COPD), lung cancer, and cystic fibrosis). Oleuropein (OLP) is a polyphenolic compound present in olive oil and widely included in the Mediterranean diet. It exerts antioxidant and anti-inflammatory activities, oxidative stress resistance, and anticarcinogenic effects with a conceivable positive impact on human health. We hypothesized that OLP positively affects the mechanisms of oxidative stress, apoptosis, DNA damage, cell viability during proliferation, and cell growth in alveolar epithelial cells and tested its effect in a human alveolar epithelial cell line (A549) in the presence of IL-17A. Our results show that OLP decreases the levels of oxidative stress (Reactive Oxygen Species, Mitochondrial membrane potential) and DNA damage (H2AX phosphorylation-ser139, Olive Tail Moment data) and increases cell apoptosis in A549 cells exposed to IL-17A. Furthermore, OLP decreases the number of viable cells during proliferation, the migratory potential (Scratch test), and the single cell capacity to grow within colonies as a cancer phenotype in A549 cells exposed to IL-17A. In conclusion, we suggest that OLP might be useful to protect lung epithelial cells from oxidative stress, DNA damage, cell growth, and cell apoptosis. This effect might be exerted in lung diseases by the downregulation of IL-17A activities. Our results suggest a positive effect of the components of olive oil on human lung health.
Assuntos
Apoptose , Proliferação de Células , Dano ao DNA , Interleucina-17 , Glucosídeos Iridoides , Iridoides , Estresse Oxidativo , Humanos , Estresse Oxidativo/efeitos dos fármacos , Interleucina-17/metabolismo , Glucosídeos Iridoides/farmacologia , Proliferação de Células/efeitos dos fármacos , Células A549 , Dano ao DNA/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Iridoides/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Antioxidantes/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Azeite de Oliva/farmacologia , Células Epiteliais Alveolares/efeitos dos fármacos , Células Epiteliais Alveolares/metabolismoRESUMO
Airway epithelium represents a physical barrier against toxic substances and pathogens but also presents pattern recognition receptors on the epithelial cells that detect pathogens leading to molecule release and sending signals that activate both the innate and adaptive immune responses. Thus, impaired airway epithelial function and poor integrity may increase the recurrence of infections. Probiotic use in respiratory diseases as adjuvant of traditional therapy is increasingly widespread. There is growing interest in the use of non-viable heat-killed bacteria, such as tyndallized bacteria (TB), due to safety concerns and to their immunomodulatory properties. This study explores in vitro the effects of a TB blend on the immune activation of airway epithelium. 16HBE bronchial epithelial cells were exposed to different concentrations of TB. Cell viability, TB internalization, TLR2 expression, IL-6, IL-8 and TGF-ßl expression/release, E-cadherin expression and wound healing were assessed. We found that TB were tolerated, internalized, increased TLR2, E-cadherin expression, IL-6 release and wound healing but decreased both IL-8 and TGF-ßl release. In conclusion, TB activate TLR2 pathway without inducing a relevant pro-inflammatory response and improve barrier function, leading to the concept that TB preserve epithelial homeostasis and could be used as strategy to prevent and to manage respiratory infection, exacerbations included.
Assuntos
Brônquios , Células Epiteliais , Imunidade Inata , Receptor 2 Toll-Like , Humanos , Receptor 2 Toll-Like/metabolismo , Células Epiteliais/imunologia , Células Epiteliais/metabolismo , Brônquios/citologia , Brônquios/imunologia , Interleucina-6/metabolismo , Probióticos , Mucosa Respiratória/imunologia , Caderinas/metabolismo , Expressão Gênica , Células Cultivadas , Interleucina-8/metabolismo , Infecções Respiratórias/imunologia , Infecções Respiratórias/microbiologia , Sobrevivência CelularRESUMO
The impact of volcanic airborne products on airway epithelium homeostasis is largely unknown. This study assessed the effects of volcanic Fumarole Condensates (FC) alone or combined with Cigarette Smoke Extracts (CSE) on airway epithelial cells (16HBE and A549). Chemical composition of FC was analyzed by gas chromatography and HPLC. Cells were exposed to FC and IL-33 and IL-8 were assessed. The effects of FC and CSE on cell injury were evaluated assessing cell metabolism/cell viability, mitochondrial stress, cell apoptosis/cell necrosis, and cell proliferation. FC contained: water vapor (70-97%), CO2 (3-30%), acid gases (H2S, SO2, HCl, HF) around 1%. FC increased the intracellular IL-33 but differently modulated IL-33 and IL-8 gene expression and IL-8 release in the tested cell lines. FC without/with CSE: (a) increased cell metabolism/cell viability in 16HBE, while decreased it in A549; (b) increased mitochondrial stress in both cell types. FC with CSE increased cell necrosis in A549 in comparison to CSE alone. CSE reduced cell proliferation in 16HB,E while increased it in A549 and FC counteracted these effects in both cell types. Overall, FC induce a pro-inflammatory profile associated to a metabolic reprogramming without a relevant toxicity also in presence of CSE in airway epithelial cells.
Assuntos
Fumar Cigarros , Interleucina-33 , Humanos , Interleucina-33/metabolismo , Interleucina-33/farmacologia , Interleucina-8/metabolismo , Células Epiteliais/metabolismo , Necrose/metabolismoRESUMO
A series of biologically unexplored substituted 1,3,4-subtituted-pyrrolo[3,2-c]quinoline derivatives (PQs) was evaluated against a panel of about 60 tumor cells (NCI). Based on the preliminary antiproliferative data, the optimizations efforts permitted us to design and synthesize a new series of derivatives allowing us to individuate a promising hit (4g). The insertion of a 4-benzo[d] [1,3]dioxol-5-yl moiety on increased and extended the activity towards five panel tumor cell lines such as leukemia, CNS, melanoma, renal and breast cancer, reaching IG50 in the low µM range. Replacement of this latter with a 4-(OH-di-Cl-Ph) group (4i) or introduction a Cl-propyl chain in position 1 (5), selectively addressed the activity against the entire leukemia sub-panel (CCRF-CEM, K-562, MOLT-4, RPMI-8226, SR). Preliminary biological assays on MCF-7 such as cell cycle, clonogenic assay, ROS content test alongside a comparison of viability between MCF-7 and non-tumorigenic MCF-10 were investigated. Among the main anticancer targets involved in breast cancer, HSP90 and ER receptors were selected for in silico studies. Docking analysis revealed a valuable affinity for HSP90 providing structural insights on the binding mode, and useful features for optimization.
Assuntos
Antineoplásicos , Neoplasias da Mama , Hidroxiquinolinas , Quinolinas , Humanos , Feminino , Estrutura Molecular , Relação Estrutura-Atividade , Proliferação de Células , Linhagem Celular Tumoral , Hidroxiquinolinas/farmacologia , Quinolinas/farmacologia , Antineoplásicos/química , Ensaios de Seleção de Medicamentos Antitumorais , Simulação de Acoplamento MolecularRESUMO
Lung cancer is one of the leading forms of cancer in developed countries. Interleukin-8 (IL-8), a pro-inflammatory cytokine, exerts relevant effects in cancer growth and progression, including angiogenesis and metastasis in lung cancer. Mesoporous silica particles, functionalized with newly extracted fish oil (Omeg@Silica), are more effective than the fish oil alone in anti-proliferative and pro-apoptotic effects in non-small cell lung cancer (NSCLC) cell lines. The mechanisms that explain this efficacy are not yet understood. The aim of the present study is therefore to decipher the anti-cancer effects of a formulation of Omeg@Silica in aqueous ethanol (FOS) in adenocarcinoma (A549) and muco-epidermoid (NCI-H292) lung cancer cells, evaluating cell migration, as well as IL-8, NF-κB, and miRNA-21 expression. Results show that in both cell lines, FOS was more efficient than oil alone, in decreasing cell migration and IL-8 gene expression. FOS reduced IL-8 protein release in both cell lines, but this effect was only stronger than the oil alone in A549. In A549, FOS was able to reduce miRNA-21 and transcription factor NF-κB nuclear expression. Taken together, these data support the potential use of the Omeg@Silica as an add-on therapy for NSCLC. Dedicated studies which prove clinical efficacy are needed.
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Carotenoids may have different effects on cancer and its progression. The safety of carotenoid supplements was evaluated in vitro on human non-small cell lung cancer (NSCLC) adenocarcinoma A549 cells by the administration of three different oleoresins containing lycopene and other lipophilic phytochemicals, such as tocochromanols. The oleoresins, obtained by the supercritical CO2 green extraction technology from watermelon (Lyc W), gac(Lyc G) and tomato (Lyc T) and chlatrated in α-cyclodextrins, were tested in comparison to synthetic lycopene (Lyc S), by cell cycle, Annexin V-FITC/PI, clonogenic test, Mytosox, intracellular ROS, Western Blot for NF-kB and RT-PCR and ELISA for IL-8. The extracts administered at the same lycopene concentration (10 µM) showed conflicting behaviors: Lyc W, with the highest lycopene/tocochromanols ratio, significantly increased cell apoptosis, mitochondrial stress, intracellular ROS, NF-kB and IL-8 expression and significantly decreased cell proliferation, whereas Lyc G and Lyc T significantly increased only cell proliferation. Lyc S treatment was ineffective. The highest amount of lycopene in Lyc W was able to counteract and revert the cell survival effect of tocochromanols supporting the importance of evaluating the lycopene bio-availability and the real effect of antioxidant tocochromanols' supplementation which may not only have no anticancer benefits but may even increase cancer aggressivity.
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Notch-1 pathway plays an important role in lung carcinoma, stem cell regulation, cellular communication, growth and differentiation. Cigarette smoke is involved in the regulation of Notch signaling. However, current data regarding the impact of cigarette smoke on the Notch pathway in lung cancer progression are limited. The present study aimed to explore whether cigarette smoke exposure altered Notch-1 pathway in ex-vivo (surgical samples of lung parenchyma from non-smoker and smoker patients with lung adenocarcinoma) and in vitro (adenocarcinoma A549 cell line) approaches. The expression of Notch-1, Jagged-1 and CD133 in surgical samples was evaluated by immunohistochemistry. A549 were exposed to cigarette smoke extracts (2.5 % and 5 % CSE for 6, 24 and 48 h) and the expression of Notch-1, Jagged-1 and Hes-1 was evaluated by Real-Time PCR and Western Blot (nuclear fractions). Expression and localization of Notch-1, Hes-1, CD133 and ABCG2 were assessed by immunofluorescence. The expression of survivin and Ki-67 was assessed by flow cytometry following CSE exposure and inhibition of Notch-1 signaling. Smokers lung parenchyma exhibited higher expression of Notch-1. CSE exposure increased Notch-1 and Hes-1 gene and nuclear protein expression in A549. Immunofluorescence confirmed higher expression of nuclear Hes-1 in CSE-stimulated A549 cells. CSE increased both survivin and Ki-67 expression and this effect was reverted by inhibition of the Notch-1 pathway. In conclusion, these data show that cigarette smoke may promote adenocarcinoma progression by activating the Notch-1 pathway thus supporting its role as hallmark of lung cancer progression and as a new target for lung cancer treatment.
Assuntos
Adenocarcinoma de Pulmão/metabolismo , Fumar Cigarros , Pulmão/metabolismo , Receptor Notch1/metabolismo , Células A549 , Antígeno AC133/genética , Antígeno AC133/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/genética , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Western Blotting , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Proteína Jagged-1/genética , Proteína Jagged-1/metabolismo , Antígeno Ki-67/genética , Antígeno Ki-67/metabolismo , Pulmão/efeitos dos fármacos , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Receptor Notch1/genética , Transdução de Sinais , Fumantes , Survivina/genética , Survivina/metabolismo , Fatores de Transcrição HES-1/genética , Fatores de Transcrição HES-1/metabolismo , Regulação para CimaRESUMO
The role of PAR-1 expression and activation was described in epithelial cells from the central and distal airways of COPD patients using an ex vivo/in vitro model. PAR-1 immunoreactivity was studied in epithelial cells from surgical specimens of the central and distal airways of COPD patients and healthy control (HC). Furthermore, PAR-1 expression and activation were measured in both the human bronchial epithelial cell line (16HBE) and normal human bronchial epithelial cells (NHBEs) exposed to cigarette smoke extract (CSE) (10%) or thrombin. Finally, cell proliferation, apoptosis, and IL-8 release were detected in stimulated NHBEs. We identified higher levels of PAR-1 expression/activation in epithelial cells from the central airways of COPD patients than in HC. Active PAR-1 increased in epithelial cells from central and distal airways of COPD, with higher levels in COPD smokers (correlated with pack-years) than in COPD ex-smokers. 16HBE and NHBEs exposed to CSE or thrombin showed increased levels of active PAR-1 (localized in the cytoplasm) than baseline conditions, while NHBEs treated with thrombin or CSE showed increased levels of IL-8 proteins, with an additional effect when used in combination. Smoking habits generate the upregulation of PAR-1 expression/activation in airway epithelial cells, and promoting IL-8 release might affect the recruitment of infiltrating cells in the airways of COPD patients.
Assuntos
Expressão Gênica , Doença Pulmonar Obstrutiva Crônica/etiologia , Doença Pulmonar Obstrutiva Crônica/metabolismo , Receptor PAR-1/genética , Receptor PAR-1/metabolismo , Mucosa Respiratória/metabolismo , Apoptose/genética , Biomarcadores , Estudos de Casos e Controles , Linhagem Celular , Proliferação de Células , Suscetibilidade a Doenças , Células Epiteliais/metabolismo , Imunofluorescência , Humanos , Interleucina-8/metabolismo , Doença Pulmonar Obstrutiva Crônica/patologia , Mucosa Respiratória/patologia , Fumar/efeitos adversosRESUMO
Adipose tissue is widely recognized as an extremely active endocrine organ producing adipokines as leptin that bridge metabolism and the immune system. Pre-B-cell leukemia homeobox (Pbx)-regulating protein-1 (PREP1) is a ubiquitous homeodomain transcription factor involved in the adipogenic differentiation and insulin-sensitivity processes. Leptin, as pleiotropic adipokine, and TGF-ß, known to be expressed by primary pre-adipocytes [adipose-derived stem cells (ASCs)] and mature differentiated adipocytes, modulate inflammatory responses. We aimed to assess for the first time if leptin and TGF-ß interfere with PREP1 expression in both ASCs and mature differentiated adipocytes. Human ASCs were isolated from subcutaneous adipose liposuction and, after expansion, fully differentiated to mature adipocytes. In both ASCs and adipocytes, leptin and TGF-ß1 significantly decreased the expression of PREP1, alone and following concurrent Toll-like receptor 4 (TLR4) activation. Moreover, in adipocytes, but not in ASCs, leptin increased TLR4 and IL-33 expression, whereas TGF-ß1 enhanced TLR4 and IL-6 expression. Taken together, we provide evidence for a direct regulation of PREP1 by leptin and TGF-ß1 in ASCs and mature adipocytes. The effects of leptin and TGF-ß1 on immune receptors and cytokines, however, are limited to mature adipocytes, suggesting that modulating immune responses depends on the differentiation of ASCs. Further studies are needed to fully understand the regulation of PREP1 expression and its potential for the development of new therapeutic approaches in obesity-related diseases.
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Melanoma is the most aggressive form of skin cancer and one of the most treatment-refractory malignancies. In metastatic melanoma cell lines, we analysed the anti-proliferative and pro-apoptotic potentials of a phenolic component of olive oil, the hydroxytyrosol. In particular, through MTS assay, DeadEnd™ Colorimetric TUNEL assay, Annexin V binding and PI uptake, western blot experiment, intracellular reactive oxygen species (ROS) analysis, and the cell colony assay, we showed that the hydroxytyrosol treatment remarkably reduces the cell viability inducing the death for apoptosis of melanoma cells. Moreover, we showed that the hydroxytyrosol treatment of melanoma cells leads to a significant increase of p53 and γH2AX expression, a significant decrease of AKT expression and the inhibition of cell colony formation ability. Finally, we propose that the increased amount of intracellular reactive oxygen species (ROS) that may be related to the regulation of the pathways involved in the activation of apoptosis and in the inhibition of melanoma growth could be the strategy used by hydroxytyrosol to exert its functions in melanoma. Therefore, for its role in melanoma growth inhibition, the hydroxytyrosol treatment could deeply interfere with melanoma progression as a promising therapeutic option for the treatment of this highly invasive tumour.
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Antioxidantes/farmacologia , Apoptose , Melanoma/patologia , Álcool Feniletílico/análogos & derivados , Espécies Reativas de Oxigênio/metabolismo , Movimento Celular , Proliferação de Células , Humanos , Melanoma/metabolismo , Álcool Feniletílico/farmacologia , Células Tumorais CultivadasRESUMO
Polybrominated diphenyl ethers (PBDEs) are widespread as flame-retardants in different types of consumer products. PBDEs present in the air or dust and their inhalation can damage human health by influencing the respiratory system. We evaluated the effects of environment relevant concentrations (0.01-1 µM) of PBDE-47, PBDE-99 and PBDE-209 on the mechanism of oxidative stress, dysregulation of cell proliferation, apoptosis, and DNA damage and repair (in term of H2AX phosphorylation ser139) in an in-vitro/ex-vivo model of bronchial epithelial cells. PBDEs (-47, -99 and -209) at the environment relevant concentrations (0.01 and 1 µM) induce oxidative stress (in term of NOX-4 expression as well as ROS and JC-1 production), activate the mechanism of DNA-damage and repair affecting Olive Tail length (comet assay) production and H2AX phosphorylation (ser139) in normal human bronchial epithelial cells. Furthermore PBDEs, although do not affect cell viability, induce cell apoptosis and single cell capacity to grow into a colony (like a cancer phenotype) in bronchial epithelial cells. Finally, PBDE-47 had a greater effect than -99 and -209. PBDE-47, -99 and -209 congeners exert cytotoxic and genotoxic effects, and play a critical role in the dysregulation of oxidative stress, damaging DNA and the related gene expression in bronchial epithelial cells. Our findings might suggest that PBDEs inhalation might have adverse effect on human health regarding pulmonary diseases in the areas of environmental pollution.
Assuntos
Brônquios/patologia , Células Epiteliais/efeitos dos fármacos , Retardadores de Chama/toxicidade , Éteres Difenil Halogenados/toxicidade , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Células Epiteliais/metabolismo , Retardadores de Chama/análise , Éteres Difenil Halogenados/análise , Humanos , Estresse Oxidativo/efeitos dos fármacosRESUMO
Melanoma is the most aggressive and deadly form of skin cancer, which is largely due to its propensity to metastasize. Therefore, with the aim to inhibit the growth and the metastatic dissemination of melanoma cells and to provide a novel treatment option, we studied the effects of the melanoma treatment with two organotin(IV) complexes of the meso-tetra(4-sulfonato-phenyl)porphine, namely (Bu2Sn)2TPPS and (Bu3Sn)4TPPS. In particular, we showed that nanomolar concentrations of (Bu2Sn)2TPPS and (Bu3Sn)4TPPS are sufficient to inhibit melanoma cell growth, to increase the expression of the full-length poly (ADP-ribose) polymerase (PARP-1), to induce the cell cycle arrest respectively at G2/M and G0/G1 through the inhibition of the Cyclin D1 expression and to inhibit cell colony formation. Nanomolar concentrations of (Bu2Sn)2TPPS and (Bu3Sn)4TPPS are also sufficient to inhibit the melanoma cell migration and the expression of some adhesion receptors. Moreover, we report that (Bu2Sn)2TPPS and (Bu3Sn)4TPPS act downstream of BRAF, mainly bypassing its functions, but targeting the STAT3 signalling protein. Finally, these results suggest that (Bu2Sn)2TPPS and (Bu3Sn)4TPPS may be effective therapeutic strategies for their role in the inhibition of melanoma growth and migration.
Assuntos
Antineoplásicos/farmacologia , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Porfirinas/farmacologia , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Quinase 1 de Adesão Focal/metabolismo , Humanos , Melanoma/genética , Melanoma/metabolismo , Estrutura Molecular , Porfirinas/química , Transdução de Sinais/efeitos dos fármacosRESUMO
[This corrects the article DOI: 10.1155/2016/8727289.].
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Cigarette smoke is a risk factor for COPD and lung cancer. In cancer, epigenetic modifications affect the expression of Enhancer of Zester Homolog 2 (EZH2), and silenced disabled homolog 2 interacting protein gene (DAB2IP) (onco-suppressor gene) by Histone H3 tri-methylation in lysine 27 (H3K27me3). In"ex vivo"studies, we assessed EZH2, H3K27me3 and DAB2IP immunoreactivity in bronchial epithelial cells from COPD patients (smokers, ex-smokers), Smoker and control subjects. In"in vitro" experiments we studied the effect of cigarette smoke extract (CSE) on EZH2/H3K27me3/DAB2IP expression, apoptosis, invasiveness, and vimentin expression in 16HBE, primary cells, and lung cancer cell lines (A549) long-term exposed to CSE. Finally, in "in vitro"studies, we tested the effect of GSK343 (selective inhibitor of EZH2). EZH2 and H3K27me3 expression was higher, while DAB2IP was lower levels, in bronchial epithelium from COPD and Smokers than in Controls. CSE increased EZH2, H3K27me3 expression and decreased DAB2IP, cell apoptosis and invasiveness in epithelial cells. GSK343 restored the effects of CSE. Cigarette smoke affects EZH2 expression, and reduced DAB2IP via H3K27me3 in COPD patients. The molecular mechanisms associated with EZH2 expression, generate a dysregulation of cell apoptosis, mesenchymal transition, and cell invasiveness in bronchial epithelial cells, encouraging the progression of airway inflammation toward lung cancer in COPD patients.
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Fumar Charutos/efeitos adversos , Proteína Potenciadora do Homólogo 2 de Zeste/genética , Neoplasias Pulmonares/genética , Doença Pulmonar Obstrutiva Crônica/genética , Proteínas Ativadoras de ras GTPase/genética , Células A549 , Células Epiteliais Alveolares/efeitos dos fármacos , Células Epiteliais Alveolares/patologia , Apoptose/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Histona Desmetilases com o Domínio Jumonji/genética , Neoplasias Pulmonares/induzido quimicamente , Neoplasias Pulmonares/patologia , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Doença Pulmonar Obstrutiva Crônica/induzido quimicamente , Doença Pulmonar Obstrutiva Crônica/patologia , Fatores de RiscoRESUMO
BACKGROUND: Allergic rhinitis is characterized by a remodeling of nasal epithelium. Since the Notch and TGF-ß signaling pathways are known to be involved in cell differentiation and remodeling processes and leptin adipokine has already been identified as a marker for homeostasis in human bronchial and nasal epithelial cells of asthmatics, roles played by these pathways have been investigated for chronic allergic rhinitis. METHODS: The leptin/leptin receptor expression has been investigated in a study with 40 biopsies from allergic (AR, nâ¯=â¯18) and non-allergic (C, nâ¯=â¯22) inferior turbinates, using immunohistochemistry, immunofluorescence staining and RT-PCR. In addition, extracts from in vitro samples prepared from primary cells of inferior turbinates as well as in vitro cultured human nasal epithelial RPMI 2650 cells (ATCC-CCL-30) were also tested for leptin expression and activation of the Notch-1 pathway. RESULTS: With regards to AR, in vivo expression levels of both leptin and its receptor significantly decreased in comparison to C. Furthermore, leptin receptor mRNA was significantly reduced in AR as compared to C. Immunofluorescence showed an apparent co-expression of leptin receptor with Notch-1, which was not seen with TGF-ß. In vitro, in primary turbinate epithelial cells, the expression of leptin receptor and Notch-1 significantly decreased in AR as compared to C. Moreover, in RPMI 2650 cells, leptin receptor expression was shown to be induced by Notch-1 ligand signaling. CONCLUSION: Thus, both the leptin and Notch-1 pathways appear to represent markers for epithelial homeostasis in allergic rhinitis.
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Leptina/genética , Mucosa Nasal/metabolismo , Receptor Notch1/genética , Receptores para Leptina/genética , Rinite Alérgica/genética , Adulto , Biópsia , Estudos de Casos e Controles , Linhagem Celular , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Feminino , Regulação da Expressão Gênica , Homeostase/genética , Humanos , Leptina/metabolismo , Masculino , Pessoa de Meia-Idade , Mucosa Nasal/patologia , Cultura Primária de Células , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor Notch1/metabolismo , Receptores para Leptina/metabolismo , Rinite Alérgica/metabolismo , Rinite Alérgica/patologia , Transdução de Sinais , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo , Conchas Nasais/metabolismo , Conchas Nasais/patologiaRESUMO
Polybrominated diphenyl ethers (PBDEs) are persistent organic pollutants that are added to numerous products to prevent accidental fires. PBDEs are present in the environment and they bio-accumulate in human and animal tissues. Recently, their presence has been correlated to several pathologies but little is known about their effect on the human innate immune system activity. In this study we investigated the effect of the congener 2,2',4,4'-Tetrabromodiphenyl ether (PBDE-47) on the functional activity of the THP-1 human macrophages cell line and on ex vivo freshly isolated human basophils. Cytotoxicity and genotoxicity studies showed that PBDE-47 was able to induce toxic effects on the THP-1â¯cell line viability at concentrations ≥25⯵M. Immune function of THP-1 was studied after stimulation with bacterial lipopolysaccharide (LPS) and PBDE-47 exposure at concentrations granting macrophage viability. Two dimensional electrophoresis showed modification of the proteome in the 3⯵M PBDE-47 treated sample and Real Time PCR and ELISA demonstrated a statistically significant reduction in the expression of IL-1ß, IL-6 and TNF-α cytokines. Furthermore, PBDE-47 was able to perturbate genes involved in cell motility upregulating CDH-1 and downregulating MMP-12 expressions. Finally, basophil activation assay showed reduced CD63 activation in PBDE-47 treated samples. In conclusion, our study demonstrated that PBDE-47 may perturb the activities of cells involved in innate immunity dampening the expression of macrophage pro-inflammatory cytokines (IL-1ß, IL-6 and TNF-α) and genes involved in cell motility (MMP-12 and E-cadherin) and interfering with basophil activation suggesting that this compound can impair innate immune response.
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Éteres Difenil Halogenados/toxicidade , Imunidade Inata/efeitos dos fármacos , Animais , Basófilos/efeitos dos fármacos , Células Cultivadas , Citocinas/metabolismo , Poluentes Ambientais/farmacologia , Éteres Difenil Halogenados/farmacologia , Humanos , Macrófagos/efeitos dos fármacos , Células THP-1RESUMO
Hepatocellular carcinoma (HCC) is one of the common malignancies and is an increasingly important cause of cancer death worldwide. Surgery, chemotherapy, and radiation therapy extend the 5-year survival limit in HCC patients by only 6%. Therefore, there is a need to develop new therapeutic approaches for the treatment of this disease. The orally bioavailable proteasome inhibitor MLN2238 (ixazomib) has been demonstrated to have anticancer activity. In the present study, we investigated the preclinical therapeutic efficacy of MLN2238 in HCC cells through in vitro and in vivo models, and examined its molecular mechanisms of action. MLN2238 inhibited cell viability in human HCC cells HepG2, Hep3B, and SNU475 in a time- and dose-dependent manner. Flow cytometry analysis demonstrated that MLN2238 induced G2/M cell cycle arrest and cellular apoptosis in HCC cells. Cell cycle arrest was associated with increased expression levels of p21 and p27. MLN2238-induced apoptosis was confirmed by caspase-3/7 activation, PARP cleavage and caspase-dependent ß-catenin degradation. In addition, MLN2238 activated ER stress genes in HCC cells and increased the expression of the stress-inducible gene nuclear protein-1. Furthermore, MLN2238 treatment induced upregulation of myeloid cell leukemia-1 (Mcl-1) protein, and Mcl-1 knockdown sensitized HCC cells to MLN2238 treatment, suggesting the contribution of Mcl-1 expression to MLN2238 resistance. This result was also confirmed using the novel Mcl-1 small molecule inhibitor A1210477. Association of A1210477 and MLN2238 determined synergistic antitumor effects in HCC cells. Finally, in vivo orally administered MLN2238 suppressed tumor growth of Hep3B cells in xenograft models in nude mice. In conclusion, our results offer hope for a new therapeutic opportunity in the treatment of HCC patients.
Assuntos
Antineoplásicos/uso terapêutico , Compostos de Boro/uso terapêutico , Carcinoma Hepatocelular/tratamento farmacológico , Glicina/análogos & derivados , Neoplasias Hepáticas/tratamento farmacológico , Inibidores de Proteassoma/uso terapêutico , Animais , Antineoplásicos/farmacologia , Compostos de Boro/farmacologia , Glicina/farmacologia , Glicina/uso terapêutico , Humanos , Camundongos , Inibidores de Proteassoma/farmacologiaRESUMO
BACKGROUND: Cigarette smoke extract (CSE) affects the expression of non-neuronal components of cholinergic system in bronchial epithelial cells and, as PEBP1/Raf-mediated MAPK1/2 and ERK1/2 pathway, promotes inflammation and oxidative stress. AIMS: We studied whether Acetylcholine (ACh) is involved in the mechanism of crosstalk between mAChRM3 and ß2Adrenergic receptors (ß2AR) promoting, via PI3/PKC/PBEP1/Raf/MEK1/2/ERK1/2 activation, ß2AR desensitization, inflammation and, oxidative stress in a bronchial epithelial cell line (16HBE) after long-term exposure to cigarette smoke extract (LECSE). METHODS: We evaluated mAChRM3 and Choline Acetyltransferase (ChAT) expression, ACh production, PEBP1, ERk1/2, and ß2AR phosphorylation, as well as NOX-4, ROS production and IL-8 release in 16HBE after LECSE. The inhibitory activity of Hemicholinium (HCh-3) (a potent choline uptake blocker), LY294002 (a highly selective inhibitor of PI3 kinase), Tiotropium (Spiriva®) (anticholinergic drug) and Olodaterol (ß2AR agonist), were tested in 16HBE after LECSE. RESULTS: mAChRM3, ChAT, ACh activity, pPEBP1, pß2AR, pERK1/2, ROS, NOX-4 and IL-8 increased after LECSE in 16HBE LECSE compared to untreated cells. HCh-3 and LY294002 (alone or in combination) as well as Tiotropium (Spiriva®) or Olodaterol (alone or in combination) all reduced the levels of pPEBP1, pß2AR, pERK1/2, ROS, NOX-4, and IL-8 in 16HBE LECSE compared to untreated cells. CONCLUSIONS: LECSE promotes ACh production which enhances PI3/PKC/PEBP1/Raf-ERK1/2 pathway activation, heterologous ß2AR desensitization, as well as release of inflammatory and oxidative mediators in bronchial epithelial cells. The use of anticholinergic drugs and long-acting ß2-agonists, alone or in combination may be dampen these inflammatory mechanisms when used in combination in some epithelial cell types.