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1.
Mar Drugs ; 18(12)2020 Dec 14.
Artigo em Inglês | MEDLINE | ID: mdl-33327522

RESUMO

The mucus layer of the nasopharynx and bronchial epithelium has a barrier function against inhaled pathogens such as the coronavirus SARS-CoV-2. We recently found that inorganic polyphosphate (polyP), a physiological, metabolic energy (ATP)-providing polymer released from blood platelets, blocks the binding of the receptor binding domain (RBD) to the cellular ACE2 receptor in vitro. PolyP is a marine natural product and is abundantly present in marine bacteria. Now, we have approached the in vivo situation by studying the effect of polyP on the human alveolar basal epithelial A549 cells in a mucus-like mucin environment. These cells express mucins as well as the ectoenzymes alkaline phosphatase (ALP) and adenylate kinase (ADK), which are involved in the extracellular production of ATP from polyP. Mucin, integrated into a collagen-based hydrogel, stimulated cell growth and attachment. The addition of polyP to the hydrogel significantly increased cell attachment and also the expression of the membrane-tethered mucin MUC1 and the secreted mucin MUC5AC. The increased synthesis of MUC1 was also confirmed by immunostaining. This morphogenetic effect of polyP was associated with a rise in extracellular ATP level. We conclude that the nontoxic and non-immunogenic polymer polyP could possibly also exert a protective effect against SARS-CoV-2-cell attachment; first, by stimulating the innate antiviral response by strengthening the mucin barrier with its antimicrobial proteins, and second, by inhibiting virus attachment to the cells, as deduced from the reduction in the strength of binding between the viral RBD and the cellular ACE2 receptor.


Assuntos
Organismos Aquáticos/metabolismo , Produtos Biológicos/farmacologia , Polifosfatos/farmacologia , Mucosa Respiratória/efeitos dos fármacos , Células A549 , Bactérias/metabolismo , Produtos Biológicos/uso terapêutico , Humanos , Imunidade Inata/efeitos dos fármacos , Mucina-5AC/metabolismo , Mucina-1/metabolismo , Polifosfatos/metabolismo , Polifosfatos/uso terapêutico , Mucosa Respiratória/imunologia , Mucosa Respiratória/metabolismo , /patogenicidade , Metabolismo Secundário , Ligação Viral/efeitos dos fármacos
2.
Biomed Environ Sci ; 33(9): 680-689, 2020 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-33106213

RESUMO

Objective: To investigate the effect of c-fos gene silencing on differentially expressed proteins (DEPs) in human bronchial epithelial (HBE) cells after exposure to fine particulate matter (PM 2.5). Methods: HBE cells and c-fos-silenced HBE cells were exposed to 50 µg/mL PM 2.5, LC-MS/MS and tandem mass tag (TMT) labeling methods were combined with bioinformatics methods, and DEPs and interaction networks were identified. Results: In the HBE group, 414 DEPs were screened, of which 227 were up-regulated and 187 down-regulated. In the c-fos silenced HBE group, 480 DEPs were screened, including 240 up-regulated proteins and 240 down-regulated proteins. KEGG annotations showed that DEPs in the HBE group are mainly concentrated in the glycolysis/gluconeogenesis pathway and those in the c-fos silenced group are concentrated mainly in endoplasmic reticulum and the processing of proteins. Additionally, the abnormal expression of GPRC5C, DKK4, and UBE2C was identified in top 15 DEPs. After constructing the protein interaction network, 20 Hub proteins including HNRNPA2B1, HNRNPL, RPS15A, and RPS25 were screened from the HBE group and the c-fos silenced HBE group. Conclusion: c-fos gene affected the expression of cancer-related proteins. Our results provided a scientific basis for further study of PM 2.5-induced carcinogenesis mechanism.


Assuntos
Poluentes Atmosféricos/toxicidade , Brônquios/efeitos dos fármacos , Regulação da Expressão Gênica , Inativação Gênica , Genes fos/genética , Material Particulado/toxicidade , Mucosa Respiratória/efeitos dos fármacos , Brônquios/metabolismo , Células Cultivadas , Humanos , Proteômica , Mucosa Respiratória/metabolismo
3.
mBio ; 11(5)2020 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-33082259

RESUMO

The emergence of severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), the etiological agent of the 2019 coronavirus disease (COVID-19), has erupted into a global pandemic that has led to tens of millions of infections and hundreds of thousands of deaths worldwide. The development of therapeutics to treat infection or as prophylactics to halt viral transmission and spread is urgently needed. SARS-CoV-2 relies on structural rearrangements within a spike (S) glycoprotein to mediate fusion of the viral and host cell membranes. Here, we describe the development of a lipopeptide that is derived from the C-terminal heptad repeat (HRC) domain of SARS-CoV-2 S that potently inhibits infection by SARS-CoV-2. The lipopeptide inhibits cell-cell fusion mediated by SARS-CoV-2 S and blocks infection by live SARS-CoV-2 in Vero E6 cell monolayers more effectively than previously described lipopeptides. The SARS-CoV-2 lipopeptide exhibits broad-spectrum activity by inhibiting cell-cell fusion mediated by SARS-CoV-1 and Middle East respiratory syndrome coronavirus (MERS-CoV) and blocking infection by live MERS-CoV in cell monolayers. We also show that the SARS-CoV-2 HRC-derived lipopeptide potently blocks the spread of SARS-CoV-2 in human airway epithelial (HAE) cultures, an ex vivo model designed to mimic respiratory viral propagation in humans. While viral spread of SARS-CoV-2 infection was widespread in untreated airways, those treated with SARS-CoV-2 HRC lipopeptide showed no detectable evidence of viral spread. These data provide a framework for the development of peptide therapeutics for the treatment of or prophylaxis against SARS-CoV-2 as well as other coronaviruses.IMPORTANCE SARS-CoV-2, the causative agent of COVID-19, continues to spread globally, placing strain on health care systems and resulting in rapidly increasing numbers of cases and mortalities. Despite the growing need for medical intervention, no FDA-approved vaccines are yet available, and treatment has been limited to supportive therapy for the alleviation of symptoms. Entry inhibitors could fill the important role of preventing initial infection and preventing spread. Here, we describe the design, synthesis, and evaluation of a lipopeptide that is derived from the HRC domain of the SARS-CoV-2 S glycoprotein that potently inhibits fusion mediated by SARS-CoV-2 S glycoprotein and blocks infection by live SARS-CoV-2 in both cell monolayers (in vitro) and human airway tissues (ex vivo). Our results highlight the SARS-CoV-2 HRC-derived lipopeptide as a promising therapeutic candidate for SARS-CoV-2 infections.


Assuntos
Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Lipopeptídeos/farmacologia , Glicoproteína da Espícula de Coronavírus/química , Internalização do Vírus/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Antivirais/química , Betacoronavirus/química , Betacoronavirus/fisiologia , Chlorocebus aethiops , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/transmissão , Células HEK293 , Humanos , Lipopeptídeos/química , Fusão de Membrana/efeitos dos fármacos , Coronavírus da Síndrome Respiratória do Oriente Médio/química , Coronavírus da Síndrome Respiratória do Oriente Médio/efeitos dos fármacos , Coronavírus da Síndrome Respiratória do Oriente Médio/fisiologia , Pandemias/prevenção & controle , Pneumonia Viral/prevenção & controle , Pneumonia Viral/transmissão , Domínios Proteicos , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/virologia , Vírus da SARS/química , Vírus da SARS/efeitos dos fármacos , Vírus da SARS/fisiologia , Células Vero
4.
Drug Discov Ther ; 14(5): 256-258, 2020 Nov 04.
Artigo em Inglês | MEDLINE | ID: mdl-33116037

RESUMO

In the ongoing coronavirus diseases-2019 (COVID-19) crisis that caused immense suffering and deaths, the choice of therapy for the prevention and life-saving conditions must be based on sound scientific evidence. Uncertainty and apprehension are exacerbated in people using angiotensin-converting enzyme (ACE) inhibitors to control their comorbidities such as hypertension and diabetes. These drugs are reported to result in unfavorable outcome as they tend to increase the levels of ACE2 which mediates the entry of SARS-CoV-2. Amiloride, a prototypic inhibitor of epithelial sodium channels (ENaC) can be an ideal candidate for COVID-19 patients, given its ACE reducing and cytosolic pH increasing effects. Moreover, its potassium-sparing and anti-epileptic activities make it a promising alternative or a combinatorial agent.


Assuntos
Amilorida/farmacologia , Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Bloqueadores do Canal de Sódio Epitelial/farmacologia , Pneumonia Viral/tratamento farmacológico , Mucosa Respiratória/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos , Células A549 , Betacoronavirus/patogenicidade , Doenças Cardiovasculares/tratamento farmacológico , Doenças Cardiovasculares/enzimologia , Infecções por Coronavirus/enzimologia , Infecções por Coronavirus/virologia , Regulação para Baixo , Interações Hospedeiro-Patógeno , Humanos , Pandemias , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/enzimologia , Pneumonia Viral/virologia , Receptores Virais/metabolismo , Mucosa Respiratória/enzimologia , Mucosa Respiratória/virologia
5.
EMBO J ; 39(21): e106057, 2020 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-32944968

RESUMO

Coronavirus disease 2019 (COVID-19) is caused by SARS-CoV-2 and has spread across the globe. SARS-CoV-2 is a highly infectious virus with no vaccine or antiviral therapy available to control the pandemic; therefore, it is crucial to understand the mechanisms of viral pathogenesis and the host immune responses to SARS-CoV-2. SARS-CoV-2 is a new member of the betacoronavirus genus like other closely related viruses including SARS-CoV and Middle East respiratory syndrome coronavirus (MERS-CoV). Both SARS-CoV and MERS-CoV have caused serious outbreaks and epidemics in the past eighteen years. Here, we report that one of the interferon-stimulated genes (ISGs), cholesterol 25-hydroxylase (CH25H), is induced by SARS-CoV-2 infection in vitro and in COVID-19-infected patients. CH25H converts cholesterol to 25-hydrocholesterol (25HC) and 25HC shows broad anti-coronavirus activity by blocking membrane fusion. Furthermore, 25HC inhibits USA-WA1/2020 SARS-CoV-2 infection in lung epithelial cells and viral entry in human lung organoids. Mechanistically, 25HC inhibits viral membrane fusion by activating the ER-localized acyl-CoA:cholesterol acyltransferase (ACAT) which leads to the depletion of accessible cholesterol from the plasma membrane. Altogether, our results shed light on a potentially broad antiviral mechanism by 25HC through depleting accessible cholesterol on the plasma membrane to suppress virus-cell fusion. Since 25HC is a natural product with no known toxicity at effective concentrations, it provides a potential therapeutic candidate for COVID-19 and emerging viral diseases in the future.


Assuntos
Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Colesterol/metabolismo , Infecções por Coronavirus/tratamento farmacológico , Pneumonia Viral/tratamento farmacológico , Mucosa Respiratória/virologia , Esteroide Hidroxilases/farmacologia , Internalização do Vírus/efeitos dos fármacos , Acetil-CoA C-Acetiltransferase/metabolismo , Animais , Linhagem Celular , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Chlorocebus aethiops , Ativação Enzimática/efeitos dos fármacos , Humanos , Coronavírus da Síndrome Respiratória do Oriente Médio/efeitos dos fármacos , Organoides/virologia , Pandemias , Mucosa Respiratória/efeitos dos fármacos , Vírus da SARS/efeitos dos fármacos , Células Vero
6.
Front Immunol ; 11: 1959, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32849655

RESUMO

The lung is the vital target organ of coronavirus disease 2019 (COVID-19) caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In the majority of patients the most active virus replication seems to be found in the upper respiratory tract, severe cases however suffer from SARS-like disease associated with virus replication in lung tissues. Due to the current lack of suitable anti-viral drugs the induction of protective immunity such as neutralizing antibodies in the lung is the key aim of the only alternative approach-the development and application of SARS-CoV-2 vaccines. However, past experience from experimental animals, livestock, and humans showed that induction of immunity in the lung is limited following application of vaccines at peripheral sides such as skin or muscles. Based on several considerations we therefore propose here to consider the application of a Modified Vaccinia virus Ankara (MVA)-based vaccine to mucosal surfaces of the respiratory tract as a favorable approach to combat COVID-19.


Assuntos
Betacoronavirus/química , Infecções por Coronavirus/prevenção & controle , Pandemias/prevenção & controle , Pneumonia Viral/prevenção & controle , Glicoproteína da Espícula de Coronavírus/imunologia , Vírus Vaccinia/imunologia , Vacinas Virais/administração & dosagem , Vacinas Virais/imunologia , Administração através da Mucosa , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Brônquios/imunologia , Infecções por Coronavirus/virologia , Humanos , Imunoglobulina A/metabolismo , Tecido Linfoide/imunologia , Plasmócitos/imunologia , Pneumonia Viral/virologia , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/imunologia , Linfócitos T/imunologia , Vacinação , Vacinas Atenuadas/imunologia
7.
Int J Mol Sci ; 21(14)2020 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-32664543

RESUMO

Recently, the world has been dealing with a devastating global pandemic coronavirus infection, with more than 12 million infected worldwide and over 300,000 deaths as of May 15th 2020, related to a novel coronavirus (2019-nCoV), characterized by a spherical morphology and identified through next-generation sequencing. Although the respiratory tract is the primary portal of entry of SARS-CoV-2, gastrointestinal involvement associated with nausea, vomiting and diarrhoea may also occur. No drug or vaccine has been approved due to the absence of evidence deriving from rigorous clinical trials. Increasing interest has been highlighted on the possible preventative role and adjunct treatment of lactoferrin, glycoprotein of human secretions part of a non-specific defensive system, known to play a crucial role against microbial and viral infections and exerting anti-inflammatory effects on different mucosal surfaces and able to regulate iron metabolism. In this review, analysing lactoferrin properties, we propose designing a clinical trial to evaluate and verify its effect using a dual combination treatment with local, solubilized intranasal spray formulation and oral administration. Lactoferrin could counteract the coronavirus infection and inflammation, acting either as natural barrier of both respiratory and intestinal mucosa or reverting the iron disorders related to the viral colonization.


Assuntos
Infecções por Coronavirus/prevenção & controle , Lactoferrina/uso terapêutico , Pandemias/prevenção & controle , Pneumonia Viral/prevenção & controle , Betacoronavirus/isolamento & purificação , Betacoronavirus/fisiologia , Infecções por Coronavirus/patologia , Infecções por Coronavirus/virologia , Humanos , Inflamação , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/virologia , Ferro/metabolismo , Lactoferrina/farmacologia , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/patologia , Pneumonia Viral/virologia , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/virologia , Internalização do Vírus/efeitos dos fármacos
8.
Nature ; 585(7826): 584-587, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32698191

RESUMO

Coronavirus disease 2019 (COVID-19) has rapidly become a global pandemic and no antiviral drug or vaccine is yet available for the treatment of this disease1-3. Several clinical studies are ongoing to evaluate the efficacy of repurposed drugs that have demonstrated antiviral efficacy in vitro. Among these candidates, hydroxychloroquine (HCQ) has been given to thousands of individuals infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-the virus that causes COVID-19-worldwide but there is no definitive evidence that HCQ is effective for treating COVID-194-7. Here we evaluated the antiviral activity of HCQ both in vitro and in SARS-CoV-2-infected macaques. HCQ showed antiviral activity in African green monkey kidney cells (Vero E6) but not in a model of reconstituted human airway epithelium. In macaques, we tested different treatment strategies in comparison to a placebo treatment, before and after peak viral load, alone or in combination with azithromycin (AZTH). Neither HCQ nor the combination of HCQ and AZTH showed a significant effect on viral load in any of the analysed tissues. When the drug was used as a pre-exposure prophylaxis treatment, HCQ did not confer protection against infection with SARS-CoV-2. Our findings do not support the use of HCQ, either alone or in combination with AZTH, as an antiviral drug for the treatment of COVID-19 in humans.


Assuntos
Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Hidroxicloroquina/uso terapêutico , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologia , Animais , Azitromicina/farmacologia , Azitromicina/uso terapêutico , Chlorocebus aethiops , Infecções por Coronavirus/patologia , Infecções por Coronavirus/fisiopatologia , Citocinas/sangue , Modelos Animais de Doenças , Feminino , Humanos , Hidroxicloroquina/farmacocinética , Hidroxicloroquina/farmacologia , Técnicas In Vitro , Cinética , Macaca fascicularis , Masculino , Pandemias , Pneumonia Viral/patologia , Pneumonia Viral/fisiopatologia , Profilaxia Pré-Exposição , Mucosa Respiratória/citologia , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/virologia , Fatores de Tempo , Falha de Tratamento , Células Vero , Carga Viral/efeitos dos fármacos
9.
Ecotoxicol Environ Saf ; 203: 110956, 2020 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-32678753

RESUMO

BACKGROUND: Atmospheric pollutants could induced over-expression of Muc5ac, which is a major pathological feature in acute exacerbation of Chronic Obstructive Pulmonary Disease (COPD) and fatal asthma. Notch signaling pathway could promote mucus cell proliferation and mucus secretion. However, the effects of Notch signaling pathway on the airway mucus secretion induced by PM2.5 remain unknown. In this study, we investigated the role of the Notch signaling pathway on Muc5ac by atmospheric PM2.5 in Beas-2B cell. METHODS: The mRNA and protein levels of the Notch1-4, downstream target gene Hes1 and Muc5ac in the Notch signaling pathway were detected by qPCR and western after Beas-2B cells were exposed to PM2.5 of different concentrations for 12h, 24h, and 48h. RESULTS: The longer the exposure time and the higher the concentration of PM2.5, the lower the survival rate of Beas-2B cells. The expressions of Hes1 and Muc5ac in mRNA and protein were significantly increased after PM2.5 exposure. Correlation analysis indicated that there was a positive correlation between the expression of Muc5ac and Hes1 in mRNA and protein. CONCLUSION: Atmospheric PM2.5 can induce the express of Muc5ac, the Notch signaling pathway may be involved in the regulation of Muc5ac by Hes1.


Assuntos
Poluentes Atmosféricos/toxicidade , Células Epiteliais/efeitos dos fármacos , Mucina-5AC/biossíntese , Material Particulado/toxicidade , Receptores Notch/metabolismo , Mucosa Respiratória/efeitos dos fármacos , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Humanos , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Transdução de Sinais
10.
Antiviral Res ; 180: 104860, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32565134

RESUMO

Middle East Respiratory Syndrome Coronavirus (MERS-CoV) causes severe respiratory in human with high mortality and it has been a challenge to determine optimum treatment for MERS-CoV-induced respiratory infection. Here, we observed the distribution of MERS-CoV receptors using human respiratory mucosa and also evaluated the contribution of interferon-lambdas (IFN-λs) in response to MERS-CoV infection using in vitro normal human nasal epithelial (NHNE) and bronchial epithelial (NHBE) cells. We found that the gene and protein expression of DPPIV, MERS-CoV receptor, were more dominantly located in nasal and bronchial epithelium although human nasal mucosa exhibited relatively lower DPPIV expression than lung parenchymal tissues. The quantitative mRNA level of the MERS-CoV envelope (upE) gene was significantly induced in MERS-CoV-infected cultured NHNE and NHBE cells until 3 days after infection. The induction of IFNs was identified in NHNE and NHBE cells after MERS-CoV infection and IFN-λs were predominantly increased in MERS-CoV-infected respiratory epithelial cells. Inoculation of IFN-λs to NHNE and NHBE cells suppressed MERS-CoV replication and in particular, IFN-λ4 showed a strong therapeutic effect in reducing MERS-CoV infection with higher induction of IFN-stimulated genes. Thus, IFN-λ has a decisive function in the respiratory epithelium that greatly limits MERS-CoV replication, and may be a key cytokine for better therapeutic outcomes against MERS-CoV infection in respiratory tract.


Assuntos
Antivirais/uso terapêutico , Interferons/uso terapêutico , Interleucinas/uso terapêutico , Coronavírus da Síndrome Respiratória do Oriente Médio/efeitos dos fármacos , Mucosa Respiratória/virologia , Replicação Viral/efeitos dos fármacos , Brônquios/virologia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Citocinas/metabolismo , Células Epiteliais/virologia , Regulação Viral da Expressão Gênica , Humanos , Imunidade Inata/imunologia , Interferons/biossíntese , Interleucinas/biossíntese , Mucosa Laríngea/virologia , Pulmão/virologia , Coronavírus da Síndrome Respiratória do Oriente Médio/genética , Reação em Cadeia da Polimerase , Mucosa Respiratória/efeitos dos fármacos
11.
J Allergy Clin Immunol ; 146(1): 80-88.e8, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32422146

RESUMO

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has dramatically changed our world, country, communities, and families. There is controversy regarding risk factors for severe COVID-19 disease. It has been suggested that asthma and allergy are not highly represented as comorbid conditions associated with COVID-19. OBJECTIVE: Our aim was to extend our work in IL-13 biology to determine whether airway epithelial cell expression of 2 key mediators critical for SARS-CoV-2 infection, namely, angiotensin-converting enzyme 2 (ACE2) and transmembrane protease, serine 2 (TMPRSS2), are modulated by IL-13. METHODS: We determined effects of IL-13 treatment on ACE2 and TMPRSS2 expression ex vivo in primary airway epithelial cells from participants with and without type 2 asthma obtained by bronchoscopy. We also examined expression of ACE2 and TMPRSS2 in 2 data sets containing gene expression data from nasal and airway epithelial cells from children and adults with asthma and allergic rhinitis. RESULTS: IL-13 significantly reduced ACE2 and increased TMPRSS2 expression ex vivo in airway epithelial cells. In 2 independent data sets, ACE2 expression was significantly reduced and TMPRSS2 expression was significantly increased in the nasal and airway epithelial cells in type 2 asthma and allergic rhinitis. ACE2 expression was significantly negatively associated with type 2 cytokines, whereas TMPRSS2 expression was significantly positively associated with type 2 cytokines. CONCLUSION: IL-13 modulates ACE2 and TMPRSS2 expression in airway epithelial cells in asthma and atopy. This deserves further study with regard to any effects that asthma and atopy may render in the setting of COVID-19 infection.


Assuntos
Asma/imunologia , Infecções por Coronavirus/imunologia , Hipersensibilidade Imediata/imunologia , Interleucina-13/imunologia , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/imunologia , Serina Endopeptidases/metabolismo , Adulto , Asma/metabolismo , Betacoronavirus/imunologia , Criança , Infecções por Coronavirus/metabolismo , Feminino , Humanos , Hipersensibilidade Imediata/metabolismo , Inflamação/imunologia , Inflamação/virologia , Interleucina-13/farmacologia , Masculino , Pandemias , Pneumonia Viral/metabolismo , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/imunologia , Mucosa Respiratória/metabolismo
12.
Med Sci (Paris) ; 36(4): 382-388, 2020 Apr.
Artigo em Francês | MEDLINE | ID: mdl-32356715

RESUMO

As burden of chronic respiratory diseases is constantly increasing, improving in vitro lung models is essential in order to reproduce as closely as possible the complex pulmonary architecture, responsible for oxygen uptake and carbon dioxide clearance. The study of diseases that affect the respiratory system has benefited from in vitro reconstructions of the respiratory epithelium with inserts in air/liquid interface (2D) or in organoids able to mimic up to the arborescence of the respiratory tree (3D). Recent development in the fields of pluripotent stem cells-derived organoids and genome editing technologies has provided new insights to better understand pulmonary diseases and to find new therapeutic perspectives.


Assuntos
Técnicas de Cultura de Células , Pulmão/citologia , Organoides/citologia , Células Epiteliais Alveolares/citologia , Células Epiteliais Alveolares/fisiologia , Animais , Bioengenharia/métodos , Bioengenharia/tendências , Dióxido de Carbono/farmacologia , Dióxido de Carbono/fisiologia , Técnicas de Cultura de Células/métodos , Técnicas de Cultura de Células/tendências , Células Cultivadas , Edição de Genes/métodos , Edição de Genes/tendências , Humanos , Pulmão/patologia , Pulmão/fisiologia , Modelos Biológicos , Organoides/patologia , Organoides/fisiologia , Oxigênio/farmacologia , Oxigênio/fisiologia , Troca Gasosa Pulmonar/fisiologia , Mucosa Respiratória/citologia , Mucosa Respiratória/efeitos dos fármacos , Tecidos Suporte/química
13.
Med Hypotheses ; 140: 109751, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32344304

RESUMO

COVID-19 pandemic is a major challenge for global and national healthcare providers. Number of new cases is continuously increasing with an emerging trend showing worse prognosis in males in comparison to females. Based on this observation, our proposed hypothesis is that 5-alpha-reductase inhibitors, that are commonly used for BPH treatment, may be one of the factors contributing to poorer prognosis in males.


Assuntos
Inibidores de 5-alfa Redutase/efeitos adversos , Infecções por Coronavirus/complicações , Pneumonia Viral/complicações , Animais , Betacoronavirus , Finasterida/efeitos adversos , Humanos , Pulmão/efeitos dos fármacos , Pulmão/embriologia , Masculino , Pandemias , Prognóstico , Hiperplasia Prostática/complicações , Hiperplasia Prostática/tratamento farmacológico , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/embriologia , Fatores Sexuais
14.
Am J Respir Cell Mol Biol ; 63(2): 172-184, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32275839

RESUMO

Air pollution particulate matter <2.5 µm (PM2.5) exposure is associated with poor respiratory outcomes. Mechanisms underlying PM2.5-induced lung pathobiology are poorly understood but likely involve cellular and molecular changes to the airway epithelium. We extracted and chemically characterized the organic and water-soluble components of air pollution PM2.5 samples, then determined the whole transcriptome response of human nasal mucociliary airway epithelial cultures to a dose series of PM2.5 extracts. We found that PM2.5 organic extract (OE), but not water-soluble extract, elicited a potent, dose-dependent transcriptomic response from the mucociliary epithelium. Exposure to a moderate OE dose modified the expression of 424 genes, including activation of aryl hydrocarbon receptor signaling and an IL-1 inflammatory program. We generated an OE-response gene network defined by eight functional enrichment groups, which exhibited high connectivity through CYP1A1, IL1A, and IL1B. This OE exposure also robustly activated a mucus secretory expression program (>100 genes), which included transcriptional drivers of mucus metaplasia (SPDEF and FOXA3). Exposure to a higher OE dose modified the expression of 1,240 genes and further exacerbated expression responses observed at the moderate dose, including the mucus secretory program. Moreover, the higher OE dose significantly increased the MUC5AC/MUC5B gel-forming mucin expression ratio and strongly downregulated ciliated cell expression programs, including key ciliating cell transcription factors (e.g., FOXJ1 and MCIDAS). Chronic OE stimulation induced mucus metaplasia-like remodeling characterized by increases in MUC5AC+ secretory cells and MUC5AC mucus secretions. This epithelial remodeling may underlie poor respiratory outcomes associated with high PM2.5 exposure.


Assuntos
Mucosa Nasal/diagnóstico por imagem , Material Particulado/efeitos adversos , Mucosa Respiratória/efeitos dos fármacos , Poluentes Atmosféricos/efeitos adversos , Poluição do Ar/efeitos adversos , Asma/induzido quimicamente , Asma/genética , Estudo de Associação Genômica Ampla/métodos , Humanos , Inflamação/induzido quimicamente , Inflamação/genética , Mucina-5AC/genética , Mucina-5B/genética , Fatores de Transcrição/genética
15.
Am J Physiol Lung Cell Mol Physiol ; 318(5): L888-L899, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32130032

RESUMO

We have previously demonstrated that upregulation of Sonic hedgehog (SHH) expression in allergic airway epithelia essentially contributes to the goblet cell metaplasia and mucous hypersecretion. However, the mechanism underlying the upregulation of SHH expression remains completely unknown. In cultured human airway epithelial cells, IL-4/IL-13 but not IL-5 robustly induces the mRNA and protein expression of SHH and in turn activates SHH signaling by promoting the JAK/STAT6-controlling transcription of SHH gene. Moreover, intratracheal instillation of IL-4 and/or IL-13 robustly activates STAT6 and concomitantly upregulates SHH expression in mouse airway epithelia, whereas, in Club cell 10-kDa protein (CC10)-positive airway epithelial cells of children with asthma, activated STAT6 closely correlates with the increased expression of SHH and high activity of SHH signaling. Finally, intratracheal inhibition of STAT6 by AS-1517499 significantly diminished the allergen-induced upregulation of SHH expression, goblet cell phenotypes, and airway hyperresponsiveness, in an ovalbumin- or house dust mite-induced mouse model with allergic airway inflammation,. Together, upregulation of SHH expression by IL-4/IL-13-induced JAK/STAT6 signaling contributes to allergic airway epithelial remodeling, and this study thus provides insight into how morphogen signaling is coordinated with Th2 cytokine pathways to regulate tissue remodeling in chronic airway diseases.


Assuntos
Asma/genética , Proteínas Hedgehog/genética , Interleucina-13/genética , Interleucina-4/genética , Mucosa Respiratória/imunologia , Animais , Antiasmáticos/farmacologia , Asma/induzido quimicamente , Asma/tratamento farmacológico , Asma/patologia , Linhagem Celular , Criança , Feminino , Regulação da Expressão Gênica , Células Caliciformes/efeitos dos fármacos , Células Caliciformes/imunologia , Células Caliciformes/patologia , Proteínas Hedgehog/imunologia , Humanos , Interleucina-13/imunologia , Interleucina-13/farmacologia , Interleucina-4/imunologia , Interleucina-4/farmacologia , Interleucina-5/genética , Interleucina-5/imunologia , Janus Quinases/genética , Janus Quinases/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Ovalbumina/administração & dosagem , Cultura Primária de Células , Pirimidinas/farmacologia , Pyroglyphidae/química , Pyroglyphidae/imunologia , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/patologia , Fator de Transcrição STAT6/antagonistas & inibidores , Fator de Transcrição STAT6/genética , Fator de Transcrição STAT6/imunologia , Transdução de Sinais , Transcrição Genética , Uteroglobina/genética , Uteroglobina/imunologia
16.
Toxicol Lett ; 325: 25-33, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32112875

RESUMO

RATIONALE: Diacetyl (DA; 2,3-butanedione) is a chemical found commonly in foods and e-cigarettes. When inhaled, DA causes epithelial injury, though the mechanism of repair remain poorly understood. The objective of this study was to evaluate airway basal cell repair after DA vapor exposure. METHODS: Primary human bronchial epithelial cells were exposed to DA or PBS for 1 h. Lactate dehydrogenase, cleaved caspase 3/7 and trans-epithelial electrical resistance were measured prior to and following exposure. Exposed cultures were analyzed for the airway basal cell markers keratin 5 and p63 as well as ubiquitin and proteasome activity. Cultures were also treated with a proteasome inhibitor (MG132). RESULTS: DA vapor exposure caused a transient decrease in trans-epithelial electrical resistance in all DA-exposed cultures. Supernatant lactate dehydrogenase and cleaved caspase 3/7 increased significantly at the highest DA concentration but not at lower DA concentrations. Increased keratin 5 ubiquitination occurred after DA exposure but resolved by day 3. Damage to airway basal cells persisted at day 3 in the presence of MG132. CONCLUSIONS: Diacetyl exposure results in airway basal cell injury with keratin 5 ubiquitination and decreased p63 expression. The ubiquitin-proteasome-pathway partially mediates airway basal cell repair after acute DA exposure.


Assuntos
Diacetil/toxicidade , Mucosa Respiratória/patologia , Biomarcadores , Brônquios/citologia , Brônquios/patologia , Caspases/metabolismo , Diacetil/administração & dosagem , Impedância Elétrica , Sistemas Eletrônicos de Liberação de Nicotina , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Humanos , Exposição por Inalação , Queratina-5/metabolismo , L-Lactato Desidrogenase/metabolismo , Leupeptinas/farmacologia , Proteínas de Membrana , Complexo de Endopeptidases do Proteassoma/efeitos dos fármacos , Mucosa Respiratória/efeitos dos fármacos , Ubiquitinação/efeitos dos fármacos
17.
Ecotoxicol Environ Saf ; 195: 110455, 2020 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-32199215

RESUMO

Inhalation of neodymium oxide, a type of rare earth metal, can induce various respiratory diseases, such as lung tissue inflammation, but the associated mechanisms underlying this induction are still unclear. In this study, we explored the role and mechanisms of circular RNA in neodymium oxide-induced airway inflammation. Our study showed that treating human bronchial epithelial (16HBE) cells with neodymium oxide caused an inflammatory response by upregulating the expression of interleukin-8 (IL-8) and interleukin-1 beta (IL-1ß). Quantitative real-time polymerase chain reaction (qRT-PCR) analyses revealed significant downregulation of circRNA_0000638 in neodymium oxide-treated 16HBE cells. Data from functional analyses further showed that circ_0000638 inhibited inflammation induced by neodymium oxide in 16HBE cells. circ_0000638 targeted miR-498-5p and further increased the expression of NKRF (NF-κB repressing factor), which inhibited the activation of NF-κB (nuclear factor κB). Moreover, circ_0000638 reduced the expression of IL-8 and IL-1ß by inhibiting NF-κB activation in neodymium oxide-treated 16HBE cells. These results suggest that circ_0000638 can inhibit NF-κB activation by competitively binding to miR-498-5p, further downregulating the expression of IL-8 and IL-1ß in neodymium oxide-treated 16HBE cells.


Assuntos
MicroRNAs/metabolismo , NF-kappa B/metabolismo , Neodímio/farmacologia , Óxidos/farmacologia , RNA Circular/metabolismo , Mucosa Respiratória/efeitos dos fármacos , Linhagem Celular , Regulação para Baixo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Regulação da Expressão Gênica , Humanos , Mediadores da Inflamação/metabolismo , Interleucina-1beta/metabolismo , Proteínas Repressoras/metabolismo , Mucosa Respiratória/metabolismo , Transdução de Sinais
18.
Am J Physiol Lung Cell Mol Physiol ; 318(5): L1063-L1073, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32208929

RESUMO

The human bronchial epithelium is an important barrier tissue that is damaged or pathologically altered in various acute and chronic respiratory conditions. To represent the epithelial component of respiratory disease, it is essential to use a physiologically relevant model of this tissue. The human bronchial epithelium is a highly organized tissue consisting of a number of specialized cell types. Primary human bronchial epithelial cells (HBEC) can be differentiated into a mucociliated tissue in air-liquid interface (ALI) cultures using appropriately supplemented media under optimized growth conditions. We compared the histology, ciliary length, and function, diffusion, and barrier properties of HBEC from donors with no respiratory disease grown in two different media, PneumaCult-ALI or Bronchial Epithelial Differentiation Medium (BEDM). In the former group, HBEC have a more physiological pseudostratified morphology and mucociliary differentiation, including increased epithelial thickness, intracellular expression of airway-specific mucin protein MUC5AC, and total expression of cilia basal-body protein compared with cells from the same donor grown in the other medium. Baseline expression levels of inflammatory mediators, thymic stromal lymphopoietin (TSLP), soluble ST2, and eotaxin-3 were lower in PneumaCult-ALI. Additionally, the physiological cilia beat frequency and electrical barrier properties with transepithelial electrical resistance were significantly different between the two groups. Our study has shown that these primary cell cultures from the same donor grown in the two media possess variable structural and functional characteristics. Therefore, it is important to objectively validate primary epithelial cell cultures before experimentation to ensure they are appropriate to answer a specific scientific question.


Assuntos
Meios de Cultura/farmacologia , Células Epiteliais/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Mucosa Respiratória/efeitos dos fármacos , Ar , Brônquios/citologia , Brônquios/metabolismo , Diferenciação Celular/efeitos dos fármacos , Quimiocina CCL26/genética , Quimiocina CCL26/metabolismo , Cílios/efeitos dos fármacos , Cílios/metabolismo , Meios de Cultura/química , Citocinas/genética , Citocinas/metabolismo , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Voluntários Saudáveis , Humanos , Proteína 1 Semelhante a Receptor de Interleucina-1/genética , Proteína 1 Semelhante a Receptor de Interleucina-1/metabolismo , Modelos Biológicos , Mucina-5AC/genética , Mucina-5AC/metabolismo , Cultura Primária de Células , Mucosa Respiratória/citologia , Mucosa Respiratória/metabolismo
19.
Toxicol Appl Pharmacol ; 394: 114959, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32201329

RESUMO

Arsenic is a ubiquitous environmental toxicant, found in high concentrations worldwide. Although abundant research has dealt with arsenic-induced cancers, studies on mechanisms of non-malignant lung diseases have not been complete. In addition, decades of research have mostly concentrated on high-dose arsenic exposure, which has very limited use in modeling the biological effects of today's low-dose exposures. Indeed, accumulated evidence has shown that low-dose arsenic exposure (i.e. ≤100 ppb) may also alter lung homeostasis by causing host susceptibility to viral infection. However, the underlying mechanism of this alteration is unknown. In this study, we found that low-dose sodium arsenite (As (III)) repressed major airway mucins-MUC5AC and MUC5B at both mRNA and protein levels. We further demonstrated that this repression was not caused by cellular toxicity or mediated by the reduction of a common mucin-inducing pathway-EGFR. Other established mucin activators- dsRNA, IL1ß or IL17 were not able to override As (III)-induced mucin repression. Interestingly, the suppressing effect of As (III) appeared to be partially reversible, and supplementation of all trans retinoic acid (t-RA) doses dependently restored mucin gene expression. Further analyses indicated that As (III) treatment significantly reduced the protein level of retinoic acid receptors (RARα, γ and RXRα) as well as RARE promoter reporter activity. Therefore, our study fills in an important knowledge gap in the field of low-dose arsenic exposure. The interference of RA signaling, and mucin gene expression may be important pathogenic factors in low-dose arsenic induced lung toxicity.


Assuntos
Arsênico/toxicidade , Mucinas/biossíntese , Mucosa Respiratória/metabolismo , Transdução de Sinais/efeitos dos fármacos , Tretinoína , Arsenitos/toxicidade , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Receptores ErbB/antagonistas & inibidores , Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Mucina-5AC/antagonistas & inibidores , Mucina-5AC/genética , Mucina-5B/antagonistas & inibidores , Mucina-5B/genética , Mucosa Respiratória/efeitos dos fármacos , Compostos de Sódio/toxicidade
20.
Am J Physiol Lung Cell Mol Physiol ; 318(5): L873-L887, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-32160007

RESUMO

Tenacious mucus produced by tracheal and bronchial submucosal glands is a defining feature of several airway diseases, including cystic fibrosis (CF). Airway acidification as a driving force of CF airway pathology has been controversial. Here we tested the hypothesis that transient airway acidification produces pathologic mucus and impairs mucociliary transport. We studied pigs challenged with intra-airway acid. Acid had a minimal effect on mucus properties under basal conditions. However, cholinergic stimulation in acid-challenged pigs revealed retention of mucin 5B (MUC5B) in the submucosal glands, decreased concentrations of MUC5B in the lung lavage fluid, and airway obstruction. To more closely mimic a CF-like environment, we also examined mucus secretion and transport following cholinergic stimulation under diminished bicarbonate and chloride transport conditions ex vivo. Under these conditions, airways from acid-challenged pigs displayed extensive mucus films and decreased mucociliary transport. Pretreatment with diminazene aceturate, a small molecule with ability to inhibit acid detection through blockade of the acid-sensing ion channel (ASIC) at the doses provided, did not prevent acid-induced pathologic mucus or transport defects but did mitigate airway obstruction. These findings suggest that transient airway acidification early in life has significant impacts on mucus secretion and transport properties. Furthermore, they highlight diminazene aceturate as an agent that might be beneficial in alleviating airway obstruction.


Assuntos
Ácido Acético/administração & dosagem , Bloqueadores do Canal Iônico Sensível a Ácido/farmacologia , Canais Iônicos Sensíveis a Ácido/genética , Obstrução das Vias Respiratórias/induzido quimicamente , Fibrose Cística/induzido quimicamente , Diminazena/análogos & derivados , Canais Iônicos Sensíveis a Ácido/metabolismo , Obstrução das Vias Respiratórias/tratamento farmacológico , Obstrução das Vias Respiratórias/metabolismo , Obstrução das Vias Respiratórias/patologia , Animais , Animais Recém-Nascidos , Bicarbonatos/metabolismo , Brônquios/efeitos dos fármacos , Brônquios/metabolismo , Brônquios/patologia , Líquido da Lavagem Broncoalveolar/química , Cloretos/metabolismo , Fibrose Cística/tratamento farmacológico , Fibrose Cística/metabolismo , Fibrose Cística/patologia , Diminazena/farmacologia , Modelos Animais de Doenças , Feminino , Expressão Gênica , Humanos , Concentração de Íons de Hidrogênio , Masculino , Mucina-5AC/genética , Mucina-5AC/metabolismo , Mucina-5B/genética , Mucina-5B/metabolismo , Depuração Mucociliar/efeitos dos fármacos , Muco/metabolismo , Mucosa Respiratória/efeitos dos fármacos , Mucosa Respiratória/metabolismo , Mucosa Respiratória/patologia , Suínos , Traqueia/efeitos dos fármacos , Traqueia/metabolismo , Traqueia/patologia
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