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1.
Ecotoxicol Environ Saf ; 205: 111283, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-32977282

RESUMO

Fine particulate matter (PM2.5) airborne pollution increases the risk of chronic respiratory diseases, such as idiopathic pulmonary fibrosis (IPF), which is characterized by non-specific inflammation of the interstitial lung and extensive deposition of collagen fibers. Type 2 alveolar epithelial cells (AEC2s) are alveolar stem cells in the adult lung that contribute to the lung repair process through complex signaling. Our previous studies demonstrated that OGG1, a kind of DNA repair enzyme, have a critical role in protecting cells from oxidative damage and apoptosis induced by PM2.5, but the contribution of OGG1 in proliferation and self-renewal of AEC2s is not known. Here, we constructed OGG1-/-mice to test the effect and mechanism of OGG1 on PM2.5-induced pulmonary fibrosis and injury in vivo. We detected proliferation and self-renewal of OGG1 overexpression or OGG1 knockout AEC2s after PM2.5 injury by flow cytometry and clone formation. We observed that knockout of OGG1 aggravated pulmonary fibrosis, oxidative stress, and AEC2 cell death in PM2.5-injured mice. In addition, OGG1 is required for the proliferation and renewal of AEC2s after PM2.5 injury. Overexpression of OGG1 promotes the proliferation and self-renewal of AEC2s by inhibiting PM2.5-mediated oxidative stress and NF-κB signaling hyperactivation in vitro. Furthermore, NF-κB inhibitors promoted proliferation and self-renewal of OGG1-deficient AEC2s cells after PM2.5 injury, and attenuated PM2.5-induced pulmonary fibrosis and injury in mice. These data establish OGG1 as a regulator of NF-κB signal that serves to regulate AEC2 cell proliferation and self-renewal, and suggest a mechanism that inhibition of the NF-κB signaling pathway may represent a potential therapeutic strategy for IPF patients with low-expression of OGG1.


Assuntos
Poluentes Atmosféricos/toxicidade , Células Epiteliais Alveolares/efeitos dos fármacos , Autorrenovação Celular/genética , DNA Glicosilases/metabolismo , Material Particulado/toxicidade , Fibrose Pulmonar/induzido quimicamente , Células-Tronco/efeitos dos fármacos , Células Epiteliais Alveolares/metabolismo , Células Epiteliais Alveolares/patologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , DNA Glicosilases/genética , Humanos , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/genética , Fibrose Pulmonar/genética , Fibrose Pulmonar/patologia , Transdução de Sinais , Células-Tronco/metabolismo , Células-Tronco/patologia
2.
Adv Exp Med Biol ; 1255: 51-62, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32949389

RESUMO

DNA methylations, including global methylation pattern and specific gene methylation, are associated with pathogenesis and progress of pulmonary fibrosis. This chapter illustrates alteration of DNA methylation in pulmonary fibrosis as a predictive or prognostic factor. Treatment with the DNA methylation inhibitors will be an emerging anti-fibrosis therapy, although we are still in the pre-clinical stage of using epigenetic markers as potential targets for biomarkers and therapeutic interventions.


Assuntos
Metilação de DNA , Fibrose Pulmonar/genética , Epigenômica , Humanos
3.
Nat Cell Biol ; 22(8): 934-946, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32661339

RESUMO

Stem cells undergo dynamic changes in response to injury to regenerate lost cells. However, the identity of transitional states and the mechanisms that drive their trajectories remain understudied. Using lung organoids, multiple in vivo repair models, single-cell transcriptomics and lineage tracing, we find that alveolar type-2 epithelial cells undergoing differentiation into type-1 cells acquire pre-alveolar type-1 transitional cell state (PATS) en route to terminal maturation. Transitional cells undergo extensive stretching during differentiation, making them vulnerable to DNA damage. Cells in the PATS show an enrichment of TP53, TGFß, DNA-damage-response signalling and cellular senescence. Gain and loss of function as well as genomic binding assays revealed a direct transcriptional control of PATS by TP53 signalling. Notably, accumulation of PATS-like cells in human fibrotic lungs was observed, suggesting persistence of the transitional state in fibrosis. Our study thus implicates a transient state associated with senescence in normal epithelial tissue repair and its abnormal persistence in disease conditions.


Assuntos
Células Epiteliais Alveolares , Diferenciação Celular , Fibrose Pulmonar/patologia , Células-Tronco Adultas/patologia , Células Epiteliais Alveolares/patologia , Animais , Linhagem da Célula , Forma Celular , Senescência Celular , Dano ao DNA , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Organoides , Fibrose Pulmonar/genética , Transdução de Sinais , Proteína Supressora de Tumor p53/metabolismo
4.
Respir Res ; 21(1): 182, 2020 Jul 14.
Artigo em Inglês | MEDLINE | ID: mdl-32664949

RESUMO

BACKGROUND: Severe acute respiratory syndrome (SARS)-CoV-2-induced coronavirus disease-2019 (COVID-19) is a pandemic disease that affects > 2.8 million people worldwide, with numbers increasing dramatically daily. However, there is no specific treatment for COVID-19 and much remains unknown about this disease. Angiotensin-converting enzyme (ACE)2 is a cellular receptor of SARS-CoV-2. It is cleaved by type II transmembrane serine protease (TMPRSS)2 and disintegrin and metallopeptidase domain (ADAM)17 to assist viral entry into host cells. Clinically, SARS-CoV-2 infection may result in acute lung injury and lung fibrosis, but the underlying mechanisms of COVID-19 induced lung fibrosis are not fully understood. METHODS: The networks of ACE2 and its interacting molecules were identified using bioinformatic methods. Their gene and protein expressions were measured in human epithelial cells after 24 h SARS-CoV-2 infection, or in existing datasets of lung fibrosis patients. RESULTS: We confirmed the binding of SARS-CoV-2 and ACE2 by bioinformatic analysis. TMPRSS2, ADAM17, tissue inhibitor of metalloproteinase (TIMP)3, angiotensinogen (AGT), transformation growth factor beta (TGFB1), connective tissue growth factor (CTGF), vascular endothelial growth factor (VEGF) A and fibronectin (FN) were interacted with ACE2, and the mRNA and protein of these molecules were expressed in lung epithelial cells. SARS-CoV-2 infection increased ACE2, TGFB1, CTGF and FN1 mRNA that were drivers of lung fibrosis. These changes were also found in lung tissues from lung fibrosis patients. CONCLUSIONS: Therefore, SARS-CoV-2 binds with ACE2 and activates fibrosis-related genes and processes to induce lung fibrosis.


Assuntos
Infecções por Coronavirus/genética , Regulação da Expressão Gênica , Peptidil Dipeptidase A/genética , Pneumonia Viral/genética , Fibrose Pulmonar/genética , Síndrome do Desconforto Respiratório do Adulto/genética , Vírus da SARS/genética , China , Infecções por Coronavirus/epidemiologia , Infecções por Coronavirus/fisiopatologia , Progressão da Doença , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Feminino , Humanos , Masculino , Pandemias/estatística & dados numéricos , Pneumonia Viral/epidemiologia , Pneumonia Viral/fisiopatologia , Prevalência , Fibrose Pulmonar/epidemiologia , Fibrose Pulmonar/etiologia , Receptores Virais/metabolismo , Síndrome do Desconforto Respiratório do Adulto/diagnóstico , Síndrome do Desconforto Respiratório do Adulto/epidemiologia , Medição de Risco , Análise de Sobrevida , Transcrição Genética , Ativação Transcricional/genética
5.
Presse Med ; 49(2): 104024, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32437840

RESUMO

At least 10% of patients with pulmonary fibrosis, whether idiopathic or secondary, present heritable pulmonary fibrosis suspected on familial aggregation of pulmonary fibrosis, specific syndromes or early age of diagnosis. Approximately 30% of those patients have an identified mutation mostly in telomere related genes (TRG) more rarely in surfactant homeostasis or other genes. TRG mutation may be associated with hematological and hepatic diseases that may worsen after lung transplantation requiring a specific care and adapted immunosuppression. Surfactant genes mutations are usually associated with ground-glass opacities and cysts on CT scan and may improve with steroids, hydroxychloroquine or azithromycin. Moreover relatives should benefit from a genetic analysis associated with a clinical evaluation according to the gene involved. Genetics of pulmonary fibrosis raise specific problems from diagnosis, therapy or genetic counseling varying from one gene to another.


Assuntos
Doenças Pulmonares Intersticiais/genética , Proteínas Associadas a Surfactantes Pulmonares/genética , Telômero/genética , Adulto , Feminino , Predisposição Genética para Doença , Humanos , Incidência , Doenças Pulmonares Intersticiais/diagnóstico por imagem , Doenças Pulmonares Intersticiais/tratamento farmacológico , Doenças Pulmonares Intersticiais/epidemiologia , Masculino , Mutação , Linhagem , Fibrose Pulmonar/diagnóstico por imagem , Fibrose Pulmonar/epidemiologia , Fibrose Pulmonar/genética , Fatores de Risco , Esteroides/uso terapêutico , Tomografia Computadorizada por Raios X
6.
Artigo em Inglês | MEDLINE | ID: mdl-32209025

RESUMO

Alveolar epithelial cell (AEC) apoptosis, arising from mitochondrial dysfunction and mitophagy defects, is important in mediating idiopathic pulmonary fibrosis (IPF). Our group established a role for the mitochondrial (mt) DNA base excision repair enzyme, 8-oxoguanine-DNA glycosylase 1 (mtOGG1), in preventing oxidant-induced AEC mtDNA damage and apoptosis and showed that OGG1-deficient mice have increased lung fibrosis. Herein, we determined whether mice overexpressing the mtOGG1 transgene (mtOgg1tg) are protected against lung fibrosis and whether AEC mtOGG1 preservation of mtDNA integrity mitigates phosphatase and tensin homolog-induced putative kinase 1 (PINK1) deficiency and apoptosis. Compared with wild type (WT), mtOgg1tg mice have diminished asbestos- and bleomycin-induced pulmonary fibrosis that was accompanied by reduced lung and AEC mtDNA damage and apoptosis. Asbestos and H2O2 promote the MLE-12 cell PINK1 deficiency, as assessed by reductions in the expression of PINK1 mRNA and mitochondrial protein expression. Compared with WT, Pink1-knockout (Pink1-KO) mice are more susceptible to asbestos-induced lung fibrosis and have increased lung and alveolar type II (AT2) cell mtDNA damage and apoptosis. AT2 cells from Pink1-KO mice and PINK1-silenced (siRNA) MLE-12 cells have increased mtDNA damage that is augmented by oxidative stress. Interestingly, mtOGG1 overexpression attenuates oxidant-induced MLE-12 cell mtDNA damage and apoptosis despite PINK1 silencing. mtDNA damage is increased in the lungs of patients with IPF as compared with controls. Collectively, these findings suggest that mtOGG1 maintenance of AEC mtDNA is crucial for preventing PINK1 deficiency that promotes apoptosis and lung fibrosis. Given the key role of AEC apoptosis in pulmonary fibrosis, strategies aimed at preserving AT2 cell mtDNA integrity may be an innovative target.


Assuntos
Células Epiteliais Alveolares/efeitos dos fármacos , Asbestose/genética , DNA Glicosilases/genética , Pulmão/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Proteínas Quinases/genética , Fibrose Pulmonar/genética , Células Epiteliais Alveolares/metabolismo , Células Epiteliais Alveolares/patologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Asbestos/administração & dosagem , Asbestose/etiologia , Asbestose/metabolismo , Asbestose/patologia , Bleomicina/administração & dosagem , Dano ao DNA , DNA Glicosilases/deficiência , DNA Mitocondrial/genética , DNA Mitocondrial/metabolismo , Feminino , Regulação da Expressão Gênica , Peróxido de Hidrogênio/farmacologia , Pulmão/metabolismo , Pulmão/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mitocôndrias/metabolismo , Cultura Primária de Células , Proteínas Quinases/metabolismo , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/patologia , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Titânio/administração & dosagem
7.
Immunity ; 52(3): 542-556.e13, 2020 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-32187520

RESUMO

Fibrosis is an incurable disorder of unknown etiology. Segregated-nucleus-containing atypical monocytes (SatMs) are critical for the development of fibrosis. Here we examined the mechanisms that recruit SatMs to pre-fibrotic areas. A screen based on cytokine expression in the fibrotic lung revealed that the chemokine Cxcl12, which is produced by apoptotic nonhematopoietic cells, was essential for SatM recruitment. Analyses of lung tissues at fibrosis onset showed increased expression of Rbm7, a component of the nuclear exosome targeting complex. Rbm7 deletion suppressed bleomycin-induced fibrosis and at a cellular level, suppressed apoptosis of nonhematopoietic cells. Mechanistically, Rbm7 bound to noncoding (nc)RNAs that form subnuclear bodies, including Neat1 speckles. Dysregulated expression of Rbm7 resulted in the nuclear degradation of Neat1 speckles, the dispersion of the DNA repair protein BRCA1, and the triggering of apoptosis. Thus, Rbm7 in epithelial cells plays a critical role in the development of fibrosis by regulating ncRNA decay and thereby the production of chemokines that recruit SatMs.


Assuntos
Apoptose/imunologia , Núcleo Celular/imunologia , Exossomos/imunologia , Fibrose Pulmonar/imunologia , Proteínas de Ligação a RNA/imunologia , Animais , Apoptose/genética , Núcleo Celular/genética , Núcleo Celular/metabolismo , Quimiocina CXCL12/imunologia , Quimiocina CXCL12/metabolismo , Exossomos/genética , Exossomos/metabolismo , Regulação da Expressão Gênica/imunologia , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Monócitos/imunologia , Monócitos/metabolismo , Células NIH 3T3 , Fibrose Pulmonar/genética , Fibrose Pulmonar/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo
8.
FASEB J ; 34(2): 2011-2023, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31907997

RESUMO

Src Homology 2-containing Inositol Phosphatase-1 (SHIP-1) is a target of miR-155, a pro-inflammatory factor. Deletion of the SHIP-1 gene in mice caused spontaneous lung inflammation and fibrosis. However, the role and function of endothelial miR-155 and SHIP-1 in lung fibrosis remain unknown. Using whole-body miR-155 knockout mice and endothelial cell-specific conditional miR-155 (VEC-Cre-miR-155 or VEC-miR-155) or SHIP-1 (VEC-SHIP-1) knockout mice, we assessed endothelial-mesenchymal transition (EndoMT) and fibrotic responses in bleomycin (BLM) induced lung fibrosis models. Primary mouse lung endothelial cells (MLEC) and human umbilical vein endothelial cells (HUVEC) with SHIP-1 knockdown were analyzed in TGF-ß1 or BLM, respectively, induced fibrotic responses. Fibrosis and EndoMT were significantly reduced in miR-155KO mice and changes in EndoMT markers in MLEC after TGF-ß1 stimulation confirmed the in vivo findings. Furthermore, lung fibrosis and EndoMT responses were reduced in VEC-miR-155 mice but significantly enhanced in VEC-SHIP-1 mice after BLM challenge. SHIP-1 knockdown in HUVEC cells resulted in enhanced EndoMT induced by BLM. Meanwhile, these changes involved the PI3K/AKT, JAK/STAT3, and SMAD/STAT signaling pathways. These studies demonstrate that endothelial miR-155 plays an important role in fibrotic responses in the lung through EndoMT. Endothelial SHIP-1 is essential in controlling fibrotic responses and SHIP-1 is a target of miR-155. Endothelial cells are an integral part in lung fibrosis.


Assuntos
Células Endoteliais da Veia Umbilical Humana/metabolismo , Sistema de Sinalização das MAP Quinases , MicroRNAs/metabolismo , Fosfatidilinositol-3,4,5-Trifosfato 5-Fosfatases/metabolismo , Fibrose Pulmonar/metabolismo , Animais , Modelos Animais de Doenças , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos , Camundongos , Camundongos Knockout , MicroRNAs/genética , Fosfatidilinositol-3,4,5-Trifosfato 5-Fosfatases/genética , Fibrose Pulmonar/genética , Fibrose Pulmonar/patologia , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo
9.
Mediators Inflamm ; 2019: 4851431, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31772503

RESUMO

The pathophysiology of the acute lung injury (ALI) is characterized by the damage of alveolar epithelial cells, which can be repaired by exogenous bone marrow-derived mesenchymal stem cells (BMSCs). However, the migration and differentiation abilities of BMSCs are not sufficient for the purpose, and a new approach that could strengthen the repair effects of BMSCs in ALI still needs to be clarified. We have previously proved that in vitro large tumor suppressor kinase 2- (Lats2-) underexpressing BMSCs may enhance their tissue repair effects in ALI; thus, in the present study, we tried to explore whether Lats2-underexpressing BMSCs could rescue lipopolysaccharide- (LPS-) induced ALI in vivo. BMSCs from C57BL/6 mice transfected with Lats2-interfering lentivirus vector or lentivirus blank controls were transplanted intratracheally into LPS-induced ALI mice. The retention and differentiation of BMSCs in the lung were evaluated by in vivo imaging, immunofluorescence staining, and Western blotting. The lung edema and permeability were assessed by lung wet weight/body weight ratio (LWW/BW) and measurements of proteins in bronchoalveolar lavage fluid (BALF) using ELISA. Acute lung inflammation was measured by the cytokines in the lung homogenate and BALF using RT-qPCR and ELISA, respectively. Lung injury was evaluated by HE staining and lung injury scoring. Pulmonary fibrosis was evaluated by Picrosirius red staining, immunohistochemistry for α-SMA and TGF-ß1, and hydroxyproline assay and RT-qPCR for Col1α1 and Col3α1. Lats2-mediated inhibition of the Hippo pathway increased the retention of BMSCs and their differentiation toward type II alveolar epithelial cells in the lung. Furthermore, Lats2-underexpressing BMSCs improved lung edema, permeability of the lung epithelium, and lung inflammation. Finally, Lats2-underexpressing BMSCs alleviated lung injury and early pulmonary fibrosis. Our studies suggest that underexpression of Lats2 could further enhance the repair effects of BMSCs against epithelial impair and the therapeutic potential of BMSCs in ALI mice.


Assuntos
Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/terapia , Células da Medula Óssea/citologia , Lipopolissacarídeos/toxicidade , Células-Tronco Mesenquimais/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Actinas/genética , Actinas/metabolismo , Animais , Western Blotting , Líquido da Lavagem Broncoalveolar/química , Células Cultivadas , Edema/metabolismo , Edema/terapia , Masculino , Células-Tronco Mesenquimais/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Microscopia de Fluorescência , Proteínas Serina-Treonina Quinases/genética , Fibrose Pulmonar/genética , Fibrose Pulmonar/metabolismo , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo , Proteínas Supressoras de Tumor/genética
10.
BMC Pulm Med ; 19(1): 178, 2019 Oct 16.
Artigo em Inglês | MEDLINE | ID: mdl-31619213

RESUMO

BACKGROUND: Hermansky-Pudlak syndrome (HPS) is an autosomal recessive disorder that is associated with oculocutaneous albinism, bleeding diathesis, granulomatous colitis, and highly penetrant pulmonary fibrosis in some subtypes. Homozygous or compound heterozygous pathological variants in HPS1, HPS3, HPS4, and several other genes lead to clinical manifestation of the disease. CASE PRESENTATION: A 57-year-old female was admitted with congenital oculocutaneous albinism, thrombocytopathy and late-onset accelerated pulmonary fibrosis (first symptoms from age 50 onwards). Chest high-resolution computed tomography identified thickening of peribronchovascular interstitium, bronchiectasis, reticulations, honeycombing, ground glass opacities and lung parenchyma consolidations. HPS was clinically suspected. We performed whole exome sequencing (WES), a form of massive parallel sequencing, of proband-parents trio. Whole exome libraries were processed using KAPA Hyper Prep Kit, SeqCap EZ MedExome Enrichment Kit and HyperCap Bead Kit according to the SeqCap EZ HyperCap Workflow. The paired-end 2 × 75 bp sequencing was performed on the Illumina NextSeq 500 Sequencer (Illumina Inc., USA). Furthermore, obtained variants by WES were evaluated using a virtual panel of genes: HPS1, AP3B1, HPS3, HPS4, HPS5, HPS6, DTNBP1, BLOC1S3, and PLDN. We identified a compound heterozygous genotype in HPS1 gene in the proband. We identified a pathogenic frameshift variant c.1189delC; p.(Gln397Serfs*2), resulting in a premature stop codon. This variant has been previously associated with HPS. Furthermore, we characterized previously undescribed nonsense variant c.1507C > T; p.(Gln503*), resulting in a premature stop codon and mRNA degradation through nonsense-mediated decay. Sanger sequencing validated the presence of both variants and simultaneously confirmed the heterozygous carrier status of parents. Unfortunately, the patient died due to fulminant progression of pulmonary fibrosis 2 months after diagnostics. CONCLUSIONS: Compound heterozygous mutations in HPS1 in the proband lead to disruption of HPS1 gene and clinical manifestation of HPS with severe pulmonary fibrosis. This case illustrates the need to consider HPS in differential diagnostics of pulmonary fibrosis. Pulmonary fibrosis is a common cause of death in HPS patients. Earlier diagnosis may enable better treatment for these patients.


Assuntos
Síndrome de Hermanski-Pudlak , Pulmão/diagnóstico por imagem , Proteínas de Membrana/genética , Fibrose Pulmonar , Progressão da Doença , Evolução Fatal , Feminino , Síndrome de Hermanski-Pudlak/diagnóstico , Síndrome de Hermanski-Pudlak/genética , Síndrome de Hermanski-Pudlak/fisiopatologia , Humanos , Pulmão/patologia , Pessoa de Meia-Idade , Mutação , Fibrose Pulmonar/diagnóstico , Fibrose Pulmonar/genética , Fibrose Pulmonar/fisiopatologia , Índice de Gravidade de Doença , Tomografia Computadorizada por Raios X/métodos , Sequenciamento Completo do Exoma/métodos
11.
BMC Vet Res ; 15(1): 379, 2019 Oct 29.
Artigo em Inglês | MEDLINE | ID: mdl-31664993

RESUMO

BACKGROUND: Canine idiopathic pulmonary fibrosis (CIPF) is a progressive interstitial lung disease mainly affecting old West Highland white terriers (WHWTs). The aetiology of CIPF is currently unknown and pathogenesis poorly understood. A genetic basis is strongly suspected based on the breed predisposition. CIPF shares clinical and pathological features with human IPF. In human IPF, coagulation disorders favouring a local and systemic pro-thrombotic state have been demonstrated in association with disease severity and outcome. The aim of this study was to compare the systemic haemostatic, fibrinolytic and inflammatory profiles of WHWTs affected with CIPF with breed-matched controls (CTRLs). Additionally, data collected in both groups were interpreted with regard to the reference intervals (when available) to assess possible pro-thrombotic features of the WHWT breed that may be related to CIPF predisposition. A total of 14 WHWTs affected with CIPF and 20 CTRLs were included. RESULTS: WHWTs affected with CIPF had prolonged activated partial thromboplastine time in comparison with CTRLs (12.2 ± 0.9 s vs. 11.5 ± 0.7 s, P = 0.028), whereas results obtained in both groups were all within reference ranges. There was no significant difference between groups for the other factors assessed including plasmatic concentrations of fibrinogen, D-dimers concentration, antithrombin III activity, protein S and protein C activities, anti-factor Xa activity, activated protein C ratio, serum C-reactive protein concentration, and rotational thromboelastometry indices. Platelet count and plasmatic fibrinogen concentration were found to be above the upper limit of the reference range in almost half of the WHWTs included, independently of the disease status. CONCLUSIONS: Results of this study provide no clear evidence of an altered systemic haemostatic, fibrinolytic or inflammatory state in WHWTs affected with CIPF compared with CTRLs. The higher platelet counts and fibrinogen concentrations found in the WHWT breed may serve as predisposing factors for CIPF or simply reflect biological variation in this breed.


Assuntos
Doenças do Cão/sangue , Fibrose Pulmonar/veterinária , Animais , Proteínas Sanguíneas , Estudos de Casos e Controles , Doenças do Cão/genética , Cães , Contagem de Eritrócitos , Feminino , Predisposição Genética para Doença , Hematócrito , Hemoglobinas , Hemostáticos/sangue , Contagem de Leucócitos , Masculino , Contagem de Plaquetas , Fibrose Pulmonar/sangue , Fibrose Pulmonar/genética , Tromboelastografia
12.
In Vivo ; 33(6): 1773-1784, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31662502

RESUMO

BACKGROUND/AIM: Ionizing radiation induces pulmonary fibrosis, which is a common dose-limiting complication in patients receiving radiotherapy. Fibrosis occurs through the accumulation of large amounts of ECM components, synthesized by myofibroblasts in damaged lung tissue. Epithelial cells serve as one of the cellular sources of myofibroblasts via the epithelial-to-mesenchymal transition (EMT) process. In this study, we investigated the role of TGF-ß-secreting M2 macrophages in association with ionizing radiation-induced EMT. MATERIALS AND METHODS: The lung epithelial cell line MLE12, was irradiated and the expression of EMT markers and chemokines was examined. Moreover, the mouse lung macrophage MH-S cell line was cultured with conditioned media from irradiated MLE12 cells, to examine the effects of the secreted factors on the migration ability of macrophages. For the murine pulmonary fibrosis model, mice were locally irradiated and the levels of M1 or M2 macrophage-related markers and cytokines were measured in bronchoalvelolar lavage (BAL) fluid and lung tissue. RESULTS: In MLE12 cells, irradiation directly induced expression of EMT-related markers and secretion of various chemokines, which lead to macrophage migration. Interestingly, the sub-population of macrophages recruited in the lung of mice after thoracic irradiation was M2 macrophages that expressed Arg-1 and CD206. M2 macrophages induced the MLE12 to undergo phenotypic conversion to form fibroblast-like cells, which leads to a down-regulation of epithelial markers and an up-regulation of new EMT-related markers. In thoracic irradiated mice, pro-inflammatory cytokines such as IL-1ß, IL-4 and IL-10 were increased at 2 weeks, but returned to normal levels from 16 weeks or 24 weeks after irradiation. However, thoracic irradiation led to a rapid increase of TGF-ß and IGF-1 levels, which lasted up to 24 weeks. It was confirmed that M2 macrophages secreted the high levels of TGF-ß. Moreover, the elimination of TGF-ß from M2 macrophages attenuated mesenchymal transition of MLE12. CONCLUSION: TGF-ß-secreting M2 macrophages play an important regulatory role in mesenchymal transition of epithelial cells in the lung of irradiated mice, thus contributing to radiation-induced pulmonary fibrosis.


Assuntos
Células Epiteliais/metabolismo , Transição Epitelial-Mesenquimal/efeitos da radiação , Pulmão/metabolismo , Macrófagos/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Animais , Linhagem Celular , Citocinas/metabolismo , Células Epiteliais/efeitos da radiação , Feminino , Fibroblastos/metabolismo , Fibroblastos/efeitos da radiação , Pulmão/efeitos da radiação , Macrófagos/efeitos da radiação , Camundongos , Camundongos Endogâmicos C57BL , Fibrose Pulmonar/genética , Fibrose Pulmonar/metabolismo , Pneumonite por Radiação/etiologia , Pneumonite por Radiação/metabolismo , Radiação Ionizante , Transdução de Sinais/efeitos da radiação
13.
Int J Biochem Cell Biol ; 116: 105595, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31473260

RESUMO

Fibroblasts are considered major contributors to the process of fibrogenesis and the progression of matrix deposition and tissue distortion in fibrotic diseases such as Pulmonary Fibrosis. Recent discovery of the fibrocyte, a circulating possible precursor cell to the tissue fibroblast in fibrosis, has raised issues regarding the characterization of fibrocytes with respect to their morphology, growth characteristics in vitro, their biological role in vivo and their potential utility as a biomarker and/ or treatment target in various human diseases. Characterization studies of the fibrocyte continue as does emerging conflicting data concerning the relationship to or with the lung fibroblast. The source of signals that direct the traffic of these cells, as well as their response to therapeutic intervention with newly available drugs, bring new insights to the understanding of this cell type. The identification of exosomes from fibrocytes that can affect resident fibroblast activities suggest mechanisms of their influence on pathogenesis. Moreover, interesting comparisons with other pathologies are emerging involving the influence of circulating mesenchymal precursor cells on tissue responses.


Assuntos
Exossomos/metabolismo , Matriz Extracelular/metabolismo , Fibroblastos/metabolismo , Células-Tronco Mesenquimais/metabolismo , Fibrose Pulmonar/genética , Animais , Biomarcadores/metabolismo , Diferenciação Celular , Citocinas/genética , Citocinas/metabolismo , Exossomos/química , Exossomos/patologia , Matriz Extracelular/química , Matriz Extracelular/patologia , Fibroblastos/patologia , Expressão Gênica , Humanos , Pulmão/metabolismo , Pulmão/patologia , Células-Tronco Mesenquimais/patologia , Camundongos , MicroRNAs/genética , MicroRNAs/metabolismo , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/patologia
14.
Redox Biol ; 26: 101307, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31473487

RESUMO

Idiopathic pulmonary fibrosis (IPF) is a progressive disease with an increased mortality. Metabolic reprogramming has a critical role in multiple chronic diseases. Lung macrophages expressing the mitochondrial calcium uniporter (MCU) have a critical role in fibrotic repair, but the contribution of MCU in macrophage metabolism is not known. Here, we show that MCU regulates peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) and metabolic reprogramming to fatty acid oxidation (FAO) in macrophages. MCU regulated PGC-1α expression by increasing the phosphorylation of ATF-2 by the p38 MAPK in a redox-dependent manner. The expression and activation of PGC-1α via the p38 MAPK was regulated by MCU-mediated mitochondrial calcium uptake, which is linked to increased mitochondrial ROS (mtROS) production. Mice harboring a conditional expression of dominant-negative MCU in macrophages had a marked reduction in mtROS and FAO and were protected from pulmonary fibrosis. Moreover, IPF lung macrophages had evidence of increased MCU and mitochondrial calcium, increased phosphorylation of ATF2 and p38, as well as increased expression of PGC-1α. These observations suggest that macrophage MCU-mediated metabolic reprogramming contributes to fibrotic repair after lung injury.


Assuntos
Canais de Cálcio/metabolismo , Metabolismo Energético , Regulação da Expressão Gênica , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/genética , Fibrose Pulmonar/genética , Fibrose Pulmonar/metabolismo , Adulto , Idoso , Animais , Cálcio/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Macrófagos Alveolares/metabolismo , Macrófagos Alveolares/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Mitocôndrias/genética , Mitocôndrias/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-NH2/metabolismo , Consumo de Oxigênio , Fenótipo , Fibrose Pulmonar/patologia , Espécies Reativas de Oxigênio/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
15.
Int J Biochem Cell Biol ; 116: 105598, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31499176

RESUMO

Chronic obstructive pulmonary disease (COPD) is characterised by an accelerated decline in airway function with age compared to age-matched non-smokers. There is increasing evidence that this is due to small airway disease rather than from emphysema, especially in the early stages of the disease. Small airways (<2 mm internal diameter) are narrowed in COPD with thickening and distortion of the airway wall and peribronchiolar fibrosis. In addition, loss of elasticity in alveolar attachments and mucus hypersecretion contribute to the airway narrowing and closure, leading to air trapping. The mechanisms of peribronchiolar fibrosis are poorly understood and small airway fibroblasts have not been characterised. In small airways of COPD patients the fibroblasts are profibrotic, pro-inflammatory and senescent. There is a reduction in the anti-ageing molecules sirtuin-1 and -6, which are regulated by specific microRNAs that are increased in COPD cells. It is plausible that extracellular vesicles from senescent airway epithelium transmit senescent signals to airway fibroblasts to stimulate fibrosis and inflammation. Small airways fibrosis is a target for new drug development that inhibit growth factor receptors, new antioxidants and particularly those that are targeted to mitochondria and inhibitors of cellular senescence or senolytic therapies.


Assuntos
Doença Pulmonar Obstrutiva Crônica/genética , Enfisema Pulmonar/genética , Fibrose Pulmonar/genética , Sistema Respiratório/metabolismo , Sirtuína 1/genética , Sirtuínas/genética , Anti-Inflamatórios/uso terapêutico , Antioxidantes/uso terapêutico , Senescência Celular , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Vesículas Extracelulares/efeitos dos fármacos , Vesículas Extracelulares/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Regulação da Expressão Gênica , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológico , Doença Pulmonar Obstrutiva Crônica/metabolismo , Doença Pulmonar Obstrutiva Crônica/patologia , Enfisema Pulmonar/tratamento farmacológico , Enfisema Pulmonar/metabolismo , Enfisema Pulmonar/patologia , Fibrose Pulmonar/tratamento farmacológico , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/patologia , Receptores de Fatores de Crescimento/genética , Receptores de Fatores de Crescimento/metabolismo , Sistema Respiratório/efeitos dos fármacos , Sistema Respiratório/patologia , Transdução de Sinais , Sirtuína 1/metabolismo , Sirtuínas/metabolismo
16.
J Agric Food Chem ; 67(35): 9789-9795, 2019 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-31373816

RESUMO

Pulmonary fibrosis is a chronic lung disease characterized by abnormal accumulation of the extracellular matrix (ECM). Chronic damage of the alveolar epithelium leads to a process called "epithelial-mesenchymal transition" (EMT) and increases synthesis and deposition of ECM proteins. Therefore, inhibition of EMT might be a promising therapeutic approach for the treatment of pulmonary fibrosis. ß-Sitosterol is one of the most abundant phytosterols in the plant kingdom and the major constituent in corn silk, which is derived from the stigma and style of maize (Zea mays). In this study, we elucidated that ß-sitosterol inhibited transforming growth factor-ß1 (TGF-ß1)-induced EMT and consequently had an antifibrotic effect. ß-Sitosterol (1-10 µg/mL) significantly downregulated the TGF-ß1-induced fibrotic proteins, such as collagen, fibronectin, and α-smooth muscle actin in human alveolar epithelial cells (p < 0.01). After 24 h, relative wound density (RWD) was increased in TGF-ß1 treated group (82.16 ± 5.70) compare to the control group (64.63 ± 2.21), but RWD was decreased in ß-sitosterol cotreated group (10 µg/mL: 71.54 ± 7.39; 20 µg/mL: 65.69 ± 6.42). In addition, the changes of the TGF-ß1-induced morphological shape and protein expression of EMT markers, N-cadherin, vimentin, and E-cadherin, were significantly blocked by ß-sitosterol treatment (p < 0.01). The effects of ß-sitosterol on EMT were found to be associated with the TGF-ß1/Snail pathway, which is regulated by Smad and non-Smad signaling pathways. Taken together, these findings suggest that ß-sitosterol can be used to attenuate pulmonary fibrosis through suppression of EMT by inhibiting the TGF-ß1/Snail pathway.


Assuntos
Células Epiteliais Alveolares/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Extratos Vegetais/farmacologia , Alvéolos Pulmonares/efeitos dos fármacos , Fibrose Pulmonar/fisiopatologia , Sitosteroides/farmacologia , Zea mays/química , Actinas/genética , Actinas/metabolismo , Células Epiteliais Alveolares/citologia , Células Epiteliais Alveolares/metabolismo , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular , Fibronectinas/genética , Fibronectinas/metabolismo , Humanos , Extratos Vegetais/química , Alvéolos Pulmonares/metabolismo , Alvéolos Pulmonares/fisiopatologia , Fibrose Pulmonar/tratamento farmacológico , Fibrose Pulmonar/genética , Fibrose Pulmonar/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo
17.
Ann Rheum Dis ; 78(10): 1379-1387, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31405848

RESUMO

OBJECTIVES: Myofibroblasts are key effector cells in the extracellular matrix remodelling of systemic sclerosis-associated interstitial lung disease (SSc-ILD); however, the diversity of fibroblast populations present in the healthy and SSc-ILD lung is unknown and has prevented the specific study of the myofibroblast transcriptome. We sought to identify and define the transcriptomes of myofibroblasts and other mesenchymal cell populations in human healthy and SSc-ILD lungs to understand how alterations in fibroblast phenotypes lead to SSc-ILD fibrosis. METHODS: We performed droplet-based, single-cell RNA-sequencing with integrated canonical correlation analysis of 13 explanted lung tissue specimens (56 196 cells) from four healthy control and four patients with SSc-ILD, with findings confirmed by cellular indexing of transcriptomes and epitopes by sequencing in additional samples. RESULTS: Examination of gene expression in mesenchymal cells identified two major, SPINT2hi and MFAP5hi, and one minor, WIF1hi, fibroblast populations in the healthy control lung. Combined analysis of control and SSc-ILD mesenchymal cells identified SPINT2hi, MFAP5hi, few WIF1hi fibroblasts and a new large myofibroblast population with evidence of actively proliferating myofibroblasts. We compared differential gene expression between all SSc-ILD and control mesenchymal cell populations, as well as among the fibroblast subpopulations, showing that myofibroblasts undergo the greatest phenotypic changes in SSc-ILD and strongly upregulate expression of collagens and other profibrotic genes. CONCLUSIONS: Our results demonstrate previously unrecognised fibroblast heterogeneity in SSc-ILD and healthy lungs, and define multimodal transcriptome-phenotypes associated with these populations. Our data indicate that myofibroblast differentiation and proliferation are key pathological mechanisms driving fibrosis in SSc-ILD.


Assuntos
Fibroblastos/metabolismo , Heterogeneidade Genética , Doenças Pulmonares Intersticiais/genética , Miofibroblastos/metabolismo , Escleroderma Sistêmico/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adulto , Estudos de Casos e Controles , Proliferação de Células/genética , Colágeno/genética , Proteínas Contráteis/metabolismo , Epitopos , Feminino , Perfilação da Expressão Gênica , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Pulmão/citologia , Masculino , Glicoproteínas de Membrana/metabolismo , Mesoderma/citologia , Pessoa de Meia-Idade , Fenótipo , Fibrose Pulmonar/genética , Escleroderma Sistêmico/complicações , Análise de Célula Única , Transcriptoma
18.
EMBO Mol Med ; 11(9): e10061, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31468711

RESUMO

Pulmonary hypertension secondary to pulmonary fibrosis (PF-PH) is one of the most common causes of PH, and there is no approved therapy. The molecular signature of PF-PH and underlying mechanism of why pulmonary hypertension (PH) develops in PF patients remains understudied and poorly understood. We observed significantly increased vascular wall thickness in both fibrotic and non-fibrotic areas of PF-PH patient lungs compared to PF patients. The increased vascular wall thickness in PF-PH patients is concomitant with a significantly increased expression of the transcription factor Slug within the macrophages and its target prolactin-induced protein (PIP), an extracellular matrix protein that induces pulmonary arterial smooth muscle cell proliferation. We developed a novel translational rat model of combined PF-PH that is reproducible and shares similar histological features (fibrosis, pulmonary vascular remodeling) and molecular features (Slug and PIP upregulation) with human PF-PH. We found Slug inhibition decreases PH severity in our animal model of PF-PH. Our study highlights the role of Slug/PIP axis in PF-PH.


Assuntos
Proteínas de Membrana Transportadoras/metabolismo , Fibrose Pulmonar/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Adulto , Idoso , Animais , Feminino , Humanos , Hipertensão Pulmonar/genética , Hipertensão Pulmonar/metabolismo , Hipertensão Pulmonar/patologia , Pulmão/metabolismo , Pulmão/patologia , Macrófagos/metabolismo , Masculino , Proteínas de Membrana Transportadoras/genética , Pessoa de Meia-Idade , Miócitos de Músculo Liso/metabolismo , Artéria Pulmonar/metabolismo , Artéria Pulmonar/patologia , Fibrose Pulmonar/genética , Fibrose Pulmonar/patologia , Ratos Wistar , Fatores de Transcrição da Família Snail/genética , Adulto Jovem
19.
Am J Physiol Lung Cell Mol Physiol ; 317(4): L434-L444, 2019 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-31364370

RESUMO

Pulmonary hypertension complicates the care of many patients with chronic lung diseases (defined as Group 3 pulmonary hypertension), yet the mechanisms that mediate the development of pulmonary vascular disease are not clearly defined. Despite being the most prevalent form of pulmonary hypertension, to date there is no approved treatment for patients with disease. Myeloid-derived suppressor cells (MDSCs) and endothelial cells in the lung express the chemokine receptor CXCR2, implicated in the evolution of both neoplastic and pulmonary vascular remodeling. However, precise cellular contribution to lung disease is unknown. Therefore, we used mice with tissue-specific deletion of CXCR2 to investigate the role of this receptor in Group 3 pulmonary hypertension. Deletion of CXCR2 in myeloid cells attenuated the recruitment of polymorphonuclear MDSCs to the lungs, inhibited vascular remodeling, and protected against pulmonary hypertension. Conversely, loss of CXCR2 in endothelial cells resulted in worsened vascular remodeling, associated with increased MDSC migratory capacity attributable to increased ligand availability, consistent with analyzed patient sample data. Taken together, these data suggest that CXCR2 regulates MDSC activation, informing potential therapeutic application of MDSC-targeted treatments.


Assuntos
Células Endoteliais/metabolismo , Hipertensão Pulmonar/metabolismo , Hipóxia/metabolismo , Células Supressoras Mieloides/metabolismo , Fibrose Pulmonar/metabolismo , Receptores de Interleucina-8B/genética , Transdução de Sinais , Animais , Bleomicina/administração & dosagem , Comunicação Celular , Movimento Celular , Células Endoteliais/patologia , Feminino , Expressão Gênica , Humanos , Hipertensão Pulmonar/induzido quimicamente , Hipertensão Pulmonar/genética , Hipertensão Pulmonar/patologia , Hipóxia/etiologia , Hipóxia/genética , Hipóxia/patologia , Pulmão/metabolismo , Pulmão/patologia , Masculino , Camundongos , Camundongos Knockout , Células Supressoras Mieloides/patologia , Cultura Primária de Células , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/genética , Fibrose Pulmonar/patologia , Receptores de Interleucina-8B/deficiência , Remodelação Vascular
20.
J Dermatol ; 46(11): 1014-1018, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31392773

RESUMO

Hereditary fibrosing poikiloderma with tendon contractures, myopathy and pulmonary fibrosis (POIKTMP) is a recently identified autosomal dominant genetic syndrome with mutations in FAM111B. Herein, we report a 14-month-old girl who presented with progressive poikiloderma on the face. Her 24-year-old mother had an identical facial poikiloderma, hyperpigmentation, mottling and Blaschko line hypopigmentation on the trunk and limbs, as well as severe tendon contractures. Next-generation sequencing based on a targeted gene capture panel revealed a missense mutation in the FAM111B gene p.Phe416Ser (c.1247T>C). Her mother had the same mutation as the proband. Moreover, this mutation was absent in the unaffected father and maternal grandparents. Based on the clinical manifestations and genetic analysis, the proband and her mother were diagnosed with POIKTMP. Protein modeling indicated that the mutation p.Phe416Ser dramatically changed the protein structure, especially its structural stability, and affected the protein function. This is the first report of POIKTMP in a Chinese family due to a novel FAM111B mutation. Furthermore, we have reviewed the genotype-phenotype correlation, differential diagnoses and management of POIKTMP.


Assuntos
Proteínas de Ciclo Celular/genética , Esclerose/genética , Anormalidades da Pele/genética , Dermatopatias Genéticas/genética , China , Contratura/genética , Feminino , Humanos , Lactente , Doenças Musculares/genética , Mutação de Sentido Incorreto , Fibrose Pulmonar/genética , Tendinopatia/genética , Adulto Jovem
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