RESUMO
In lung disease, persistence of KRT8-expressing aberrant basaloid cells in the alveolar epithelium is associated with impaired tissue regeneration and pathological tissue remodeling. We analyzed single cell RNA sequencing datasets of human interstitial lung disease and found the profibrotic Interleukin-11 (IL11) cytokine to be highly and specifically expressed in aberrant KRT8+ basaloid cells. IL11 is similarly expressed by KRT8+ alveolar epithelial cells lining fibrotic lesions in a mouse model of interstitial lung disease. Stimulation of alveolar epithelial cells with IL11 causes epithelial-to-mesenchymal transition and promotes a KRT8-high state, which stalls the beneficial differentiation of alveolar type 2 (AT2)-to-AT1 cells. Inhibition of IL11-signaling in AT2 cells in vivo prevents the accumulation of KRT8+ cells, enhances AT1 cell differentiation and blocks fibrogenesis, which is replicated by anti-IL11 therapy. These data show that IL11 inhibits reparative AT2-to-AT1 differentiation in the damaged lung to limit endogenous alveolar regeneration, resulting in fibrotic lung disease.
Assuntos
Células Epiteliais Alveolares , Diferenciação Celular , Interleucina-11 , Regeneração , Interleucina-11/metabolismo , Interleucina-11/genética , Animais , Regeneração/genética , Humanos , Camundongos , Células Epiteliais Alveolares/metabolismo , Células Epiteliais Alveolares/patologia , Alvéolos Pulmonares/patologia , Alvéolos Pulmonares/metabolismo , Transição Epitelial-Mesenquimal/genética , Modelos Animais de Doenças , Doenças Pulmonares Intersticiais/patologia , Doenças Pulmonares Intersticiais/genética , Doenças Pulmonares Intersticiais/metabolismo , Camundongos Endogâmicos C57BL , Masculino , Transdução de Sinais , FemininoRESUMO
Severe viral pneumonia can induce rapid expansion of KRT5+ basal-like cells in small airways and alveoli; this forms a scar-like structure that persists in the injured alveoli and impedes normal alveolar epithelium regeneration. In this study, we investigated the mechanism by which viral infection induced this remodeling response. Through comparing different lung-injury models, we demonstrated that infection induced strong IFN-γ signal-stimulated dysplastic KRT5+ cell formation. Inactivation of interferon receptor 1 (Ifngr1) reduced dysplastic cell formation, ameliorated lung fibrosis, and improved lung-function recovery. Mechanistically, IFN-γ regulated dysplastic cell formation via the focal adhesion kinase (FAK)/Yes-associated protein 1 (YAP) pathway. Inhibiting FAK/Src diminished IFN-γ-induced YAP nuclear translocation and dysplastic cell formation. Inhibiting YAP during viral infection prevented dysplastic cell formation, whereas inhibiting YAP in persistent KRT5+ cells led to their conversion into distal club cells. Importantly, human dysplastic cells exhibited elevated FAK and YAP activity, and IFN-γ treatment promoted the transformation of human alveolar progenitor cells into dysplastic cells. These findings uncover the role of infection-induced inflammatory response in alveolar remodeling and may provide potential therapeutic avenues for the treatment of alveolar remodeling in patients with severe viral pneumonia.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Quinase 1 de Adesão Focal , Interferon gama , Alvéolos Pulmonares , Proteínas de Sinalização YAP , Proteínas de Sinalização YAP/metabolismo , Animais , Camundongos , Humanos , Interferon gama/metabolismo , Quinase 1 de Adesão Focal/metabolismo , Quinase 1 de Adesão Focal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Alvéolos Pulmonares/patologia , Alvéolos Pulmonares/metabolismo , Alvéolos Pulmonares/virologia , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Transdução de Sinais , Camundongos Knockout , Inflamação/patologia , Inflamação/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genéticaRESUMO
Alveolar capillary dysplasia with misalignment of pulmonary veins (ACDMPV) is an interstitial lung disease. In ACDMPV, respiratory impairment with severe pulmonary hypertension occurs from the early hours of life. Anomalies in the cardiovascular, gastrointestinal and genitourinary systems have been reported. However, little is known about upper airway abnormalities. We encountered a genetically diagnosed ACDMPV infant who presented with subglottic and bronchial stenosis. The prenatal diagnosis was hypoplastic left heart syndrome. Her respiratory condition worsened at 16 hours of life. We found subglottic stenosis when intubating. She died on day 7. Autopsy imaging with CT scan showed bilateral main bronchial stenosis. Chromosomal microarray revealed a 531 kb deletion in chromosome 16q24.1, including FOXF1.
Assuntos
Laringoestenose , Síndrome da Persistência do Padrão de Circulação Fetal , Alvéolos Pulmonares , Humanos , Feminino , Recém-Nascido , Síndrome da Persistência do Padrão de Circulação Fetal/complicações , Síndrome da Persistência do Padrão de Circulação Fetal/genética , Laringoestenose/etiologia , Evolução Fatal , Alvéolos Pulmonares/anormalidades , Alvéolos Pulmonares/patologia , Constrição Patológica , Veias Pulmonares/anormalidades , Veias Pulmonares/diagnóstico por imagem , Fatores de Transcrição ForkheadRESUMO
Current pharmacotherapy remains futile in acute alveolar inflammation induced by Gram-negative bacteria (GNB), eliciting consequent respiratory failure. The release of lipid polysaccharides after antibiotic treatment and subsequent progress of proinflammatory cascade highlights the necessity to apply effective inflammation management simultaneously. This work describes modular self-assembling peptides for rapid anti-inflammatory programming (SPRAY) to form nanoparticles targeting macrophage specifically, having anti-inflammation and bactericidal functions synchronously. SPRAY nanoparticles accelerate the self-delivery process in macrophages via lysosomal membrane permeabilization, maintaining anti-inflammatory programming in macrophages with efficacy close to T helper 2 cytokines. By pulmonary deposition, SPRAY nanoparticles effectively suppress inflammatory infiltration and promote alveoli regeneration in murine aseptic acute lung injury. Moreover, SPRAY nanoparticles efficiently eradicate multidrug-resistant GNB in alveoli by disrupting bacterial membrane. The universal molecular design of SPRAY nanoparticles provides a robust and clinically unseen local strategy in reverse acute inflammation featured by a high accumulation of proinflammatory cellularity and drug-resistant bacteria.
Assuntos
Infecções por Bactérias Gram-Negativas , Nanopartículas , Animais , Camundongos , Nanopartículas/química , Infecções por Bactérias Gram-Negativas/tratamento farmacológico , Peptídeos/química , Peptídeos/farmacologia , Peptídeos/administração & dosagem , Bactérias Gram-Negativas/efeitos dos fármacos , Alvéolos Pulmonares/efeitos dos fármacos , Alvéolos Pulmonares/patologia , Alvéolos Pulmonares/metabolismo , Administração por Inalação , Antibacterianos/farmacologia , Antibacterianos/química , Antibacterianos/administração & dosagem , Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/administração & dosagem , Anti-Inflamatórios/química , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Modelos Animais de Doenças , Inflamação/tratamento farmacológico , Inflamação/patologiaRESUMO
BACKGROUND: The alveolar epithelium is protected by a heparan sulfate-rich, glycosaminoglycan layer called the epithelial glycocalyx. It is cleaved in patients with acute respiratory distress syndrome (ARDS) and in murine models of influenza A (IAV) infection, shedding fragments into the airspace from the cell surface. Glycocalyx shedding results in increased permeability of the alveolar-capillary barrier, amplifying acute lung injury. The mechanisms underlying alveolar epithelial glycocalyx shedding in IAV infection are unknown. We hypothesized that induction of host sheddases such as matrix metalloproteinases (MMPs) during IAV infection results in glycocalyx shedding and increased lung injury. MATERIALS AND METHODS: We measured glycocalyx shedding and lung injury during IAV infection with and without treatment with the pan-MMP inhibitor Ilomastat (ILO) and in an MMP-7 knock out (MMP-7KO) mouse. C57BL/6 or MMP-7KO male and female mice were given IAV A/PR/8/34 (H1N1) at 30,000 PFU/mouse or PBS intratracheally. For some experiments, C56BL/6 mice were infected in the presence of ILO (100mg/kg) or vehicle given daily by IP injection. Bronchoalveolar lavage (BAL) and lung tissue were collected on day 1, 3, and 7 for analysis of glycocalyx shedding (BAL Syndecan-1) and lung injury (histology, BAL protein, BAL cytokines, BAL immune cell infiltrates, BAL RAGE). Expression and localization of the sheddase MMP-7 and its inhibitor TIMP-1 was examined by RNAScope. For in vitro experiments, MLE-12 mouse lung epithelial cells were cultured and treated with active or heat-inactivated heparinase (2.5 U/mL) prior to infection with IAV (MOI 1) and viral load and MMP-7 and TIMP-1 expression analyzed. RESULTS: IAV infection caused shedding of the epithelial glycocalyx into the BAL. Inhibition of MMPs with ILO reduced glycocalyx shedding by 36% (p = 0.0051) and reduced lung epithelial injury by 40% (p = 0.0404). ILO also reduced viral load by 68% (p = 0.027), despite having no significant effect on lung cytokine production. Both MMP-7 and its inhibitor TIMP-1 were upregulated in IAV infected mice: MMP-7 colocalized with IAV, while TIMP-1 was limited to cells adjacent to infection. However, MMP-7KO mice had similar glycocalyx shedding, epithelial injury, and viral load compared to WT littermates, suggesting redundancy in MMP sheddase function in the lung. In vitro, heparinase treatment before infection led to a 52% increase in viral load (p = 0.0038) without altering MMP-7 or TIMP-1 protein levels. CONCLUSIONS: Glycocalyx shedding and MMPs play key roles in IAV-induced epithelial injury, with significant impact on IAV viral load. Further studies are needed to understand which specific MMPs regulate lung epithelial glycocalyx shedding.
Assuntos
Glicocálix , Metaloproteinase 7 da Matriz , Camundongos Endogâmicos C57BL , Infecções por Orthomyxoviridae , Animais , Glicocálix/metabolismo , Camundongos , Feminino , Masculino , Metaloproteinase 7 da Matriz/metabolismo , Metaloproteinase 7 da Matriz/genética , Infecções por Orthomyxoviridae/metabolismo , Infecções por Orthomyxoviridae/virologia , Infecções por Orthomyxoviridae/patologia , Vírus da Influenza A Subtipo H1N1/fisiologia , Camundongos Knockout , Alvéolos Pulmonares/virologia , Alvéolos Pulmonares/metabolismo , Alvéolos Pulmonares/patologia , Ácidos Hidroxâmicos/farmacologia , Inibidores de Metaloproteinases de Matriz/farmacologia , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Metaloproteinases da Matriz/metabolismo , IndóisAssuntos
Anestésicos Inalatórios , Hemorragia , Alvéolos Pulmonares , Sevoflurano , Humanos , Sevoflurano/efeitos adversos , Sevoflurano/administração & dosagem , Anestésicos Inalatórios/efeitos adversos , Anestésicos Inalatórios/administração & dosagem , Hemorragia/induzido quimicamente , Alvéolos Pulmonares/efeitos dos fármacos , Alvéolos Pulmonares/patologia , Masculino , Pneumopatias/induzido quimicamente , FemininoRESUMO
Using a 50-compartment Python-coded mathematical lung model, we compared mixed venous blood flow (Q) distributions and arterial oxygen tension/inspired oxygen fraction (PaO2/FiO2) relationships in lungs modeled with log normal distributions (LND) of inspired (VI) versus expired (VA) alveolar gas volumes. In lungs with normal V/Q heterogeneity, Q versus VA/Q and Q versus VI/Q distributions were similar with either approach, and PaO2/FiO2 sequences remained indistinguishable. In V/Q heterogeneous lungs at high FiO2, VILND generated low Q versus VA/Q shoulders and some negative VA units, while VALND preserved Q versus VA/Q log normality by blood flow diversion from low VI/Q units. We managed VILND-induced negative VA units either by shunt conversion (VI decreased to 0) or VI redistribution simulating collateral ventilation (VI increased till VA = 0). Comparing oxygen transfer: VALND > VILND (redistribution) > VILND (shunt). In V/Q heterogeneous lungs VALND and VILND (redistribution) regained near optimal oxygen transfer on 100% oxygen, while impairment persisted with VILND (shunt). Unlike VALND, VILND (redistribution) produced Q versus VA/Q distributions in V/Q heterogeneity compatible with multiple inert gas (MIGET) reports. VILND (redistribution) is a physiologically-based MIGET-compatible alternative to West's original VALND lung modeling approach.
Assuntos
Pulmão , Troca Gasosa Pulmonar , Humanos , Troca Gasosa Pulmonar/fisiologia , Pulmão/fisiologia , Pulmão/metabolismo , Pulmão/irrigação sanguínea , Modelos Biológicos , Oxigênio/metabolismo , Oxigênio/sangue , Alvéolos Pulmonares/fisiologia , Alvéolos Pulmonares/metabolismo , AnimaisRESUMO
BACKGROUND: Chronic lung disease of prematurity, called bronchopulmonary dysplasia (BPD), lacks effective therapies, stressing the need for preclinical testing systems that reflect human pathology for identifying causal pathways and testing novel compounds. Alveolar organoids derived from human pluripotent stem cells (hPSC) are promising test platforms for studying distal airway diseases like BPD, but current protocols do not accurately replicate the distal niche environment of the native lung. Herein, we investigated the contributions of cellular constituents of the alveolus and fetal respiratory movements on hPSC-derived alveolar organoid formation. METHODS: Human PSCs were differentiated in 2D culture into lung progenitor cells (LPC) which were then further differentiated into alveolar organoids before and after removal of co-developing mesodermal cells. LPCs were also differentiated in Transwell® co-cultures with and without human fetal lung fibroblast. Forming organoids were subjected to phasic mechanical strain using a Flexcell® system. Differentiation within organoids and Transwell® cultures was assessed by flow cytometry, immunofluorescence, and qPCR for lung epithelial and alveolar markers of differentiation including GATA binding protein 6 (GATA 6), E-cadherin (CDH1), NK2 Homeobox 1 (NKX2-1), HT2-280, surfactant proteins B (SFTPB) and C (SFTPC). RESULTS: We observed that co-developing mesenchymal progenitors promote alveolar epithelial type 2 cell (AEC2) differentiation within hPSC-derived lung organoids. This mesenchymal effect on AEC2 differentiation was corroborated by co-culturing hPSC-NKX2-1+ lung progenitors with human embryonic lung fibroblasts. The stimulatory effect did not require direct contact between fibroblasts and NKX2-1+ lung progenitors. Additionally, we demonstrate that episodic mechanical deformation of hPSC-derived lung organoids, mimicking in situ fetal respiratory movements, increased AEC2 differentiation without affecting proximal epithelial differentiation. CONCLUSION: Our data suggest that biophysical and mesenchymal components promote AEC2 differentiation within hPSC-derived distal organoids in vitro.
Assuntos
Diferenciação Celular , Pulmão , Organoides , Humanos , Organoides/citologia , Organoides/metabolismo , Pulmão/citologia , Pulmão/metabolismo , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Mesoderma/citologia , Mesoderma/metabolismo , Técnicas de Cocultura/métodos , Alvéolos Pulmonares/citologia , Alvéolos Pulmonares/metabolismoRESUMO
Micro-scale models of lung tissue have been employed by researchers to investigate alveolar mechanics; however, they have been limited by the lack of biofidelic material properties for the alveolar wall. To address this challenge, a finite element model of an alveolar cluster was developed comprising a tetrakaidecahedron array with the nominal characteristics of human alveolar structure. Lung expansion was simulated in the model by prescribing a pressure and monitoring the volume, to produce a pressure-volume (PV) response that could be compared to experimental PV data. The alveolar wall properties in the model were optimized to match experimental PV response of lungs filled with saline, to eliminate surface tension effects and isolate the alveolar wall tissue response. When simulated in uniaxial tension, the model was in agreement with reported experimental properties of uniaxial tension on excised lung tissue. The work presented herein was able to link micro-scale alveolar response to two disparate macroscopic experimental datasets (stress-stretch and PV response of lung) and presents hyperelastic properties of the alveolar wall for use in alveolar scale finite element models and multi-scale models. Future research will incorporate surface tension effects, and investigate alveolar injury mechanisms.
Assuntos
Elasticidade , Análise de Elementos Finitos , Pressão , Alvéolos Pulmonares , Estresse Mecânico , Alvéolos Pulmonares/fisiologia , Humanos , Modelos Biológicos , Fenômenos BiomecânicosRESUMO
The evolution of T cell molecular signatures in the distal lung of patients with severe pneumonia is understudied. Here, we analyzed T cell subsets in longitudinal bronchoalveolar lavage fluid samples from 273 patients with severe pneumonia, including unvaccinated patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) or with respiratory failure not linked to pneumonia. In patients with SARS-CoV-2 pneumonia, activation of interferon signaling pathways, low activation of the NF-κB pathway and preferential targeting of spike and nucleocapsid proteins early after intubation were associated with favorable outcomes, whereas loss of interferon signaling, activation of NF-κB-driven programs and specificity for the ORF1ab complex late in disease were associated with mortality. These results suggest that in patients with severe SARS-CoV-2 pneumonia, alveolar T cell interferon responses targeting structural SARS-CoV-2 proteins characterize individuals who recover, whereas responses against nonstructural proteins and activation of NF-κB are associated with poor outcomes.
Assuntos
COVID-19 , NF-kappa B , SARS-CoV-2 , Humanos , COVID-19/imunologia , SARS-CoV-2/imunologia , Masculino , Feminino , Pessoa de Meia-Idade , NF-kappa B/metabolismo , Idoso , Líquido da Lavagem Broncoalveolar/imunologia , Adulto , Transdução de Sinais/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Interferons/metabolismo , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Linfócitos T/imunologia , Alvéolos Pulmonares/imunologia , Alvéolos Pulmonares/patologiaRESUMO
A middle-aged man presented with inferior wall ST-elevation myocardial infarction and underwent primary percutaneous coronary intervention with tirofiban administered due to extensive thrombus. He developed sudden-onset dyspnoea, bilateral crepitations, haemoptysis, desaturation and hypotension an hour after starting tirofiban infusion. The tirofiban, antiplatelet medications and heparin were stopped immediately. Chest X-ray showed patchy opacities in the left upper, middle and lower zones. High-resolution CT showed confluent areas of consolidation with surrounding ground glass opacities and interlobular septal thickening (crazy pavement appearance) representing diffuse alveolar haemorrhage (DAH). He was managed with inotropes, non-invasive ventilation and intravenous furosemide. He was asymptomatic with complete resolution of lung opacities in chest X-ray done 2 months follow-up. DAH is a rare but potentially life-threatening complication which is often misidentified with other respiratory syndromes. Treatment includes stopping tirofiban and anticoagulant medication, blood transfusion, and institution of mechanical ventilation.
Assuntos
Hemorragia , Alvéolos Pulmonares , Tirofibana , Humanos , Masculino , Pessoa de Meia-Idade , Hemorragia/induzido quimicamente , Pneumopatias/induzido quimicamente , Intervenção Coronária Percutânea/efeitos adversos , Inibidores da Agregação Plaquetária/administração & dosagem , Inibidores da Agregação Plaquetária/efeitos adversos , Alvéolos Pulmonares/patologia , Infarto do Miocárdio com Supradesnível do Segmento ST/tratamento farmacológico , Infarto do Miocárdio com Supradesnível do Segmento ST/cirurgia , Tirofibana/administração & dosagem , Tirofibana/efeitos adversos , Tomografia Computadorizada por Raios XRESUMO
Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal fibrotic disease. Recent studies have highlighted the persistence of an intermediate state of alveolar stem cells in IPF lungs. In this study, we discovered a close correlation between the distribution pattern of intermediate alveolar stem cells and the progression of fibrotic changes. We showed that amphiregulin (AREG) expression is significantly elevated in intermediate alveolar stem cells of mouse fibrotic lungs and IPF patients. High levels of serum AREG correlate significantly with profound deteriorations in lung function in IPF patients. We demonstrated that AREG in alveolar stem cells is both required and sufficient for activating EGFR in fibroblasts, thereby driving lung fibrosis. Moreover, pharmacological inhibition of AREG using a neutralizing antibody effectively blocked the initiation and progression of lung fibrosis in mice. Our study underscores the therapeutic potential of anti-AREG antibodies in attenuating IPF progression, offering a promising strategy for treating fibrotic diseases.
Assuntos
Anfirregulina , Progressão da Doença , Fibrose Pulmonar Idiopática , Anfirregulina/metabolismo , Animais , Humanos , Fibrose Pulmonar Idiopática/patologia , Fibrose Pulmonar Idiopática/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Células-Tronco/metabolismo , Receptores ErbB/metabolismo , Fibroblastos/metabolismo , Fibroblastos/patologia , Células Epiteliais Alveolares/metabolismo , Células Epiteliais Alveolares/patologia , Masculino , Alvéolos Pulmonares/patologia , Alvéolos Pulmonares/metabolismo , Anticorpos Neutralizantes/farmacologia , FemininoRESUMO
Type 2 alveolar epithelial (AT2) cells of the lung are fundamental in regulating alveolar inflammation in response to injury. Impaired mitochondrial long-chain fatty acid ß-oxidation (mtLCFAO) in AT2 cells is assumed to aggravate alveolar inflammation in acute lung injury (ALI), yet the importance of mtLCFAO to AT2 cell function needs to be defined. Here we show that expression of carnitine palmitoyltransferase 1a (CPT1a), a mtLCFAO rate limiting enzyme, in AT2 cells is significantly decreased in acute respiratory distress syndrome (ARDS). In mice, Cpt1a deletion in AT2 cells impairs mtLCFAO without reducing ATP production and alters surfactant phospholipid abundance in the alveoli. Impairing mtLCFAO in AT2 cells via deleting either Cpt1a or Acadl (acyl-CoA dehydrogenase long chain) restricts alveolar inflammation in ALI by hindering the production of the neutrophilic chemokine CXCL2 from AT2 cells. This study thus highlights mtLCFAO as immunometabolism to injury in AT2 cells and suggests impaired mtLCFAO in AT2 cells as an anti-inflammatory response in ARDS.
Assuntos
Lesão Pulmonar Aguda , Células Epiteliais Alveolares , Carnitina O-Palmitoiltransferase , Ácidos Graxos , Mitocôndrias , Oxirredução , Síndrome do Desconforto Respiratório , Animais , Carnitina O-Palmitoiltransferase/metabolismo , Carnitina O-Palmitoiltransferase/genética , Mitocôndrias/metabolismo , Células Epiteliais Alveolares/metabolismo , Ácidos Graxos/metabolismo , Lesão Pulmonar Aguda/metabolismo , Lesão Pulmonar Aguda/patologia , Lesão Pulmonar Aguda/imunologia , Lesão Pulmonar Aguda/genética , Camundongos , Síndrome do Desconforto Respiratório/metabolismo , Síndrome do Desconforto Respiratório/imunologia , Síndrome do Desconforto Respiratório/patologia , Síndrome do Desconforto Respiratório/genética , Masculino , Humanos , Quimiocina CXCL2/metabolismo , Quimiocina CXCL2/genética , Camundongos Endogâmicos C57BL , Neutrófilos/imunologia , Neutrófilos/metabolismo , Camundongos Knockout , Acil-CoA Desidrogenase de Cadeia Longa/metabolismo , Acil-CoA Desidrogenase de Cadeia Longa/genética , Inflamação/metabolismo , Inflamação/patologia , Alvéolos Pulmonares/metabolismo , Alvéolos Pulmonares/patologia , Alvéolos Pulmonares/imunologia , Trifosfato de Adenosina/metabolismo , Pneumonia/metabolismo , Pneumonia/imunologia , Pneumonia/patologia , Pneumonia/genéticaRESUMO
BACKGROUND: During pseudoglandular stage of the human lung development the primitive bronchial buds are initially conformed by simple tubules lined by endoderm-derived epithelium surrounded by mesenchyme, which will progressively branch into airways and start to form distal epithelial saculles. For first time alveolar type II (AT2) pneumocytes appears. This study aims to characterize the genes and microRNAs involved in this differentiation process and decipher its role in the starting alveolar differentiation. METHODS: Gene and microRNA profiling was performed in human embryonic lungs from 7 to 12 post conception weeks (pcw). Protein expression location of candidate genes were analyzed by immunofluorescense in embryonic lung tissue sections. mRNA/miRNA target pairs were identified using computational approaches and their expression was studied in purified epithelial/mesenchymal cell populations and in isolated tips and stalks from the bronchial tree. Additionally, silencing experiments in human embryonic lung mesenchymal cells and in human embryonic tip-derived lung organoids were performed, as well as organoid differentiation studies. AT2 cell markers were studied by qRT-PCR and by immunofluorescence. The TGFB-ß phosphorylated pathways was analyzed with membrane protein arrays. Lung explants were cultured in air/liquid interface with/without peptides. RESULTS: We identified 88 differentially expressed genes, including IGFBP3. Although IGFBP3 mRNA was detected in both epithelial and mesenchymal populations, the protein was restricted to the epithelium, indicating post-transcriptional regulation preventing IGFBP3 protein expression in the mesenchyme. MicroRNA profiling identified miR-34a as an IGFBP3 regulator. miR-34a was up-regulated in mesenchymal cells, and its silencing in human embryonic lung mesenchymal cells increased IGFBP3 levels. Additionally, IGFBP3 expression showed a marked downregulation from 7 to 12 pcw, suggesting its involvement in the differentiation process. The differentiation of human tip-derived lung embryonic organoids showed a drastic reduction in IGFBP3, supported by the scRNAseq data. IGFBP3 silencing in organoids activated an alveolar-like differentiation process characterized by stem cell markers downregulation and upregulation of AT2 markers. This process was mediated by TGFß signalling inhibition and BMP pathway activation. CONCLUSIONS: The IGFBP3/miR-34a axis restricts IGFBP3 expression in the embryonic undifferentiated lung epithelium, and the progressive downregulation of IGFBP3 during the pseudoglandular stage is required for alveolar differentiation.
Assuntos
Diferenciação Celular , Proteína 3 de Ligação a Fator de Crescimento Semelhante à Insulina , Pulmão , MicroRNAs , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Proteína 3 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Proteína 3 de Ligação a Fator de Crescimento Semelhante à Insulina/genética , Pulmão/metabolismo , Pulmão/embriologia , Pulmão/citologia , Células Epiteliais Alveolares/metabolismo , Células Epiteliais Alveolares/citologia , Alvéolos Pulmonares/metabolismo , Alvéolos Pulmonares/citologia , Regulação da Expressão Gênica no Desenvolvimento , Células-Tronco Mesenquimais/metabolismo , Células-Tronco Mesenquimais/citologiaRESUMO
Diffuse alveolar hemorrhage (DAH) is a life-threatening pulmonary toxicity that can arise after hematopoietic cell transplantation (HCT). Risk factors and outcomes are not well understood owing to a sparsity of cases spread across multiple centers. The objectives of this epidemiologic study were to characterize the incidence, outcomes, transplantation-related risk factors and comorbid critical care diagnoses associated with post-HCT DAH. Retrospective analysis was performed in a multicenter cohort of 6995 patients age ≤21 years who underwent allogeneic HCT between 2008 and 2014 identified through the Center for International Blood and Marrow Transplant Research registry and cross-matched with the Virtual Pediatric Systems database to obtain critical care characteristics. A multivariable Cox proportional hazard model was used to determine risk factors for DAH. Logistic regression models were used to determine critical care diagnoses associated with DAH. Survival outcomes were analyzed using both a landmark approach and Cox regression, with DAH as a time-varying covariate. DAH occurred in 81 patients at a median of 54 days post-HCT (interquartile range, 23 to 160 days), with a 1-year post-transplantation cumulative incidence probability of 1.0% (95% confidence interval [CI], .81% to 1.3%) and was noted in 7.6% of all pediatric intensive care unit patients. Risk factors included receipt of transplantation for nonmalignant hematologic disease (reference: malignant hematologic disease; hazard ratio [HR], 1.98; 95% CI, 1.22 to 3.22; P = .006), use of a calcineurin inhibitor (CNI) plus mycophenolate mofetil (MMF) as graft-versus-host disease (GVHD) prophylaxis (referent: CNI plus methotrexate; HR, 1.89; 95% CI, 1.07 to 3.34; P = .029), and grade III-IV acute GVHD (HR, 2.67; 95% CI, 1.53-4.66; P < .001). Critical care admitted patients with DAH had significantly higher rates of systemic hypertension, pulmonary hypertension, pericardial disease, renal failure, and bacterial/viral/fungal infections (P < .05) than those without DAH. From the time of DAH, median survival was 2.2 months, and 1-year overall survival was 26% (95% CI, 17% to 36%). Among all HCT recipients, the development of DAH when considered was associated with a 7-fold increase in unadjusted all-cause post-HCT mortality (HR, 6.96; 95% CI, 5.42 to 8.94; P < .001). In a landmark analysis of patients alive at 2 months post-HCT, patients who developed DAH had a 1-year overall survival of 33% (95% CI, 18% to 49%), compared to 82% (95% CI, 81% to 83%) for patients without DAH (P < .001). Although DAH is rare, it is associated with high mortality in the post-HCT setting. Our data suggest that clinicians should have a heightened index of suspicion of DAH in patients with pulmonary symptoms in the context of nonmalignant hematologic indication for HCT, use of CNI + MMF as GVHD prophylaxis, and severe acute GVHD. Further investigations and validation of modifiable risk factors are warranted given poor outcomes.
Assuntos
Transplante de Células-Tronco Hematopoéticas , Hemorragia , Humanos , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Criança , Feminino , Masculino , Adolescente , Pré-Escolar , Hemorragia/epidemiologia , Hemorragia/etiologia , Hemorragia/mortalidade , Estudos Retrospectivos , Fatores de Risco , Lactente , Transplante Homólogo/efeitos adversos , Incidência , Alvéolos Pulmonares , Adulto Jovem , Pneumopatias/epidemiologia , Pneumopatias/etiologia , Doença Enxerto-Hospedeiro/epidemiologia , Doença Enxerto-Hospedeiro/etiologiaRESUMO
INTRODUCTION: Diffuse alveolar hemorrhage (DAH) is a rare complication with high mortality in patients with systemic lupus erythematosus (SLE). Early diagnosis and treatment are essential to improve patient prognosis. To determine the characteristics of patients with DAH and their mortality in a Spanish cohort of patients with SLE. METHODS: Patients from the RELESSER (Spanish Society of Rheumatology Lupus Register) who had had at least one confirmed episode of DAH were included. Epidemiological, clinical, and laboratory characteristics were analyzed. RESULTS: 4024 patients were included in the RELESSER register, 37 (0.9%), had at least one recorded episode of DAH. Only further data for 14 patients could be analyzed. In total, 92.9% were women, and for 4 (28.6%) DAH coincided with the debut of SLE. More than 80% of patients had renal involvement and thrombocytopenia. The most frequent manifestations were dyspnea (85.7%) and hypoxemia (100%), with the classic triad of hemoptysis, anemia and pulmonary infiltrates, appearing in 6 (46.2%) patients. The most frequently used treatments were glucocorticoids (85.7%) and cyclophosphamide (69.2%); plasmapheresis was utilized in 5 patients (35.7%) and 8, (57.1%) received intravenous immunoglobulins; 12 (85.7%) patients required admission to the ICU and 5 (35.7%) died. Tobacco use, history of lupus nephritis (LN), concomitant infection, and treatment with cyclophosphamide were more frequent in patients who died. CONCLUSIONS: DAH is rare in patients with SLE; in up to one-third of patients, it may appear at the onset of the disease. Some factors, such as smoking, a history of LN, treatment with cyclophosphamide, or concomitant infection, are more prevalent in patients with an unfavorable outcome.
Assuntos
Hemorragia , Pneumopatias , Lúpus Eritematoso Sistêmico , Sistema de Registros , Humanos , Lúpus Eritematoso Sistêmico/complicações , Lúpus Eritematoso Sistêmico/epidemiologia , Feminino , Adulto , Espanha/epidemiologia , Masculino , Hemorragia/epidemiologia , Hemorragia/etiologia , Pneumopatias/epidemiologia , Pneumopatias/etiologia , Pneumopatias/terapia , Pessoa de Meia-Idade , Alvéolos Pulmonares/patologia , Glucocorticoides/uso terapêutico , Ciclofosfamida/uso terapêutico , Adulto Jovem , Imunossupressores/uso terapêutico , PlasmafereseRESUMO
The lung is constantly exposed to airborne pathogens and particles that can cause alveolar damage. Hence, appropriate repair responses are essential for gas exchange and life. Here, we deciphered the spatiotemporal trajectory and function of an atypical population of macrophages after lung injury. Post-influenza A virus (IAV) infection, short-lived monocyte-derived Ly6G-expressing macrophages (Ly6G+ Macs) were recruited to the alveoli of lung perilesional areas. Ly6G+ Macs engulfed immune cells, exhibited a high metabolic potential, and clustered with alveolar type 2 epithelial cells (AT2s) in zones of active epithelial regeneration. Ly6G+ Macs were partially dependent on granulocyte-macrophage colony-stimulating factor and interleukin-4 receptor signaling and were essential for AT2-dependent alveolar regeneration. Similar macrophages were recruited in other models of injury and in the airspaces of lungs from patients with suspected pneumonia. This study identifies perilesional alveolar Ly6G+ Macs as a spatially restricted, short-lived macrophage subset promoting epithelial regeneration postinjury, thus representing an attractive therapeutic target for treating lung damage.