Regenerative Signatures in Bronchioalveolar Lavage of Acute Respiratory Distress Syndrome.

Background: In patients with severe acute respiratory distress syndrome (ARDS) associated with sepsis, lung recovery is considerably delayed, and mortality is much high. More insight into the process of lung regeneration in ARDS patients is needed. Exosomes are important cargos for intercellular communication by serving as autocrine and/or paracrine. Cutting-edge exomics (exosomal proteomics) makes it possible to study the mechanisms of re-alveolarization in ARDS lungs. Aims: This study aimed to identify potential regenerative niches by characterizing differentially expressed proteins in the exosomes of bronchioalveolar lavage (BAL) in ARDS patients. Methods: We purified exosomes from BAL samples collected from ARDS patients by NIH-supported ALTA and SPIROMICS trials. The abundance of exosomal proteins/peptides was quantified using liquid chromatography-mass spectrometry (LC-MS). Differentially expressed exosomal proteins between healthy controls and ARDS patients were profiled for functional annotations, cell origins, signaling pathways, networks, and clinical correlations. Results: Our results show that more exosomal proteins were identified in the lungs of late-stage ARDS patients. Immune cells and lung epithelial stem cells were major contributors to BAL exosomes in addition to those from other organs. We enriched a wide range of functions, stem cell signals, growth factors, and immune niches in both mild and severe patients. The differentially expressed proteins that we identified were associated with key clinical variables. The severity-associated differences in protein-protein interaction, RNA crosstalk, and epigenetic network were observed between mild and severe groups. Moreover, alveolar type 2 epithelial cells could serve as both exosome donors and recipients via autocrine and paracrine mechanisms. Conclusions: This study identifies novel exosomal proteins associated with diverse functions, signaling pathways, and cell origins in ARDS lavage samples. These differentiated proteins may serve as regenerative niches for re-alveolarization in injured lungs.

Alveolar Type 2 Epithelial Cell Organoids: Focus on Culture Methods.

Lung diseases rank third in terms of mortality and represent a significant economic burden globally. Scientists have been conducting research to better understand respiratory diseases and find treatments for them. An ideal in vitro model must mimic the in vivo organ structure, physiology, and pathology. Organoids are self-organizing, three-dimensional (3D) structures originating from adult stem cells, embryonic lung bud progenitors, embryonic stem cells (ESCs), and induced pluripotent stem cells (iPSCs). These 3D organoid cultures may provide a platform for exploring tissue development, the regulatory mechanisms related to the repair of lung epithelia, pathophysiological and immunomodulatory responses to different respiratory conditions, and screening compounds for new drugs. To create 3D lung organoids in vitro, both co-culture and feeder-free methods have been used. However, there exists substantial heterogeneity in the organoid culture methods, including the sources of AT2 cells, media composition, and feeder cell origins. This article highlights the currently available methods for growing AT2 organoids and prospective improvements to improve the available culture techniques/conditions. Further, we discuss various applications, particularly those aimed at modeling human distal lung diseases and cell therapy.

PAI-1 regulates AT2-mediated re-alveolarization and ion permeability.

BACKGROUND: Acute lung injury is characterized by overwhelmingly elevated PAI-1 in both lung edema fluid and the circulating system. The role of increased PAI-1, encoded by Serpine1 gene, in the regeneration of injured lung epithelium has not been understood completely. This study aimed to investigate the role of Serpine1 in the regulation of alveolar type 2 epithelial cell (AT2) fate in a humanized mouse line carrying diseased mutants (Serpine1Tg). METHODS: Wild-type (wt) and Serpine1Tg AT2 cells were either cultured as monolayers or 3D alveolospheres. Colony-forming assay and total surface area of organoids were analyzed. AT1 and AT2 cells in organoids were counted by immunohistochemistry and fluorescence-activated cell sorting (FACS). To test the potential effects of elevated PAI-1 on the permeability in the epithelial monolayers, we digitized the biophysical properties of polarized AT2 monolayers grown at the air-liquid interface. RESULTS: A significant reduction in total AT2 cells harvested in Serpine1Tg mice was observed compared with wt controls. AT2 cells harvested from Serpine1Tg mice reduced significantly over the wt controls. Spheroids formed by Serpine1Tg AT2 cells were lesser than wt control. Similarly, the corresponding surface area, a readout of re-alveolarization of injured epithelium, was markedly reduced in Serpine1Tg organoids. FACS analysis revealed a significant suppression in the number of AT2 cells, in particular, the CD44+ subpopulation, in Serpine1Tg organoids. A lesser ratio of AT1:AT2 cells in Serpine1Tg organoids was observed compared with wt cultures. There was a significant increase in transepithelial resistance but not amiloride inhibition. CONCLUSIONS: Our study suggests elevated PAI-1 in injured lungs downregulates alveolar epithelial regeneration by reducing the AT2 self-renewal, particularly in the CD44+ cells.

PAI-1 regulates AT2-mediated re-alveolarization and ion permeability.

Background Acute lung injury is characterized by overwhelmingly elevated PAI-1 in both lung edema fluid and the circulating system. The role of increased PAI-1, encoded by Serpine1 gene, in the regeneration of injured lung epithelium has not been understood completely. This study aimed to investigate the role of Serpine1 in the regulation of alveolar type 2 epithelial cell (AT2) fate in a humanized mouse line carrying diseased mutants (Serpine1Tg). Methods Wild type (wt) and Serpine1Tg AT2 cells were either cultured as monolayers or 3D alveolospheres. Colony forming assay and total surface area of organoids were analyzed. AT1 and AT2 cells in organoids were counted by immunohistochemistry and fluorescence-activated cell sorting (FACS). To test the potential effects of elevated PAI-1 on the permeability in the epithelial monolayers, we digitized the biophysical properties of polarized AT2 monolayers grown at the air-liquid interface. Results A significant reduction in total AT2 cells harvested in Serpine1Tg mice was observed compared with wt controls. AT2 cells harvested from Serpine1Tg mice reduced significantly over the wt controls. Spheroids formed by Serpine1Tg AT2 cells were lesser than wt control. Similarly, the corresponding surface area, a readout of realveolarization of injured epithelium, was markedly reduced in Serpine1Tg organoids. FACS analysis revealed a significant suppression in the number of AT2 cells, in particular, the CD44+ subpopulation, in Serpine1Tg organoids. A lesser ratio of AT1:AT2 cells in Serpine1Tg organoids was observed compared with wt cultures. There was a significant increase in transepithelial resistance but not amiloride inhibition. Conclusions Our study suggests elevated PAI-1 in injured lungs downregulates alveolar epithelial regeneration by reducing the AT2 self-renewal, particularly in the CD44+ cells.

Biomarkers and molecular endotypes of sarcoidosis: lessons from omics and non-omics studies.

Sarcoidosis is a chronic granulomatous disorder characterized by unknown etiology, undetermined mechanisms, and non-specific therapies except TNF blockade. To improve our understanding of the pathogenicity and to predict the outcomes of the disease, the identification of new biomarkers and molecular endotypes is sorely needed. In this study, we systematically evaluate the biomarkers identified through Omics and non-Omics approaches in sarcoidosis. Most of the currently documented biomarkers for sarcoidosis are mainly identified through conventional "one-for-all" non-Omics targeted studies. Although the application of machine learning algorithms to identify biomarkers and endotypes from unbiased comprehensive Omics studies is still in its infancy, a series of biomarkers, overwhelmingly for diagnosis to differentiate sarcoidosis from healthy controls have been reported. In view of the fact that current biomarker profiles in sarcoidosis are scarce, fragmented and mostly not validated, there is an urgent need to identify novel sarcoidosis biomarkers and molecular endotypes using more advanced Omics approaches to facilitate disease diagnosis and prognosis, resolve disease heterogeneity, and facilitate personalized medicine.

Wnt5a/ß-catenin axis is involved in the downregulation of AT2 lineage by PAI-1.

Failure to regenerate injured alveoli functionally and promptly causes a high incidence of fatality in coronavirus disease 2019 (COVID-19). How elevated plasminogen activator inhibitor-1 (PAI-1) regulates the lineage of alveolar type 2 (AT2) cells for re-alveolarization has not been studied. This study aimed to examine the role of PAI-1-Wnt5a-ß catenin cascades in AT2 fate. Dramatic reduction in AT2 yield was observed in Serpine1Tg mice. Elevated PAI-1 level suppressed organoid number, development efficiency, and total surface area in vitro. Anti-PAI-1 neutralizing antibody restored organoid number, proliferation and differentiation of AT2 cells, and ß-catenin level in organoids. Both Wnt family member 5A (Wnt5a) and Wnt5a-derived N-butyloxycarbonyl hexapeptide (Box5) altered the lineage of AT2 cells. This study demonstrates that elevated PAI-1 regulates AT2 proliferation and differentiation via the Wnt5a/ß catenin cascades. PAI-1 could serve as autocrine signaling for lung injury repair.

Prkg2 regulates alveolar type 2-mediated re-alveolarization.

BACKGROUND: The cGMP-dependent type 2 protein kinase, encoded by the prkg2 gene, is highly expressed in alveolar type 2 epithelial (AT2) cells. It is unclear whether prkg2 regulates AT2 cell homeostasis and re-alveolarization of injured lungs. This study aimed to investigate the role of prkg2 in the regulation of the fate of AT2 in vitro. METHODS: Primary AT2 cells of wild-type (wt) and prkg2-/- mice were co-cultured with fibroblasts as three-dimensional organoids. The colony formation was analyzed between days 4 and 12 post-seeding. EdU assay was used to detect cells with active DNA synthesis. AT1 and AT2 cells in organoids were visualized with anti-podoplanin and anti-surfactant protein C antibodies, respectively. RESULTS: Prkg2-/- AT2 cells developed a greater number of organoids than wt controls. However, compared to wt organoids, a lower number of AT2 but a greater number of AT1 cells were visualized. In addition, a lower number of proliferated cells (EdU+) were observed in prkg2-/- organoids compared to wt controls. The numbers of organoids and EdU+ cells were significantly reduced in protein kinase A (PKA) inhibitor H89-treated wt and prkg2-/- cultures. Organoids and EdU+ cells were increased by lipopolysaccharides (LPS) in both wt and prkg2-/- groups. The increase in the proportion of AT1 and AT2 cells in organoids was only seen in wt controls. CONCLUSIONS: Prkg2 may regulate the lineage of AT2 cells, which is affected by endotoxins and the interactive PKA signaling pathway.

Associations of D-Dimer on Admission and Clinical Features of COVID-19 Patients: A Systematic Review, Meta-Analysis, and Meta-Regression.

Background: Dynamic D-dimer level is a key biomarker for the severity and mortality of COVID-19 (coronavirus disease 2019). How aberrant fibrinolysis influences the clinical progression of COVID-19 presents a clinicopathological dilemma challenging intensivists. Methods: We performed meta-analysis and meta regression to analyze the associations of plasma D-dimer with 106 clinical variables to identify a panoramic view of the derangements of fibrinolysis in 14,862 patients of 42 studies. There were no limitations of age, gender, race, and country. Raw data of each group were extracted separately by two investigators. Individual data of case series, median and interquartile range, and ranges of median or mean were converted to SDM (standard deviation of mean). Findings: The weighted mean difference of D-dimer was 0.97 µg/mL (95% CI 0.65, 1.29) between mild and severe groups, as shown by meta-analysis. Publication bias was significant. Meta-regression identified 58 of 106 clinical variables were associated with plasma D-dimer levels. Of these, 11 readouts were negatively related to the level of plasma D-dimer. Further, age and gender were confounding factors. There were 22 variables independently correlated with the D-dimer level, including respiratory rate, dyspnea plasma K+, glucose, SpO2, BUN (blood urea nitrogen), bilirubin, ALT (alanine aminotransferase), AST (aspartate aminotransferase), systolic blood pressure, and CK (creatine kinase). Interpretation: These findings support elevated D-dimer as an independent predictor for both mortality and complications. The identified D-dimer-associated clinical variables draw a landscape integrating the aggregate effects of systemically suppressive and pulmonary hyperactive derangements of fibrinolysis, and the D-dimer-associated clinical biomarkers, and conceptually parameters could be combined for risk stratification, potentially for tracking thrombolytic therapy or alternative interventions.

Insufficient hyperfibrinolysis in COVID-19: a systematic review of thrombolysis based on meta-analysis and meta-regression.

Background How aberrant fibrinolysis influences the clinical progression of COVID-19 presents a clinicopathological dilemma challenging intensivists. To investigate whether abnormal fibrinolysis is a culprit or protector or both, we associated elevated plasma D-dimer with clinical variables to identify a panoramic view of the derangements of fibrinolysis that contribute to the pathogenesis of COVID-19 based on studies available in the literature. Methods We performed this systematic review based on both meta-analysis and meta-regression to compute the correlation of D-dimer at admission with clinical features of COVID-19 patients in retrospective studies or case series. We searched the databases until Aug 18, 2020, with no limitations by language. The first hits were screened, data extracted, and analyzed in duplicate. We did the random-effects meta-analyses and meta-regressions (both univariate and multivariate). D-dimer associated clinical variables and potential mechanisms were schematically reasoned and graphed. Findings Our search identified 42 observational, or retrospective, or case series from six countries (n=14,862 patients) with all races and ages from 1 to 98-year-old. The weighted mean difference of D-dimer was 0.97 µg/mL (95% CI 0.65, 1.29) between relatively mild (or healthy control) and severely affected groups with significant publication bias. Univariate meta-regression identified 58 of 106 clinical variables were associated with plasma D-dimer levels, including 3 demographics, 5 comorbidies, 22 laboratory tests, 18 organ injury biomarkers, 8 severe complications, and 2 outcomes (discharge and death). Of these, 11 readouts were negatively associated with the level of plasma D-dimer. Further, age and gender were confounding factors for the identified D-dimer associated variables. There were 22 variables independently correlated with the D-dimer level, including respiratory rate, dyspnea plasma K+, glucose, SpO2, BUN, bilirubin, ALT, AST, systolic blood pressure, and CK. We thus propose that "insufficient hyperfibrinolysis (fibrinolysis is accelerated but unable to prevent adverse clinical impact for clinical deterioration COVID-19)" as a peculiar mechanism. Interpretation The findings of this meta-analysis- and meta-regression-based systematic review supports elevated D-dimer as an independent predictor for mortality and severe complications. D-dimer-associated clinical variables draw a landscape integrating the aggregate effects of systemically suppressive and locally (i.e., in the lung) hyperactive derangements of fibrinolysis. D-dimer and associated clinical biomarkers and conceptually parameters could be combined for risk stratification, potentially for tracking thrombolytic therapy or alternative interventions.

Stem cell therapy for COVID-19 and other respiratory diseases: Global trends of clinical trials.

Respiratory diseases, including coronavirus disease 2019 and chronic obstructive pulmonary disease (COPD), are leading causes of global fatality. There are no effective and curative treatments, but supportive care only. Cell therapy is a promising therapeutic strategy for refractory and unmanageable pulmonary illnesses, as proved by accumulating preclinical studies. Stem cells consist of totipotent, pluripotent, multipotent, and unipotent cells with the potential to differentiate into cell types requested for repair. Mesenchymal stromal cells, endothelial progenitor cells, peripheral blood stem cells, and lung progenitor cells have been applied to clinical trials. To date, the safety and feasibility of stem cell and extracellular vesicles administration have been confirmed by numerous phase I/II trials in patients with COPD, acute respiratory distress syndrome, bronchial dysplasia, idiopathic pulmonary fibrosis, pulmonary artery hypertension, and silicosis. Five routes and a series of doses have been tested for tolerance and advantages of different regimes. In this review, we systematically summarize the global trends for the cell therapy of common airway and lung diseases registered for clinical trials. The future directions for both new clinical trials and preclinical studies are discussed.

Ion transport mechanisms for smoke inhalation-injured airway epithelial barrier.

Smoke inhalation injury is the leading cause of death in firefighters and victims. Inhaled hot air and toxic smoke are the predominant hazards to the respiratory epithelium. We aimed to analyze the effects of thermal stress and smoke aldehyde on the permeability of the airway epithelial barrier. Transepithelial resistance (RTE) and short-circuit current (ISC) of mouse tracheal epithelial monolayers were digitized by an Ussing chamber setup. Zonula occludens-1 tight junctions were visualized under confocal microscopy. A cell viability test and fluorescein isothiocyanate-dextran assay were performed. Thermal stress (40 °C) decreased RTE in a two-phase manner. Meanwhile, thermal stress increased ISC followed by its decline. Na+ depletion, amiloride (an inhibitor for epithelial Na+ channels [ENaCs]), ouabain (a blocker for Na+/K+-ATPase), and CFTRinh-172 (a blocker of cystic fibrosis transmembrane regulator [CFTR]) altered the responses of RTE and ISC to thermal stress. Steady-state 40 °C increased activity of ENaCs, Na+/K+-ATPase, and CFTR. Acrolein, one of the main oxidative unsaturated aldehydes in fire smoke, eliminated RTE and ISC. Na+ depletion, amiloride, ouabain, and CFTRinh-172 suppressed acrolein-sensitive ISC, but showed activating effects on acrolein-sensitive RTE. Thermal stress or acrolein disrupted zonula occludens-1 tight junctions, increased fluorescein isothiocyanate-dextran permeability but did not cause cell death or detachment. The synergistic effects of thermal stress and acrolein exacerbated the damage to monolayers. In conclusion, the paracellular pathway mediated by the tight junctions and the transcellular pathway mediated by active and passive ion transport pathways contribute to impairment of the airway epithelial barrier caused by thermal stress and acrolein. Graphical abstract Thermal stress and acrolein are two essential determinants for smoke inhalation injury, impairing airway epithelial barrier. Transcellular ion transport pathways via the ENaC, CFTR, and Na/K-ATPase are interrupted by both thermal stress and acrolein, one of the most potent smoke toxins. Heat and acrolein damage the integrity of the airway epithelium through suppressing and relocating the tight junctions.

Elevated Plasmin(ogen) as a Common Risk Factor for COVID-19 Susceptibility.

Patients with hypertension, diabetes, coronary heart disease, cerebrovascular illness, chronic obstructive pulmonary disease, and kidney dysfunction have worse clinical outcomes when infected with SARS-CoV-2, for unknown reasons. The purpose of this review is to summarize the evidence for the existence of elevated plasmin(ogen) in COVID-19 patients with these comorbid conditions. Plasmin, and other proteases, may cleave a newly inserted furin site in the S protein of SARS-CoV-2, extracellularly, which increases its infectivity and virulence. Hyperfibrinolysis associated with plasmin leads to elevated D-dimer in severe patients. The plasmin(ogen) system may prove a promising therapeutic target for combating COVID-19.

Plasmin improves blood-gas barrier function in oedematous lungs by cleaving epithelial sodium channels.

BACKGROUND AND PURPOSE: Lung oedema in association with suppressed fibrinolysis is a hallmark of lung injury. Here, we have tested whether plasmin cleaves epithelial sodium channels (ENaC) to resolve lung oedema fluid. EXPERIMENTAL APPROACH: Human lungs and airway acid-instilled mice were used for analysing fluid resolution. In silico prediction, mutagenesis, Xenopus oocytes, immunoblotting, voltage clamp, mass spectrometry, and protein docking were combined for identifying plasmin cleavage sites. KEY RESULTS: Plasmin improved lung fluid resolution in both human lungs ex vivo and injured mice. Plasmin activated αßγENaC channels in oocytes in a time-dependent manner. Deletion of four consensus proteolysis tracts (αΔ432-444, γΔ131-138, γΔ178-193, and γΔ410-422) eliminated plasmin-induced activation significantly. Further, immunoblotting assays identified 7 cleavage sites (K126, R135, K136, R153, K168, R178, K179) for plasmin to trim both furin-cleaved C-terminal fragments and full-length human γENaC proteins. In addition, 9 new sites (R122, R137, R138, K150, K170, R172, R180, K181, K189) in synthesized peptides were found to be cleaved by plasmin. These cleavage sites were located in the finger and the thumb, particularly the GRIP domain of human ENaC 3D model composed of two proteolytic centres for plasmin. Novel uncleaved sites beyond the GRIP domain in both α and γ subunits were identified to interrupt the plasmin cleavage-induced conformational change in ENaC channel complexes. Additionally, plasmin could regulate ENaC activity via the G protein signal. CONCLUSION AND IMPLICATIONS: Plasmin can cleave ENaC to improve blood-gas exchange by resolving oedema fluid and could be a potent therapy for oedematous lungs.

Proliferative regulation of alveolar epithelial type 2 progenitor cells by human Scnn1d gene.

Lung epithelial sodium channel (ENaC) encoded by Scnn1 genes is essential for maintaining transepithelial salt and fluid homeostasis in the airway and the lung. Compared to α, ß, and γ subunits, the role of respiratory δ-ENaC has not been studied in vivo due to the lack of animal models. Methods: We characterized full-length human δ802-ENaC expressed in both Xenopus oocytes and humanized transgenic mice. AT2 proliferation and differentiation in 3D organoids were analysed with FACS and a confocal microscope. Both two-electrode voltage clamp and Ussing chamber systems were applied to digitize δ802-ENaC channel activity. Immunoblotting was utilized to analyse δ802-ENaC protein. Transcripts of individual ENaC subunits in human lung tissues were quantitated with qPCR. Results: The results indicate that δ802-ENaC functions as an amiloride-inhibitable Na+ channel. Inhibitory peptide α-13 distinguishes δ802- from α-type ENaC channels. Modified proteolysis of γ-ENaC by plasmin and aprotinin did not alter the inhibition of amiloride and α-13 peptide. Expression of δ802-ENaC at the apical membrane of respiratory epithelium was detected with biophysical features similar to those of heterologously expressed channels in oocytes. δ802-ENaC regulated alveologenesis through facilitating the proliferation of alveolar type 2 epithelial cells. Conclusion: The humanized mouse line conditionally expressing human δ802-ENaC is a novel model for studying the expression and function of this protein in vivo .

Preclinical and clinical studies of smoke-inhalation-induced acute lung injury: update on both pathogenesis and innovative therapy.

Smoke-inhalation-induced acute lung injury (SI-ALI) is a leading cause of morbidity and mortality in victims of fire tragedies. SI-ALI contributes to an estimated 30% of burn-caused patient deaths, and recently, more attention has been paid to the specific interventions for this devastating respiratory illness. In the last decade, much progress has been made in the understanding of SI-ALI patho-mechanisms and in the development of new therapeutic strategies in both preclinical and clinical studies. This article reviews the recent progress in the treatment of SI-ALI, based on pathophysiology, thermal damage, airway obstruction, the nuclear-factor kappa-B signaling pathway, and oxidative stress. Preclinical therapeutic strategies include use of mesenchymal stem cells, hydrogen sulfide, peroxynitrite decomposition catalysts, and proton-pump inhibitors. Clinical interventions include high-frequency percussive ventilation, perfluorohexane, inhaled anticoagulants, and nebulized epinephrine. The animal model, dose, clinical application, and pharmacology of these medications are summarized. Future directions and further needs for developing innovative therapies are discussed.

Meta-Analysis of Preclinical Studies of Fibrinolytic Therapy for Acute Lung Injury.

Background: Acute lung injury (ALI) is characterized by suppressed fibrinolytic activity in bronchoalveolar lavage fluid (BALF) attributed to elevated plasminogen activator inhibitor-1 (PAI-1). Restoring pulmonary fibrinolysis by delivering tissue-type plasminogen activator (tPA), urokinase plasminogen activator (uPA), and plasmin could be a promising approach. Objectives: To systematically analyze the overall benefit of fibrinolytic therapy for ALI reported in preclinical studies. Methods: We searched PubMed, Embase, Web of Science, and CNKI Chinese databases, and analyzed data retrieved from 22 studies for the beneficial effects of fibrinolytics on animal models of ALI. Results: Both large and small animals were used with five routes for delivering tPA, uPA, and plasmin. Fibrinolytics significantly increased the fibrinolytic activity both in the plasma and BALF. Fibrin degradation products in BALF had a net increase of 408.41 ng/ml vs controls (P < 0.00001). In addition, plasma thrombin-antithrombin complexes increased 1.59 ng/ml over controls (P = 0.0001). In sharp contrast, PAI-1 level in BALF decreased 21.44 ng/ml compared with controls (P < 0.00001). Arterial oxygen tension was improved by a net increase of 15.16 mmHg, while carbon dioxide pressure was significantly reduced (11.66 mmHg, P = 0.0001 vs controls). Additionally, fibrinolytics improved lung function and alleviated inflammation response: the lung wet/dry ratio was decreased 1.49 (P < 0.0001 vs controls), lung injury score was reduced 1.83 (P < 0.00001 vs controls), and BALF neutrophils were lesser (3 × 104/ml, P < 0.00001 vs controls). The mortality decreased significantly within defined study periods (6 h to 30 days for mortality), as the risk ratio of death was 0.2-fold of controls (P = 0.0008). Conclusion: We conclude that fibrinolytic therapy may be effective pharmaceutic strategy for ALI in animal models.

Gene editing as a promising approach for respiratory diseases.

Respiratory diseases, which are leading causes of mortality and morbidity in the world, are dysfunctions of the nasopharynx, the trachea, the bronchus, the lung and the pleural cavity. Symptoms of chronic respiratory diseases, such as cough, sneezing and difficulty breathing, may seriously affect the productivity, sleep quality and physical and mental well-being of patients, and patients with acute respiratory diseases may have difficulty breathing, anoxia and even life-threatening respiratory failure. Respiratory diseases are generally heterogeneous, with multifaceted causes including smoking, ageing, air pollution, infection and gene mutations. Clinically, a single pulmonary disease can exhibit more than one phenotype or coexist with multiple organ disorders. To correct abnormal function or repair injured respiratory tissues, one of the most promising techniques is to correct mutated genes by gene editing, as some gene mutations have been clearly demonstrated to be associated with genetic or heterogeneous respiratory diseases. Zinc finger nucleases (ZFN), transcription activator-like effector nucleases (TALEN) and clustered regulatory interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) systems are three innovative gene editing technologies developed recently. In this short review, we have summarised the structure and operating principles of the ZFNs, TALENs and CRISPR/Cas9 systems and their preclinical and clinical applications in respiratory diseases.