African lungfish genome sheds light on the vertebrate water-to-land transition.

Lungfishes are the closest extant relatives of tetrapods and preserve ancestral traits linked with the water-to-land transition. However, their huge genome sizes have hindered understanding of this key transition in evolution. Here, we report a 40-Gb chromosome-level assembly of the African lungfish (Protopterus annectens) genome, which is the largest genome assembly ever reported and has a contig and chromosome N50 of 1.60 Mb and 2.81 Gb, respectively. The large size of the lungfish genome is due mainly to retrotransposons. Genes with ultra-long length show similar expression levels to other genes, indicating that lungfishes have evolved high transcription efficacy to keep gene expression balanced. Together with transcriptome and experimental data, we identified potential genes and regulatory elements related to such terrestrial adaptation traits as pulmonary surfactant, anxiolytic ability, pentadactyl limbs, and pharyngeal remodeling. Our results provide insights and key resources for understanding the evolutionary pathway leading from fishes to humans.

The anti-inflammatory and antiviral properties of anionic pulmonary surfactant phospholipids.

The pulmonary surfactant system of the lung is a lipid and protein complex, which regulates the biophysical properties of the alveoli to prevent lung collapse and the innate immune system in the lung. Pulmonary surfactant is a lipoprotein complex consisting of 90% phospholipids and 10% protein, by weight. Two minor components of pulmonary surfactant phospholipids, phosphatidylglycerol (PG) and phosphatidylinositol (PI), exist at very high concentrations in the extracellular alveolar compartments. We have reported that one of the most dominant molecular species of PG, palmitoyl-oleoyl-phosphatidylglycerol (POPG) and PI inhibit inflammatory responses induced by multiple toll-like receptors (TLR2/1, TLR3, TLR4, and TLR2/6) by interacting with subsets of multiprotein receptor components. These lipids also exert potent antiviral effects against RSV and influenza A, in vitro, by inhibiting virus binding to host cells. POPG and PI inhibit these viral infections in vivo, in multiple animal models. Especially noteworthy, these lipids markedly attenuate SARS-CoV-2 infection including its variants. These lipids are natural compounds that already exist in the lung and, thus, are less likely to cause adverse immune responses by hosts. Collectively, these data demonstrate that POPG and PI have strong potential as novel therapeutics for applications as anti-inflammatory compounds and preventatives, as treatments for broad ranges of RNA respiratory viruses.

Factors influencing airway smooth muscle tone: a comprehensive review with a special emphasis on pulmonary surfactant.

A thin film of pulmonary surfactant lines the surface of the airways and alveoli, where it lowers the surface tension in the peripheral lungs, preventing collapse of the bronchioles and alveoli and reducing the work of breathing. It also possesses a barrier function for maintaining the blood-gas interface of the lungs and plays an important role in innate immunity. The surfactant film covers the epithelium lining both large and small airways, forming the first line of defense between toxic airborne particles/pathogens and the lungs. Furthermore, surfactant has been shown to relax airway smooth muscle (ASM) after exposure to ASM agonists, suggesting a more subtle function. Whether surfactant masks irritant sensory receptors or interacts with one of them is not known. The relaxant effect of surfactant on ASM is absent in bronchial tissues denuded of an epithelial layer. Blocking of prostanoid synthesis inhibits the relaxant function of surfactant, indicating that prostanoids might be involved. Another possibility for surfactant to be active, namely through ATP-dependent potassium channels and the cAMP-regulated epithelial chloride channels [cystic fibrosis transmembrane conductance regulators (CFTRs)], was tested but could not be confirmed. Hence, this review discusses the mechanisms of known and potential relaxant effects of pulmonary surfactant on ASM. This review summarizes what is known about the role of surfactant in smooth muscle physiology and explores the scientific questions and studies needed to fully understand how surfactant helps maintain the delicate balance between relaxant and constrictor needs.

Alveolar macrophage lipid burden correlates with clinical improvement in patients with pulmonary alveolar proteinosis.

Pulmonary alveolar proteinosis (PAP) is a life-threatening, rare lung syndrome for which there is no cure and no approved therapies. PAP is a disease of lipid accumulation characterized by alveolar macrophage foam cell formation. While much is known about the clinical presentation, there is a paucity of information regarding temporal changes in lipids throughout the course of disease. Our objectives were to define the detailed lipid composition of alveolar macrophages in PAP patients at the time of diagnosis and during treatment. We performed comprehensive mass spectrometry to profile the lipid signature of alveolar macrophages obtained from three independent mouse models of PAP and from PAP and non-PAP patients. Additionally, we quantified changes in macrophage-associated lipids during clinical treatment of PAP patients. We found remarkable variations in lipid composition in PAP patients, which were consistent with data from three independent mouse models. Detailed lipidomic analysis revealed that the overall alveolar macrophage lipid burden inversely correlated with clinical improvement and response to therapy in PAP patients. Specifically, as PAP patients experienced clinical improvement, there was a notable decrease in the total lipid content of alveolar macrophages. This crucial observation suggests that the levels of these macrophage-associated lipids can be utilized to assess the efficacy of treatment. These findings provide valuable insights into the dysregulated lipid metabolism associated with PAP, offering the potential for lipid profiling to serve as a means of monitoring therapeutic interventions in PAP patients.

Comparative biophysical study of clinical surfactants using constrained drop surfactometry.

Surfactant replacement therapy is crucial in managing neonatal respiratory distress syndrome (RDS). Currently licensed clinical surfactants in the United States and Europe, including Survanta, Infasurf, Curosurf, and Alveofact, are all derived from bovine or porcine sources. We conducted a comprehensive examination of the biophysical properties of these four clinical surfactant preparations under physiologically relevant conditions, using constrained drop surfactometry (CDS). The assessed biophysical properties included the adsorption rate, quasi-static and dynamic surface activity, resistance to surfactant inhibition by meconium, and the morphology of the adsorbed surfactant films. This comparative study unveiled distinct in vitro biophysical properties of these clinical surfactants and revealed correlations between their chemical composition, lateral film structure, and biophysical functionality. Notably, at 1 mg/mL, Survanta exhibited a significantly lower adsorption rate compared with the other preparations at the same surfactant concentration. At 10 mg/mL, Infasurf, Curosurf, and Survanta all demonstrated excellent dynamic surface activity, whereas Alveofact exhibited the poorest quasi-static and dynamic surface activity. The suboptimal surface activity of Alveofact is found to be correlated with its unique monolayer-predominant morphology, in contrast to other surfactants forming multilayers. Curosurf, in particular, showcased superior resistance to biophysical inhibition by meconium compared with other preparations. Understanding the diverse biophysical behaviors of clinical surfactants provides crucial insights for precision and personalized design in treating RDS and other respiratory conditions. The findings from this study contribute valuable perspectives for the development of more efficacious and fully synthetic surfactant preparations.NEW & NOTEWORTHY A thorough investigation into the biophysical properties of four animal-derived clinical surfactant preparations was conducted through constrained drop surfactometry under physiologically relevant conditions. This comparative study unveiled unique in vitro biophysical characteristics among these clinical surfactants, establishing correlations between their chemical composition, lateral film structure, and biophysical functionality. The acquired knowledge offers essential insights for the precise and personalized design of clinical surfactant for the treatment of respiratory distress syndrome and other respiratory conditions.

Activation of alveolar epithelial ER stress by ß-coronavirus infection disrupts surfactant homeostasis in mice: implications for COVID-19 respiratory failure.

COVID-19 syndrome is characterized by acute lung injury, hypoxemic respiratory failure, and high mortality. Alveolar type 2 (AT2) cells are essential for gas exchange, repair, and regeneration of distal lung epithelium. We have shown that the causative agent, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and other members of the ß-coronavirus genus induce an endoplasmic reticulum (ER) stress response in vitro; however, the consequences for host AT2 cell function in vivo are less understood. To study this, two murine models of coronavirus infection were used-mouse hepatitis virus-1 (MHV-1) in A/J mice and a mouse-adapted SARS-CoV-2 strain. MHV-1-infected mice exhibited dose-dependent weight loss with histological evidence of distal lung injury accompanied by elevated bronchoalveolar lavage fluid (BALF) cell counts and total protein. AT2 cells showed evidence of both viral infection and increased BIP/GRP78 expression, consistent with activation of the unfolded protein response (UPR). The AT2 UPR included increased inositol-requiring enzyme 1α (IRE1α) signaling and a biphasic response in PKR-like ER kinase (PERK) signaling accompanied by marked reductions in AT2 and BALF surfactant protein (SP-B and SP-C) content, increases in surfactant surface tension, and emergence of a reprogrammed epithelial cell population (Krt8+ and Cldn4+). The loss of a homeostatic AT2 cell state was attenuated by treatment with the IRE1α inhibitor OPK-711. As a proof-of-concept, C57BL6 mice infected with mouse-adapted SARS-CoV-2 demonstrated similar lung injury and evidence of disrupted surfactant homeostasis. We conclude that lung injury from ß-coronavirus infection results from an aberrant host response, activating multiple AT2 UPR stress pathways, altering surfactant metabolism/function, and changing AT2 cell state, offering a mechanistic link between SARS-CoV-2 infection, AT2 cell biology, and acute respiratory failure.NEW & NOTEWORTHY COVID-19 syndrome is characterized by hypoxemic respiratory failure and high mortality. In this report, we use two murine models to show that ß-coronavirus infection produces acute lung injury, which results from an aberrant host response, activating multiple epithelial endoplasmic reticular stress pathways, disrupting pulmonary surfactant metabolism and function, and forcing emergence of an aberrant epithelial transition state. Our results offer a mechanistic link between SARS-CoV-2 infection, AT2 cell biology, and respiratory failure.

First vertebrate BRICHOS antimicrobial peptides: ß-hairpin host defense peptides in limbless amphibia lung resemble those of marine worms.

Innate immunity of invertebrates offers potent antimicrobial peptides (AMPs) against drug-resistant infections. To identify new worm ß-hairpin AMPs, we explored the sequence diversity of proteins with a BRICHOS domain, which comprises worm AMP precursors. Strikingly, we discovered new BRICHOS AMPs not in worms, but in caecilians, the least studied clade of vertebrates. Two precursor proteins from Microcaecilia unicolor and Rhinatrema bivittatum resemble SP-C lung surfactants and bear worm AMP-like peptides at C-termini. The analysis of M. unicolor tissue transcriptomes shows that the AMP precursor is highly expressed in the lung along with regular SP-C, suggesting a different, protective function. The peptides form right-twisted ß-hairpins, change conformation upon lipid binding, and rapidly disrupt bacterial membranes. Both peptides exhibit broad-spectrum activity against multidrug-resistant ESKAPE pathogens with 1-4 µM MICs and remarkably low toxicity, giving 40-70-fold selectivity towards bacteria. These BRICHOS AMPs, previously unseen in vertebrates, reveal a novel lung innate immunity mechanism and offer a promising antibiotics template.

Evaluating Novel SP-B and SP-C Synthetic Analogues for Pulmonary Surfactant Efficacy.

Pulmonary surfactants, a complex assembly of phospholipids and surfactant proteins such as SP-B and SP-C, are critical for maintaining respiratory system functionality by lowering surface tension (ST) and preventing alveolar collapse. Our study introduced five synthetic SP-B peptides and one SP-C peptide, leading to the synthesis of CHAsurf candidates (CHAsurf-1 to CHAsurf-5) for evaluation. We utilized a modified Wilhelmy balance test to assess the surface tension properties of the surfactants, measuring spreading rate, surface adsorption, and ST-area diagrams to comprehensively evaluate their performance. Animal experiments were performed on New Zealand white rabbits to test the efficacy of CHAsurf-4B, a variant chosen for its economic viability and promising ST reduction properties, comparable to Curosurf®. The study confirmed that higher doses of SP-B in CHAsurf-4 are associated with improved ST reduction. However, due to cost constraints, CHAsurf-4B was selected for in vivo assessment. The animal model revealed that CHAsurf-4B could restore alveolar structure and improve lung elasticity, akin to Curosurf®. Our research highlights the significance of cysteine residues and disulfide bonds in the structural integrity and function of synthetic SP-B analogues, offering a foundation for future surfactant therapy in respiratory disorders. This study's findings support the potential of CHAsurf-4B as a therapeutic agent, meriting further investigation to solidify its role in clinical applications.

The Synthetic Surfactant CHF5633 Restores Lung Function and Lung Architecture in Severe Acute Respiratory Distress Syndrome in Adult Rabbits.

PURPOSE: Acute respiratory distress syndrome (ARDS) is a major cause of hypoxemic respiratory failure in adults. In ARDS extensive inflammation and leakage of fluid into the alveoli lead to dysregulation of pulmonary surfactant metabolism and function. Altered surfactant synthesis, secretion, and breakdown contribute to the clinical features of decreased lung compliance and alveolar collapse. Lung function in ARDS could potentially be restored with surfactant replacement therapy, and synthetic surfactants with modified peptide analogues may better withstand inactivation in ARDS alveoli than natural surfactants. METHODS: This study aimed to investigate the activity in vitro and the bolus effect (200 mg phospholipids/kg) of synthetic surfactant CHF5633 with analogues of SP-B and SP-C, or natural surfactant Poractant alfa (Curosurf®, both preparations Chiesi Farmaceutici S.p.A.) in a severe ARDS model (the ratio of partial pressure arterial oxygen and fraction of inspired oxygen, P/F ratio ≤ 13.3 kPa) induced by hydrochloric acid instillation followed by injurious ventilation in adult New Zealand rabbits. The animals were ventilated for 4 h after surfactant treatment and the respiratory parameters, histological appearance of lung parenchyma and levels of inflammation, oxidative stress, surfactant dysfunction, and endothelial damage were evaluated. RESULTS: Both surfactant preparations yielded comparable improvements in lung function parameters, reductions in lung injury score, pro-inflammatory cytokines levels, and lung edema formation compared to untreated controls. CONCLUSIONS: This study indicates that surfactant replacement therapy with CHF5633 improves lung function and lung architecture, and attenuates inflammation in severe ARDS in adult rabbits similarly to Poractant alfa. Clinical trials have so far not yielded conclusive results, but exogenous surfactant may be a valid supportive treatment for patients with ARDS given its anti-inflammatory and lung-protective effects.

Pulmonary Surfactant Proteins Are Inhibited by Immunoglobulin A Autoantibodies in Severe COVID-19.

Rationale: Coronavirus disease 2019 (COVID-19) can lead to acute respiratory distress syndrome with fatal outcomes. Evidence suggests that dysregulated immune responses, including autoimmunity, are key pathogenic factors. Objectives: To assess whether IgA autoantibodies target lung-specific proteins and contribute to disease severity. Methods: We collected 147 blood, 9 lung tissue, and 36 BAL fluid samples from three tertiary hospitals in Switzerland and one in Germany. Severe COVID-19 was defined by the need to administer oxygen. We investigated the presence of IgA autoantibodies and their effects on pulmonary surfactant in COVID-19 using the following methods: immunofluorescence on tissue samples, immunoprecipitations followed by mass spectrometry on BAL fluid samples, enzyme-linked immunosorbent assays on blood samples, and surface tension measurements with medical surfactant. Measurements and Main Results: IgA autoantibodies targeting pulmonary surfactant proteins B and C were elevated in patients with severe COVID-19 but not in patients with influenza or bacterial pneumonia. Notably, pulmonary surfactant failed to reduce surface tension after incubation with either plasma or purified IgA from patients with severe COVID-19. Conclusions: Our data suggest that patients with severe COVID-19 harbor IgA autoantibodies against pulmonary surfactant proteins B and C and that these autoantibodies block the function of lung surfactant, potentially contributing to alveolar collapse and poor oxygenation.

Investigating the effects of dexamethasone on pulmonary surfactant lipids based on lipidomics studies.

Dexamethasone, a glucocorticoid commonly used in pediatric patients, has potent anti-inflammatory and immunosuppressive properties. However, it is associated with side effects such as reduced lung function and decreased immunity. Pulmonary surfactant lipids are closely linked to lung disease and play a role in reducing surface tension, immune response and antiviral activity. The dysregulation of lipid metabolism is closely associated with lung disease. Hence, untargeted lipidomics may be instrumental in elucidating the effects of dexamethasone on pulmonary surfactant lipids. We obtained surfactant lipid samples from the bronchoalveolar lavage fluid of young mice injected subcutaneously with dexamethasone and conducted a comprehensive lipidomic analysis, comparing them with a control group. We observed a decrease in lipids, such as phosphatidylcholine, phosphatidylglycerol and phosphatidylethanolamine, and an increase in ceramide, fatty acid, diacylglycerol and monoglyceride, which may impact lung health. This study revealed the influence of dexamethasone on pulmonary surfactant lipids, offering new insights into adverse reactions in clinical settings.

Validation of a high-performance liquid chromatography method for determining lysophosphatidylcholine content in bovine pulmonary surfactant medication.

Pulmonary surfactant replacement therapy is a promising improvement in neonatal care for infants with respiratory distress syndrome. Lysophosphatidylcholine (LPC) is an undesirable component that can hinder surfactant proteins from enhancing the adsorption of surfactant lipids to balance surface tensions by creating a saturated coating on the interior of the lungs. A novel normal-phase liquid chromatography method utilizing UV detection and non-toxic solvents was developed and validated for the first time to analyze LPC in the complex matrix of pulmonary surfactant medication. The analytical method validation included evaluation of system suitability, repeatability, intermediate precision, linearity, accuracy, limit of detection (LOD), limit of quantification (LOQ), stability and robustness. The method yielded detection and quantification limits of 4.4 and 14.5 µg/ml, respectively. The calibration curve was modified linearly within the LOQ to 1.44 mg/ml range, with a determination coefficient of 0.9999 for standards and 0.9997 for sample solutions. Given the lack of reliable published data on LPC analysis in pulmonary surfactant medications, this newly developed method demonstrates promising results and offers advantages of HPLC methodology, including simplicity, accuracy, specificity, sensitivity and an exceptionally low LOD and LOQ. These attributes contribute to considering this achievement as an innovative method.

Liraglutide alleviates sepsis-induced acute lung injury by regulating pulmonary surfactant through inhibiting autophagy.

BACKGROUND: Pulmonary surfactant (PS) plays an important role in the treatment of sepsis-induced acute lung injury (ALI). Liraglutide, a glucagon-like peptide-1 (GLP-1) analog, improves the secretion and function of PS in ALI, but the underlying mechanism remains unknown. This study aimed to investigate how liraglutide regulates PS secretion in ALI. METHODS: C57BL/6 mice were injected subcutaneously with normal saline containing different concentrations of liraglutide after the establishment of the ALI model. MLE-12 cells were treated with liraglutide after LPS stimulation. The survival rate of mice, wet/dry weight ratio, inflammatory factors in bronchoalveolar lavage fluid (BALF), pulmonary injury, and apoptosis were analyzed. Cell viability, proliferation, apoptosis, the expression of SP-A, SP-B, and expression of autophagy-related proteins in cells were measured. RESULTS: ALI mice showed reduced pulmonary injury, less apoptosis, and less inflammation compared to the controls. Liraglutide prolonged survival, decreased the wet/dry weight ratio, reduced inflammatory responses, and attenuated pulmonary edema compared with the ALI group. Moreover, LPS-induced cell damage and reduction of SP-A and SP-B expression were markedly reversed by liraglutide in MLE-12 cells. Furthermore, the protective effects of liraglutide were reversed by rapamycin. CONCLUSION: Liraglutide alleviate sepsis-induced ALI by inhibiting autophagy and regulating PS.

Stratification of asthma by lipidomic profiling of induced sputum supernatant.

BACKGROUND: Asthma is a chronic respiratory disease with significant heterogeneity in its clinical presentation and pathobiology. There is need for improved understanding of respiratory lipid metabolism in asthma patients and its relation to observable clinical features. OBJECTIVE: We performed a comprehensive, prospective, cross-sectional analysis of the lipid composition of induced sputum supernatant obtained from asthma patients with a range of disease severities, as well as from healthy controls. METHODS: Induced sputum supernatant was collected from 211 adults with asthma and 41 healthy individuals enrolled onto the U-BIOPRED (Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes) study. Sputum lipidomes were characterized by semiquantitative shotgun mass spectrometry and clustered using topologic data analysis to identify lipid phenotypes. RESULTS: Shotgun lipidomics of induced sputum supernatant revealed a spectrum of 9 molecular phenotypes, highlighting not just significant differences between the sputum lipidomes of asthma patients and healthy controls, but also within the asthma patient population. Matching clinical, pathobiologic, proteomic, and transcriptomic data helped inform the underlying disease processes. Sputum lipid phenotypes with higher levels of nonendogenous, cell-derived lipids were associated with significantly worse asthma severity, worse lung function, and elevated granulocyte counts. CONCLUSION: We propose a novel mechanism of increased lipid loading in the epithelial lining fluid of asthma patients resulting from the secretion of extracellular vesicles by granulocytic inflammatory cells, which could reduce the ability of pulmonary surfactant to lower surface tension in asthmatic small airways, as well as compromise its role as an immune regulator.

Descriptive and Functional Genomics in Neonatal Respiratory Distress Syndrome: From Lung Development to Targeted Therapies.

In this up-to-date study, we first aimed to highlight the genetic and non-genetic factors associated with respiratory distress syndrome (RDS) while also focusing on the genomic aspect of this condition. Secondly, we discuss the treatment options and the progressing therapies based on RNAs or gene therapy. To fulfill this, our study commences with lung organogenesis, a highly orchestrated procedure guided by an intricate network of conserved signaling pathways that ultimately oversee the processes of patterning, growth, and differentiation. Then, our review focuses on the molecular mechanisms contributing to both normal and abnormal lung growth and development and underscores the connections between genetic and non-genetic factors linked to neonatal RDS, with a particular emphasis on the genomic aspects of this condition and their implications for treatment choices and the advancing therapeutic approaches centered around RNAs or gene therapy.

Amyloid-Forming Corpora Amylacea and Spheroid-Type Amyloid Deposition: Comprehensive Analysis Using Immunohistochemistry, Proteomics, and a Literature Review.

This study aimed to elucidate the similarities and differences between amyloid-forming corpora amylacea (CA) in the prostate and lung, examine the nature of CAs in cystic tumors of the atrioventricular node (CTAVN), and clarify the distinctions between amyloid-forming CA and spheroid-type amyloid deposition. We conducted proteomics analyses using liquid chromatography-tandem mass spectrometry with laser microdissection and immunohistochemistry to validate the characteristics of CAs in the lung and prostate. Our findings revealed that the CAs in these organs primarily consisted of common proteins (ß2-microglobulin and lysozyme) and locally produced proteins. Moreover, we observed a discrepancy between the histopathological and proteomic analysis results in CTAVN-associated CAs. In addition, while the histopathological appearance of the amyloid-forming CAs and spheroid-type amyloid deposits were nearly identical, the latter deposition lacked ß2-microglobulin and lysozyme and exhibited evident destruction of the surrounding tissue. A literature review further supported these findings. These results suggest that amyloid-forming CAs in the lung and prostate are formed through a shared mechanism, serving as waste containers (wasteosomes) and/or storage for excess proteins (functional amyloids). In contrast, we hypothesize that while amyloid-forming CA and spheroid-type amyloid deposits are formed, in part, through common mechanisms, the latter are pathological.

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OBJECTIVES: To investigate the effects of different angles of pulmonary surfactant (PS) administration on the incidence of bronchopulmonary dysplasia and intracranial hemorrhage in preterm infants. METHODS: A prospective study was conducted on 146 preterm infants (gestational age <32 weeks) admitted to the Department of Neonatology, Provincial Hospital Affiliated to Anhui Medical University from January 2019 to May 2023. The infants were randomly assigned to different angles for injection of pulmonary surfactant groups: 0° group (34 cases), 30° group (36 cases), 45° group (38 cases), and 60° group (38 cases). Clinical indicators and outcomes were compared among the groups. RESULTS: The oxygenation index was lower in the 60° group compared with the other three groups, with shorter invasive ventilation time and oxygen use time, and a lower incidence of bronchopulmonary dysplasia than the other three groups (P<0.05). The incidence of intracranial hemorrhage was lower in the 60° group compared to the 0° group (P<0.05). The cure rate in the 60° group was higher than that in the 0° group and the 30° group (P<0.05). CONCLUSIONS: The clinical efficacy of injection of pulmonary surfactant at a 60° angle is higher than other angles, reducing the incidence of intracranial hemorrhage and bronchopulmonary dysplasia in preterm infants.

Biophysical function of pulmonary surfactant in liquid ventilation.

Liquid ventilation is a mechanical ventilation technique in which the entire or part of the lung is filled with oxygenated perfluorocarbon (PFC) liquids rather than air in conventional mechanical ventilation. Despite its many ideal biophysicochemical properties for assisting liquid breathing, a general misconception about PFC is to use it as a replacement for pulmonary surfactant. Because of the high PFC-water interfacial tension (59 mN/m), pulmonary surfactant is indispensable in liquid ventilation to increase lung compliance. However, the biophysical function of pulmonary surfactant in liquid ventilation is still unknown. Here, we have studied the adsorption and dynamic surface activity of a natural surfactant preparation, Infasurf, at the PFC-water interface using constrained drop surfactometry. The constrained drop surfactometry is capable of simulating the intra-alveolar microenvironment of liquid ventilation under physiologically relevant conditions. It was found that Infasurf adsorbed to the PFC-water interface reduces the PFC-water interfacial tension from 59 mN/m to an equilibrium value of 9 mN/m within seconds. Atomic force microscopy revealed that after de novo adsorption, Infasurf forms multilayered structures at the PFC-water interface with an average thickness of 10-20 nm, depending on the adsorbing surfactant concentration. It was found that the adsorbed Infasurf film is capable of regulating the interfacial tension of the PFC-water interface within a narrow range, between ∼12 and ∼1 mN/m, during dynamic compression-expansion cycles that mimic liquid ventilation. These findings have novel implications in understanding the physiological and biophysical functions of the pulmonary surfactant film at the PFC-water interface, and may offer new translational insights into the development of liquid ventilation and liquid breathing techniques.

Constrained drop surfactometry for studying adsorbed pulmonary surfactant at physiologically relevant high concentrations.

Exogenous surfactant therapy has been used as a standard clinical intervention for treating premature newborns with respiratory distress syndrome. The phospholipid concentrations of exogenous surfactants used in clinical practice are consistently higher than 25 mg/mL; while it was estimated that the phospholipid concentration of endogenous surfactant is approximately in the range between 15 and 50 mg/mL. However, most in vitro biophysical simulations of pulmonary surfactants were only capable of studying surfactant concentrations up to 3 mg/mL, one order of magnitude lower than the physiologically relevant concentration. Using a new in vitro biophysical model, called constrained drop surfactometry, in conjunction with atomic force microscopy and other technological advances, we have investigated the biophysical properties, ultrastructure, and topography of the pulmonary surfactant film adsorbed from the subphase at physiologically relevant high surfactant concentrations of 10-35 mg/mL. It was found that the effect of surfactant concentration on the dynamic surface activity of the surfactant film was only important when the surface area of the surfactant film varied no more than 15%, mimicking normal tidal breathing. The adsorbed surfactant film depicts a multilayer conformation consisting of a layer-by-layer assembly of stacked bilayers with the height of the multilayers proportional to the surfactant concentration. Our experimental data suggest that the biophysical function of these multilayer structures formed after de novo adsorption is to act as a buffer zone to store surface-active materials ejected from the interfacial monolayer under extreme conditions such as deep breathing.NEW & NOTEWORTHY An in vitro biophysical model, called constrained drop surfactometry, was developed to study the biophysical properties, ultrastructure, and topography of the pulmonary surfactant film adsorbed from the subphase at physiologically relevant high surfactant concentrations of 10-35 mg/mL. These results suggest that the biophysical function of multilayers formed after de novo adsorption is to act as a buffer zone to store surface-active materials ejected from the interfacial monolayer under extreme conditions such as deep breathing.

Decreased expression of surfactant Protein-C and CD74 in alveolar epithelial cells during influenza virus A(H1N1)pdm09 and H3N2 infection.

The primary replication site of Influenza A virus (IAV) is type II alveolar epithelial cells (AECII), which are central to normal lung function and present important immune functions. Surfactant components are synthesized primarily by AECII, which play a crucial role in host defense against infection. The aim of this study was to analyze if the impact of influenza infection is differential between A(H1N1)pdm09 and A/Victoria/3/75 (H3N2) on costimulatory molecules and ProSP-C expression in AECII from BALB/c mice infected and A549 cell line infected with both strains. Pandemic A(H1N1)pdm09 and A/Victoria/3/75 (H3N2) were used to infect BALB/c mice and the A549 cell line. We evaluated the surface expression of co-stimulatory molecules (CD45/CD31/CD74/ProSP-C) in AECII and A549 cell lines. Our results showed a significant decrease in ProSP-C+ CD31- CD45- and CD74+ CD31- CD45- expression in AECII and A549 cell line with the virus strain A(H1N1)pdm09 versus A/Victoria/3/75 (H3N2) and controls (non-infection conditions). Our findings indicate that changes in the expression of ProSP-C in AECII and A549 cell lines in infection conditions could result in dysfunction leading to decreased lung compliance, increased work of breathing and increased susceptibility to injury.