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1.
Cell ; 172(6): 1336-1336.e1, 2018 03 08.
Article in English | MEDLINE | ID: mdl-29522751

ABSTRACT

Hydrocarbon-degrading bacteria are phylogenetically and physiologically diverse and employ layered strategies to sense hydrocarbons, respond transcriptionally, and then move toward an oil source. They then produce biopolymers that increase hydrocarbon bioavailability. This SnapShot highlights how these bacteria respond to and then remove hydrocarbon contaminants from the environment. To view this SnapShot, open or download the PDF.


Subject(s)
Bacteria/metabolism , Hydrocarbons/metabolism , Soil Microbiology , Water Microbiology , Biodegradation, Environmental , Cell Membrane/metabolism , Hydrocarbons/chemistry , Hydrocarbons/classification , Hydrophobic and Hydrophilic Interactions , Metabolic Networks and Pathways , Models, Biological , Quorum Sensing , Surface-Active Agents/chemistry , Surface-Active Agents/metabolism
2.
Annu Rev Biochem ; 86: 585-608, 2017 06 20.
Article in English | MEDLINE | ID: mdl-28125290

ABSTRACT

Many critical biological processes take place at hydrophobic:hydrophilic interfaces, and a wide range of organisms produce surface-active proteins and peptides that reduce surface and interfacial tension and mediate growth and development at these boundaries. Microorganisms produce both small lipid-associated peptides and amphipathic proteins that allow growth across water:air boundaries, attachment to surfaces, predation, and improved bioavailability of hydrophobic substrates. Higher-order organisms produce surface-active proteins with a wide variety of functions, including the provision of protective foam environments for vulnerable reproductive stages, evaporative cooling, and gas exchange across airway membranes. In general, the biological functions supported by these diverse polypeptides require them to have an amphipathic nature, and this is achieved by a diverse range of molecular structures, with some proteins undergoing significant conformational change or intermolecular association to generate the structures that are surface active.


Subject(s)
Caseins/chemistry , Glycoproteins/chemistry , Membrane Proteins/chemistry , Neoplasm Proteins/chemistry , Phosphoproteins/chemistry , Pulmonary Surfactants/chemistry , Surface-Active Agents/chemistry , Animals , Bacteria/chemistry , Bacteria/genetics , Bacteria/metabolism , Caseins/genetics , Caseins/metabolism , Fungi/chemistry , Fungi/genetics , Fungi/metabolism , Glycoproteins/genetics , Glycoproteins/metabolism , Humans , Hydrophobic and Hydrophilic Interactions , Mammals , Membrane Proteins/genetics , Membrane Proteins/metabolism , Neoplasm Proteins/genetics , Neoplasm Proteins/metabolism , Phosphoproteins/genetics , Phosphoproteins/metabolism , Protein Conformation , Pulmonary Surfactants/metabolism , Surface Properties , Surface-Active Agents/metabolism , Water/chemistry , Water/metabolism
3.
Proc Natl Acad Sci U S A ; 121(22): e2403013121, 2024 May 28.
Article in English | MEDLINE | ID: mdl-38781207

ABSTRACT

Biomolecular condensates are cellular compartments that concentrate biomolecules without an encapsulating membrane. In recent years, significant advances have been made in the understanding of condensates through biochemical reconstitution and microscopic detection of these structures. Quantitative visualization and biochemical assays of biomolecular condensates rely on surface passivation to minimize background and artifacts due to condensate adhesion. However, the challenge of undesired interactions between condensates and glass surfaces, which can alter material properties and impair observational accuracy, remains a critical hurdle. Here, we introduce an efficient, broadly applicable, and simple passivation method employing self-assembly of the surfactant Pluronic F127 (PF127). The method greatly reduces nonspecific binding across a range of condensates systems for both phase-separated droplets and biomolecules in dilute phase. Additionally, by integrating PF127 passivation with the Biotin-NeutrAvidin system, we achieve controlled multipoint attachment of condensates to surfaces. This not only preserves condensate properties but also facilitates long-time fluorescence recovery after photobleaching imaging and high-precision single-molecule analyses. Using this method, we have explored the dynamics of polySIM molecules within polySUMO/polySIM condensates at the single-molecule level. Our observations suggest a potential heterogeneity in the distribution of available polySIM-binding sites within the condensates.


Subject(s)
Avidin , Biomolecular Condensates , Biotin , Poloxamer , Biomolecular Condensates/chemistry , Biomolecular Condensates/metabolism , Poloxamer/chemistry , Biotin/chemistry , Biotin/metabolism , Avidin/chemistry , Avidin/metabolism , Fluorescence Recovery After Photobleaching/methods , Surface Properties , Surface-Active Agents/chemistry , Surface-Active Agents/metabolism , Single Molecule Imaging/methods
4.
Proc Natl Acad Sci U S A ; 120(3): e2211092120, 2023 01 17.
Article in English | MEDLINE | ID: mdl-36634141

ABSTRACT

Recent experimental and computational investigations have shown that trace amounts of surfactants, unavoidable in practice, can critically impair the drag reduction of superhydrophobic surfaces (SHSs), by inducing Marangoni stresses at the air-liquid interface. However, predictive models for realistic SHS geometries do not yet exist, which has limited the understanding and mitigation of these adverse surfactant effects. To address this issue, we derive a model for laminar, three-dimensional flow over SHS gratings as a function of geometry and soluble surfactant properties, which together encompass 10 dimensionless groups. We establish that the grating length g is the key geometric parameter and predict that the ratio between actual and surfactant-free slip increases with g2. Guided by our model, we perform synergistic numerical simulations and microfluidic experiments, finding good agreement with the theory as we vary surfactant type and SHS geometry. Our model also enables the estimation, based on velocity measurements, of a priori unknown properties of surfactants inherently present in microfluidic systems. For SHSs, we show that surfactant effects can be predicted by a single parameter, representing the ratio between the grating length and the interface length scale beyond which the flow mobilizes the air-water interface. This mobilization length is more sensitive to the surfactant chemistry than to its concentration, such that even trace-level contaminants may significantly increase drag if they are highly surface active. These findings advance the fundamental understanding of realistic interfacial flows and provide practical strategies to maximize superhydrophobic drag reduction.


Subject(s)
Pulmonary Surfactants , Surface-Active Agents , Surface-Active Agents/chemistry , Microfluidics , Lipoproteins , Hydrophobic and Hydrophilic Interactions
5.
Proc Natl Acad Sci U S A ; 120(51): e2309900120, 2023 Dec 19.
Article in English | MEDLINE | ID: mdl-38085774

ABSTRACT

How acute respiratory distress syndrome progresses from underlying disease or trauma is poorly understood, and there are no generally accepted treatments resulting in a 40% mortality rate. However, during the inflammation that accompanies this disease, the phospholipase A2 concentration increases in the alveolar fluids leading to the hydrolysis of bacterial, viral, and lung surfactant phospholipids into soluble lysolipids. We show that if the lysolipid concentration in the subphase reaches or exceeds its critical micelle concentration, the surface tension, γ, of dipalmitoyl phosphatidylcholine (DPPC) or Curosurf monolayers increases and the dilatational modulus, [Formula: see text], decreases to that of a pure lysolipid interface. This is consistent with DPPC being solubilized in lysolipid micelles and being replaced by lysolipid at the interface. These changes lead to [Formula: see text] which is the criterion for the Laplace instability that can lead to mechanical instabilities during lung inflation, potentially causing alveolar collapse. These findings provide a mechanism behind the alveolar collapse and uneven lung inflation during ARDS.


Subject(s)
Pulmonary Surfactants , Respiratory Distress Syndrome , Humans , Lung , Phospholipases A2 , Surface-Active Agents
6.
Annu Rev Biochem ; 79: 737-75, 2010.
Article in English | MEDLINE | ID: mdl-20307193

ABSTRACT

Membrane proteins (MPs) are usually handled in aqueous solutions as protein/detergent complexes. Detergents, however, tend to be inactivating. This situation has prompted the design of alternative surfactants that can be substituted for detergents once target proteins have been extracted from biological membranes and that keep them soluble in aqueous buffers while stabilizing them. The present review focuses on three such systems: Amphipols (APols) are amphipathic polymers that adsorb onto the hydrophobic transmembrane surface of MPs; nanodiscs (NDs) are small patches of lipid bilayer whose rim is stabilized by amphipathic proteins; fluorinated surfactants (FSs) resemble detergents but interfere less than detergents do with stabilizing protein/protein and protein/lipid interactions. The structure and properties of each of these three systems are described, as well as those of the complexes they form with MPs. Their respective usefulness, constraints, and prospects for functional and structural studies of MPs are discussed.


Subject(s)
Biochemistry/methods , Membrane Proteins/chemistry , Detergents/metabolism , Humans , Hydrophobic and Hydrophilic Interactions , Membrane Proteins/metabolism , Nanoparticles , Protein Conformation , Surface-Active Agents/metabolism
7.
Dev Biol ; 506: 64-71, 2024 Feb.
Article in English | MEDLINE | ID: mdl-38081502

ABSTRACT

INTRODUCTION: After birth, the lungs must resorb the fluid they contain. This process involves multiple actors such as surfactant, aquaporins and ENaC channels. Preterm newborns often exhibit respiratory distress syndrome due to surfactant deficiency, and transitory tachypnea caused by a delay in lung liquid resorption. Our hypothesis is that surfactant, ENaC and aquaporins are involved in respiratory transition to extrauterine life and altered by preterm birth. We compared these candidates in preterm and term fetal sheeps. MATERIALS AND METHODS: We performed cesarean sections in 8 time-dated pregnant ewes (4 at 100 days and 4 at 140 days of gestation, corresponding to 24 and 36 weeks of gestation in humans), and obtained 13 fetal sheeps in each group. We studied surfactant synthesis (SP-A, SP-B, SP-C), lung liquid resorption (ENaC, aquaporins) and corticosteroid regulation (glucocorticoid receptor, mineralocorticoid receptor and 11-betaHSD2) at mRNA and protein levels. RESULTS: The mRNA expression level of SFTPA, SFTPB and SFTPC was higher in the term group. These results were confirmed at the protein level for SP-B on Western Blot analysis and for SP-A, SP-B and SP-C on immunohistochemical analysis. Regarding aquaporins, ENaC and receptors, mRNA expression levels for AQP1, AQP3, AQP5, ENaCα, ENaCß, ENaCγ and 11ßHSD2 mRNA were also higher in the term group. DISCUSSION: Expression of surfactant proteins, aquaporins and ENaC increases between 100 and 140 days of gestation in an ovine model. Further exploring these pathways and their hormonal regulation could highlight some new explanations in the pathophysiology of neonatal respiratory diseases.


Subject(s)
Aquaporins , Premature Birth , Pregnancy , Humans , Animals , Sheep , Female , Surface-Active Agents/metabolism , Premature Birth/metabolism , Lung/metabolism , Aquaporins/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism
8.
Hum Mol Genet ; 32(18): 2842-2855, 2023 09 05.
Article in English | MEDLINE | ID: mdl-37471639

ABSTRACT

Pulmonary surfactant is a lipoprotein synthesized and secreted by alveolar type II cells in lung. We evaluated the associations between 200,139 single nucleotide polymorphisms (SNPs) of 40 surfactant-related genes and lung cancer risk using genotyped data from two independent lung cancer genome-wide association studies. Discovery data included 18,082 cases and 13,780 controls of European ancestry. Replication data included 1,914 cases and 3,065 controls of European descent. Using multivariate logistic regression, we found novel SNPs in surfactant-related genes CTSH [rs34577742 C > T, odds ratio (OR) = 0.90, 95% confidence interval (CI) = 0.89-0.93, P = 7.64 × 10-9] and SFTA2 (rs3095153 G > A, OR = 1.16, 95% CI = 1.10-1.21, P = 1.27 × 10-9) associated with overall lung cancer in the discovery data and validated in an independent replication data-CTSH (rs34577742 C > T, OR = 0.88, 95% CI = 0.80-0.96, P = 5.76 × 10-3) and SFTA2 (rs3095153 G > A, OR = 1.14, 95% CI = 1.01-1.28, P = 3.25 × 10-2). Among ever smokers, we found SNPs in CTSH (rs34577742 C > T, OR = 0.89, 95% CI = 0.85-0.92, P = 1.94 × 10-7) and SFTA2 (rs3095152 G > A, OR = 1.20, 95% CI = 1.14-1.27, P = 4.25 × 10-11) associated with overall lung cancer in the discovery data and validated in the replication data-CTSH (rs34577742 C > T, OR = 0.88, 95% CI = 0.79-0.97, P = 1.64 × 10-2) and SFTA2 (rs3095152 G > A, OR = 1.15, 95% CI = 1.01-1.30, P = 3.81 × 10-2). Subsequent transcriptome-wide association study using expression weights from a lung expression quantitative trait loci study revealed genes most strongly associated with lung cancer are CTSH (PTWAS = 2.44 × 10-4) and SFTA2 (PTWAS = 2.32 × 10-6).


Subject(s)
Lung Neoplasms , Pulmonary Surfactants , Humans , Genome-Wide Association Study , Lung/metabolism , Genotype , Pulmonary Surfactants/metabolism , Surface-Active Agents/metabolism , Polymorphism, Single Nucleotide , Genetic Predisposition to Disease , Cathepsin H/genetics , Cathepsin H/metabolism
9.
Chem Rev ; 123(23): 13209-13290, 2023 12 13.
Article in English | MEDLINE | ID: mdl-37862151

ABSTRACT

Pulmonary surfactant is a critical component of lung function in healthy individuals. It functions in part by lowering surface tension in the alveoli, thereby allowing for breathing with minimal effort. The prevailing thinking is that low surface tension is attained by a compression-driven squeeze-out of unsaturated phospholipids during exhalation, forming a film enriched in saturated phospholipids that achieves surface tensions close to zero. A thorough review of past and recent literature suggests that the compression-driven squeeze-out mechanism may be erroneous. Here, we posit that a surfactant film enriched in saturated lipids is formed shortly after birth by an adsorption-driven sorting process and that its composition does not change during normal breathing. We provide biophysical evidence for the rapid formation of an enriched film at high surfactant concentrations, facilitated by adsorption structures containing hydrophobic surfactant proteins. We examine biophysical evidence for and against the compression-driven squeeze-out mechanism and propose a new model for surfactant function. The proposed model is tested against existing physiological and pathophysiological evidence in neonatal and adult lungs, leading to ideas for biophysical research, that should be addressed to establish the physiological relevance of this new perspective on the function of the mighty thin film that surfactant provides.


Subject(s)
Pulmonary Surfactants , Infant, Newborn , Humans , Pulmonary Surfactants/chemistry , Pulmonary Surfactants/metabolism , Phospholipids/chemistry , Surface-Active Agents , Surface Tension , Chemical Phenomena
10.
Nature ; 572(7770): 507-510, 2019 08.
Article in English | MEDLINE | ID: mdl-31435058

ABSTRACT

The ability to manipulate droplets on a substrate using electric signals1-known as digital microfluidics-is used in optical2,3, biomedical4,5, thermal6 and electronic7 applications and has led to commercially available liquid lenses8 and diagnostics kits9,10. Such electrical actuation is mainly achieved by electrowetting, with droplets attracted towards and spreading on a conductive substrate in response to an applied voltage. To ensure strong and practical actuation, the substrate is covered with a dielectric layer and a hydrophobic topcoat for electrowetting-on-dielectric (EWOD)11-13; this increases the actuation voltage (to about 100 volts) and can compromise reliability owing to dielectric breakdown14, electric charging15 and biofouling16. Here we demonstrate droplet manipulation that uses electrical signals to induce the liquid to dewet, rather than wet, a hydrophilic conductive substrate without the need for added layers. In this electrodewetting mechanism, which is phenomenologically opposite to electrowetting, the liquid-substrate interaction is not controlled directly by electric field but instead by field-induced attachment and detachment of ionic surfactants to the substrate. We show that this actuation mechanism can perform all the basic fluidic operations of digital microfluidics using water on doped silicon wafers in air, with only ±2.5 volts of driving voltage, a few microamperes of current and about 0.015 times the critical micelle concentration of an ionic surfactant. The system can also handle common buffers and organic solvents, promising a simple and reliable microfluidic platform for a broad range of applications.


Subject(s)
Electrowetting/methods , Microfluidics/methods , Surface-Active Agents/chemistry , Acetonitriles/chemistry , Buffers , Dimethyl Sulfoxide/chemistry , Ethylene Glycol/chemistry , Hydrophobic and Hydrophilic Interactions , Ions/chemistry , Microfluidics/instrumentation , Silicon/chemistry
11.
Proc Natl Acad Sci U S A ; 119(25): e2123564119, 2022 06 21.
Article in English | MEDLINE | ID: mdl-35696565

ABSTRACT

In the context of the rapid increase of antibiotic-resistant infections, in particular of pneumonia, antimicrobial photodynamic therapy (aPDT), the microbiological application of photodynamic therapy (PDT), comes in as a promising treatment alternative since the induced damage and resultant death are not dependent on a specific biomolecule or cellular pathway. The applicability of aPDT using the photosensitizer indocyanine green with infrared light has been successfully demonstrated for different bacterial agents in vitro, and the combination of pulmonary delivery using nebulization and external light activation has been shown to be feasible. However, there has been little progress in obtaining sufficient in vivo efficacy results. This study reports the lung surfactant as a significant suppressor of aPDT in the lungs. In vitro, the clinical surfactant Survanta® reduced the aPDT effect of indocyanine green, Photodithazine®, bacteriochlorin-trizma, and protoporphyrin IX against Streptococcus pneumoniae. The absorbance and fluorescence spectra, as well as the photobleaching profile, suggested that the decrease in efficacy is not a result of singlet oxygen quenching, while a molecular dynamics simulation showed an affinity for the polar head groups of the surfactant phospholipids that likely impacts uptake of the photosensitizers by the bacteria. Methylene blue is the exception, likely because its high water solubility confers a higher mobility when interacting with the surfactant layer. We propose that the interaction between lung surfactant and photosensitizer must be taken into account when developing pulmonary aPDT protocols.


Subject(s)
Anti-Bacterial Agents , Bacteria , Photochemotherapy , Photosensitizing Agents , Surface-Active Agents , Anti-Bacterial Agents/pharmacology , Bacteria/drug effects , Indocyanine Green/pharmacology , Lung/microbiology , Molecular Dynamics Simulation , Photochemotherapy/methods , Photosensitizing Agents/pharmacology , Surface-Active Agents/metabolism
12.
Biophys J ; 123(13): 1781-1791, 2024 Jul 02.
Article in English | MEDLINE | ID: mdl-38783603

ABSTRACT

Controlling the reduction midpoint potential of heme B is a key factor in many bioelectrochemical reactions, including long-range electron transport. Currently, there are a number of globular model protein systems to study this biophysical parameter; however, there are none for large polymeric protein model systems (e.g., the OmcS protein from G. sulfurreducens). Peptide amphiphiles, short peptides with a lipid tail that polymerize into fibrous structures, fill this gap. Here, we show a peptide amphiphile model system where one can tune the electrochemical potential of heme B by changing the loading ratio and peptide sequence. Changing the loading ratio resulted in the most significant increase, with values as high as -22 mV down to -224 mV. Circular dichroism spectra of certain sequences show Cotton effects at lower loading ratios that disappear as more heme B is added, indicating an ordered environment that becomes disrupted if heme B is overpacked. These findings can contribute to the design of functional self-assembling biomaterials.


Subject(s)
Heme , Oxidation-Reduction , Peptides , Heme/chemistry , Peptides/chemistry , Amino Acid Sequence , Surface-Active Agents/chemistry
13.
J Proteome Res ; 23(8): 3542-3551, 2024 Aug 02.
Article in English | MEDLINE | ID: mdl-38973097

ABSTRACT

Trypsin digestion plays a pivotal role in successful bottom-up peptide characterization and quantitation. While denaturants are often incorporated to enhance protein solubility, surfactants are recognized to inhibit enzyme activity. However, several reports have suggested that incorporating surfactants or other solvent additives may enhance digestion and MS detection. Here, we assess the impacts of ionic surfactants on cumulative trypsin activity and subsequently evaluate the total digestion efficiency of a proteome mixture by quantitative MS. Although low surfactant concentrations, such as 0.01% SDS or 0.2% SDC, significantly enhanced the initial trypsin activity (by 14 or 42%, respectively), time course assays revealed accelerated enzyme deactivation, evident by 10- or 40-fold reductions in trypsin activity half-life at these respective surfactant concentrations. Despite enhanced initial tryptic activity, quantitative MS analysis of a common liver proteome extract, digested with various surfactants (0.01 or 0.1% SDS, 0.5% SDC), consistently revealed decreased peptide counts and signal intensity, indicative of a lower digestion efficiency compared to a nonsurfactant control. Furthermore, including detergents for digestion did not improve the detection of membrane proteins, nor hydrophobic peptides. These results stress the importance of assessing cumulative enzyme activity when optimizing the digestion of a proteome mixture, particularly in the presence of denaturants.


Subject(s)
Proteome , Proteomics , Surface-Active Agents , Trypsin , Trypsin/metabolism , Trypsin/chemistry , Surface-Active Agents/pharmacology , Surface-Active Agents/chemistry , Proteome/analysis , Proteomics/methods , Animals , Sodium Dodecyl Sulfate/pharmacology , Sodium Dodecyl Sulfate/chemistry , Liver/metabolism , Liver/enzymology , Liver/drug effects
14.
J Proteome Res ; 23(8): 2908-2918, 2024 Aug 02.
Article in English | MEDLINE | ID: mdl-38315831

ABSTRACT

The extracellular matrix (ECM) is a complex assembly of proteins that provide interstitial scaffolding and elastic recoil for human lungs. The pulmonary extracellular matrix is increasingly recognized as an independent bioactive entity, by creating biochemical and mechanical signals that influence disease pathogenesis, making it an attractive therapeutic target. However, the pulmonary ECM proteome ("matrisome") remains challenging to analyze by mass spectrometry due to its inherent biophysical properties and relatively low abundance. Here, we introduce a strategy designed for rapid and efficient characterization of the human pulmonary ECM using the photocleavable surfactant Azo. We coupled this approach with trapped ion mobility MS with diaPASEF to maximize the depth of matrisome coverage. Using this strategy, we identify nearly 400 unique matrisome proteins with excellent reproducibility that are known to be important in lung biology, including key core matrisome proteins.


Subject(s)
Extracellular Matrix , Lung , Proteomics , Humans , Proteomics/methods , Lung/metabolism , Extracellular Matrix/metabolism , Azo Compounds/chemistry , Extracellular Matrix Proteins/metabolism , Surface-Active Agents/chemistry , Proteome/analysis , Mass Spectrometry/methods
15.
Am J Physiol Lung Cell Mol Physiol ; 327(4): L535-L546, 2024 Oct 01.
Article in English | MEDLINE | ID: mdl-39159363

ABSTRACT

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.


Subject(s)
Pulmonary Surfactants , Respiratory Distress Syndrome, Newborn , Pulmonary Surfactants/chemistry , Pulmonary Surfactants/metabolism , Animals , Humans , Respiratory Distress Syndrome, Newborn/drug therapy , Swine , Cattle , Adsorption , Biological Products/chemistry , Biological Products/pharmacology , Phospholipids/chemistry , Phospholipids/metabolism , Meconium/chemistry , Biophysical Phenomena , Surface Properties , Surface-Active Agents/chemistry , Surface-Active Agents/pharmacology , Infant, Newborn
16.
Am J Physiol Lung Cell Mol Physiol ; 326(3): L330-L343, 2024 Mar 01.
Article in English | MEDLINE | ID: mdl-38252635

ABSTRACT

Extremely preterm infants are often exposed to long durations of mechanical ventilation to facilitate gas exchange, resulting in ventilation-induced lung injury (VILI). New lung protective strategies utilizing noninvasive ventilation or low tidal volumes are now common but have not reduced rates of bronchopulmonary dysplasia. We aimed to determine the effect of 24 h of low tidal volume ventilation on the immature lung by ventilating preterm fetal sheep in utero. Preterm fetal sheep at 110 ± 1(SD) days' gestation underwent sterile surgery for instrumentation with a tracheal loop to enable in utero mechanical ventilation (IUV). At 112 ± 1 days' gestation, fetuses received either in utero mechanical ventilation (IUV, n = 10) targeting 3-5 mL/kg for 24 h, or no ventilation (CONT, n = 9). At necropsy, fetal lungs were collected to assess molecular and histological markers of lung inflammation and injury. IUV significantly increased lung mRNA expression of interleukin (IL)-1ß, IL-6, IL-8, IL-10, and tumor necrosis factor (TNF) compared with CONT, and increased surfactant protein (SP)-A1, SP-B, and SP-C mRNA expression compared with CONT. IUV produced modest structural changes to the airways, including reduced parenchymal collagen and myofibroblast density. IUV increased pulmonary arteriole thickness compared with CONT but did not alter overall elastin or collagen content within the vasculature. In utero ventilation of an extremely preterm lung, even at low tidal volumes, induces lung inflammation and injury to the airways and vasculature. In utero ventilation may be an important model to isolate the confounding mechanisms of VILI to develop effective therapies for preterm infants requiring prolonged respiratory support.NEW & NOTEWORTHY Preterm infants often require prolonged respiratory support, but the relative contribution of ventilation to the development of lung injury is difficult to isolate. In utero mechanical ventilation allows for mechanistic investigations into ventilation-induced lung injury without confounding factors associated with sustaining extremely preterm lambs ex utero. Twenty-four hours of in utero ventilation, even at low tidal volumes, increased lung inflammation and surfactant protein expression and produced structural changes to the lung parenchyma and vasculature.


Subject(s)
Pneumonia , Ventilator-Induced Lung Injury , Humans , Infant, Newborn , Sheep , Animals , Infant, Extremely Premature , Lung/metabolism , Fetus/metabolism , Respiration, Artificial/adverse effects , Respiration, Artificial/methods , Ventilator-Induced Lung Injury/metabolism , Collagen/metabolism , Pneumonia/pathology , Surface-Active Agents/metabolism , RNA, Messenger/metabolism
17.
Anal Chem ; 96(39): 15797-15807, 2024 Oct 01.
Article in English | MEDLINE | ID: mdl-39285721

ABSTRACT

Clustered regularly interspaced short palindromic repeats (CRISPR) technology has opened a new path for molecular diagnostics based on RNA programmed trans-cleavage activity. However, their accessibility for highly sensitive clinical diagnostics remains insufficient. In this study, we systematically investigated the impact of various surfactants on the CRISPR-Cas12a system and found that poly(vinylpyrrolidone) (PVP), a nonionic surfactant, showed the highest enhancement effect among these tested surfactants. Additionally, the enhancement effects of PVP are compatible and versatile to CRISPR-Cas12b and Cas13a systems, improving the sensitivity of these CRISPR-Cas systems toward synthetic targets by 1-2 orders of magnitude. By integrating the PVP-enhanced CRISPR system with isothermal nucleic acid amplification, both the two- and one-step assays exhibited comparable sensitivity and specificity to gold-standard quantitative polymerase chain reaction (qPCR) in the assay of clinical human papillomavirus (HPV) samples, thereby holding significant promise for advancing clinical diagnostics and biomedical research.


Subject(s)
CRISPR-Cas Systems , Povidone , CRISPR-Cas Systems/genetics , Povidone/chemistry , Humans , Nucleic Acid Amplification Techniques/methods , Surface-Active Agents/chemistry , Papillomaviridae/genetics
18.
Anal Chem ; 96(17): 6746-6755, 2024 04 30.
Article in English | MEDLINE | ID: mdl-38632675

ABSTRACT

Nonionic surfactant excipients (NISEs) are commonly added to biologics formulations to mitigate the effects of stress incurred by the active biotherapeutic during manufacturing, transport, and storage. During manufacturing, NISEs are added by dilution of a stock solution directly into a protein formulation, and their accurate addition is critical in maintaining the quality and integrity of the drug product and thus ensuring patient safety. This is especially true for the common NISEs, polysorbates 20 and 80 (PS20 and PS80, respectively) and poloxamer 188 (P188). With the increasing diversity of biologic modalities within modern pharmaceutical pipelines, there is thus a critical need to develop and deploy convenient and user-accessible analytical techniques that can rapidly and reliably quantify these NISEs under biopharmaceutically relevant conditions. We thus pursued 60 MHz benchtop quantitative NMR (qNMR) as a nondestructive and user-friendly analytical technique for the quantification of PS20, PS80, and P188 under such conditions. We demonstrated the ability of benchtop qNMR (1) to quantify simulated PS20, PS80, and P188 stock solutions representative of those used during the drug substance (DS) formulation step in biomanufacturing and (2) to quantify these NISEs at and below their target concentrations (≤0.025% w/v) directly in biologics formulations containing histidine, sucrose, and one of three biotherapeutic modalities (monoclonal antibody, antibody-drug conjugate, and Fc-fusion protein). Our results demonstrate that benchtop qNMR offers a fit-for-purpose, reliable, user-friendly, and green analytical route by which NISE of interest to the biopharmaceutical industry may be readily and reliably quantified. We conclude that benchtop qNMR has the potential to be applied to other excipient formulation components in the presence of various biological modalities as well as the potential for routine integration within analytical and QC laboratories across pharmaceutical development and manufacturing sites.


Subject(s)
Excipients , Magnetic Resonance Spectroscopy , Surface-Active Agents , Surface-Active Agents/chemistry , Excipients/chemistry , Excipients/analysis , Magnetic Resonance Spectroscopy/methods , Polysorbates/chemistry , Poloxamer/chemistry , Biological Products/chemistry , Biological Products/analysis
19.
Anal Chem ; 96(18): 7257-7264, 2024 05 07.
Article in English | MEDLINE | ID: mdl-38664861

ABSTRACT

Confocal fluorescence imaging of fine structures of the cell membrane is important for understanding their biofunctions but is often neglected due to the lack of an effective method. Herein, we develop new amphiphilic rhodamine fluorescent probe RMGs in combination with basal imaging for this purpose. The probes show high signal-to-noise ratio and brightness and low internalization rate, making them suitable for imaging the fine substructures of the cell membrane. Using the representative probe RMG3, we not only observed the cell pseudopodia and intercellular nanotubes but also monitored the formation of migrasomes in real time. More importantly, in-depth imaging studies on more cell lines revealed for the first time that hepatocellular carcinoma cells secreted much more adherent extracellular vesicles than other cell lines, which might serve as a potential indicator of liver cells. We believe that RMGs may be useful for investigating the fine structures of the cell membrane.


Subject(s)
Cell Membrane , Fluorescent Dyes , Rhodamines , Fluorescent Dyes/chemistry , Rhodamines/chemistry , Humans , Cell Membrane/chemistry , Optical Imaging , Microscopy, Confocal/methods , Surface-Active Agents/chemistry
20.
Biochem Biophys Res Commun ; 733: 150572, 2024 Nov 12.
Article in English | MEDLINE | ID: mdl-39191187

ABSTRACT

Fungal lipolytic enzymes play crucial roles in various lipid bio-industry processes. Here, we elucidated the biochemical and structural characteristics of an unexplored fungal lipolytic enzyme (TaLip) from Thermoascus aurantiacus var. levisporus, a strain renowned for its significant industrial relevance in carbohydrate-active enzyme production. TaLip belongs to a poorly understood phylogenetic branch within the class 3 lipase family and prefers to hydrolyze mainly short-chain esters. Nonetheless, it also displays activity against natural long-chain triacylglycerols. Furthermore, our analyses revealed that the surfactant sodium dodecyl sulfate (SDS) enhances the hydrolytic activity of TaLip on pNP butyrate by up to 5.0-fold. Biophysical studies suggest that interactions with SDS may prevent TaLip aggregation, thereby preserving the integrity and stability of its monomeric form and improving its performance. These findings highlight the resilience of TaLip as a lipolytic enzyme capable of functioning in tandem with surfactants, offering an intriguing enzymatic model for further exploration of surfactant tolerance and activation in biotechnological applications.


Subject(s)
Esterases , Lipase , Surface-Active Agents , Surface-Active Agents/chemistry , Surface-Active Agents/pharmacology , Lipase/metabolism , Lipase/chemistry , Esterases/metabolism , Esterases/chemistry , Sodium Dodecyl Sulfate/chemistry , Substrate Specificity , Hydrolysis , Fungal Proteins/chemistry , Fungal Proteins/metabolism , Anions/chemistry , Anions/metabolism , Enzyme Stability
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