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Single-walled carbon nanotubes (SWCNTs) are at present synthesized on a large scale with a variety of applications. The increasing likelihood of exposure to SWCNTs, however, puts human health at a high risk. As the front line of the innate host defense system, the pulmonary surfactant monolayer (PSM) at the air–water interface of the lungs interacts with the inhaled SWCNTs, which in turn inevitably perturb the ultrastructure of the PSM and affect its biophysical functions. Here, using molecular dynamics simulations, we demonstrate how the diameter and length of SWCNTs critically regulate their interactions with the PSM. Compared to their diameters, the inhalation toxicity of SWCNTs was found to be largely affected by their lengths. Short SWCNTs with lengths comparable to the monolayer thickness are found to vertically insert into the PSM with no indication of translocation, possibly leading to accumulation of SWCNTs in the PSM with prolonged retention and increased inflammation potentials. The perturbation also comes from the forming water pores across the PSM. Longer SWCNTs are found to horizontally insert into the PSM during inspiration, and they can be wrapped by the PSM during deep expiration via a tube diameter-dependent self-rotation. The potential toxicity of longer SWCNTs comes from severe lipid depletion and the PSM-rigidifying effect. Our findings could help reveal the inhalation toxicity of SWCNTs, and pave the way for the safe use of SWCNTs as vehicles for pulmonary drug delivery.

Environmental exposure and health risk upon engineered nanomaterials are increasingly concerned. The family of mesoporous carbon nanomaterials (MCNs) is a rising star in nanotechnology for multidisciplinary research with versatile applications in electronics, energy and gas storage, and biomedicine. Meanwhile, there is mounting concern on their environmental health risks due to the growing production and usage of MCNs. The lung is the primary site for particle invasion under environmental exposure to nanomaterials. Here, we studied the comprehensive toxicological profile of MCNs in the lung under the scenario of moderate environmental exposure. It was found that at a low concentration of 10μg/mL MCNs induced biophysical inhibition of natural pulmonary surfactant. Moreover, MCNs at similar concentrations reduced viability of J774A.1 macrophages and lung epithelial A549 cells. Incubating with nature pulmonary surfactant effectively reduced the cytotoxicity of MCNs. Regarding the pro-inflammatory responses, MCNs activated macrophages in vitro, and stimulated lung inflammation in mice after inhalation exposure, associated with lung fibrosis. Moreover, we found that the size of MCNs played a significant role in regulating cytotoxicity and pro-inflammatory potential of this nanomaterial. In general, larger MCNs induced more pronounced cytotoxic and pro-inflammatory effects than their smaller counterparts. Our results provided valuable information on the toxicological profile and environmental health risks of MCNs, and suggested that fine-tuning the size of MCNs could be a practical precautionary design strategy to increase safety and biocompatibility of this nanomaterial.

Alterations to the pulmonary surfactant system have been observed consistently in ventilation-induced lung injury (VILI) including composition changes and impairments in the surface tension reducing ability of the isolated extracellular surfactant. However, there is limited information about the effects of VILI on the intracellular form of surfactant, the lamellar body. It is hypothesized that VILI leads to alterations of lamellar body numbers and function. To test this hypothesis, rats were randomized to one of three groups, nonventilated controls, control ventilation, and high tidal volume ventilation (VILI). Following physiological assessment to confirm lung injury, isolated lamellar bodies were tested for surfactant function on a constrained sessile drop surfactometer. A separate cohort of animals was used to fix the lungs followed by examination of lamellar body numbers and morphology using transmission electron microscopy. The results showed an impaired ability of reducing surface tension for the lamellar bodies isolated from the VILI group as compared with the two other groups. The morphological assessment revealed that the number, and the relative area covered by, lamellar bodies were significantly decreased in animals with VILI animals as compared with the other groups. It is concluded that VILI causes significant alterations to lamellar bodies. It is speculated that increased secretion causes a depletion of lamellar bodies that cannot be compensated by de novo synthesis of surfactant in these injured lungs.

Vascular plants transport water under negative pressure without constantly creating gas bubbles that would disable their hydraulic systems. Attempts to replicate this feat in artificial systems almost invariably result in bubble formation, except under highly controlled conditions with pure water and only hydrophilic surfaces present. In theory, conditions in the xylem should favor bubble nucleation even more: there are millions of conduits with at least some hydrophobic surfaces, and xylem sap is saturated or sometimes supersaturated with atmospheric gas and may contain surface-active molecules that can lower surface tension. So how do plants transport water under negative pressure? Here, we show that angiosperm xylem contains abundant hydrophobic surfaces as well as insoluble lipid surfactants, including phospholipids, and proteins, a composition similar to pulmonary surfactants. Lipid surfactants were found in xylem sap and as nanoparticles under transmission electron microscopy in pores of intervessel pit membranes and deposited on vessel wall surfaces. Nanoparticles observed in xylem sap via nanoparticle-tracking analysis included surfactant-coated nanobubbles when examined by freeze-fracture electron microscopy. Based on their fracture behavior, this technique is able to distinguish between dense-core particles, liquid-filled, bilayer-coated vesicles/liposomes, and gas-filled bubbles. Xylem surfactants showed strong surface activity that reduces surface tension to low values when concentrated as they are in pit membrane pores. We hypothesize that xylem surfactants support water transport under negative pressure as explained by the cohesion-tension theory by coating hydrophobic surfaces and nanobubbles, thereby keeping the latter below the critical size at which bubbles would expand to form embolisms.

Microalgae are one of the most promising renewable energy sources with environmental sustainability. The surface free energy of microalgal cells determines their biofouling and bioflocculation behavior and hence plays an important role in microalgae cultivation and harvesting. To date, the surface energetic properties of microalgal cells are still rarely studied. We developed a novel spectrophotometric method for directly determining the surface free energy of microalgal cells. The principles of this method are based on analyzing colloidal stability of microalgae suspensions. We have shown that this method can effectively differentiate the surface free energy of four microalgal strains, i.e., marine Chlorella sp., marine Nannochloris oculata, freshwater autotrophic Chlorella sp., and freshwater heterotrophic Chlorella sp. With advantages of high-throughput and simplicity, this new spectrophotometric method has the potential to evolve into a standard method for measuring the surface free energy of cells and abiotic particles.

Hydroxyapatite/polyamide-66 (HA/PA66) composite scaffolds were prepared using injection-molding technique, and also analyzed by means of scanning electron microscopy (SEM), dynamic mechanical analysis (DMA), Fourier transform infrared spectroscopy (FTIR) and mechanical testing. Compared with common methods including solvent casting/particulate leaching, phase separation and so forth to fabricate scaffolds, this process is of a rapid and convenient manner. The increase of HA content can increase the stiffness of composite scaffolds accompanied by the reduction of impact strength, pore size and porosity. The storage modulus of composite scaffolds increases with increasing HA content and with decreasing porosity. The damping (tan δ) of PA66 decreases with the increase of HA content, and the α and β relaxation peaks of PA66 for the foamed HA/PA66 composites slightly shift to lower value, indicating that both HA and gas in the foamed HA/PA66 composites have an effect on the chain mobility of the polymer and the interaction between the polymer chains. The microstructure of the 30 wt% and 40 wt% HA/PA66 composite scaffolds with porosity more than 59 % and pore size ranging from 100 to 500 μm is similar to that of dry human trabecular bone. The obtained composite scaffolds with 30 wt% and 40 wt% HA have a compressive modulus of 232 to 443 MPa, and a compressive strength of 9.3 to 9.8 MPa, similar to or a little higher than those of trabecular bone, and close to those of the cancellous bone.

Polymeric nanoparticles (NPs) have had much focus on their ability to penetrate deep pulmonary structures as potential drug carriers. However, research on the toxicological effects of NPs is in its infancy, and interaction mechanisms are largely unknown. Studies have shown that the interactions with pulmonary structures are heavily dependent on the physicochemical properties of the NPs. Here, we studied how hydrophobicity of polymeric NPs affect pulmonary surfactant biophysics in vitro. We investigated a naturally derived lung surfactant, Infasurf, mixed with three polymeric NPs with varying hydrophobicities through the use of a Langmuir trough and atomic force microscopy to probe the intricacies at the air–water interface. In addition, a novel technique, constrained drop surfactometer (CDS), was used to gain insight on how NPs affect surfactant under physiological conditions. We found that the CDS can be used as a sensitive precautionary tool for identifying surfactant inhibition by NPs. Our data suggest...

Nanoparticles (NPs) show great promises in biomedical applications as the respiratory drug carrier system. Once reaching the alveolar region, NPs first interact with the pulmonary surfactant (PS) film, which serves as the first biological barrier and plays an important role in maintaining the normal respiratory mechanics. Therefore, understanding the interactions between NPs and PS can help promote the NP-based respiratory drug carrier systems. Using coarse-grained molecular dynamics simulations, we studied the effect of rigid spherical NPs with different hydrophobicity and sizes on a dipalmitoylphosphatidylcholine (DPPC) monolayer at the air-water interface. Four different NPs were considered, including hydrophilic and hydrophobic NPs, each with two diameters of 3 nm and 5 nm (the sizes are comparable to that of generation 3 and 5 PAMAM dendrimers, which have been widely used for nanoscale drug carrier systems). Our simulations showed that hydrophilic NPs can readily penetrate into the aqueous phase with little or no disturbance on the DPPC monolayer. However, hydrophobic NPs tend to induce large structural disruptions, thus inhibiting the normal phase transition of the DPPC monolayer upon film compression. Our simulations also showed that this inhibitory effect of hydrophobic NPs can be mitigated through PEGylation. Our results provide useful guidelines for molecular design of NPs as carrier systems for pulmonary drug delivery.

Interaction with the pulmonary surfactant film, being the first line of host defense, represents the initial bio-nano interaction in the lungs. Such interaction determines the fate of the inhaled nanoparticles and their potential therapeutic or toxicological effect. Despite considerable progress in optimizing physicochemical properties of nanoparticles for improved delivery and targeting, the mechanisms by which inhaled nanoparticles interact with the pulmonary surfactant film are still largely unknown. Here, using combined in vitro and in silico methods, we show how hydrophobicity and surface charge of nanoparticles differentially regulate the translocation and interaction with the pulmonary surfactant film. While hydrophilic nanoparticles generally translocate quickly across the pulmonary surfactant film, a significant portion of hydrophobic nanoparticles are trapped by the surfactant film and encapsulated in lipid protrusions upon film compression. Our results support a novel model of pulmonary surfactant lipoprotein corona associated with inhaled nanoparticles of different physicochemical properties. Our data suggest that the study of pulmonary nanotoxicology and nanoparticle-based pulmonary drug delivery should consider this lipoprotein corona.

A self-assembled phospholipid monolayer at an air–water interface is a well-defined model system for studying surface thermodynamics, membrane biophysics, thin-film materials, and colloidal soft matter. Here we report a study of two-dimensional phase transitions in the dipalmitoylphosphatidylcholine (DPPC) monolayer at the air–water interface using a newly developed methodology called constrained drop surfactometry (CDS). CDS is superior to the classical Langmuir balance in its capacity for rigorous temperature control and leak-proof environments, thus making it an ideal alternative to the Langmuir balance for studying lipid polymorphism. In addition, we have developed a novel Langmuir–Blodgett (LB) transfer technique that allows the direct transfer of lipid monolayers from the droplet surface under well-controlled conditions. This LB transfer technique permits the direct visualization of phase coexistence in the DPPC monolayer. With these technological advances, we found that the two-dimensional phase behavior of the DPPC monolayer is analogous to the three-dimensional phase transition of a pure substance. This study has implications in the fundamental understanding of surface thermodynamics as well as applications such as self-assembled monolayers and pulmonary surfactant biophysics.

Droplet manipulation plays an important role in a wide range of scientific and industrial applications, such as synthesis of thin-film materials, control of interfacial reactions, and operation of digital microfluidics. Compared to micron-sized droplets, which are commonly considered as spherical beads, millimeter-sized droplets are generally deformable by gravity, thus introducing nonlinearity into control of droplet properties. Such a nonlinear drop shape effect is especially crucial for droplet manipulation, even for small droplets, at the presence of surfactants. In this paper, we have developed a novel closed-loop axisymmetric drop shape analysis (ADSA), integrated into a constrained drop surfactometer (CDS), for manipulating millimeter-sized droplets. The closed-loop ADSA generalizes applications of the traditional drop shape analysis from a surface tension measurement methodology to a sophisticated tool for manipulating droplets in real time. We have demonstrated the feasibility and advantages of the closed-loop ADSA in three applications, including control of drop volume by automatically compensating natural evaporation, precise control of surface area variations for high-fidelity biophysical simulations of natural pulmonary surfactant, and steady control of surface pressure for in situ Langmuir–Blodgett transfer from droplets. All these applications have demonstrated the accuracy, versatility, applicability, and automation of this new ADSA-based droplet manipulation technique. Combining with CDS, the closed-loop ADSA holds great promise for advancing droplet manipulation in a variety of material and surface science applications, such as thin-film fabrication, self-assembly, and biophysical study of pulmonary surfactant.

Corticosteroids have been widely used in clinical medicine as a first-line therapy to modify the inflammatory response in many pulmonary and systemic diseases. Inhaled and intratracheally administered corticosteroids have a particular interest in that their use allows the clinician to circumvent systemic steroid side effects. However, it is vital that corticosteroids delivered via the lungs not interfere with surface activity of the pulmonary surfactant lining layer.We found differential effects of cholesterol and budesonide on the biophysical properties of a cholesterol-free clinical surfactant preparation, Curosurf. At a low concentration up to 1%, both steroids play a similar role of fluidizing the surfactant film. However, when steroid concentration is increased to 10%, cholesterol induces a unique phase transition that abolishes the surface activity of the Curosurf film. By contrast, 10% budesonide simply fluidizes the film, thus having only limited effects on surface activity.Together with those of a previous study using a cholesterol-containing surfactant, our findings suggest that cholesterol-free surfactant preparations may be more advantageous than cholesterol-containing preparations as a carrier of budesonide because a larger amount of the drug may be delivered to the lungs without significantly compromising the surface activity of pulmonary surfactant.Langmuir balance was used to study the effect of cholesterol and budesonide added at different concentrations on surface activity of Curosurf. Atomic force microscopy (AFM) was used to reveal their effects on the interfacial molecular organization and lateral structure of Curosurf films.

Pulmonary surfactant facilitates breathing by forming a surface tension reducing film at the air–liquid interface of the alveoli. The objective was to characterize the structure of surfactant films using endogenous rat surfactant. Solid-support surfactant films, at different surface pressures, were obtained using a Langmuir balance and were analyzed using atomic force microscopy. The results showed a lipid film structure with three distinct phases: liquid expanded, liquid ordered and liquid condensed. The area covered by the liquid condensed domains increased as surface pressure increased. The presence of liquid ordered phase within these structures correlated with the cholesterol content. At a surface pressure of 50 mN/m, stacks of bilayers appeared. Several structural details of these films differ from previous observations made with goat and exogenous surfactants. Overall, the data indicate that surfactant films demonstrate phase separation at low surface pressures and multilayer formation at higher pressure, features likely important for normal surfactant function.

Clinical pulmonary surfactant is routinely used to treat premature newborns with respiratory distress syndrome, and has shown great potential in alleviating a number of neonatal and adult respiratory diseases. Despite extensive study of chemical composition, surface activity, and clinical performance of various surfactant preparations, a direct comparison of surfactant films is still lacking. In this study, we use atomic force microscopy to characterize and compare four animal-derived clinical surfactants currently used throughout the world, i.e., Survanta, Curosurf, Infasurf and BLES. These modified-natural surfactants are further compared to dipalmitoyl phosphatidylcholine (DPPC), a synthetic model surfactant of DPPC:palmitoyl-oleoyl phosphatidylglycerol (POPG) (7:3), and endogenous bovine natural surfactant. Atomic force microscopy reveals significant differences in the lateral structure and molecular organization of these surfactant preparations. These differences are discussed in terms of DPPC and cholesterol contents. We conclude that all animal-derived clinical surfactants assume a similar structure of multilayers of fluid phospholipids closely attached to an interfacial monolayer enriched in DPPC, at physiologically relevant surface pressures. This study provides the first comprehensive survey of the lateral structure of clinical surfactants at various surface pressures. It may have clinical implications on future application and development of surfactant preparations.

Natural lung surfactant contains less than 40% disaturated phospholipids, mainly dipalmitoylphosphatidylcholine (DPPC). The mechanism by which lung surfactant achieves very low near-zero surface tensions, well below its equilibrium value, is not fully understood. To date, the low surface tension of lung surfactant is usually explained by a squeeze-out model which predicts that upon film compression non-DPPC components are gradually excluded from the air-water interface into a surface-associated surfactant reservoir. However, detailed experimental evidence of the squeeze-out within the physiologically relevant high surface pressure range is still lacking. In the present work, we studied four animal-derived clinical surfactant preparations, including Survanta, Curosurf, Infasurf, and BLES. By comparing compression isotherms and lateral structures of these surfactant films obtained by atomic force microscopy within the physiologically relevant high surface pressure range, we have derived an updated squeeze-out model. Our model suggests that the squeeze-out originates from fluid phases of a phase-separated monolayer. The squeeze-out process follows a nucleation-growth model and only occurs within a narrow surface pressure range around the equilibrium spreading pressure of lung surfactant. After the squeeze-out, three-dimensional nuclei stop growing, thereby resulting in a DPPC-enriched interfacial monolayer to reduce the air-water surface tension to very low values.

Current therapies for nerve regeneration within injured tissues have had limited success due to complicated neural anatomy and inhibitory barriers in situ. Recent advancements in 3D bioprinting technologies have enabled researchers to develop novel 3D scaffolds with complex architectures in an effort to mitigate the challenges that beset reliable and defined neural tissue regeneration. Among several possible neuroregenerative treatment approaches that are being explored today, 3D bioprinted scaffolds have the unique advantage of being highly modifiable, which promotes greater resemblance to the native biological architecture of in vivo systems. This high architectural similarity between printed constructs and in vivo structures is thought to facilitate a greater capacity for repair of damaged nerve tissues. In this review, advances of several 3D bioprinting methods are introduced, including laser bioprinting, inkjet bioprinting, and extrusion-based printing. In addition, the emergence of 4D printing is discussed, which adds a dimension of transformation over time to traditional 3D printing. Finally, an overview of emerging trends in advanced bioprinting materials is provided and their therapeutic potential for application in neural tissue regeneration is evaluated in both the central nervous system and the peripheral nervous system.

Pulmonary surfactant (PS) dysfunction because of the leakage of serum proteins into the alveolar space could be an operative pathogenesis in acute respiratory distress syndrome. Albumin-inhibited PS is a commonly used in vitro model for studying surfactant abnormality in acute respiratory distress syndrome. However, the mechanism by which PS is inhibited by albumin remains controversial. This study investigated the film organization of albumin-inhibited bovine lipid extract surfactant (BLES) with and without surfactant protein A (SP-A), using atomic force microscopy. The BLES and albumin (1:4w/w) were cospread at an air-water interface from aqueous media. Cospreading minimized the adsorption barrier for phospholipid vesicles imposed by preadsorbed albumin molecules, i.e., inhibition because of competitive adsorption. Atomic force microscopy revealed distinct variations in film organization, persisting up to 40mN/m, compared with pure BLES monolayers. Fluorescence confocal microscopy confirmed that albumin remained within the liquid-expanded phase of the monolayer at surface pressures higher than the equilibrium surface pressure of albumin. The remaining albumin mixed with the BLES monolayer so as to increase film compressibility. Such an inhibitory effect could not be relieved by repeated compression-expansion cycles or by adding surfactant protein A. These experimental data indicate a new mechanism of surfactant inhibition by serum proteins, complementing the traditional competitive adsorption mechanism.

Monolayers of a functional pulmonary surfactant (PS) can reach very low surface tensions well below their equilibrium value. The mechanism by which PS monolayers reach such low surface tensions and maintain film stability remains unknown. As shown previously by fluorescence microscopy, phospholipid phase transition and separation seem to be important for the normal biophysical properties of PS. This work studied phospholipid phase transitions and separations in monolayers of bovine lipid extract surfactant using atomic force microscopy. Atomic force microscopy showed phospholipid phase separation on film compression and a monolayer-to-multilayer transition at surface pressure 40–50mN/m. The tilted-condensed phase consisted of domains not only on the micrometer scale, as detected previously by fluorescence microscopy, but also on the nanometer scale, which is below the resolution limits of conventional optical methods. The nanodomains were embedded uniformly within the liquid-expanded phase. On compression, the microdomains broke up into nanodomains, thereby appearing to contribute to tilted-condensed and liquid-expanded phase remixing. Addition of surfactant protein A altered primarily the nanodomains and promoted the formation of multilayers. We conclude that the nanodomains play a predominant role in affecting the biophysical properties of PS monolayers and the monolayer-to-multilayer transition.

The interaction between a cationic polyelectrolyte, chitosan, and an exogenous bovine lung extract surfactant (BLES) was studied using dynamic compression/expansion cycles of dilute BLES preparations in a Constrained Sessile Drop (CSD) device equipped with an environmental chamber conditioned at 37 °C and 100% R.H. air. Under these conditions, dilute BLES preparations tend to produce variable and relatively high minimum surface tensions. Upon addition of “low” chitosan to BLES ratios, the minimum surface tension of BLES–chitosan preparations were consistently low (i.e. 15 mJ/m2). The zeta potential of the lung surfactant aggregates in the subphase suggests that chitosan binds to the anionic lipids (phosphatidyl glycerols) in BLES, and that this binding is ultimately responsible for the changes in the surface activity (elasticity and stability) of these surfactant–polyelectrolyte mixtures. Furthermore the transition from “low” to “high” chitosan to BLES ratios correlates with the flocculation and de-flocculation of surfactant aggregates in the subphase. It is proposed that the aggregation/segregation of “patches” of anionic lipids in the surfactant monolayer produced at different chitosan to BLES ratios explains the enhancing/inhibitory effects of chitosan. These observations highlight the importance of electrostatic interactions in lung surfactant systems.

Automatic surface tension measurement of turbid liquids using a bubble method (e.g., pendant/captive bubble) or interfacial tension measurement of some liquid–liquid systems necessitates an image analysis scheme robust against noise. Measuring the surface tension of lung surfactant–polymer systems using the combination of axisymmetric drop shape analysis (ADSA) and a captive bubble is such an example. We have recently developed a sophisticated image analysis scheme featuring the combination of the Canny edge detector and a novel edge smoothing technique called axisymmetric liquid fluid interfaces-smoothing (ALFI-S). The Canny edge detector is highly noise-resistant and ALFI-S is efficient in further removing noise close to the bubble profile (i.e., adhering noise). However, the procedure of eliminating noise far away from the bubble profile (i.e., isolated noise) is still not satisfactory in the previous scheme. Here a component labeling method is developed to automatically remove isolated noise. Component labeling refers to the process of detecting connected objects in a digital image. Once labeling is completed, information, such as area, location and pixel intensity, of these separate objects can be obtained. Hence, component labeling can be used as a region-based segmentation technique that is capable of differentiating the bubble and the isolated noise. The new component labeling-based noise reduction method is used to analyze captive bubble images with different amounts of noise. It is found that with the help of the labeling procedure smooth edges can be detected with a simple set of user-specified parameters even from images with extensive noise. Combined with the previous image analysis scheme, the component labeling method significantly improves the effectiveness and reliability of ADSA in automatically measuring surface tension from noisy images.

The surface activity of bovine lipid extracted surfactant (BLES) preparations used in surfactant replacement therapy is studied in dynamic film compression/expansion cycles as a function of relative humidity, surfactant concentration, compression rate, and compression periodicity. BLES droplets were formed in a constrained sessile droplet configuration (CSD). Images obtained during cycling were analyzed using axisymmetric drop shape analysis (ADSA) to yield surface tension, surface area, and drop volume data. The experiments were conducted in a chamber that allowed both humid (100% RH), and "dry" air (i.e. less than 20% RH) environments. It was observed that in humid environments BLES films are not stable and tend to have poor surface activity compared to BLES films exposed to dry air. Further analysis of the data reveal that if BLES films are compressed fast enough (i.e. at physiological conditions) to avoid film hydration, lower minimum surface tensions are achieved. A film hydration-relaxation mechanism is proposed to explain these observations.

Surfactant protein B (SP-B) is essential for lung function. Previous studies have indicated that a SP-B 1580C/T polymorphism (SNP rs1130866) was associated with lung diseases including pneumonia. The SNP causes an altered N-linked glycosylation modification at Asn129 of proSP-B, e.g. the C allele with this glycosylation site but not in the T allele. This study aimed to generate humanized SP-B transgenic mice carrying either SP-B C or T allele without a mouse SP-B background and then examine functional susceptibility to bacterial pneumonia in vivo . A total of 18 transgenic mouse founders were generated by the DNA microinjection method. These founders were back-crossed with SP-B KO mice to eliminate mouse SP-B background. Four founder lines expressing similar SP-B levels to human lung were chosen for further investigation. After intratracheal infection with 50 μl of Pseudomonas aeruginosa solution (1 × 10 6 CFU/mouse) or saline in SP-B-C, SP-B-T mice the mice were sacrificed 24 h post-infection and tissues were harvested. Analysis of surfactant activity revealed differential susceptibility between SP-B-C and SP-B-T mice to bacterial infection, e.g. higher minimum surface tension in infected SP-B-C versus infected SP-B-T mice. These results demonstrate for the first time that human SP-B C allele is more susceptible to bacterial pneumonia than SP-B T allele in vivo .

Graphene oxide (GO) is increasingly used in biomedical applications because it possesses not only the unique properties of graphene including large surface area and flexibility but also hydrophilicity and dispersibility in aqueous solutions. However, there are conflicting results on its biocompatibility and biosafety partially due to large variations in physicochemical properties of GO, and the role of these properties including lateral size in the biological or toxicological effects of GO is still unclear. In this study, we focused on the role of lateral size by preparing a panel of GO samples with differential lateral sizes using the same starting material. We found that, in comparison to its smaller counterpart, larger GO showed a stronger adsorption onto the plasma membrane with less phagocytosis, which elicited more robust interaction with toll-like receptors and more potent activation of NF-κB pathways. By contrast, smaller GO sheets were more likely taken up by cells. As a result, larger GO promoted greater M1 polarization, associated with enhanced production of inflammatory cytokines and recruitment of immune cells. The in vitro results correlated well with local and systemic inflammatory responses after GO administration into the abdominal cavity, lung, or bloodstream through the tail vein. Together, our study delineated the size-dependent M1 induction of macrophages and pro-inflammatory responses of GO in vitro and in vivo. Our data also unearthed the detailed mechanism underlying these effects: a size-dependent interaction between GO and the plasma membrane.

Inhalation of nanoparticles (NP), including lightweight airborne carbonaceous nanomaterials (CNM), poses a direct and systemic health threat to those who handle them. Inhaled NP penetrate deep pulmonary structures in which they first interact with the pulmonary surfactant (PS) lining at the alveolar air-water interface. In spite of many research efforts, there is a gap of knowledge between in vitro biophysical study and in vivo inhalation toxicology since all existing biophysical models handle NP-PS interactions in the liquid phase. This technical limitation, inherent in current in vitro methodologies, makes it impossible to simulate how airborne NP deposit at the PS film and interact with it. Existing in vitro NP-PS studies using liquid-suspended particles have been shown to artificially inflate the no-observed adverse effect level of NP exposure when compared to in vivo inhalation studies and international occupational exposure limits (OELs). Here, we developed an in vitro methodology called the constrained drop surfactometer (CDS) to quantitatively study PS inhibition by airborne CNM. We show that airborne multiwalled carbon nanotubes and graphene nanoplatelets induce a concentration-dependent PS inhibition under physiologically relevant conditions. The CNM aerosol concentrations controlled in the CDS are comparable to those defined in international OELs. Development of the CDS has the potential to advance our understanding of how submicron airborne nanomaterials affect the PS lining of the lung.

In the lungs, oxygen transfer from the inspired air to the capillary blood needs to cross the surfactant lining layer of the alveoli. Therefore, the gas transfer characteristics of lung surfactant film are of fundamental physiological interest. However, previous in vitro studiesmost relied on the Langmuir-type balancefail to cover the low surface tension range (i.e., less than the equilibrium surface tension of approximately 25 mJ/m2) due to film leakage. We have recently developed a novel in vitro experimental strategy, the combination of axisymmetric drop shape analysis and captive bubble technique (ADSA-CB), in studying the effect of surfactant films on interfacial gas transfer (Langmuir 2005, 21, 5446). In the present work, ADSA-CB is used as a micro-film-balance to study the effect of compressed bovine lipid extract surfactant (BLES) films on oxygen transfer. A low surface tension ranging from approximately 25 mJ/m2 to 2 mJ/m2 is studied. The experimental results suggest that lung surfactant films at...

Chitosan is a natural, cationic polysaccharide derived from fully or partially deacetylated chitin. Chitosan is capable of inducing large phospholipid aggregates, closely resembling the function of nonionic polymers tested previously as additives to therapeutic lung surfactants. The effects of chitosan on improving the surface activity of a dilute lung surfactant preparation, bovine lipid extract surfactant (BLES), and on resisting albumin-induced inactivation were studied using a constrained sessile drop (CSD) method. Also studied in parallel were the effects of polyethylene glycol (PEG, 10 kD) and hyaluronan (HA, 1240 kD). Both adsorption and dynamic cycling studies showed that chitosan is able to significantly enhance the surface activity of 0.5 mg/mL BLES and to resist albumin-induced inactivation at an extremely low concentration of 0.05 mg/mL, 1000 times smaller than the usual concentration of PEG and 20 times smaller than HA. Optical microscopy found that chitosan induced large surfactant aggregates even in the presence of albumin. Cytotoxicity tests confirmed that chitosan has no deleterious effect on the viability of lung epithelial cells. The experimental results suggest that chitosan may be a more effective polymeric additive to lung surfactant than the other polymers tested so far.

The in vitro adsorption kinetics of lung surfactant at air-water interfaces is affected by both the composition of the surfactant preparations and the conditions under which the assessment is conducted. Relevant experimental conditions are surfactant concentration, temperature, subphase pH, electrolyte concentration, humidity, and gas composition of the atmosphere exposed to the interface. The effect of humidity on the adsorption kinetics of a therapeutic lung surfactant preparation, bovine lipid extract surfactant (BLES), was studied by measuring the dynamic surface tension (DST). Axisymmetric drop shape analysis (ADSA) was used in conjunction with three different experimental methodologies, i.e., captive bubble (CB), pendant drop (PD), and constrained sessile drop (CSD), to measure the DST. The experimental results obtained from these three methodologies show that for 100% relative humidity (RH) at 37 degrees C the rate of adsorption of BLES at an air-water interface is substantially slower than for low humidity. It is also found that there is a difference in the rate of surface tension decrease measured from the PD and CB/CSD methods. These experimental results agree well with an adsorption model that considers the combined effects of entropic force, electrostatic interaction, and gravity. These findings have implications for the development and evaluation of new formulations for surfactant replacement therapy.

Recent progress in the study of pulmonary surfactant is reviewed. The first half of this paper provides general background in both physiological and clinical perspectives. The second half focuses on the in vitro assessment of pulmonary surfactant using methods based on a drop shape technique, Axisymmetric Drop Shape Analysis (ADSA). Theories, experiments, and techniques of image analysis used in these ADSA methods are briefly described. Typical applications of these methods are discussed in detail. It is concluded that the accuracy, versatility, and simplicity of these ADSA methods render them suitable to the study of pulmonary surfactant.

Axisymmetric drop shape analysis (ADSA) is a method to measure surface tension using drop or bubble profiles. Combining ADSA with a captive bubble configuration (ADSA-CB) facilitates pulmonary surfactant related studies. The accuracy of ADSA-CB is crucially dependent on the quality of the bubble profile extracted from the raw image. In the previous version of ADSA-CB, a global thresholding method was used to segment the bubble profile. However, that technique is of limited accuracy for images with noise and/or lack of contrast. In this paper, a new generation of ADSA-CB using the Canny edge detector was developed. To obtain better results, a novel edge smoothing technique, termed axisymmetric liquid fluid interfaces-smoothing (ALFI-S), was introduced and incorporated with the Canny edge detector to extract bubble profiles. The performance of the new version of ADSA-CB was evaluated using captive bubble images under different conditions. The results suggest that the new methodology is capable of producing accurate surface tension values under a variety of circumstances.

The methodology combining Axisymmetric Drop Shape Analysis (ADSA) with a captive bubble (ADSA-CB) facilitates pulmonary surfactant related studies. The accuracy of ADSA-CB is crucially dependent on the quality of the bubble profile extracted from the raw image. In a previous paper, an image analysis scheme featuring a Canny edge detector and a Axisymmetric Liquid Fluid Interfaces-Smoothing (ALFI-S) algorithm was developed to process captive bubble images under a variety of conditions, including images with extensive noise and/or lack of contrast. A new version of ADSA-CB based on that image analysis scheme is developed and applied to pulmonary surfactant and pulmonary surfactant-polymer systems. The new version is found to be highly noise-resistant and well self-adjusting.

The hot/cold rolled steel coil can be considered as a periodically laminated material composed of layers of steel strips and gas gaps in the radial direction. The conduction of steel, of gas, of contact points due to the surface roughness, as well as radiation have been included in a determination of the radial effective thermal conductivity. Based on the analysis of heat transfer mechanisms in radial coils, a new formula for the radial effective thermal conductivity has been derived, which depends not only on the temperature but also on the type of atmosphere gas, the surface characteristic of coils, strip thickness, and compressive stress. Using this effective conductivity, a detailed mathematical model has been developed to predict the temperature distribution of coils in a high performance hydrogen (HPH) furnace. The calculated annealing curves are in good agreement with experimental data.

The impact of substratum surface property change on biofilm community structure was investigated using laboratory biological aerated filter (BAF) reactors and molecular microbial community analysis. Two substratum surfaces that differed in surface properties were created via surface coating and used to develop biofilms in test (modified surface) and control (original surface) BAF reactors. Microbial community analysis by 16S rRNA gene-based PCR-denaturing gradient gel electrophoresis (DGGE) showed that the surface property change consistently resulted in distinct profiles of microbial populations during replicate reactor start-ups. Pyrosequencing of the bar-coded 16S rRNA gene amplicons surveyed more than 90% of the microbial diversity in the microbial communities and identified 72 unique bacterial species within 19 bacterial orders. Among the 19 orders of bacteria detected, Burkholderiales and Rhodocyclales of the Betaproteobacteria class were numerically dominant and accounted for 90.5 to 97.4% of the sequence reads, and their relative abundances in the test and control BAF reactors were different in consistent patterns during the two reactor start-ups. Three of the five dominant bacterial species also showed consistent relative abundance changes between the test and control BAF reactors. The different biofilm microbial communities led to different treatment efficiencies, with consistently higher total organic carbon (TOC) removal in the test reactor than in the control reactor. Further understanding of how surface properties affect biofilm microbial communities and functional performance would enable the rational design of new generations of substrata for the improvement of biofilm-based biological treatment processes.

The mechanisms of heat transfer in high-performance hydrogen bell-type annealing furnaces are discussed in this paper. It examines two important parameters, the convective heat transfer coefficient and the equivalent radial thermal conductivity, which have large effects on the overall heat transfer in the furnace. The calculated annealing curves are in good agreement with experimental data. © 2001 Scripta Technica, Heat Trans Asian Res, 30(8): 615–623, 2001

The influences of algal cell size and surface charge on rheological properties of microalgae suspensions were investigated. The effective viscosity of two microalgae suspensions, i.e., the freshwater Chlorella sp . and the marine Chlorella sp . , was measured as a function of their volume fractions in the range of 0.70–4.31%. The hydrodynamic diameters of the freshwater Chlorella sp . and the marine Chlorella sp. were measured to be 3.13 and 6.00 μm, respectively. The Zeta potentials of these two algal cells were measured to be −23.73 and −81.81 mV, respectively. The intrinsic viscosities of these two microalgae suspensions were further determined to be 24.7 and 16.1, respectively. Combining with theoretical models, these results indicated that the algal cell size has a predominant effect over cell surface charge in affecting rheological properties of microalgae suspensions. Smaller algal cells result in a higher effective viscosity of the microalgae suspension.

Intratracheal administration of corticosteroids using a natural pulmonary surfactant as a delivery vehicle has recently received significant attention in hopes of treating premature newborns with or at high risk for chronic lung disease. As a new practice, both the surfactant preparation used as the carrier and the corticosteroid delivered as the anti-inflammatory agent, and their mixing ratios, have not been standardized and optimized. Given the concern that corticosteroids delivered via a pulmonary surfactant may compromise its surface activity and thus worsen lung mechanics, the present study was carried out to characterize the biophysical interaction between a natural surfactant preparation, Infasurf, and two commonly used inhaled corticosteroids, budesonide and beclomethasone dipropionate (BDP). Based on surface activity measurements by the Langmuir balance and lateral film structure studied by atomic force microscopy, our findings suggest that when Infasurf is used as a carrier, a budesonide concentration less than 1 wt% of surfactant or a BDP concentration up to 10 wt % should not significantly affect the biophysical properties of Infasurf, thus being feasible for pulmonary delivery. Increasing corticosteroid concentration beyond this range leads to early collapse of the surfactant film due to increased film fluidization. Our study further suggests that different affinities to the surfactant films are responsible for the different behavior of budesonide and BDP. In addition to the translational value in treating chronic lung disease, this study may also have implications in inhaled steroid therapy to treat asthma.

article i nfo The exact mechanism by which pulmonary surfactant films reach the very low surface tensions required to stabilize the alveoli at end expiration remains uncertain. We utilized the nanoscale sensitivity of atomic force microscopy (AFM) to examine phospholipid (PL) phase transition and multilayer formation for two Langmuir-Blodgett (LB) systems: a simple 3 PL surfactant-like mixture and the more complex bovine lipid extract surfactant (BLES). AFM height images demonstrated that both systems develop two types of liquid condensed (LC) domains (micro- and nano-sized) within a liquid expanded phase (LE). The 3 PL mixture failed to form significant multilayers at high surface pressure (π while BLES forms an extensive network of multilayer structures containing up to three bilayers. A close examination of the progression of multilayer formation reveals that multilayers start to form at the edge of the solid-like LC domains and also in the fluid-like LE phase. We used the elemental analysis capability of time-of-flight secondary ion mass spectrom- etry (ToF-SIMS) to show that multilayer structures are enriched in unsaturated PLs while the saturated PLs are concentrated in the remaining interfacial monolayer. This supports a modified squeeze-out model where film compression results in the hydrophobic surfactant protein-dependent formation of unsaturated PL-rich multilayers which remain functionally associated with a monolayer enriched in disaturated PL spe- cies. This allows the surface film to attain low surface tensions during compression and maintain values near equilibrium during expansion.

Inhaled nanoparticles (NPs) must first interact with the pulmonary surfactant (PS) lining layer that covers the entire internal surface of the respiratory tract and plays an important role in surface tension reduction and host defense. Interactions with the PS film determine the subsequent clearance, retention, and translocation of the inhaled NPs and hence their potential toxicity. To date, little is known how NPs interact with PS, and whether or not NPs have adverse effects on the biophysical function of PS. We found a time-dependent toxicological effect of hydroxyapatite NPs (HA-NPs) on a natural PS, Infasurf, and the time scale of surfactant inhibition after particle exposure was comparable to the turnover period of surfactant metabolism. Using a variety of in vitro biophysicochemical characterization techniques, we have determined the inhibition mechanism to be due to protein adsorption onto the HA-NPs. Consequently, depletion of surfactant proteins from phospholipid vesicles caused conversion of original large vesicles into much smaller vesicles with poor surface activity. These small vesicles, in turn, inhibited biophysical function of surfactant films after adsorption at the air-water interface. Cytotoxicity study found that the HA-NPs at the studied concentration were benign to human bronchial epithelial cells, thereby highlighting the importance of evaluating biophysical effect of NPs on PS. The NP-PS interaction mechanism revealed by this study may not only provide new insight into the toxicological study of nanoparticles but also shed light on the feasibility of NP-based pulmonary drug delivery.

The generalized theory of capillarity thermodynamics of axisymmetric capillary systems thermodynamic status of contact angles line tension in multiphase equilibrium systems the equation of state approach to interfacial tensions the theory of surface tension components and the equation of state approach theoretical approaches for estimating solid-liquid interfacial tensions contact angle and liquid surface tension measurements - general procedures and techniques capillary rise at a vertical plate as a contact angle technique axisymmetric drop shape analysis (ADSA) wettability and surface tension of particles behaviour of particles at solidification fronts.

This article examines the manner in which some new methodologies and novel concepts have contributed to our understanding of how pulmonary surfactant reduces alveolar surface tension. Investigations utilizing small angle X-ray diffraction, inverted interface fluorescence microscopy, time of flight-secondary ion mass spectroscopy, atomic force microscopy, two-photon fluorescence microscopy and electrospray mass spectroscopy are highlighted and a new model of ventilation-induced acute lung injury described. This contribution attempts to emphasize how these new approaches have resulted in a fuller appreciation of events presumably occurring at the alveolar interface.

Recent studies employing a variety of microscopic techniques have revealed that phospholipid (PL) phase separations and transitions may play important roles in determining the biophysical properties of pulmonary surfactant. Most of these microscopic studies used model systems which were composed of simple mixtures of PL with or without hydrophobic surfactant proteins and cholesterol. The present work compared modified natural (lipid extract) bovine and rat surfactants using atomic force microscopy (AFM). AFM revealed PL phase separation upon compression of both surfactant monolayers, and a monolayer-to-multilayer transition at surface pressure 40-50 mN/m. Similar to bovine surfactant, the tilted-condensed (TC) phase in rat surfactant consisted of domains both on micrometer and nanometer scales. Upon film compression, the microdomains were dissociated into nanodomains, thus forming a more homogeneous two-phase mixture. Differences between rat and bovine surfactants were: (1) more TC domains were formed at lower surface pressures in rat than in bovine surfactant; and (2) an interesting domain-in-domain structure was exclusively observed in rat surfactant. These structural differences were attributed to the higher cholesterol content of rat surfactant (∼ 10 vs ∼2.5 wt%). To further investigate the effects of cholesterol on the structure of surfactant films, we have studied cholesterol-depleted bovine surfactant (∼ 0%) prepared by repetitive acetone extraction. Removal of cholesterol from bovine surfactant induced significant variations in film structure. More importantly, the film structure can be effectively restored by recombining cholesterol with the cholesterol-depleted bovine surfactant. Recombinant bovine surfactant with 10% cholesterol showed domain-in-domain structures similar to those found with rat surfactant. These interspecies studies of the micro- and nano- structures of natural pulmonary surfactants add insight into the biophysical interpretation of phospholipid phase transition and separation, in particular the role of cholesterol.

Pulmonary surfactant (PS) is a complicated mixture of approximately 90% lipids and 10% proteins. It plays an important role in maintaining normal respiratory mechanics by reducing alveolar surface tension to near-zero values. Supplementing exogenous surfactant to newborns suffering from respiratory distress syndrome (RDS), a leading cause of perinatal mortality, has completely altered neonatal care in industrialized countries. Surfactant therapy has also been applied to the acute respiratory distress syndrome (ARDS) but with only limited success. Biophysical studies suggest that surfactant inhibition is partially responsible for this unsatisfactory performance. This paper reviews the biophysical properties of functional and dysfunctional PS. The biophysical properties of PS are further limited to surface activity, i.e., properties related to highly dynamic and very low surface tensions. Three main perspectives are reviewed. (1) How does PS permit both rapid adsorption and the ability to reach very low surface tensions? (2) How is PS inactivated by different inhibitory substances and how can this inhibition be counteracted? A recent research focus of using water-soluble polymers as additives to enhance the surface activity of clinical PS and to overcome inhibition is extensively discussed. (3) Which in vivo, in situ, and in vitro methods are available for evaluating the surface activity of PS and what are their relative merits? A better understanding of the biophysical properties of functional and dysfunctional PS is important for the further development of surfactant therapy, especially for its potential application in ARDS.

An easy-to-use noninvasive method was developed to simultaneously measure the thermophysical parameters and blood perfusion in cylindrically shaped living tissues. This method is based on a two-dimensional mathematical model which requires temperature measurements at only three separate points along the axial direction on the cylinder surface. A sensitivity analysis has shown that the key thermophysical parameters, such as the thermal conductivity, volumetric heat capacity, and blood perfusion can be estimated simultaneously with high accuracy. Genetic algorithm (GA) selection, crossover, and mutation operators were developed to solve this multi-parameter optimization problem. This three-point method was validated by measuring the properties of a dynamic tissue-equivalent phantom with known thermal parameters. The method has also been applied to measure the thermophysical parameters and blood perfusion in human forearms with measured results agreeing well with the literature values.

In the lungs, oxygen transfer from the inspired air to the capillary blood needs to cross the surfactant lining layer of the alveoli. Therefore, the gas transfer characteristics of lung surfactant film are of fundamental physiological interest. However, previous in vitro studies-most relied on the Langmuir-type balance-fail to cover the low surface tension range (i.e., less than the equilibrium surface tension of approximately 25 mJ/m2) due to film leakage. We have recently developed a novel in vitro experimental strategy, the combination of axisymmetric drop shape analysis and captive bubble technique (ADSA-CB), in studying the effect of surfactant films on interfacial gas transfer (Langmuir 2005, 21, 5446). In the present work, ADSA-CB is used as a micro-film-balance to study the effect of compressed bovine lipid extract surfactant (BLES) films on oxygen transfer. A low surface tension ranging from approximately 25 mJ/m2 to 2 mJ/m2 is studied. The experimental results suggest that lung surfactant films at a low surface tension near 2 mJ/m2 provide resistance to oxygen transfer, as indicated by a decrease of 30-50% in the mass transfer coefficient (kL) of oxygen in BLES suspensions with respect to water. At higher surface tension (i.e.,6 mJ/m2), the resistance to oxygen transfer is only modest, i.e., the decrease in kL is less than 20% compared to water. The experimental results suggest that lung surfactant plays a role in oxygen transfer in the pulmonary system.

A new method combining axisymmetric drop shape analysis (ADSA) and a captive bubble (CB) is proposed to study the effect of surfactant on interfacial gas transfer. In this method, gas transfer from a static CB to the surrounding quiescent liquid is continuously recorded for a short period (i.e., 5 min). By photographical analysis, ADSA-CB is capable of yielding detailed information pertinent to the surface tension and geometry of the CB, e.g., bubble area, volume, curvature at the apex, and the contact radius and height of the bubble. A steady-state mass transfer model is established to evaluate the mass transfer coefficient on the basis of the output of ADSA-CB. In this way, we are able to develop a working prototype capable of simultaneously measuring dynamic surface tension and interfacial gas transfer. Other advantages of this method are that it allows for the study of very low surface tensions (

Shortage or malfunction of pulmonary surfactant in alveolar space leads to a critical condition termed respiratory distress syndrome (RDS). Surfactant replacement therapy, the major method to treat RDS, is an expensive treatment. In this paper, the effect of poly(ethylene glycol) (PEG) to improve dynamic surface activity of a bovine lipid extract surfactant (BLES) was studied by axisymmetric drop shape analysis (ADSA) and a captive bubble method. The activity of BLES + PEG mixtures was compared to that of a natural surfactant containing surfactant proteins A and D. When PEG was added into BLES mixtures, the surface tension hysteresis of BLES films was minimized when the films were compressed by more than 50%. PEG also helps to quickly restore surfactant films after film collapse. Thus, as far as surface tension effects go, the findings suggest that PEG might be used as a substitute for surfactant-associated protein SP-A in therapeutic surfactant products, and might also be used to reduce the amount of BLES required in clinical applications.