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In spite of the considerable efforts invested to understand the environmental health and safety (EHS) impacts of ultrafine particles, such as the representative PM2.5, there are still significant knowledge gaps to be filled. No conclusive understandings have been obtained about the physicochemical determinants in accounting for differential adverse outcomes. Here we compared the cytotoxicity of four carbon black (CB) particles with similar physicochemical properties except for their oxygen contents (C824455 < C1864 < Printex U < SB4A). We found that these four CB particles manifested in vitro and in vivo cytotoxicity reversely related to their oxygen contents, namely a hierarchy of cytotoxicity: C824455 > C1864 > Printex U > SB4A. Among these CB particles, the most significant lung injury (e.g. collapses and inflammation) and macrophagic activation were found for C824455 and C1864, in particular for C824455. All these differences in toxicity profiles, including in vitro and in vivo cytotoxicity, pro-inflammatory effects and direct damages to the lung epithelia, should be (at least partially) ascribed to the oxygen content in these CB particles that in turn determined their transformation, i.e. the different aggregation states. Nonetheless, PM2.5 likewise caused severe in vivo and in vitro toxicities to the lung cells and macrophages. This study thus offers more insights into the structure-activity relationship (SAR) and opens a new avenue to elucidate the physicochemical determinants in evoking lung injuries by ultrafine airborne particles., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Amyloid formation of proteins is not only relevant for neurodegenerative diseases, but has recently emerged as a groundbreaking approach in materials science and biotechnology. However, amyloid aggregation of proteins in vitro generally requires a long incubation time under extremely harsh conditions, and the understanding of the structural motif to determine amyloid assembly is extremely limited. Herein we reveal that the integration of three important building blocks in typical globular proteins is crucial for superfast protein amyloid-like assembly including the segment required for high fibrillation propensity, abundant α-helix structures and intramolecular S-S bonds to lock the α-helix. With the reduction of the S-S bond by tris(2-carboxyethyl)phosphine (TCEP), the α-helix was rapidly unlocked from the protein chain, and the resultant unfolded monomer underwent a fast transition to β-sheet-rich amyloid oligomers and protofibrils in minutes, which further assembled into a macroscopic nanofilm at the air/water interface and microparticles in bulk solution, respectively.

P-glycoprotein [P-gp or the ATP-binding cassette transporter B1 (ABCB1)] is an important participant in multidrug resistance of cancer cells, yet the precise function of this arthropod transporter is unknown. The aim of this study was to determine the importance of P-gp for susceptibility to insecticides in the beet armyworm (Spodoptera exigua) using clustered regularly interspaced short palindromic repeats/CRISPR-associated 9 (CRISPR/Cas9) gene-editing technology. We cloned an open reading frame (ORF) encoding the S. exigua P-gp protein (SeP-gp) predicted to display structural characteristics common to P-gp and other insect ABCB1 transporters. A knockout line with a frame shift deletion of four nucleotides in the SeP-gp ORF was established using the CRISPR/Cas9 gene-editing system to test its potential role in determining susceptibility to chemical insecticides or insecticidal proteins from the bacterium Bacillus thuringiensis (Bt). Results from comparative bioassays demonstrate that knockout of SeP-gp significantly increases susceptibility of S. exigua by around threefold to abamectin and emamectin benzoate (EB), but not to spinosad, chlorfenapyr, beta-cypermethrin, carbosulfan indoxacarb, chlorpyrifos, phoxim, diafenthiuron, chlorfluazuron, chlorantraniliprole or two Bt toxins (Cry1Ca and Cry1Fa). Our data support an important role for SeP-gp in susceptibility of S. exigua to abamectin and EB and imply that overexpression of SeP-gp may contribute to abamectin and EB resistance in S. exigua., (© 2017 The Royal Entomological Society.)

Objective: Nanomaterials, such as carbon nanotubes (CNTs), have been introduced to modify the surface properties of scaffolds, thus enhancing the interaction between the neural cells and biomaterials. In addition to superior electrical conductivity, CNTs can provide nanoscale structures similar to those present in the natural neural environment. The primary objective of this study is to investigate the proliferative capability and differential potential of neural stem cells (NSCs) seeded on a CNT incorporated scaffold., Approach: Amine functionalized multi-walled carbon nanotubes (MWCNTs) were incorporated with a PEGDA polymer to provide enhanced electrical properties as well as nanofeatures on the surface of the scaffold. A stereolithography 3D printer was employed to fabricate a well-dispersed MWCNT-hydrogel composite neural scaffold with a tunable porous structure. 3D printing allows easy fabrication of complex 3D scaffolds with extremely intricate microarchitectures and controlled porosity., Main Results: Our results showed that MWCNT-incorporated scaffolds promoted neural stem cell proliferation and early neuronal differentiation when compared to those scaffolds without the MWCNTs. Furthermore, biphasic pulse stimulation with 500 µA current promoted neuronal maturity quantified through protein expression analysis by quantitative polymerase chain reaction., Significance: Results of this study demonstrated that an electroconductive MWCNT scaffold, coupled with electrical stimulation, may have a synergistic effect on promoting neurite outgrowth for therapeutic application in nerve regeneration.

Private consumers and professionals may experience acute inhalation toxicity after inhaling aerosolized impregnation products. The distinction between toxic and non-toxic products is difficult to make for producers and product users alike, as there is no clearly described relationship between the chemical composition of the products and induction of toxicity. The currently accepted method for determination of acute inhalation toxicity is based on experiments on animals; it is time-consuming, expensive and causes stress for the animals. Impregnation products are present on the market in large numbers and amounts and exhibit great variety. Therefore, an alternative method to screen for acute inhalation toxicity is needed. The aim of our study was to determine if inhibition of lung surfactant by impregnation products in vitro could accurately predict toxicity in vivo in mice. We tested 21 impregnation products using the constant flow through set-up of the constrained drop surfactometer to determine if the products inhibited surfactant function or not. The same products were tested in a mouse inhalation bioassay to determine their toxicity in vivo. The sensitivity was 100%, i.e., the in vitro method predicted all the products that were toxic for mice to inhale. The specificity of the in vitro test was 63%, i.e., the in vitro method found three false positives in the 21 tested products. Six of the products had been involved in accidental human inhalation where they caused acute inhalation toxicity. All of these six products inhibited lung surfactant function in vitro and were toxic to mice.

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., (Copyright © 2017. Published by Elsevier B.V.)

Axisymmetric drop shape analysis (ADSA) has been used in a broad range of applications for determining surface tensions of air-liquid surfaces and interfacial tensions of liquid-liquid interfaces. However, it is well-known that the accuracy of ADSA deteriorates upon the reduction of drop volume. Here, we systematically compared different criteria and parameters in evaluating the accuracy of ADSA upon reducing drop volume. By scrutinizing the dependence of ADSA accuracy on various parameters, including the capillary constant (c), the Bond number (Bo), the Worthington number (Wo), and the shape parameter (P s ), we concluded that the classical Bond number failed to predict the accuracy of drop shape analysis at very small drop volumes. Thus, we proposed a replacement of the classical Bond number, called the Neumann number Ne ≡ (ΔρgR 0 H)/γ. The design rationale of this new dimensionless number lies in the use of the geometric mean of the radius of curvature at the drop apex (R 0 ) and the drop height (H) as the new characteristic length (L) to represent the drop size, that is, L = (R 0 H) 1/2 . It is found that the Neumann number is capable of evaluating the accuracy of ADSA. Moreover, we have demonstrated the usefulness of the local Neumann number, Ne z ≡ (ΔρgR 0 /γ)z, in evaluating the contribution of the local drop profile to the accuracy of ADSA.

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., (Copyright © 2017 the American Physiological Society.)

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.

The growing risk of human exposure to airborne nanoparticles (NPs) causes a general concern on the biosafety of nanotechnology. Inhaled NPs can deposit in the deep lung at which they interact with the pulmonary surfactant (PS). Despite the increasing study of nano-bio interactions, detailed molecular mechanisms by which inhaled NPs interact with the natural PS system remain unclear. Using coarse-grained molecular dynamics simulation, we studied the interaction between NPs and the PS system in the alveolar fluid. It was found that regardless of different physicochemical properties, upon contacting the PS, both silver and polystyrene NPs are immediately coated with a biomolecular corona that consists of both lipids and proteins. Structure and molecular conformation of the PS corona depend on the hydrophobicity of the pristine NPs. Quantitative analysis revealed that lipid composition of the corona formed on different NPs is relatively conserved and is similar to that of the bulk phase PS. However, relative abundance of the surfactant-associated proteins, SP-A, SP-B, and SP-C, is notably affected by the hydrophobicity of the NP. The PS corona provides the NPs with a physicochemical barrier against the environment, equalizes the hydrophobicity of the pristine NPs, and may enhance biorecognition of the NPs. These modifications in physicochemical properties may play a crucial role in affecting the biological identity of the NPs and hence alter their subsequent interactions with cells and other biological entities. Our results suggest that all studies of inhalation nanotoxicology or NP-based pulmonary drug delivery should consider the influence of the PS corona.

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., (© 2017 American Society of Plant Biologists. All Rights Reserved.)

Desired characteristics of nanocarriers are crucial to explore its therapeutic potential. This investigation aimed to develop tunable bioresponsive newly synthesized unique arginine grafted poly(cystaminebis(acrylamide)-diaminohexane) [ABP] polymeric matrix based nanocarriers by using L9 Taguchi factorial design, desirability function, and multivariate method. The selected formulation and process parameters were ABP concentration, acetone concentration, the volume ratio of acetone to ABP solution, and drug concentration. The measured nanocarrier characteristics were particle size, polydispersity index, zeta potential, and percentage drug loading. Experimental validation of nanocarrier characteristics computed from initially developed predictive model showed nonsignificant differences (p > 0.05). The multivariate modeling based optimized cationic nanocarrier formulation of <100 nm loaded with hydrophilic acetaminophen was readapted for a hydrophobic etoposide loading without significant changes (p > 0.05) except for improved loading percentage. This is the first study focusing on ABP polymeric matrix based nanocarrier development. Nanocarrier particle size was stable in PBS 7.4 for 48 h. The increase of zeta potential at lower pH 6.4, compared to the physiological pH, showed possible endosomal escape capability. The glutathione triggered release at the physiological conditions indicated the competence of cytosolic targeting delivery of the loaded drug from bioresponsive nanocarriers. In conclusion, this unique systematic approach provides rational evaluation and prediction of a tunable bioresponsive ABP based matrix nanocarrier, which was built on selected limited number of smart experimentation.

Acute respiratory distress syndrome (ARDS) is a pulmonary disorder associated with alterations to the pulmonary surfactant system. Recent studies showed that supra-physiological levels of cholesterol in surfactant contribute to impaired function. Since cholesterol is incorporated into surfactant within the alveolar type II cells which derives its cholesterol from serum, it was hypothesized that serum hypercholesterolemia would predispose the host to the development of lung injury due to alterations of cholesterol content in the surfactant system. Wistar rats were randomized to a standard lab diet or a high cholesterol diet for 17-20 days. Animals were then exposed to one of three models of lung injury: i) acid aspiration ii) ventilation induced lung injury, and iii) surfactant depletion. Following physiological monitoring, lungs were lavaged to obtain and analyze the surfactant system. The physiological results showed there was no effect of the high cholesterol diet on the severity of lung injury in any of the three models of injury. There was also no effect of the diet on surfactant cholesterol composition. Rats fed a high cholesterol diet had a significant impairment in surface tension reducing capabilities of isolated surfactant compared to those fed a standard diet exposed to the surfactant depletion injury. In addition, only rats that were exposed to ventilation induced lung injury had elevated levels of surfactant associated cholesterol compared to non-injured rats. It is concluded that serum hypercholesterolemia does not predispose rats to altered surfactant cholesterol composition or to lung injury. Elevated cholesterol within surfactant may be a marker for ventilation induced lung damage.

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.

The acute respiratory distress syndrome (ARDS) is characterized by arterial hypoxemia accompanied by severe inflammation and alterations to the pulmonary surfactant system. Published data has demonstrated a protective effect of matrix metalloproteinase-3 (Mmp3) deficiency against the inflammatory response associated with ARDS; however, the effect of Mmp3 on physiologic parameters and alterations to surfactant have not been previously studied. It was hypothesized that Mmp3 deficient (Mmp3(-/-)) mice would be protected against lung dysfunction associated with ARDS and maintain a functional pulmonary surfactant system. Wild type (WT) and Mmp3(-/-) mice were subjected to acid-aspiration followed by mechanical ventilation. Mmp3(-/-) mice maintained higher arterial oxygenation compared with WT mice at the completion of ventilation. Significant increase in functional large aggregate surfactant forms were observed in Mmp3(-/-) mice compared with WT mice. These findings further support a role of Mmp3 as an attractive therapeutic target for drug development in the setting of ARDS.

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.

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., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Graphene oxide (GO) is the most common derivative of graphene and has been used in a large range of biomedical applications. Despite considerable progress in understanding its cytotoxicity, its potential inhalation toxicity is still largely unknown. As the pulmonary surfactant (PS) film is the first line of host defense, interaction with the PS film determines the fate of the inhaled nanomaterials and their potential toxicity. Using a coarse-grained molecular dynamics model, we reported, for the first time, a novel mechanism of toxicity caused by the inhaled GO nanosheets. Upon deposition, the GO nanosheets induce pores in the PS film and thus have adverse effects on the ultrastructure and biophysical properties of the PS film. Notably, the pores induced by GO nanosheets result in increasing the compressibility of the PS film, which is an important indication of surfactant inhibition. In vitro experiments have also been conducted to study the interactions between GO and animal-derived natural PS films, qualitatively confirming the simulation results.

The time-averaged serum potassium was more comprehensive to reflect the all-time changes of serum potassium levels during peritoneal dialysis (PD). However, the association of fluctuation of time-averaged serum potassium level with long-time survival of PD patients remains unknown. In this retrospective study, we included 357 incident PD patients in 2 centers from January 1, 2007 to October 31, 2012 with follow-up through October 31, 2014. Our data demonstrated that it was the lower time-averaged serum potassium level rather than baseline of serum potassium level that was associated with high risk of death. Patients with higher standard deviation (SD) had significantly poorer all-cause (p = 0.016) and cardiovascular mortality (p = 0.041). Among the patients with time-averaged serum potassium levels below 4.0 mEq/L, a lower mean value was more important than its SD to predict death risk. In contrast, the patients with time-averaged serum potassium levels above 4.0 mEq/L, those with serum potassium SD < 0.54 mEq/L, exhibited a higher 3-year and 5-year survival rate for both all-cause and cardiovascular mortality compared to the control groups. Our data clearly suggested both time-averaged serum potassium and its fluctuation contributed disproportionately to the high death risk in PD patients.

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.

The role of mitochondrial energy metabolism in maintaining lung function is not understood. We previously observed reduced lung function in mice lacking the fatty acid oxidation enzyme long-chain acyl-CoA dehydrogenase (LCAD). Here, we demonstrate that long-chain acylcarnitines, a class of lipids secreted by mitochondria when metabolism is inhibited, accumulate at the air-fluid interface in LCAD(-/-) lungs. Acylcarnitine accumulation is exacerbated by stress such as influenza infection or by dietary supplementation with l-carnitine. Long-chain acylcarnitines co-localize with pulmonary surfactant, a unique film of phospholipids and proteins that reduces surface tension and prevents alveolar collapse during breathing. In vitro, the long-chain species palmitoylcarnitine directly inhibits the surface adsorption of pulmonary surfactant as well as its ability to reduce surface tension. Treatment of LCAD(-/-) mice with mildronate, a drug that inhibits carnitine synthesis, eliminates acylcarnitines and improves lung function. Finally, acylcarnitines are detectable in normal human lavage fluid. Thus, long-chain acylcarnitines may represent a risk factor for lung injury in humans with dysfunctional fatty acid oxidation., (© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.)

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.

Bacterial adhesion onto solid surfaces is of importance in a wide spectrum of problems, including environmental microbiology, biomedical research, and various industrial applications. Despite many research efforts, present thermodynamic models that rely on the evaluation of the adhesion energy are often elusive in predicting the bacterial adhesion behavior. Here, we developed a new spectrophotometric method to determine the surface free energy (SFE) of bacterial cells. The adhesion behaviors of five bacterial species, Pseudomonas putida KT2440, Salmonella Typhimurium ATCC 14028, Staphylococcus epidermidis ATCC 12228, Enterococcus faecalis ATCC 29212, and Escherichia coli DH5α, onto two model substratum surfaces, i.e., clean glass and silanized glass surfaces, were studied. We found that bacterial adhesion was unambiguously mediated by the SFE difference between the bacterial cells and the solid substratum. The lower the SFE difference, the higher degree of bacterial adhesion. We therefore propose the use of the SFE difference as an accurate and simple thermodynamic measure for quantitatively predicting bacterial adhesion. The methodological advance and thermodynamic simplification in the paper have implications in controlling bacterial adhesion and biofilm formation on solid surfaces.

Antibiotic-resistant bacterial infections represent an emerging health concern in clinical settings, and a lack of novel developments in the pharmaceutical pipeline is creating a "perfect storm" for multidrug-resistant bacterial infections. Antimicrobial peptides (AMPs) have been suggested as future therapeutics for these drug-resistant bacteria, since they have potent broad-spectrum activity, with little development of resistance. Due to the unique structure of the lung, bacterial pneumonia has the additional problem of delivering antimicrobials to the site of infection. One potential solution is coadministration of AMPs with exogenous surfactant, allowing for distribution of the peptides to distal airways and opening of collapsed lung regions. The objective of this study was to test various surfactant-AMP mixtures with regard to maintaining pulmonary surfactant biophysical properties and bactericidal functions. We compared the properties of four AMPs (CATH-1, CATH-2, CRAMP, and LL-37) suspended in bovine lipid-extract surfactant (BLES) by assessing surfactant-AMP mixture biophysical and antimicrobial functions. Antimicrobial activity was tested against methillicin-resistant Staphylococcus aureus and Pseudomonas aeruginosa. All AMP/surfactant mixtures exhibited an increase of spreading compared to a BLES control. BLES+CATH-2 mixtures had no significantly different minimum surface tension versus the BLES control. Compared to the other cathelicidins, CATH-2 retained the most bactericidal activity in the presence of BLES. The BLES+CATH-2 mixture appears to be an optimal surfactant-AMP mixture based on in vitro assays. Future directions involve investigating the potential of this mixture in animal models of bacterial pneumonia., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

The interactions with the pulmonary surfactant, the initial biological barrier of respiratory pathway, determine the potential therapeutic applications and toxicological effects of inhaled nanoparticles (NPs). Although much attention has been paid to optimize the physicochemical properties of NPs for improved delivery and targeting, shape effects of the inhaled NPs on their interactions with the pulmonary surfactant are still far from clear. Here, we studied the shape effects of NPs on their penetration abilities and structural disruptions to the dipalmitoyl-phosphatidylcholine (DPPC) monolayer (being model pulmonary surfactant film) using coarse-grained molecular dynamics simulations. It is found that during the inspiration process (i.e., surfactant film expansion), shape effects are negligible. However, during the expiration process (i.e., surfactant film compression), NPs of different shapes show various penetration abilities and degrees of structural disruptions to the DPPC monolayer. We found that rod-like NPs showed the highest degree of penetration and the smallest side-effects to the DPPC monolayer. Our results may provide a useful insight into the design of NPs for respiratory therapeutics., Competing Interests: Conflict of interest The authors declare that they have no conflict of interest.

Cell sorting is an important screening process in microbiology, biotechnology, and clinical research. Existing methods are mainly based on single-cell analysis as in flow cytometric and microfluidic cell sorters. Here we report a label-free bulk method for sorting cells by differentiating their characteristic surface free energies (SFEs). We demonstrated the feasibility of this method by sorting model binary cell mixtures of various bacterial species, including Pseudomonas putida KT2440, Enterococcus faecalis ATCC 29212, Salmonella Typhimurium ATCC 14028, and Escherichia coli DH5α. This method can effectively separate 10(10) bacterial cells within 30 min. Individual bacterial species can be sorted with up to 96% efficiency, and the cell viability ratio can be as high as 99%. In addition to its capacity of sorting evenly mixed bacterial cells, we demonstrated the feasibility of this method in selecting and enriching cells of minor populations in the mixture (presenting at only 1% in quantity) to a purity as high as 99%. This SFE-activated method may be used as a stand-alone method for quickly sorting a large quantity of bacterial cells or as a prescreening tool for microbial discrimination. Given its advantages of label-free, high-throughput, low cost, and simplicity, this SFE-activated cell sorting method has potential in various applications of sorting cells and abiotic particles.

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.

Long-chain acyl-CoA dehydrogenase (LCAD) is a mitochondrial fatty acid oxidation enzyme whose expression in humans is low or absent in organs known to utilize fatty acids for energy such as heart, muscle, and liver. This study demonstrates localization of LCAD to human alveolar type II pneumocytes, which synthesize and secrete pulmonary surfactant. The physiological role of LCAD and the fatty acid oxidation pathway in lung was subsequently studied using LCAD knock-out mice. Lung fatty acid oxidation was reduced in LCAD(-/-) mice. LCAD(-/-) mice demonstrated reduced pulmonary compliance, but histological examination of lung tissue revealed no obvious signs of inflammation or pathology. The changes in lung mechanics were found to be due to pulmonary surfactant dysfunction. Large aggregate surfactant isolated from LCAD(-/-) mouse lavage fluid had significantly reduced phospholipid content as well as alterations in the acyl chain composition of phosphatidylcholine and phosphatidylglycerol. LCAD(-/-) surfactant demonstrated functional abnormalities when subjected to dynamic compression-expansion cycling on a constrained drop surfactometer. Serum albumin, which has been shown to degrade and inactivate pulmonary surfactant, was significantly increased in LCAD(-/-) lavage fluid, suggesting increased epithelial permeability. Finally, we identified two cases of sudden unexplained infant death where no lung LCAD antigen was detectable. Both infants were homozygous for an amino acid changing polymorphism (K333Q). These findings for the first time identify the fatty acid oxidation pathway and LCAD in particular as factors contributing to the pathophysiology of pulmonary disease.

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.

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.

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.

Objective: To investigate the feasibility and accuracy of bedside ultrasound measuring peak velocity variation of common carotid artery to estimate preload of the patients in surgery ICU., Methods: In this prospective cohort study, SICU patients with sinus rhythm and positive pressure ventilation were included. The peak velocity variation in common carotid artery (delta V peak) during each respiratory circle was measured by ICU resident with short-term training and experienced attending songrapher. Stroke volume before and after the fluid challenge was also measured by the experienced songrapher as the gold standard of fluid responsiveness. Then the ROC, feasibility and accuracy of the diagnosis trial were analyzed., Results: There were 46 patients included. The peak velocity variation of common carotid artery measured by the attending (delta V peak) is highly related with delta SV (r1 = 0.76, P < 0.05). Area under the receiver operating characteristic curve was 0.95 (P < 0.05). The peak velocity variation of common carotid artery (delta V peak) > 12.1% predicted fluid responsiveness with sensitivity of 90.9%, specificity of 83.3%, positive predictive value of 83.3% and negative predictive value of 90.9%. The peak velocity variation of common carotid artery measured by the resident (delta V peak2) were highly related with delta V peak1 (R2 = 0.68, P < 0.05), the AUC was 0.94., Conclusion: Measurement of peak velocity variation of common carotid artery with bedside ultrasound can accurately estimate the volume status of the patients in surgery ICU, and it is easy to be performed by the residents of ICU.

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., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Objective: To investigate the relationship between serum albumin variation within 24 hours of ICU admission and prognosis in critically ill patients with sever acute pancreatitis (SAP)., Methods: We retrospectively analyzed all the clinical data of the patients with SAP treated in ICU from Jul. 2005 to Jun. 2010. The patients were divided into two groups, survival group (n = 131) and death group (n = 108). Clinical and laboratory data, outcomes, and serum albumin within 24 hours after admission to ICU of those patients were evaluated respectively, and Logistic regression analysis was performed., Results: Significant differences were existed between two groups (P < 0.01) in average age, the score of acute physiology and chronic health evaluation II (APACHE II), Ranson criteria, the score of sequential organ failure assessment (SOFA), and albumin variation within 24 hours (Serum albumin descent degree and ratio in early stage). However, sex ratio, mean serum albumin concentration within 24 hours, and onset time did not have significant differences (P > 0.05). The area under ROC curve for albumin variation within 24 hours were 0. 728 and 0. 742, the best cut-off values were 4.25 g/L (sensitivity 61.45%, specificity 81.67%) and 13.5% (sensitivity 62.65%, specificity 78.33%), respectively. Multiple logistic regression analysis showed that albumin variation within 24 hours was associated with death., Conclusion: The albumin variation within 24 hours was the risk factors for poor prognosis of critically ill SAP in early stage.

Objective: To investigate the effect of dietary calcium on plasma lipoprotein profile in castrated and ovariectomized hamsters., Methods: Male, castrated, female and ovariectomized hamsters (n=36 each group) were randomly divided into three sub-groups (n=12) and fed one of the three diets containing 0, 2, and 8 g calcium per kg diet for a period of six weeks. Changes in plasma lipoprotein profile were monitored at the end of week 0, 3 and 6., Results: Plasma total cholesterol (TC), non-high density lipoprotein cholesterol (non-HDL-C), triacylglycerols (TG) and TC/HDL-C were decreased only in intact female and ovariectomized hamsters. In contrast, three levels of dietary calcium had no effect on lipoprotein profiles in both intact male and castrated hamsters., Conclusion: Beneficial modification of lipoprotein profile by dietary calcium was gender-dependent at least in hamsters., (Copyright © 2012 The Editorial Board of Biomedical and Environmental Sciences. Published by Elsevier B.V. All rights reserved.)

In this study, a molecular self-assembly strategy to develop a novel protein scaffold for amplifying the extent and variety of proteins displayed on the surface of Saccharomyces cerevisiae is presented. The cellulosomal scaffolding protein cohesin and its upstream hydrophilic domain (HD) were genetically fused with the yeast Ure2p N-terminal fibrillogenic domain consisting of residues 1 to 80 (Ure2p(1-80)). The resulting Ure2p(1-80)-HD-cohesin fusion protein was successfully expressed in Escherichia coli to produce self-assembled supramolecular nanofibrils that serve as a novel protein scaffold displaying multiple copies of functional cohesin domains. The amyloid-like property of the nanofibrils was confirmed via thioflavin T staining and atomic force microscopy. These cohesin nanofibrils attached themselves, via a green fluorescent protein (GFP)-dockerin fusion protein, to the cell surface of S. cerevisiae engineered to display a GFP-nanobody. The excess cohesin units on the nanofibrils provide ample sites for binding to dockerin fusion proteins, as exemplified using an mCherry-dockerin fusion protein as well as the Clostridium cellulolyticum CelA endoglucanase. More than a 24-fold increase in mCherry fluorescence and an 8-fold increase in CelA activity were noted when the cohesin nanofibril scaffold-mediated yeast display was used, compared to using yeast display with GFP-cohesin that contains only a single copy of cohesin. Self-assembled supramolecular cohesin nanofibrils created by fusion with the yeast Ure2p fibrillogenic domain provide a versatile protein scaffold that expands the utility of yeast cell surface display.

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 spectrometry (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 species. This allows the surface film to attain low surface tensions during compression and maintain values near equilibrium during expansion., (Copyright © 2011 Elsevier B.V. All rights reserved.)

A simple light microscopic technique was developed in order to quantify the damage inflicted by high-power low-frequency ultrasound (0-160 W, 20 kHz) treatment on potato starch granules in aqueous dispersions. The surface properties of the starch granules were modified using ethanol and SDS washing methods, which are known to displace proteins and lipids from the surface of the starch granules. The study showed that in the case of normal and ethanol-washed potato starch dispersions, two linear regions were observed. The number of defects first increased linearly with an increase in ultrasound power up to a threshold level. This was then followed by another linear dependence of the number of defects on the ultrasound power. The power threshold where the change-over occurred was higher for the ethanol-washed potato dispersions compared to non-washed potato dispersions. In the case of SDS-washed potato starch, although the increase in defects was linear with the ultrasound power, the power threshold for a second linear region was not observed. These results are discussed in terms of the different possible mechanisms of cavitation induced-damage (hydrodynamic shear stresses and micro-jetting) and by taking into account the hydrophobicity of the starch granule surface., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Introduction: 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., Results: 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., Discussion: 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., Methods: 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.

Purpose: The objective of this study was to retrospectively analyze the association of mean glucose level (MGL) and glycemic lability index (GLI; as a measure of glucose variability) with intensive care unit (ICU) mortality in patients with severe acute pancreatitis (SAP)., Materials and Methods: Paper-based medical records of patients with SAP who were admitted to the ICU of West China Hospital between July 1, 2005, and July 1, 2010, were analyzed. Glucose measurements, demographic characteristics, clinical features, data on the first and second 24-hour Acute Physiology and Chronic Health Evaluation (APACHE) II scores, and outcomes were obtained. Time-weighted glucose parameters were used. We statistically analyzed the relationship between these variables and both ICU and hospital mortality., Results: A total of 294 patients with 34,796 glucose measurements were included in the final analysis. The time-weighted MGL was 9.31 ± 1.91 mmol/L, and the median of GLI was 55.27 (mmol/L)(2) h-(1) wk-(1). Intensive care unit mortality was 43.5% and increased progressively as GLI increased, reaching 62.5% of patients with GLI above 115.89 (mmol/L)(2) h-(1) wk-(1). The highest odds ratio for ICU death was found in patients with the highest quartile of GLI: odds ratio, 3.47 (95% confidence interval, 1.76-6.86; P < .000). No such relationship could be found with MGL. Glycemic lability index was better able to predict ICU death than was MGL (the area under the curves were 0.642 vs 0.561, respectively; z test was 2.677; P = .0074). The logistic regression analysis showed that GLI, the second 24-hour APACHE II score, and the number of organ failures upon ICU admission contributed independently to the risk of mortality., Conclusions: We observed that GLI was a better predictor of ICU and hospital mortality than was MGL. Together with the second 24-hour APACHE II score and the number of organ failures upon ICU admission, GLI is an independent predictor of mortality in patients with SAP., (Copyright © 2012 Elsevier Inc. All rights reserved.)

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.

Objective: To investigate effects of intensive glucose control (IGC) on glucose variability (GV) and clinical outcomes in patients with severe acute pancreatitis (SAP), and to assess predictive values of different measures of GV on intensive care unit (ICU) death., Methods: A prospective, randomized, non-blinded clinical trial was conducted. All adult patients with diagnosis of SAP, who were admitted to ICU of West China Hospital between July 1st 2010 and July 1st 2011, were enrolled. Eligible patients were randomly assigned to receive either IGC to maintain glucose level between 6.1 mmol/L and 8.3 mmol/L or control group (none intervention was given). GV, ICU mortality, the incidence of infection, length of mechanical ventilation and ICU stay of the two groups were compared. The standard deviation of blood glucose level [GLU(SD)], mean amplitude of glycemic excursion [GLU(MAGE)] and glycemic lability index [GLU(GLI)] were chosen as measures of GV to analyze the multi variable correlation between them and ICU mortality, and the ability of these three parameters was assessed in predicting ICU death using area under a receiver operating characteristic curve (AUC)., Results: One hundred and twenty-two patients were eligible for the study, and 30 patients with SAP were enrolled in the study, with 15 cases in each group. The basic data of the two groups were comparable. Compared with control group, IGC showed an effect to reduce GV, including GLU(SD) [mmol/L: 1.81 (0.97, 2.65) vs. 2.48 (1.29, 2.87)], GLU(MAGE) [mmol/L: 3.76 (3.67, 5.85) vs. 5.30 (4.35, 6.80)], GLU(GLI) [306.8 (220.6, 613.3) vs. 339.5 (218.4, 423.1)], and lower ICU mortality (13.3% vs. 40.0%), but the difference showed no significant difference (all P > 0.05). It could also shorten the length of ICU stay (days: 11.3 ± 9.9 vs. 15.8 ± 7.6, P < 0.01), decrease the incidence of infection of blood stream and the lung (6.7% vs. 40.0%; 33.3% vs. 73.3%, both P < 0.05). A positive correlation between GLU(GLU) and ICU mortality was found (r = 0.371, P = 0.044), but no correlation was found between GLU(SD) or GLU(MAGE) and ICU mortality (r value was -0.144 and -0.065, P value was 0.448 and 0.731). AUC for GLU(SD), GLU(MAGE) and GLU(GLI) was 0.594 [95% confidence interval (95%CI) 0.382-0.805], 0.543 (95%CI 0.287-0.798) and 0.751 (95%CI 0.548-0.954) respectively, and GLU(GLI) was the best predictor of ICU death., Conclusions: IGC had an effect to reduce GV, decrease ICU mortality, shorten length of ICU stay, and lower the incidence of infection. Compared with GLU(SD) and GLU(MAGE), GLU(GLI) was the best predictor of ICU death.

Objective: To explore the effect of ketamine on adrenal pheochromocytoma (PC12) cell proliferation inhibition and induction of apoptosis and its mechanism., Methods: PC12 cells of rats were models for dopaminergic neuron. PC12 cells were cultured with ketamine at concentrations of 0.9, 1.2, 1.5, 1.8 and 2.1 mmol/L, respectively. The cell viability was measured by MTT method after incubation at 12, 24, 48 and 72h. Hoechst stain was used to observe the morphological changes of apoptosis. PC12 cells cultured after 48 h with different concentrations of ketamine were selected to detect apoptotic rate using flow cytometry and detect the expression of bax and bcl-2 proteins using Western blotting., Results: For different concentrations of ketamine, vitality of PC12 cells significantly decreased with increase of the incubation time. Apoptosis was obviously observed using Hoechst staining. Flow cytometry showed that apoptosis rates significantly increased with increasing ketamine concentrations., Conclusion: Ketamine can inhibit the proliferation of PC12 cell by inducing apoptosis of the PC12 cell in a concentrations-dependent manner. The underlying mechanism may be related to promoting the expression of bax and inhibiting the expression of bcl-2 in the cells.

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., (Copyright © 2011 Elsevier B.V. All rights reserved.)

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., (© 2011 American Chemical Society)

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., (© 2011 American Chemical Society)

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., (Copyright © 2011 Elsevier B.V. All rights reserved.)

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., (Copyright (c) 2010 Elsevier B.V. All rights reserved.)

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.