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Concept: Pulmonary surfactant

171

The peripheral lungs are a potential entrance portal for nanoparticles into the human body due to their large surface area. The fact that nanoparticles can be deposited in the alveolar region of the lungs is of interest for pulmonary drug delivery strategies and is of equal importance for toxicological considerations. Therefore, a detailed understanding of nanoparticle interaction with the structures of this largest and most sensitive part of the lungs is important for both nanomedicine and nanotoxicology. Astonishingly, there is still little known about the bio-nano interactions that occur after nanoparticle deposition in the alveoli. In this study, we compared the effects of surfactant-associated protein A (SP-A) and D (SP-D) on the clearance of magnetite nanoparticles (mNP) with either more hydrophilic (starch) or hydrophobic (phosphatidylcholine) surface modification by an alveolar macrophage (AM) cell line (MH-S) using flow cytometry and confocal microscopy. Both proteins enhanced the AM uptake of mNP compared with pristine nanoparticles; for the hydrophilic ST-mNP, this effect was strongest with SP-D, whereas for the hydrophobic PL-mNP it was most pronounced with SP-A. Using gel electrophoretic and dynamic light scattering methods, we were able to demonstrate that the observed cellular effects were related to protein adsorption and to protein-mediated interference with the colloidal stability. Next, we investigated the influence of various surfactant lipids on nanoparticle uptake by AM because lipids are the major surfactant component. Synthetic surfactant lipid and isolated native surfactant preparations significantly modulated the effects exerted by SP-A and SP-D, respectively, resulting in comparable levels of macrophage interaction for both hydrophilic and hydrophobic nanoparticles. Our findings suggest that because of the interplay of both surfactant lipids and proteins, the AM clearance of nanoparticles is essentially the same, regardless of different intrinsic surface properties.

Concepts: Metabolism, Lung, Lipid, Pulmonary surfactant, Cell biology, Colloid, Nanotechnology, Protein

28

A high resolution interferometric technique was used to determine swelling behavior of weakly charged polyacrylamide hydrogels in the presence of oppositely charged surfactants and subsequent exposure to cyclodextrins. Hydrogels of copolymerized acrylamide and 2-acrylamido-2-methyl-1-propanesulfonic acid (0.22, 0.44, 0.88 mol%) crosslinked with bisacrylamide (3, 6, 12 mol%) were employed. The equilibrium swelling and swelling kinetics of the hydrogels were determined with 2nm resolution of the optical length and sampled at approximately 1 Hz. These properties were determined for the hydrogels exposed to cationic surfactants dodecyltrimethylammonium bromide (DTAB) and cetyltrimethylammonium bromide (CTAB) at concentrations from 10(-7) up to 2×10(-3)M. The distribution of surfactant within one AAM-co-AMPSA hydrogel equilibrated in CTAB/perylene solution was investigated by confocal laser scanning microscopy. Hydrogels equilibrated at selected surfactant concentrations were subsequently exposed to cyclodextrins (α-CD, β-CD, methyl-β-CD and γ-CD) forming inclusion complexes with the surfactants. The results show different types of behavior for the two surfactants used, arising from the difference in the length of surfactant hydrophobic tail. The changes in the surfactant induced swelling of the hydrogels are suggested to arise from the net effect of electrostatic screening of sulfonic acid-amide group interactions and surfactant micellization. Hydrogels with the largest charge density and the lowest crosslink density yielded the most pronounced changes in swelling properties on exposure to DTAB or CTAB. The hydrogels displayed swelling kinetics on stepwise changes in surfactant concentrations that depended on the surfactant concentration range. The high resolution monitoring of hydrogel swelling associated with supramolecular complex formation in three-component systems hydrogel-amphiphilic molecule-cyclodextrin provides more details on the swelling behavior than previously disclosed.

Concepts: Pulmonary surfactant, Electric charge, Micelle, Supramolecular chemistry, Solution, Surfactant

28

Herein we report the surfactant-triggered assembly of fullerene (C60) into three dimensional (3D) flower-like microcrystals at liquid-liquid interface. C60 crystals were grown using a liquid-liquid interfacial precipitation (LLIP) method by layering surfactant solution in butanol with saturated solution of C60 in benzene. In the LLIP method, it is suggested that the crystal formation mechanism is driven by supersaturation related to the low C60 solubility in alcohol. We found that the dimensions of the synthesized C60 flowers depend on concentration and surfactant type. In the absence of surfactant, i.e., in butanol/benzene system, 1D C60 nanowhiskers (nanorods) and C60 nanotubes (diameter 400 nm  2 m and length 5  20 m) are obtained. However, when surfactants are incorporated in the system flower-like microcrystals consisting of C60 nanotubes are observed; for instance, crystals grown at the interface of 0.01% diglycerol monolaurate (C12G2) nonionic surfactant in butanol with benzene leads to the formation of flower-shaped microcrystals of average sizes in the range of 10  35 m. To the best of our knowledge, this is the first example of surfactant-assisted assembly of C60 crystals. X-ray diffraction (XRD) and transmission electron microscopy (TEM) measurements have shown that the fullerene flowers have hexagonal structure with cell dimensions a = 2.539 nm and c = 1.021 nm, which differs from the pristine C60. Mixtures of flower-shaped C60 crystals and freely standing C60 nanotubes are found in the 0.1% C12G2/butanol system. On the other hand, clusters or giant aggregates of nanowhiskers lacking any specific shape are observed in the 1% C12G2/butanol system although these crystals exhibit hexagonal close packed structures. Flower-shaped C60 microcrystals are also observed with anionic surfactants cetyltrimethylammonium bromide (CTAB) and cetyltrimethylammonium chloride (CTAC) systems. C60 flowers obtained from the 0.01% CTAB and 0.01% CTAC also exhibit hexagonal structures with cell dimensions a = 2.329 nm and c = 1.273 nm, a = 2.459 nm and c = 0.938 nm, respectively. Our C60 flowers exhibit intense photoluminescence (PL) and blue shifted PL intensity maximum compared to pristine C60, demonstrating the potential to control the optoelectronic properties of fullerene-based nanostructures.

Concepts: Pulmonary surfactant, Hexagonal crystal system, Transmission electron microscopy, Solution, Surfactant

27

The impact of Ca2+ counterions on the adsorption at the air-water interface and self-assembly in aqueous solution of the rhamnolipid biosurfactant and its mixture with the anionic surfactant sodium dodecylbenzene sulfonate, LAS, has been studied using neutron reflectometry, NR and small angle neutron scattering, SANS. The results illustrate how rhamnolipids are calcium tolerant and how their blending with conventional anionic surfactants improves the calcium tolerance of the anionic surfactant. Ca2+ has relatively little effect upon the adsorption and self-assembly of the mono-rhamnose, R1, and di-rhamnose, R2, rhamnolipids, even at high pH, due to their predominantly non-ionic nature. For R1/R2 mixtures the addition of Ca2+ has little impact upon the adsorbed amount or the surface composition. For R2/LAS mixtures the addition of Ca2+ results in an increased adsorption and a surface slightly richer in R2. The weak binding of Ca2+ to R1 and R2 does result in a change to the degree of ionization of the micelles and especially for mixed R1/R2 micelles at R1 rich solution compositions. The stronger binding of Ca2+ to LAS results in the addition of Ca2+ having a much greater impact on the self-assembly of R1/LAS and R2/LAS mixtures. For R1/LAS mixtures the addition of Ca2+ promotes the formation of more planar structures, even at low surfactant concentrations where in the absence of Ca2+ mixed globular micelle formation dominates. In R2/LAS mixtures, where there is a greater contrast between the high and low preferred curvatures associated with R2 and LAS, the addition of Ca2+ results in a more complex evolution in micellar aggregation and the degree of ionization of the micelles. This results in variations in Ca2+ binding that promotes micellar structures in which a spatial segregation of the two surfactant components within the micelle occur.

Concepts: Concentration, Surfactants, Pulmonary surfactant, Solution, Colloidal chemistry, Chemistry, Micelle, Surfactant

26

The sigh is a normal homeostatic reflex that maintains lung compliance and decreases atelectasis. General anesthesia abolishes the sigh reflex with rapid onset of atelectasis in 100% of patients. Studies show a strong correlation between atelectasis and postoperative pulmonary complications, raising health-care costs. Alveolar recruitment maneuvers recruit collapsed alveoli, increase gas exchange, and improve arterial oxygenation. There is no consensus in the literature about the benefits of alveolar recruitment maneuvers. A systematic review is necessary to delineate their usefulness.

Concepts: Pulmonary artery, Vein, Emphysema, Pulmonary surfactant, Respiratory physiology, Heart

15

The paper describes the adsorption of purified protein from seeds of Moringa oleifera to a sapphire interface and the effects of addition of the anionic surfactant sodium dodecylsulfate (SDS) and the cationic surfactant hexadecyltrimethylammonium bromide (CTAB). Neutron reflection was used to determine the structure and composition of interfacial layers adsorbed at the solid/solution interface. The maximum surface excess of protein was found to be about 5.3 mg m(-2). The protein does not desorb from the solid/liquid interface when rinsed with water. Addition of SDS increases the reflectivity indicating co-adsorption. It was observed that CTAB is able to remove the protein from the interface. The distinct differences to the behavior observed previously for the protein at the silica/water interface are identified. The adsorption of the protein to alumina in addition to other surfaces has shown why it is an effective flocculating agent for the range of impurities found in water supplies. The ability to tailor different surface layers in combination with various surfactants also offers the potential for adsorbed protein to be used in separation technologies.

Concepts: Ammonium lauryl sulfate, Pulmonary surfactant, Colloidal chemistry, Moringa oleifera, Emulsion, Surface tension, Surfactant

8

Lung immaturity is a major cause of morbidity and mortality in premature infants. Understanding the molecular mechanisms driving normal lung development could provide insights on how to ameliorate disrupted development. While transcriptomic and proteomic analyses of normal lung development have been previously reported, characterization of changes in the lipidome is lacking. Lipids play significant roles in the lung, such as dipalmitoylphosphatidylcholine in pulmonary surfactant; however, many of the roles of specific lipid species in normal lung development, as well as in disease states, are not well defined. In this study, we used liquid chromatography-mass spectrometry (LC-MS/MS) to investigate the murine lipidome during normal postnatal lung development. Lipidomics analysis of lungs from post-natal day 7, day 14 and 6-8 week mice (adult) identified 924 unique lipids across 21 lipid subclasses, with dramatic alterations in the lipidome across developmental stages. Our data confirmed previously recognized aspects of post-natal lung development and revealed several insights, including in sphingolipid-mediated apoptosis, inflammation and energy storage/usage. Complementary proteomics, metabolomics and chemical imaging corroborated these observations. This multi-omic view provides a unique resource and deeper insight into normal pulmonary development.

Concepts: Pneumonia, Lipidomics, Pulmonary surfactant, Metabolism, Childbirth, Systems biology, Lung, Proteomics

3

Thin liquid films are central to everyday life. They are ubiquitous in modern technology (pharmaceuticals, coatings), consumer products (foams, emulsions) and also serve vital biological functions (tear film of the eye, pulmonary surfactants in the lung). A common feature in all these examples is the presence of surface-active molecules at the air-liquid interface. Though they form only molecular-thin layers, these surfactants produce complex surface stresses on the free surface, which have important consequences for the dynamics and stability of the underlying thin liquid film. Here we conduct simple thinning experiments to explore the fundamental mechanisms that allow the surfactant molecules to slow the gravity-driven drainage of the underlying film. We present a simple model that works for both soluble and insoluble surfactant systems in the limit of negligible adsorption-desorption dynamics. We show that surfactants with finite surface rheology influence bulk flow through viscoelastic interfacial stresses, while surfactants with inviscid surfaces achieve stability through opposing surface-tension induced Marangoni flows.

Concepts: Fluid mechanics, Pulmonary surfactant, Surface tension, Solution, Emulsion, Viscosity, Fluid dynamics, Surfactant

2

Nanobody (Nb) is a promising vector for targeted drug delivery. This study aims to identify an Nb that can specifically target the lung by binding human pulmonary surfactant protein A (SP-A). Human lung frozen tissue sections were used for 3 rounds of biospanning of our previously constructed Nb library for rat SP-A to establish a sub-library of Nb, which specifically bound human lung tissues. Phage-ELISA was performed to screen the sub-library to identify Nb4, which specifically bound human SP-A. The binding affinity Kd of Nb4 to recombinant human SP-A was 7.48 × 10(-7) M. Nb4 (19 kDa) was stable at 30 °C-37 °C and pH 7.0-7.6 and specifically bound the SP-A in human lung tissue homogenates, human lung A549 cells, and human lung tissues, whereas didn’t react with human liver L-02 cells, kidney 293T cells, and human tissues from organs other than the lung. Nb4 accumulated in the lung of nude mice 5 minutes after a tail vein injection of Nb4 and was excreted 3 hours. Short-term exposure (one month) to Nb4 didn’t cause apparent liver and kidney toxicity in rats, whereas 3-month exposure resulted in mild liver and kidney injuries. Nb4 may be a promising vector to specifically deliver drugs to the lung.

Concepts: Pulmonary surfactant, Red blood cell, Hemoglobin, Liver, Organs, Glucose, Organ, Lung

2

The short palate, lung and nasal epithelial clone 1 (SPLUNC1) protein is a member of the palate, lung, and nasal epithelium clone (PLUNC) family, also known as bactericidal/permeability-increasing (BPI) fold-containing protein, family A, member 1 (BPIFA1). SPLUNC1 is an abundant protein in human airways, but its function remains poorly understood. The lipid ligands of SPLUNC1 as well as other PLUNC family members are largely unknown, although some reports provide evidence that lipopolysaccharide (LPS) could be a lipid ligand. Unlike previous hypotheses, we found significant structural differences between SPLUNC1 and BPI. Recombinant SPLUNC1 produced in HEK 293 cells harbored several molecular species of sphingomyelin and phosphatidylcholine as its ligands. Significantly, in vitro lipid-binding studies failed to demonstrate interactions between SPLUNC1 and LPS, lipoteichoic acid, or polymyxin B. Instead, one of the major and most important pulmonary surfactant phospholipids, dipalmitoylphosphatidylcholine (DPPC), bound to SPLUNC1 with high affinity and specificity. We found that SPLUNC1 could be the first protein receptor for DPPC. These discoveries provide insight into the specific determinants governing the interaction between SPLUNC1 and lipids and also shed light on novel functions that SPLUNC1 and other PLUNC family members perform in host defense.-Ning, F., Wang, C., Berry, K. Z., Kandasamy, P., Liu, H., Murphy, R. C., Voelker, D. R., Nho, C. W., Pan, C.-H., Dai, S., Niu, L., Chu, H-W., Zhang, G. Structural characterization of the pulmonary innate immune protein SPLUNC1 and identification of lipid ligands.

Concepts: Cell culture, Cell biology, Epithelium, Lipid, Receptor, HEK cell, Protein, Pulmonary surfactant