Artificial Lung Surfactant Research Articles

Staphylococcus aureus Biofilm Destabilization by Tween-80 and Lung Surfactants to Overcome Biofilm-Imposed Drug Resistance.

This study is aimed to evaluating the potential of tween-80 and artificial lung surfactant (ALS) to destabilize S. aureus biofilm. The biofilm destabilization was studied by crystal violet staining, bright field microscopy, and scanning electron microscopy (SEM). During the study, S. aureus biofilm was exposed with tween-80 along various concentrations (1%, 0.1%, and 0.05%) or LS (lung surfactant) at (2.5%, 5%, and 15%) for 2 hrs. It was observed that 0.1% of tween-80 destabilized 63.83 ± 4.35% and 15% ALS 77 ± 1.7% biofilm in comparison to without treatment. The combination of tween-80 and ALS was used and showed a synergistic effect to destabilize 83.4 ± 1.46% biofilm. These results showed the potential of tween-80 and ALS as biofilm disruptors, which further needs to explore in an in-vivo animal model to access the actual potential of biofilm disruption in natural conditions. This study could play a pivotal role to overcome the problem of antibiotic resistance imposed due to biofilm formation to combat antibiotic resistance imposed by bacteria.

Behavior of surfactants in aqueous dispersions of single-walled carbon nanotubes

Industrial surfactants were strongly adsorbed on the surface of the SWCNTs, and did not readily exchange with artificial lung surfactant.

Thermodynamic and structural characterization of a mixed perfluorocarbon–phospholipid ternary monolayer surfactant system

Pulmonary lung surfactant is a mixture of surfactants that reduces surface tension during respiration. Perfluorinated surfactants have potential applications for artificial lung surfactant formulations, but the interactions that exist between these compounds and phospholipids in surfactant monolayer mixtures are poorly understood. We report here, for the first time, a detailed thermodynamic and structural characterization of a minimal pulmonary lung surfactant model system that is based on a ternary phospholipid–perfluorocarbon mixture. Langmuir and Langmuir–Blodgett monolayers of binary and ternary mixtures of the surfactants 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), 1,2-dipalmitoyl-sn-glycero-3-phosphoglycerol (DPPG) and perfluorooctadecanoic acid (C18F) have been studied in terms of miscibility, elasticity and film structure. The extent of surfactant miscibility and elasticity has been evaluated via Gibbs excess free energies of mixing and isothermal compressibilities. Film structure has been studied by a combination of atomic force microscopy and fluorescence microscopy. Combined thermodynamic and microscopy data indicate that the ternary monolayer films were fully miscible, with the mixed films being more stable than their pure individual components alone, and that film compressibility is minimally improved by the addition of perfluorocarbons to the phospholipids. The importance of these results is discussed in context of these mixtures’ potential applications in pulmonary lung surfactant formulations.

Compatible solutes: Ectoine and hydroxyectoine improve functional nanostructures in artificial lung surfactants

Ectoine and hydroxyectoine belong to the family of compatible solutes and are among the most abundant osmolytes in nature. These compatible solutes protect biomolecules from extreme conditions and maintain their native function. In the present study, we have investigated the effect of ectoine and hydroxyectoine on the domain structures of artificial lung surfactant films consisting of dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylglycerol (DPPG) and the lung surfactant specific surfactant protein C (SP-C) in a molar ratio of 80:20:0.4. The pressure–area isotherms are found to be almost unchanged by both compatible solutes. The topology of the fluid domains shown by scanning force microscopy, which is thought to be responsible for the biophysical behavior under compression, however, is modified giving rise to the assumption that ectoine and hydroxyectoine are favorable for a proper lung surfactant function. This is further evidenced by the analysis of the insertion kinetics of lipid vesicles into the lipid–peptide monolayer, which is clearly enhanced in the presence of both compatible solutes. Thus, we could show that ectoine and hydroxyectoine enhance the function of lung surfactant in a simple model system, which might provide an additional rationale to inhalative therapy.

Nanosized Aluminum Altered Immune Function

On the basis of their uses in jet fuels and munitions, the most likely scenario for aluminum nanoparticle (NP) exposure is inhalation. NPs have been shown to be capable of penetrating deep into the alveolar regions of the lung, and therefore human alveolar macrophages (U937) with human type II pneumocytes (A549) were cultured together and exposed to NPs dispersed in an artificial lung surfactant to more accurately mimic the lung microenvironment. Two types of NPs were evaluated: aluminum (Al) and aluminum oxide (Al2O3). Following a 24-h incubation, cell viability was assessed using MTS, and mild toxicity was observed at higher doses with the U937 cells affected more than the A549. Since the U937 cells provided protection from NP toxicity, the cocultures were exposed to a benign concentration of NPs and infected with the respiratory pathogen community-associated methicillin-resistant Staphylococcus aureus (ca-MRSA) to determine any changes in cellular function. Phagocytosis assays demonstrated that the NPs impaired phagocytic function, and bacterial growth curves confirmed that this reduction in phagocytosis was not related to NP-bacteria interactions. Furthermore, NFkappaB PCR arrays and an IL-6 and TNF-alpha real time PCR demonstrated that both types of NPs altered immune response activation. This change was confirmed by ELISA assays that evaluated the secretion of IL-6, IL-8, IL-10, IL-1beta, and TNF-alpha and illustrated that the NPs repressed secretion of these cytokines. Therefore, although the NPs were not toxic to the cells, they did impair the cell's natural ability to respond to a respiratory pathogen regardless of NP composition.

Effects of preparation methods for multi-wall carbon nanotube (MWCNT) suspensions on MWCNT induced rat pulmonary toxicity

Since there is a possibility of inhaling the fibers of multi-wall carbon nanotube (MWCNT) without any agglomeration, it is important that the pulmonary toxicity is evaluated by intratracheal instillation without agglomeration. MWCNT suspended in an artificial lung surfactant (ALS) with or without grinding in an agate mortar was instilled once intratracheally to rats to determine whether differences of the effects to pulmonary toxicity by different amounts of agglomerated MWCNT particle. The MWCNT suspension preparation method with grinding was effective at reducing agglomerates and in increasing uniform dispersion of the fibers. The ground MWCNT induced higher LDH levels and neutrophil ratios in the bronchoalveolar lavage fluid (BALF). There were no remarkable responses in rats in the non-ground MWCNT group, with the exception of inflammatory responses in the early phase. Some histopathological findings varied between rats given the ground MWCNT and non-ground MWCNT. A major difference was an MWCNT-laden macrophage infiltration site in the lung, which were in the alveolus in the ground MWCNT group, and in the interstitium in non-ground MWCNT group. Accordingly, the preparation method with grinding is considered to be effective at reducing agglomerates and ensuring uniform dispersion of the fibers. These findings lead us to conclude that the amount of agglomerates in the suspension is an important factor affecting the pulmonary toxicity of MWCNT.

Comparison of Free Radical Generation by Pre- and Post-Sintered Cemented Carbide Particles

Rapid generation of reactive oxygen species (ROS) may occur in response to cellular contact with metal particles. Generation of ROS by cobalt and/or tungsten carbide is implicated in causing hard metal lung disease (HMD) and allergic contact dermatitis (ACD). In this study, ROS generation and particle properties that influence radical generation were assessed for three sizes of tungsten, tungsten carbide, cobalt, admixture (tungsten carbide and cobalt powders), spray dryer, and post-sintered chamfer grinder powders using chemical (H2O2 plus phosphate buffered saline, artificial lung surfactant, or artificial sweat) and cellular (RAW 264.7 mouse peritoneal monocytes plus artificial lung surfactant) reaction systems. For a given material, on a mass basis, hydroxyl (·OH) generation generally increased as particle size decreased; however, on a surface area basis, radical generation levels were more, but not completely, similar. Chamfer grinder powder, polycrystalline aggregates of tungsten carbide in a metallic cobalt matrix, generated the highest levels of ·OH radicals (p < 0.05). Radical generation was not dependent on the masses of metals, rather, it involved surface-chemistry-mediated reactions that were limited to a biologically active fraction of the total available surface area of each material. Improved understanding of particle surface chemistry elucidated the importance of biologically active surface area in generation of ROS by particle mixtures.

Effect of SP-C on surface potential distribution in pulmonary surfactant: Atomic force microscopy and Kelvin probe force microscopy study

The air–lung interface is covered by a molecular film of pulmonary surfactant (PS). The major function of the film is to reduce the surface tension of the lung's air–liquid interface, providing stability to the alveolar structure and reducing the work of breathing. Earlier we have shown that function of bovine lipid extract surfactant (BLES) is related to the specific molecular architecture of surfactant films. Defined molecular arrangement of the lipids and proteins of the surfactant film also give rise to a local highly variable electrical surface potential of the interface. In this work we investigated a simple model of artificial lung surfactant consisting of DPPC, eggPG, and surfactant protein C (SP-C). Effects of surface compression and the presence of SP-C on the monolayer structure and surface potential distribution were investigated using atomic force microscopy (AFM) and Kelvin probe force microscopy (KPFM). We show that topography and locally variable surface potential of DPPC–eggPG lipid mixture are similar to those of pulmonary surfactant BLES in the presence of SP-C and differ in surface potential when SP-C is absent.

Structure and dynamics of interfacial water in model lung surfactants

The last decade has seen a transformation in understanding of the role of membrane-bound interfacial water. Whereas until recently water was treated principally as a continuum (primarily screening charges of lipids and proteins), it has become apparent recently that consideration of water's molecular-level properties is critical in understanding a variety of biochemical and biophysical processes. Here we investigate the structure and dynamics of water in contact with a monolayer of artificial lung surfactant, composed of four types of lipids and one protein. We probe this water using frequency-domain sum-frequency generation (SFG) spectroscopy, and a newly developed time-domain, three-pulse technique, in which the vibrational relaxation of interfacial water molecules is followed in real time. We characterize interfacial water in three systems: a monolayer of the pure lipid that is the majority of the lung surfactant mixture, a monolayer of the four lipids constituting the mixture, and a monolayer of the four lipids and the protein. We find subtle differences in the water structure and dynamics that depend on the mixture density and composition. In particular, frequency-domain measurements suggest that in the lipid mixture and the lipid mixture + protein, the relatively bulky lipids (those that have either three or unsaturated hydrocarbon tails) tend to be squeezed out at higher pressure. Measurements using the time-domain, three-pulse technique make clear that structural relaxation of interfacial water is significantly slowed down upon adding small amounts of protein to the lipids. Both results are consistent with prior measurements using other techniques in which more fluid lipids were shown to be 'squeezed out' of lung surfactant at high compression and the role of protein in the mixture was demonstrated to be a catalyst for the formation of multilayers under compression that are subsequently reintegrated into the monolayer on expansion.

Mixing of Saturated and Unsaturated Phosphatidylcholines and Phosphatidylglycerols in Monolayers at the Air/Water Interface

In the context of understanding the properties of model lung surfactant systems, surface phase behavior of spread monolayers containing mixtures of dipalmitoylphosphatidylcholine (DPPC), with various unsaturated phosphatidylcholine (PC) and phosphatidylglycerol (PG) derivatives, has been determined using surface pressure area isotherms on Tris buffer (pH 7.4, 15 mMNaCl) and fluorescence microscopy. DPPC, at all surface pressures below collapse, is completely miscible with dipalmitoylphosphatidylglycerol and with 1-palmitoyl-2-oleoylphosphatidylcholine dioleoylphosphatidylcholine up to mole fractions of at least 0.70 DPPC. On the other hand, DPPC is partially immiscible at mole fractions of 0.50 and greater when mixed with 1-palmitoyl-2-oleoylphosphatidylglycerol and dioleoylphosphatidylglycerol, with immiscibility increasing as the surface pressure is increased and with the replacement of 15 mMNaCl of the buffered subphase by 5 mMCa2+. The substitution of one unsaturated acyl chain into PG appears to produce phase separation equivalent to that produced by two saturated acyl chains of PG with about four carbons less than saturated acyl chains of DPPC. When DPPC/unsaturated PG mixtures are considered in model lung surfactant studies or as artificial lung surfactant systems, such strong tendencies for surface phase separation may be important and should be taken into account.

Minimal surface tension, squeeze-out and transition temperatures of binary mixtures of dipalmitoylphosphatidylcholine and unsaturated phospholipids

Fluoresence polarization (FP) measurements and surface tension (ST) experiments were performed to determine the gel-to-liquid-crystal transition or melting temperature of phospholipid mixtures. The FP-temperature diagrams showed main transition temperatures of 41°C for dipalmitoylphosphatidylcholine (DPPC). The 7:3 and 9:1 binary mixtures of DPPC and phosphatidylinositol (PI), phosphatidylglycerol (PG) and phosphatidylcholine (PC) had main transition temperatures of, respectively, 32–36°C and 37–30°C. The minimal surface tension of DPPC monolayers increased rapidly at 40°C, suggesting that this was the transition temperature for the melting of these monolayers. This value was in close accordance with the main transition temperature of DPPC, observed with the fluorescence polarization measurements. Melting temperatures of monolayers were higher for almost all mixtures than the temperatures at which the transition started, indicating preferential squeeze out of the unsaturated component and enrichment of the monolayer with DPPC. However, neither the 7:3 DPPC/PC nor the DPPC/PG mixtures could withstand high surface pressures at temperatures above 30°C, whereas monolayers of DPPC/PG (9:1) became fluid at temperatures above 35°C. Preferential squeeze-out of the unsaturated phospholipid was especially effective in both the 7:3 and 9:1 DPPC/PI mixtures. These monolayers started to melt at 39–40°C, which is above their main transition temperatures of, respectively, 32 and 37°C, and which approximate the melting temperature of DPPC. Preferential squeeze-out is essential for an artificial lung surfactant. The estimation of this phenomenon by determining the monolayer melting temperatures is therefore useful for distinguishing between mixtures which are effective surfactants at body temperature and those which are less effective.

Development of synthetic lung surfactants.

We have previously reported the development of a reconstituted lung surfactant consisting of an organic solvent extract of natural bovine lung surfactant supplemented with synthetic lipids. This "artificial" surfactant was used successfully to treat surfactant deficiency states both in animals and humans. We now report on the successful testing of a synthetic lung surfactant consisting of a lipid-bound protein isolated from natural lung surfactant and the lipids present in the "artificial" lung surfactant and now used in the same concentration but in a synthetic, commercially available form. The synthetic lung surfactant possessed the in vitro and in vivo surface properties characterizing the "artificial" lung surfactant. In order to identify the components of the synthetic lung surfactant that are responsible for the required surface properties, a series of 25 simple mixtures was prepared. Of these, three possessed surface properties very similar to those of the "artificial" lung surfactant and the synthetic lung surfactant, in vitro as well as in vivo. These three mixtures had four components in common. Besides dipalmitoyl phosphatidylcholine and the lipid-bound protein, they each had a saturated fatty acid, palmitic or stearic, and they each had an acidic phospholipid, phosphatidylglycerol or phosphatidylserine.

Late hypertriglyceridemia in very low birth weight infants fed human milk exclusively

efficacy in respiratory distress syndrome. Pediatrics 1983; 71:473-82. 4. Wilkinson A, ,lenkins PA, Jeffrey JA. Two controlled trials of dry artificial surfactant: early effects and later outcome in babies with surfactant deficiency. Lancet 1985;2:287-90. 5. Enhorning G, Grossman G, Robertson B. Tracheal deposition of surfactant before the first breath. Am Rev Respir Dis 1973;107:921. 6. Weibel ER, Stereological methods, vol 1. Practical methods for biological morphometry. New York: Academic Press, 1979. 7. Enhorning G, Robertson B. Lung expansion in the premature rabbit fetus after tracheal deposition of surfactant. Pediatrics 1972;50:58-62. 8. Egan EA, Notter RH, Kwong MS, et al. Natural and artificial lung surfactant replacement therapy in premature lambs. ,l Appl Physiol 1983;55:875-83. 9. Robertson B, Enhorning G. The alveolar lining of the premature newborn rabbit after pharyngeal deposition of surfactant. Lab Invest 1974;31:54-9. 10..lobe A, Ikegami M, Jacobs H, et al. Surfactant and pulmonary blood flow distribution following treatment of premature lambs with natural surfactant. J Clin Invest 1984;73:84856. 11. Hulbert WC, Forster BB, Laird W, et al. An improved method for fixation of the respiratory epithelial surface with the mucous and surfactant layers. Lab Invest 1982;47:35463.

Nonselective squeeze-out of dioleoylphosphatidylcholine and dioleoylphosphatidylglycerol from binary mixed monolayers with dipalmitoylphosphatidylcholine

In order to enable the possible use of dipalmitoylphosphatidylcholine as an artificial lung surfactant, the addition of dioleoylphosphatidylcholine or dioleoylphosphatidylglycerol has been suggested. A preferential loss of molecules of the second component during compression of the interfacial layer was proposed. In this study two types of measurement were carried out in order to verify such a preferential squeeze-out. In the first type, electron micrographs of a pure dipalmitoylphosphatidylcholine monolayer and of mixed monolayers of dipalmitoylphosphatidylcholine and egg phosphatidylglycerol were taken in order to study the nature of the structures formed during compression of the monolayer. The electron microscopy photos show horizontally stacked layers in the collapse phase of dipalmitoylphosphatidylcholine, and long vertical ridges in the mixed monolayers up to 20% second component. At higher concentrations of the second component no such structures can be detected. The second type involved monolayer studies with binary mixtures of dipalmitoylphosphatidylcholine and dioleoylphosphatidylcholine or dioleoylphosphatidylglycerol, one of the pair being always radioactively labelled. Counting the radioactivities in bulk phase and monolayer after compression revealed nonselective squeeze-out of either component.

Natural and artificial lung surfactant replacement therapy in premature lambs.

The effect of tracheal instillation of surface-active mixtures in premature lambs was studied as an animal model of exogenous surfactant replacement therapy for the respiratory distress syndrome (RDS). Specific mixtures studied were 7:3 (molar ratio) dipalmitoyl phosphatidylcholine (DPPC):egg phosphatidylglycerol (PG) and extracted mixed lipids (with 1% protein) from cow lung lavage (CLL). Preventilatory tracheal instillation of greater than 15 mg/kg of CLL in 10 ml 0.15 M NaCl to premature lambs gave improved alveolar-arterial O2 gradient and blood gases and increased lung compliance, compared with control lambs over a 15-h period. Lambs receiving 7:3 DPPC:PG dispersions were not improved over controls with regard to pressure-volume characteristics and were worse than controls in arterial oxygenation. In terms of in vitro surface properties, both extracted natural CLL and 7:3 DPPC:egg PG were able to lower aqueous surface tension to 1 dyn/cm under dynamic compression. However, the dynamic respreading of CLL films on successive surface cycles was superior to that of 7:3 DPPC:PG. Moreover, after dispersal in 0.15 M NaCl by vortexing (5 mg/80 ml), CLL adsorbed to surface pressure (tau values of 45 dyn/cm within 10 min. 7:3 DPPC:PG adsorbed to significantly lower tau values after subphase dispersal by a variety of methods.

DRY ARTIFICIAL LUNG SURFACTANT AND ITS EFFECT ON VERY PREMATURE BABIES

Artificial lung surfactant prepared with pure dipalmitoylphosphatidylcholine and phosphatidylglycerol in a ratio of 7:3 was made as a dry powder and then blown down an endotracheal tube into the lungs of 22 very premature babies at birth. Only one dose was given. The treated babies did better than their 33 controls. Fewer needed ventilation, and those who did required lower pressures in the first six hours of life. None of the treated babies died, compared with 8 of the controls.