This study focuses on the structural organization of surfactant protein B (SP-B) containing lipid monolayers. The artificial system is composed of the saturated phospholipids dipalmitoylphosphatidylcholine (DPPC) and dipalmitoylphosphatidylglycerol (DPPG) in a molar ratio of 4:1 with 0.2 mol% SP-B. The different "squeeze-out" structures of SP-B were visualized by scanning probe microscopy and compared with structures formed by SP-C. Particularly, the morphology and material properties of mixed monolayers containing 0.2 mol% SP-B in a wide pressure range of 10 to 54 mN/m were investigated revealing that filamentous domain boundaries occur at intermediate surface pressure (15-30 mN/m), while disc-like protrusions prevail at elevated pressure (50-54 mN/m). In contrast, SP-C containing lipid monolayers exhibit large flat protrusions composed of stacked bilayers in the plateau region (app. 52 mN/m) of the pressure-area isotherm. By using different scanning probe techniques (lateral force microscopy, force modulation, phase imaging) it was shown that SP-B is dissolved in the liquid expanded rather than in the liquid condensed phase of the monolayer. Although artificial, the investigation of this system contributes to further understanding of the function of lung surfactant in the alveolus.

In this study, we explored the calcium-induced formation of phosphatidylserine-enriched lipid domains in Langmuir−Blodgett (LB) monolayers composed of the binary mixture of the two saturated lipids 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and 1,2-dipalmitoyl-sn-glycero-3-phosphoserine (DPPS) in a molar ratio of 4:1 by means of surface analysis tools. We employed fluorescence microscopy in conjunction with pressure−area isotherms, time-of-flight secondary ion mass spectrometry (TOF-SIMS) imaging, scanning electron microscopy (SEM), and lateral force microscopy to visualize and analyze lipid domains by their chemical and physical properties. TOF-SIMS and SEM images of DPPC/DPPS LB monolayers transferred onto gold surfaces clearly reveal that small amounts of calcium ions in the aqueous phase are sufficient to trigger the formation of circular DPPS-enriched domains, but domain formation is abolished in the presence of ethylene glycol-bis(β-aminoethyl) ether-N,N,N ‘ ,N ‘-tetraacetic acid (EGTA). Lateral force microscopy of solid supported lipid bilayers transferred from the air/water interface to mica surfaces and imaged in water also allowed visualization of these DPPS-enriched domains despite the absence of a topographic contrast.

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An often-used model lung surfactant containing dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylglycerol (DPPG), and the surfactant protein C (SP-C) was analyzed as Langmuir-Blodgett film by spatially resolved time-of-flight secondary ion mass spectrometry (TOF-SIMS) to directly visualize the formation and composition of domains. Binary lipid and lipid/SP-C systems were probed for comparison. TOF-SIMS spectra revealed positive secondary ions (SI) characteristic for DPPC and SP-C, but not for DPPG. SI mapping results in images with domain structures in DPPC/DPPG and DPPG/SP-C, but not in DPPC/SP-C films. We are able to distinguish between the fluid and condensed areas probably due to a matrix effect. These findings correspond with other imaging techniques, fluorescence light microscopy (FLM), scanning force microscopy (SFM), and silver decoration. The ternary mixture DPPC/DPPG/SP-C transferred from the collapse region exhibited SP-C-rich domains surrounding pure lipid areas. The results obtained are in full accordance with our earlier SFM picture of layered protrusions that serve as a compressed reservoir for surfactant material during expansion. Our study demonstrates once more that SP-C plays a unique role in the respiration process.

By means of the quartz crystal microbalance (QCM) technique, the interaction of annexin A1 with lipid membranes was quantified using solid-supported bilayers immobilized on gold electrodes deposited on 5 MHz quartz plates. Solid-supported lipid bilayers were composed of a first octanethiol monolayer chemisorbed on gold and a physisorbed phospholipid monolayer obtained from vesicle fusion. This experimental setup enabled us to determine for the first time rate constants and affinity constants of annexin A1 binding to phosphatidylserine-containing layers as a function of the calcium ion concentration in solution and the cholesterol content within the outer leaflet of the solid-supported bilayer. The results reveal that a decrease in Ca(2+) concentration from 1 mM to 100 microM significantly increases the rate of annexin A1 binding to the membrane independent of the cholesterol content. However, the presence of cholesterol in the membrane altered the affinity constants considerably. While the association constant decreases with decreasing Ca(2+) concentration in the case of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoserine (POPS) membranes lacking cholesterol, it remains high in the presence of cholesterol.