Biomedical subjects

Stephen B Hall

Publications and source records attributed to Stephen B Hall.

15 recordsLinked to original sources

PubMed2006

The melting of pulmonary surfactant monolayers.

Monomolecular films of phospholipids in the liquid-expanded (LE) phase after supercompression to high surface pressures (pi), well above the equilibrium surface pressure (pi(e)) at which fluid films collapse from the interface to form a three-dimensional bulk phase, and in the tilted-condensed (TC) phase both replicate the resistance to collapse that is characteristic of alveolar films in the lungs. To provide the basis for determining which film is present in the alveolus, we measured the melting characteristics of monolayers containing TC dipalmitoyl phosphatidylcholine (DPPC), as well as supercompressed 1-palmitoyl-2-oleoyl phosphatidylcholine and calf lung surfactant extract (CLSE). Films generated by appropriate manipulations on a captive bubble were heated from < or =27 degrees C to > or =60 degrees C at different constant pi above pi(e). DPPC showed the abrupt expansion expected for the TC-LE phase transition, followed by the contraction produced by collapse. Supercompressed CLSE showed no evidence of the TC-LE expansion, arguing that supercompression did not simply convert the mixed lipid film to TC DPPC. For both DPPC and CLSE, the melting point, taken as the temperature at which collapse began, increased at higher pi, in contrast to 1-palmitoyl-2-oleoyl phosphatidylcholine, for which higher pi produced collapse at lower temperatures. For pi between 50 and 65 mN/m, DPPC melted at 48-55 degrees C, well above the main transition for bilayers at 41 degrees C. At each pi, CLSE melted at temperatures >10 degrees C lower. The distinct melting points for TC DPPC and supercompressed CLSE provide the basis by which the nature of the alveolar film might be determined from the temperature-dependence of pulmonary mechanics.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2006

Distribution of coexisting solid and fluid phases alters the kinetics of collapse from phospholipid monolayers.

To determine how coexistence of liquid-expanded (LE) and tilted-condensed (TC) phases in phospholipid monolayers affects collapse from the air/water interface, we studied binary films containing dioleoyl phosphatidylcholine-dipalmitoyl phosphatidylcholine (DPPC) mixtures between 10 and 100% DPPC. Previously published results established that this range of compositions represents the LE-TC coexistence region at the equilibrium spreading pressure of 47 mN/m. When held at 49.5 mN/m on a captive bubble, the extent of total collapse fit with the LE area predicted by the phase diagram. The kinetics of collapse, however, when normalized for changes in the LE area, slowed with increasing mole fraction of DPPC. Surface area expressed as stretched exponential functions of time yielded an Avrami exponent that decreased from 1 for the homogeneously LE film to 0.3 for DPPC > or = 70%. Microscopic studies showed that the largest changes in kinetics occurred when either alterations of the initial composition or the process of collapse induced the films to cross the percolation threshold, so that the LE phase became divided into isolated domains. Our results show that although coexisting solid and fluid phases collapse to extents that are independent, the kinetics of collapse, corrected for differences in LE area, depend on the distribution of the two phases.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2006

Differential effects of lysophosphatidylcholine on the adsorption of phospholipids to an air/water interface.

To determine how the hydrophobic surfactant proteins promote insertion of the surfactant lipids into an air/water interface, we measured the effect of lysophosphatidylcholine (LPC) on adsorption. Existing models contend that the proteins function either by disordering the lipids or by stabilizing a negatively curved structure located between the adsorbing vesicle and the interface. Because LPC produces greater disorder but positive curvature, the models predict opposite effects. With vesicles containing either dioleoyl phosphatidylcholine (DOPC) or the neutral and phospholipids isolated from calf surfactant, LPC increased the initial rate at which surface tension fell. The final surface tension, however, remained well above the value of approximately 25 mN/m expected for a saturated surface. With two preparations, dioleoyl phosphatidylethanolamine and gramicidin A-DOPC, which form the negatively curved hexagonal-II (H(II)) phase and adsorb rapidly, LPC instead had little effect on initial adsorption but delayed the fall of surface tension below approximately 30 mN/m. LPC produced a similar inhibition of the late adsorption for extracted calf surfactant. Unlike dioleoyl phosphatidylethanolamine and gramicidin A-DOPC, small-angle x-ray scattering and (31)P-nuclear magnetic resonance for extracted calf surfactant detected no evidence for the H(II) phase. Our results indicate that although LPC can promote the initial adsorption of vesicles containing only lamellar lipids, it inhibits the facilitation by the hydrophobic proteins of late adsorption. Our findings support a model in which the surfactant proteins accelerate adsorption by producing a focal tendency to stabilize a negatively curved kinetic intermediate without a general shift to the H(II) phase.

Adsorption↗

PubMed2005

Effects of gramicidin-A on the adsorption of phospholipids to the air-water interface.

Prior studies suggest that the hydrophobic surfactant proteins, SP-B and SP-C, promote adsorption of the lipids in pulmonary surfactant to an air-water interface by stabilizing a negatively curved rate-limiting structure that is intermediate between bilayer vesicles and the surface film. This model predicts that other peptides capable of stabilizing negative curvature should also promote lipid adsorption. Previous reports have shown that under appropriate conditions, gramicidin-A (GrA) induces dioleoyl phosphatidylcholine (DOPC), but not dimyristoyl phosphatidylcholine (DMPC), to form the negatively curved hexagonal-II (H(II)) phase. The studies reported here determined if GrA would produce the same effects on adsorption of DMPC and DOPC that the hydrophobic surfactant proteins have on the surfactant lipids. Small angle X-ray scattering and (31)P-nuclear magnetic resonance confirmed that at the particular conditions used to study adsorption, GrA induced DOPC to form the H(II) phase, but DMPC remained lamellar. Measurements of surface tension showed that GrA in vesicles produced a general increase in the rate of adsorption for both phospholipids. When restricted to the interface, however, in preexisting films, GrA with DOPC, but not with DMPC, replicated the ability of the surfactant proteins to promote adsorption of vesicles containing only the lipids. The correlation between the structural and functional effects of GrA with the two phospholipids, and the similar effects on adsorption of GrA with DOPC and the hydrophobic surfactant proteins with the surfactant lipids fit with the model in which SP-B and SP-C facilitate adsorption by stabilizing a rate-limiting intermediate with negative curvature.

Adsorption↗

PubMed2005

Kinetics for the collapse of trilayer liquid-crystalline disks from a monolayer at an air-water interface.

Unlike surfactants considered in previous studies, when phosphatidylcholine (PC) monolayers collapse at constant surface tension to form a 3D bulk phase, surface area decreases at rates that slow. The different kinetics could result from collapse by a distinct mechanism. Rather than the transfer of molecules all along the interface between the monolayer and bulk phase, PC films can collapse by the folding and subsequent sliding of a bilayer over the monolayer. By this mechanism, molecules can transfer to collapsed trilayers through a locus of constant size. In this article, we use the theory of nucleation and growth to show analytically that during collapse, the area can decrease at rates that decelerate when each individual structure grows through a region of fixed dimensions. We also show that binary films of 30% dihydrocholesterol (dchol) and dipalmitoyl phosphatidylcholine (DPPC), which have previously been shown to form a homogeneous monolayer from which trilayer disks grow through a point, collapse with rates of area decay that slow, in agreement with our analytical expressions.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2005

The collapse of monolayers containing pulmonary surfactant phospholipids is kinetically determined.

Prior studies have shown that during and after slow compressions of monomolecular films containing the complete set of purified phospholipids (PPL) from calf surfactant at an air/water interface, surface pressures (pi) reach and sustain values that are remarkably high relative to expectations from simple systems with model lipids. Microscopy shows that the liquid-expanded, tilted-condensed, and collapsed phases are present together in the PPL films between 45 and 65 mN/m. The Gibbs phase rule restricts equilibrium coexistence of three phases to a single pi for films with two components but not for more constituents. We therefore determined if the surprising stability of PPL reflects release from the thermodynamic restrictions of simple model systems by the presence of multiple components. Experiments with binary films containing dioleoyl phosphatidylcholine and dipalmitoyl phosphatidylcholine first tested the predictions of the phase rule. The onset of three-phase coexistence, determined by fluorescence microscopy, and its termination, established by relaxation of collapsing films on a captive bubble, occurred at similar pi. Experiments for PPL using the same methods suggested that the three phases might coexist over a range of pi, but limited to approximately 2 mN/m, and extending below rather than above the coexistence pi for the binary films. Our results show that the PPL films at high pi must deviate from equilibrium and that they must then be metastable.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2004

Transformation diagrams for the collapse of a phospholipid monolayer.

The kinetics of phase transitions in three-dimensional bulk materials are commonly presented in transformation diagrams. Time-temperature transformation (TTT) and continuous-cooling-transformation (CCT) diagrams plot the time required to transform specific fractions of the material to the new phase by cooling below a transition temperature. Transformation occurs isothermally for the TTT diagrams and during continuous cooling through a range of temperatures for CCT curves. Here we present analogous transformation diagrams for two-dimensional monolayers, which collapse at the equilibrium spreading pressure (pi e) to form a three-dimensional bulk phase. Time-surface pressure-transformation (TpiT) diagrams give the time required for specific fractions of the film to collapse when surface pressure is constant, and continuous-compression-transformation diagrams give the same information when surface pressure varies continuously. The diagrams, constructed here from previously reported data for 1-palmitoyl-2-oleoyl phosphatidylcholine, provide insights into the behavior of the films. The TpiT diagrams successfully predict the existence and approximate magnitude of a threshold rate for compressing the films to high surface pressures above pi e and the approximate shape of isotherms obtained with different rates of interfacial compression. The diagrams also caution that the behavior of mixed monolayers, explained previously in terms of compositional changes, can instead result from collapse that varies with surface pressure. Finally, the similarity between the shapes of the TTT and TpiT diagrams, with the time for transformation passing through a minimum and then increasing as the systems deviate further from equilibrium, suggests that analogous mechanisms determine the behavior of both systems.

In Vitro Techniques↗

PubMed2004

Persistence of metastability after expansion of a supercompressed fluid monolayer.

Fluid monolayers of 1-palmitoyl-2-oleoyl-phosphatidylcholine collapse from an air/water interface to form a three-dimensional bulk phase at the equilibrium spreading pressure (pie) of approximately 47 mN/m. This phase transition limits access to higher surface pressures under equilibrium conditions or during slow continuous compressions. We have shown previously that these films avoid collapse and become metastable when compressed on a captive bubble to surface pressures above 60 mN/m and that the metastability persists during expansion at least to pie. Here, we first documented the extent of this persistent metastability. Rates of isobaric collapse during expansion of the metastable films were up to 3 orders of magnitude slower than those during the initial compression to high surface pressures. Recovery of the ability to collapse depended on the surface pressure to which the films were expanded and how long they were held there. Films reverted after brief exposure to 20 mN/m and after 1 h at 35 mN/m. At pie, films remained capable of reaching high surface pressures during slow compressions after 65 h, although an increase in compressibility above 55 mN/m suggested somewhat increased rates of collapse. We also determined if the films remained metastable when they acquired sufficient free area to allow reinsertion of collapsed material. Faster isobaric expansion in the presence of more collapsed material and with further deviation below pie supported the existence of reinsertion. The persistence of metastability to pie shows that films with sufficient free area to allow reinsertion remain resistant to collapse. Observations that suggest heterogeneous reinsertion, however, argue that free area may be distributed heterogeneously and leave open the possibility that metastability persists because significant regions retain a restricted free area.

Phosphatidylcholines↗

PubMed2004

SP-B and SP-C alter diffusion in bilayers of pulmonary surfactant.

The hydrophobic proteins SP-B and SP-C promote rapid adsorption of pulmonary surfactant to an air/water interface by an unknown mechanism. We tested the hypothesis that these proteins accelerate adsorption by disrupting the structure of the lipid bilayer, either by a generalized increase in fluidity or by a focal induction of interfacial boundaries within the bilayer. We used fluorescence recovery after photobleaching to measure diffusion of nitrobenzoxadiazolyl-dimyristoyl-phosphatidylethanolamine between 11 and 54 degrees C in multilayers containing the complete set of lipids and proteins in calf lung surfactant extract (CLSE), or the complete set of neutral and phospholipids without the proteins. Above 35 degrees C, Arrhenius plots of diffusion were parallel for CLSE and neutral and phospholipids, but shifted to lower values for CLSE, suggesting that the proteins rigidify the lipid bilayer rather than producing the proposed increase in membrane fluidity. The slopes of the Arrhenius plots for CLSE were steeper below 35 degrees C, suggesting that the proteins induce phase separation at that temperature. The mobile fraction fell below 27 degrees C, consistent with a percolation threshold of coexisting gel and liquid-crystal phases. The induction of lateral phase separation in CLSE, however, does not correlate with apparent changes in adsorption kinetics at this temperature. Our results suggest that SP-B and SP-C accelerate adsorption through a mechanism other than the disruption of surfactant bilayers, possibly by stabilizing a high-energy, highly curved adsorption intermediate.

4-Chloro-7-nitrobenzofurazan↗

PubMed2003

Non-cooperative effects of lung surfactant proteins on early adsorption to an air/water interface.

Two small hydrophobic proteins, SP-B and SP-C, are responsible for rapid adsorption of pulmonary surfactant to the air/water interface. Despite their physiological importance, the number of protein molecules required to trigger an absorption event remains unknown. To investigate this issue, we varied the protein content of calf lung surfactant extract (CLSE) by dilution with protein-depleted surfactant lipids (neutral and phospholipids, N&PL). Vesicles of a constant size and of composition ranging between 100% N&PL and 100% CLSE were generated by probe sonication. Their adsorption kinetics to an air/water interface were monitored at different temperatures using a Wilhelmy plate to measure surface tension. When plotted versus protein concentration, the adsorption rates during the initial change in surface tension exhibit a diphasic behavior, first increasing rapidly and linearly between 0% and 25% CLSE, and then more slowly at higher concentrations. Direct linearity at low protein content (0-5% CLSE ratio) was confirmed at 37 degrees C. These observations argue against cooperative behavior, for which the adsorption rate would first rise slowly with the protein content, and then increase suddenly once the critical number of proteins on each vesicle is reached. The apparent activation energy E(a) and the free energy of activation DeltaG(0)*, calculated from the temperature dependence of adsorption, further support the view that at least the early stages of protein-induced surfactant adsorption proceeds through a sequence of events involving not several, but a single surfactant protein.

Adsorption↗

PubMed2003

Rapid compressions in a captive bubble apparatus are isothermal.

Captive bubbles are commonly used to determine how interfacial films of pulmonary surfactant respond to changes in surface area, achieved by varying hydrostatic pressure. Although assumed to be isothermal, the gas phase temperature (Tg) would increase by >100 degrees C during compression from 1 to 3 atm if the process were adiabatic. To determine the actual change in temperature, we monitored pressure (P) and volume (V) during compressions lasting <1 s for bubbles with and without interfacial films and used P x V to evaluate Tg. P x V fell during and after the rapid compressions, consistent with reductions in n, the moles of gas phase molecules, because of increasing solubility in the subphase at higher P. As expected for a process with first-order kinetics, during 1 h after the rapid compression P x V decreased along a simple exponential curve. The temporal variation of n moles of gas was determined from P x V >10 min after the compression when the two phases should be isothermal. Back extrapolation of n then allowed calculation of Tg from P x V immediately after the compression. Our results indicate that for bubbles with or without interfacial films compressed to >3 atm within 1 s, the change in Tg is <2 degrees C.

Gases↗

PubMed2003

Metastability of a supercompressed fluid monolayer.

Previous studies showed that monomolecular films of extracted calf surfactant collapse at the equilibrium spreading pressure during quasi-static compressions but become metastable at much higher surface pressures when compressed faster than a threshold rate. To determine the mechanism by which the films become metastable, we studied single-component films of 1-palmitoyl-2-oleoyl phosphatidylcholine (POPC). Initial experiments confirmed similar metastability of POPC if compressed above a threshold rate. Measurements at different surface pressures then showed that rates of collapse, although initially increasing above the equilibrium spreading pressure, reached a sharply defined maximum and then slowed considerably. When heated, rapidly compressed films recovered their ability to collapse with no discontinuous change in area, arguing that the metastability does not reflect transition of the POPC film to a new phase. These observations indicate that in several respects, the supercompression of POPC monolayers resembles the supercooling of three-dimensional liquids toward a glass transition.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2003

Liquid-crystalline collapse of pulmonary surfactant monolayers.

During exhalation, the surfactant film of lipids and proteins that coats the alveoli in the lung is compressed to high surface pressures, and can remain metastable for prolonged periods at pressures approaching 70 mN/m. Monolayers of calf lung surfactant extract (CLSE), however, collapse in vitro, during an initial compression at approximately 45 mN/m. To gain information on the source of this discrepancy, we investigated how monolayers of CLSE collapse from the interface. Observations with fluorescence, Brewster angle, and light scattering microscopies show that monolayers containing CLSE, CLSE-cholesterol (20%), or binary mixtures of dipalmitoyl phosphatidylcholine(DPPC)-dihydrocholesterol all form bilayer disks that reside above the monolayer. Upon compression and expansion, lipids flow continuously from the monolayer into the disks, and vice versa. In several respects, the mode of collapse resembles the behavior of other amphiphiles that form smectic liquid-crystal phases. These findings suggest that components of surfactent films must collapse collectively rather than being squeezed out individually.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed2002

Effect of neutral lipids on coexisting phases in monolayers of pulmonary surfactant.

We previously established that compression of monolayers containing the lipids in pulmonary surfactant, with or without the surfactant proteins, initially leads to phase separation. On further compression, however, phase coexistence terminates at a critical point that requires the presence of cholesterol. The studies reported here address the changes in the phospholipid phase diagram produced by cholesterol. We used the two systems of the lipids from calf surfactant with and without the surfactant proteins. For both systems, we began with the postulate that cholesterol had no effect on the composition of other constituents in the two phases, and then used the known behavior of interfacial tension at a critical point to test the two extreme cases in which the cholesterol partitions exclusively into condensed domains or into the surrounding film. Measurements of surface potential along with the fraction of the nonfluorescent area and the radius of the domains, both obtained by fluorescence microscopy, for films with and without cholesterol allowed calculation of the interfacial tension between the two phases. Only the model that assumes the presence of cholesterol within the domains accurately predicts a decreasing line tension during film compression toward the critical point. That model, however, also predicts an unlikely decrease during compression of the dipole moment density for the condensed phase. Our results are best explained in terms of cholesterol partitioning predominantly into the condensed domains, with a resulting partial redistribution of the phospholipids between the two phases.

Animals↗

PubMed2002

Pulmonary surfactant: phase behavior and function.

Pulmonary surfactant functions by first flowing rapidly into the alveolar air/water interface, but then resisting collapse from the surface when the adsorbed interfacial film is compressed during exhalation. Widely accepted models emphasize the importance of phase behavior in both processes. Recent studies show, however, that fluidity is a relatively minor determinant of adsorption and that solid films, which resist collapse, can form by kinetic processes unrelated to equilibrium phase behavior.

1,2-Dipalmitoylphosphatidylcholine↗