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A J Mautone

Publications and source records attributed to A J Mautone.

At least 19 recordsLinked to original sources
PubMed2001

Dosage regimens of intranasal aerosolized surfactant on otitis media with effusion in an animal model.

OBJECTIVE: To determine optimal dosage regimens of intranasal metered dose aerosolized surfactant with and without other medications in the treatment of otitis media with effusion (OME). STUDY DESIGN: Resolution of experimental OME in gerbils was determined based on otomicroscopy and tympanometry. Experimental intranasal drugs were: surfactant, surfactant with betamethasone, surfactant with phenylephrine, and a normal saline solution placebo. Medications were administered once or twice daily via a metered dose inhaler. RESULTS: For twice-daily dosing, mean days to OME resolution were 8.5 for the aerosolized surfactant, 6.3 for the surfactant with betamethasone, 18.7 for the surfactant with phenylephrine, and 16 each for control and placebo. Resolution with the once-daily dosage was longer for all conditions. Results were comparable using tympanometry. CONCLUSION: OME resolved faster than the natural course when treated with intranasal surfactant with and without steroids. Twice-daily dosing was statistically superior. SIGNIFICANCE: This study reiterates the effectiveness of OME treatment with an aerosolized synthetic surfactant with and without steroids and establishes a superior twice-daily dosage schedule.

Acoustic Impedance Tests↗
PubMed2000

Intranasal surfactant aerosol therapy for otitis media with effusion.

OBJECTIVE: To determine the effect of surfactant alone and with other medications delivered intranasally as a metered dose inhaler (MDI) aerosol on the resolution of experimentally induced otitis media with effusion (OME). BACKGROUND: Eustachian tube dysfunction is a primary factor in the pathogenesis of OME. Intranasal surfactant via MDI has been shown in this laboratory to reduce passive opening pressure of the eustachian tube in normal gerbils and mice. STUDY DESIGN: OME was developed in 35 gerbils by transtympanic injection of 10 microg lipopolysaccharide from Klebsiella pneumoniae. Pretreatment otomicroscopy and tympanometry were performed to exclude pre-existing middle ear disease, and postinfection evaluations were performed on alternate days for a period of 30 days. Five animals received no treatment (control group); four were treated with propellant only (placebo); seven received surfactant alone; eight received surfactant and betamethasone; and six received surfactant with phenylephrine. All medications were sprayed intranasally as an aerosolized MDI and administered daily from postinfection day 2 onward. RESULTS: OME resolved after 16.0 +/- 0.44 days (mean + SD) in controls. There was no difference seen in the placebo or the surfactant with phenylephrine groups. Treatment with surfactant yielded resolution in 10.57 +/- 0.37 days; this was reduced to 8.57 +/- 0.37 days with surfactant plus betamethasone. These differences are statistically significant. There was no recurrence of OME in any group. CONCLUSION: This study demonstrates that using an aerosolized MDI surfactant with and without betamethasone decreases the duration of OME in this in vivo gerbil model.

Administration, Intranasal↗
PubMed1997

Intraalveolar bubbles and bubble films: III. Vulnerability and preservation in the laboratory.

BACKGROUND: Having confirmed (Scarpelli et al. 1996. Anat. Rec. 244:344-357 and 246:245-270) the discovery of intraalveolar bubbles and films as the normal anatomical infrastructure of aerated alveoli at all ages, we now address three questions. Why have these structures been so elusive? Visible in fresh lungs from the in vivo state, can they be preserved by known laboratory methods? Can they be preserved intact for study in tissue sections? METHODS: Lungs of adult rabbits and pups were examined in thorax directly from the in vivo state to confirm normal bubbles both at functional residual capacity and at maximal volume; other lungs were permitted to deflate naturally to minimal volume. The fate of bubbles in situ (either intact, transected, or diced lung tissue) and of isolated bubbles was assessed (1) during conventional histopreparative processing, (2) during inflation-deflation after degassing, (3) after drying in air, (4) during and after quick freezing in liquid N2, and (5) after preservation in fixed and stained tissue sections prepared by a new double-impregnation procedure in which glutaraldehyde-fixed tissue was preembedded in agar, dehydrated and clarified chemically, embedded in paraffin, sectioned, and stained. Control studies included both blocking of bubble formation by rinsing the air spaces with Tween 20 prior to double impregnation and preparation of normal tissue without preembedding in agar. RESULTS: (1) Each of the following procedures in conventional processing dislocated and disrupted bubbles and films: osmium tetroxide and glutaraldehyde:formaldehyde:tannic acid mixture fixation; chemical dehydration (70-100% ethanol) and clarification (xylene and acetone); and embedding in paraffin or epoxy resin. Transection and dicing of the tissue aggravated the untoward effects. In contrast, bubbles and films remained stable in either glutaraldehyde or formaldehyde, which, however, did not protect against the other agents. (2) Degassing destroyed all bubbles as expected; however, bubbles and films re-formed immediately with reinflation. (3) Topography of fixed bubbles and films was retained after air drying. The dry polygonal configuration reverted to spherical-oval either in saline solution or in 50% ethanol, whereas vulnerability to upgraded ethanol concentrations was unchanged. (4) Normal topography and shape appeared to be retained during quick freezing and after thawing. (5) Intraalveolar and intraductal bubbles and films were preserved and photographed in sections from tissue prepared by the double-impregnation procedure; they were not seen either when bubble formation had been blocked (double-impregnation procedure) or when preembedding in agar had been omitted. CONCLUSIONS: (1) Whether or not fixed in glutaraldehyde or formaldehyde, preservation of intraalveolar and intraductal bubbles and films is not to be expected in tissue prepared by conventional histopreparative procedures, whereas product artifacts may be expected from bubble rupture in situ. (2) Degassing cannot be recommended for studies of alveolar structure-function interrelations because all natural bubbles are disrupted in the process, and bubble re-formation may not parallel their "natural history" in vivo. (3) Compared with glutaraldehyde or formaldehyde fixation, air drying offers no added protection against the untoward effects of conventional processing. (4) Quick-frozen tissue is equally at risk. (5) A new double-impregnation procedure does preserve bubbles and films during processing, sectioning, and staining.

Air↗
PubMed1996

Intraalveolar bubbles and bubble films. I. formation and development during the first 48 hours of extrauterine life in rabbits.

BACKGROUND: Aeration of mature lungs at birth depends on formation of intraalveolar bubbles and bubble films (Scarpelli 1978. Pediatr. Res., 12:1070-1076). Bubbles establish immediately structural stability and pulmonary gas exchange. Given that air spaces are cleared in minutes of fetal liquid (the initial substrate for bubble formation), in formation possible beyond this period? If so, is this related to early development of pulmonary function and structure? METHODS: Mature, spontaneously breathing rabbit pups at 1-10 min and 1, 3, 8, 24, and 48 h after vaginal birth were anesthetized, trachea was occluded at "resting volume" (approximately functional residual capacity), and lungs were rapidly exposed to preserve in vivo intrapulmonary status. The entire lung was examined by stereomicroscopy. Other determinations included resting volume, lung wet weight, volume-pressure curves, histological sections, lung dry weight, tissue hydroxyproline (OH-Pro), and lavage phospholipids (PL). Bubble mobility in situ was tested. Bubbles were released into bathing liquid by incision of peripheral units and monitored over time. RESULTS: Pup activity and gross appearance of the lungs, together with septal thinning, secondary septal development, clearance of intraluminal liquid, increasing tissue OH-Pro, and PL distribution indicated normal postnatal development. Each aerated unit examined at resting volume (all lobes, all ages) contained intraalveolar bubbles. Transition to free gas exclusively in conducting airways and bubbles/bubble films in peripheral gas exchange units occurred within 1 h. Bubbles appeared to be exclusively within alveoli at 4 h and thereafter. Bubbles persisted and new bubbles were formed during subsequent inflation to maximal volume and deflation to atmospheric pressure (P0). Volume of intact lungs at P0 was maintained by the counterforce of rigid bubble films against tissue retraction. When bubbles were released either at resting volume or at P0, the bubble-free loci became airless. Constant size and stability of released bubbles support preferential incorporation of surfactants into bubble films and constant "near-zero surface tension" (Scarpelli 1978. Pediatr. Res., 12:1070-1076). CONCLUSIONS: We show the ubiquitous presence of intraalveolar bubbles and bubble films in vivo throughout the first 48 h of postnatal life. Bubble film rigidity sustains aeration and prevents collapse, while low surface tension of the films facilitate liquid removal from the air spaces. Bubbles in situ are stable and, within apparent limits, mobile; after birth they are quickly restricted to the alveolar spaces, leaving airways bubble free.

Air↗
PubMed1996

Intraalveolar bubbles and bubble films: II. Formation in vivo through adulthood.

BACKGROUND: Intraalveolar bubbles and bubble films have been shown to be part of the normal alveolar architecture in vivo from birth through the first 2 days of extrauterine life of rabbit pups (Scarpelli et al., 1996a. Anat. Rec. 244:344-357). The intraluminal boundary between air-way free gas and alveolar bubbles at the level of respiratory bronchioles is established within 1 hour after birth. We now examine the lung through the rest of development, namely, 2 weeks, 1, 2, and 3 months, and adulthood. METHODS: In quick succession in anesthetized spontaneously breathing rabbits, the abdominal aorta was transected and trachea was occluded either after an end-tidal exhalation at functional residual capacity (FRC) or after volume expansion in vivo by a single inflation from FRC to 20 or 25 cm H2O pressure (V20, V25). Immediately the thorax was opened and lungs were examined (anterior, anterolateral) through a dissecting stereomicroscope while still in the chest, unperturbed (pleural surface temperature 34 degrees C). Heart and lungs were then removed en bloc and re-examined (anterior, lateral, posterior) to confirm that architecture had not changed (22-27 degrees C). After these immediate examinations, lungs were entered into one of the protocols enumerated in Results. RESULTS: Immediate examination revealed bubbles in all aerated subpleural and deep ("central") alveoli from apex to base at all ages and temperatures. Bubbles were confirmed from two views (top and tangential) and from their individual mobility in response to gentle microprobe pressure. A "common bubble" (> 30 microns to approximately 120 microns inside diameter at FRC) appeared to occupy a single alveolus, sometimes arranged in clusters and collectively accounting for approximately 84% of the total bubble population. Few "large bubbles" appeared to be intraductal. We concluded that "small bubbles" (< or = 30 microns; approximately 16% of the total population) were contracted common bubbles. The free gas-bubble film boundary of the airways was at the level of respiratory bronchioles. Subsequent protocols: (1) Common bubbles moved out of adjoining tissue following subpleural incision. Adjacent bubbles either moved into vacated spaces or into the outside liquid medium. Large bubble(s) followed common bubbles out of the tissue. Small bubbles were less mobile and distal common bubbles did not move. The sequence of bubble movement at V25 was the same. Isolated bubbles had normal surfactant content and surface tension according to "Pattle's stability ratio." Transection revealed analogous conditions in central alveoli. (2) Bubble size increased during inflation from FRC to V25. Airless spaces were aerated with bubbles during inflation. (3) The bubble surface was compressed during deflation to 81% of maximal volume (Vmax) and below, including deflation to minimal volume (Vmin). (4) Bubble/alveolar shape changed from spherical-oval to polygonal when the pleural surface dried at FRC and V25. The original shape was restored when the surface was re-wet. Dry tissue showed but did not emit bubbles when cut; re-wet tissue did. (5) Lung liquid content and volume-pressure were normal at FRC. (6) As expected, conventionally fixed, dehydrated, and embedded sections showed no bubbles. CONCLUSIONS: Bubbles and bubble films are fundamental to normal architecture of aerated alveoli at all lung volumes from birth through adulthood. As infrastructure, they sustain aeration and resist deformation. With ductal films, they may be expected to form an alveolar surface liquid (foam film) network (Scarpelli, 1988. Surfactants and the Lining of the Lung) that modulates liquid balance principally at Plateau borders. They expand and contract respectively during inflation and deflation, maintaining their closed film integrity. Films are compressed to "film collapse" in situ during deflation from volumes well above FRC to Vmin. At these volumes, intact films sustain aeration; some may disperse into t

Age Factors↗
PubMed1996

Rapid in vitro tests of surfactant film formation: advantages of the Exerowa black film method.

Based on our research, the natural configuration of surfactant from birth through adulthood takes the form of intraalveolar bubbles. Thus, bubble film analysis would seem to be the specific in vitro testing method for lung surfactant. In the present study we report a battery of five in vitro tests for assessing structural and functional properties of surfactant bubbles and bubble films from hydrophobic extracts, namely, the therapeutic surfactants Survanta (SU) and Infasurf (IN) (full strength and diluted to 3 mg phospholipid/mL) and from aqueous extracts from rabbit lung lavage at 3 mg phospholipid/mL (SAM). Each substrate was assessed by: 1) Shake test: stable bubbles from SU, IN, and SAM (50/50, v/v in 95% ethanol) covered the peripheral surfaces, indicating positive response; bubble production by IN and SAM always exceeded SU; 2) Click test: bubble clicking began immediately in all preparations except for undiluted SU, in which the onset of clicking was delayed more than 40 sec; 3) Pattle's stability test: diameters of SU, IN, and SAM bubbles were unchanged for more than 20 min in aerated solution, indicating stable very low surface tension; 4) bubble generation by gas dispersion from a single capillary: full-strength concentration of SU and IN produced relatively large bubbles-bubble rate (number/min) and size were comparable; all SU bubbles rupture in < 25 min, whereas IN bubbles were stable for > 30 min; and 5) Exerowa black film method: in contrast with each of the preceding methods for studying intact bubbles, the Exerowa method focuses on the contact between bubble films and permits direct observation of film formation and determination of film structure. Stable black films were formed spontaneously by both IN (full strength and diluted) and SAM. Conversely, SU (full strength) formed no black films but stable rheological films. Diluted SU films ruptured in 50% of trials. Since methods 1, 2, and 3 were nondiscriminatory and method 4 produced unphysiologically large bubbles for most mammals, we concluded that the black film method of Exerowa is the most discriminating of the tests studied here. It provides a unique visual record of foam film formation and stability and clearly defines differences relative to both the nature and concentration of the preparations.

Animals↗
PubMed1996

Whole cell potassium currents in fetal rat alveolar type II cells cultured on Matrigel matrix.

Patch-clamp studies were performed on fetal rat alveolar type II cells isolated at 19 days of gestation and cultured on either plastic for 7 days or Matrigel matrix (40-50 microliters/cm2) for 10 days. Before study, cells cultured on Matrigel matrix were dissociated from alveolar-like structures with enzymes, replated, and washed with cold buffer at a constant flow rate to remove residual gel. This wash significantly improved obtaining of successful seals. Potassium current-voltage relationships and maximum whole cell K+ conductance (normalized to membrane capacitance) were significantly changed with time in cells cultured on plastic, but no significant change occurred in cells cultured on Matrigel matrix. Application of 20 mM tetraet hyl ammonium, 2mM 4-aminopyridine, and 5mM BaCl2 significantly inhibited K+ currents, showing differences in channel sensitivity to these agents and a voltage-dependent blockage between culture groups or with time in culture. To conclude, we have developed a new method by which epithelial cells cultured on Matrigel matrix can be successfully studied with the use of patch-clamp techniques. Furthermore, these studies show that fetal type II cells have voltage-activated K+ channels and that channel density and their sensitivity to channel blockers are modulated by the substratum on which the cells are cultured.

Animals↗
PubMed1995

External reflection absorption infrared spectroscopy study of lung surfactant proteins SP-B and SP-C in phospholipid monolayers at the air/water interface.

The interactions of the hydrophobic pulmonary surfactant proteins SP-B and SP-C with 1,2-dipalmitoylphosphatidylcholine in mixed, spread monolayer films have been studied in situ at the air/water interface with the technique of external reflection absorption infrared spectroscopy (IRRAS). SP-C has a mostly alpha-helical secondary structure both in the pure state and in the presence of lipids, whereas SP-B secondary structure is a mixture of alpha-helical and disordered forms. When films of SP-B/1,2-dipalmitoylphosphatidylcholine are compressed to surface pressures (pi) greater than approximately 40-43 mN/m, the protein is partially (15-35%) excluded from the surface, as measured by intensity ratios of the peptide bond amide l/lipid C==O stretching vibrations. The extent of exclusion increases as the protein/lipid ratio in the film increases. In contrast, SP-C either remains at the surface at high pressures or leaves accompanied by lipids. The amide l peak of SP-C becomes asymmetric as a result of the formation of intermolecular sheet structures (1615-1630 cm-1) suggestive of peptide aggregation. The power of the IRRAS experiment for determination of film composition and molecular structure, i.e., as a direct test of the squeeze-out hypothesis of pulmonary surfactant function, is evident from this work.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1994

A direct test of the "squeeze-out" hypothesis of lung surfactant function. External reflection FT-IR at the air/water interface.

The current theory of pulmonary surfactant function requires that very low surface tension be achieved and maintained in the alveolar surface film during compression (expiration). To effect this condition, it has been hypothesized that the unsaturated and/or fluid components of surfactant are selectively excluded or "squeezed out" from mixed monolayers containing both saturated and unsaturated phospholipids, leaving a surface film of essentially pure 1,2-dipalmitoylphosphatidylcholine (DPPC). External reflection Fourier transform infrared (FT-IR) spectroscopy has been employed to quantitatively test this hypothesis. Mixed monolayer films of acyl chain-perdeuterated 1,2-dipalmitoylphosphatidylcholine (DPPC-d62) with 1,2-dioleoylphosphatidylglycerol (DOPG), 1-palmitoyl, 2-oleoylPG (POPG), 1,2-dipalmitoylPG (DPPG) were examined in situ at the air/water interface as a function of surface pressure. The relative intensities of CD2 (CH2) stretching vibrations of the deuterated (proteated) components permitted quantitative determination of the relative concentrations of each in the film. For 7:1 (mol:mol) mixtures of DPPC-d62/DOPG, progressive, selective squeeze out of up to about 90% of the PG component is observed over a range of surface pressures from about 51 to 68 mN/m. The extent of maximal PG squeeze out was reduced to 61% for a 7:1 (mol:mol) mixture of DPPC-d62/POPG. This phenomenon, which is at least partially reversible, appears to require relatively high rates of film compression. Squeeze out was reduced (< 20%) for 7:1 (mol:mol) mixtures of DPPC-d62/DPPG or for 7:3 mixtures of DPPC-d62/POPG. Squeeze out requires that the lipid mixture achieve surface pressures greater than about 50-60 mN/m along with unsaturation (or at least conformational disorder) in the acyl chains of the non-DPPC component.(ABSTRACT TRUNCATED AT 250 WORDS)

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1994

Surface biophysics of the surface monolayer theory is incompatible with regional lung function.

The surface monolayer theory of Clements was tested on open surface films of calf lung surfactant extract in a leak-free vertical film surface balance in which alveolar area (A) changes in each lung zone were simulated in accordance with the theory. We found that: 1) physiologically necessary low surface tension (gamma), < 4 dyn/cm, was sustained only by continuous film compression ("expiration"); 2) compression from A equivalent to total lung capacity to functional residual capacity produced fleeting gamma reduction in all zones and quick reversal to high gamma with A changes that simulated tidal volume (VT) breathing at both 14 (adult) and 40 (neonatal) cpm; 3) phase differences between gamma and A axes of VT loops that indicate mixed surface film composition may be attributable to film inertia and viscoelasticity; 4) estimated alveolar retraction pressure due to gamma (P gamma) exceeds "net" transpulmonary pressure, i.e., favors alveolar collapse, under virtually all conditions of the theory in all zones; 5) return to transient, fleeting low gamma in successive VT cycles was determined by the inherent difference in compression and decompression rates, which results in exhaustion of available A in very few cycles; 6) the "sigh", which restores stable low gamma according to the theory, actually produced unstable high gamma during virtually all phases of the maneuver. In contrast, closed bubble films of the surfactant were structurally stable and produce stable near 0 gamma and P gamma.

Animals↗
PubMed1993

Surface chemistry of binary mixtures of phospholipids in monolayers. Infrared studies of surface composition at varying surface pressures in a pulmonary surfactant model system.

Phospholipid monomolecular films at the air/water interface were studied using Langmuir-Blodgett (L-B) surface chemistry, 31P NMR spectroscopy, and infrared (IR) spectroscopy. These monolayers were composed of binary mixtures of acyl chain perdeuterated 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (i.e., DPPC-d62) with 1,2-dipalmitoyl-sn-glycero-3-phosphoglycerol (i.e., DPPG). This particular PC-PG binary mixture was chosen for study since this lipid system has been used as a model for pulmonary surfactant, especially in conjunction with the so-called "squeezing-out" hypothesis of pulmonary mechanics. This theory predicts that upon successive compression-expansion cycles, a surfactant surface film will reorganize to exclude all components except DPPC, thus resulting in a stable, low surface tension film. Several general results were obtained from these experiments. First, we have developed a combined spectroscopic assay using high-resolution 31P NMR spectroscopy in combination with the C-H and C-D vibrational intensities obtained from the IR spectroscopy of binary mixtures in which one component is acyl chain perdeuterated. Using attenuated total reflectance IR spectroscopy of transferred L-B films, this combined spectroscopic approach allows us to quantitatively describe the fractional composition of each component in the binary monomolecular film. Second, when these methods are applied to transferred monolayer films of DPPC-d62 and DPPG (at an initial PC:PG mole ratio of 7:1), we find no evidence for a "squeezing-out" of the DPPG monolayer component at high surface pressure resulting in an enrichment of the DPPC component in the transferred monolayer film.(ABSTRACT TRUNCATED AT 250 WORDS)

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1993

Identification of phosphocholine plasmalogen as a lipid component in mammalian pulmonary surfactant using high-resolution 31P NMR spectroscopy.

High-resolution 31P NMR spectroscopy was used to analyze the phospholipid composition of mammalian pulmonary surfactant from two different sources. Under conditions which considerably narrow the usually broad 31P phospholipid signals, solution-phase NMR spectra of these surfactant preparations unequivocally demonstrate that a phosphocholine plasmalogen (i.e., 1-O-(1'-alkenyl)-2-acyl-sn-glycero-3-phosphocholine) exists as a major secondary component (approximately 4 mol%) in mammalian pulmonary surfactant. Phosphocholine (PC) plasmalogen was identified in preparations obtained from both adult cow lung surfactant extract as well as in ovine (lamb) fetal pulmonary liquid. PC plasmalogens have not previously been identified any mammalian pulmonary surfactant preparation. The amount of PC plasmalogen in these preparations occurs at fractional levels that are comparable to that of phosphoglycerol (PG), which previously had been thought of as the second-most common phospholipid class in pulmonary surfactant. The presence of PC plasmalogen in pulmonary surfactant may have important physiological ramifications and immediately suggests new directions for biochemical and biophysical investigations of pulmonary surfactant.

Animals↗
PubMed1993

Fluid dynamics during initial aeration of mature fetal lung and effect of temperature.

Air volume-pressure (VP) curves were recorded simultaneously on pairs of mature rabbit fetuses from the same litter with one member of the pair at 37 degrees C and the other at 22 degrees C. Intrasaccular bubbles, formed primarily during inflation, were assessed for stability and surface tension (gamma). Average air flow rates (dV/dt) were calculated from the VP data. In separate experiments, liquid VP curves were recorded at 37 degrees and 22 degrees C: maximal liquid V was matched to maximal air V at 37 degrees and 22 degrees C, respectively. Fetal pulmonary liquid (FPL) viscosity (eta) and density (rho) were determined by standard methods. Both the effect of temperature on lung mechanics as reported previously, and the reliability of the rabbit model were confirmed in the paired fetuses. Analysis of fluid dynamics revealed that of the six parameters relevant to initial inflation-deflation of FPL-filled lungs, liquid rho, distensibility (recoil), and gamma were not altered significantly by temperature increase from 22 degrees to 37 degrees C. Enhanced lung mechanics at 37 degrees C (including enhanced inflation at lower P, higher maximal V, increased production of intrasaccular bubbles, and higher V at end-deflation) was primarily due to lowering of FPL eta at the higher temperature which appears to have an effect by augmenting bulk liquid flow and liquid drainage. Lower eta increases bulk flow through airways directly. Consequent recruitment and distention of these conducting units effectively increases radius (r) and further enhances flow. (The ultimate "brake" to airways flow at both temperatures is counter P from gamma at air/liquid menisci.).(ABSTRACT TRUNCATED AT 250 WORDS)

Air Pressure↗
PubMed1992

Temperature affects mechanics and stability during initial inflation-deflation of mature fetal lung.

Volume-pressure (VP) curves of initial aeration of mature (0.94-0.97 term) rabbit fetuses were compared in three groups, respectively, at 37 degrees C, with maximal inflation pressure of 25 cm H2O (P25); 22 degrees C, P25; and 22 degrees C, P30. Anesthetized fetuses were delivered through uterotomy; chest was opened; trachea of fetal pulmonary liquid (FPL)-filled lungs cannulated; and lungs inflated-deflated in 5 cm H2O, 2 min steps under continuous microscopic observation. As distending pressure was increased, FPL moved peripherally with airways inflation by free gas and with saccular recruitment by free gas and bubbles. Saccular aeration continued during initial reduction of P from Pmax. At end-deflation, air was retained in saccules virtually exclusively as bubbles. Airways inflation required less P at 37 degrees C, though airways volume (V) was the same at both temperatures. Opening P was lower, and saccular aeration was larger and more rapid at 37 degrees C. The apparently higher distensibility at 37 degrees C was most likely due to temperature effects on fluid dynamics rather than on tissue elasticity. Maximal V attained during early P reduction in all groups, was total lung capacity (TLC) at 37 degrees C and less than TLC at 22 degrees C. Air retention at end-deflation, with films of near-zero surface tension, was greatest at 37 degrees C and least at 22 degrees C, P25. Lung stability, greater at 37 degrees C than at 22 degrees C, was best discriminated when V at P0 was taken into account.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Fourier transform infrared studies of secondary structure and orientation of pulmonary surfactant SP-C and its effect on the dynamic surface properties of phospholipids.

SP-C, a highly hydrophobic, 3.7-kDa protein constituent of lung surfactant, has been isolated from bovine lung lavage, purified, and reconstituted into binary lipid mixtures of 1,2-dipalmitoyl-phosphatidylcholine (DPPC) and 1,2-dipalmitoylphosphatidylglycerol (DPPG). Fourier transform infrared (FT-IR) spectroscopy has been applied to examine SP-C secondary structure, the average orientation of alpha-helical segments relative to the bilayer normal in membrane films, and the effect of protein on the thermotropic properties of the phospholipid acyl chains. In addition, dynamic surface measurements were made on phospholipid films at the A/W interface in the presence and absence of SP-C. SP-C (0.5 mol %) was found to possess about 60% alpha-helical secondary structure in lipid vesicles. Higher levels (1.5 mol %) of SP-C resulted in a slight increase of beta-forms, possibly resulting from protein aggregation. The helical segments exhibited an average angle of orientation of about 24 degrees with respect to the bilayer normal, suggesting a trans-bilayer orientation of the peptide. The observation that 70% of the peptide bond hydrogens are hard to exchange in D2O further reflects the hydrophobic nature of the molecule. SP-C produced little effect on the thermotropic properties of the binary lipid mixture, as measured from acyl chain C-H and C-D stretching frequencies. However, the presence of 1 mol % protein markedly reduced the viscance and increased the elasticity of surface films suggesting a mechanism by which SP-C facilitates the spreading of phospholipids on an aqueous surface. The possible physiological consequences of these observations are discussed.

Amino Acid Sequence↗
PubMed1989

Fourier-transform infrared spectroscopy studies of lipid/protein interaction in pulmonary surfactant.

The thermotropic behavior of intact bovine lung surfactant and its hydrophobic extract has been monitored via the temperature dependence of the 2850 cm-1 phospholipid acyl chain CH2 symmetric stretching frequencies in the IR spectrum. A broad, reversible, melting event was noted from about 15 to 40 degrees C in both the lipid extract and the native surfactant. Slight protein-induced disordering of the lipid acyl chains was evident. The melting event was confirmed by differential scanning calorimetry. The major surfactant protein, a 30-36-kDa class of glycoprotein (SP-A), has been isolated from bovine lung lavage and purified by affinity chromatography. SP-A was reconstituted into a binary lipid mixture of acyl chain perdeuterated dipalmitoylphosphatidylcholine/dipalmitoylphosphatidylglycerol (DPPC-d62/DPPG, 85:15 w/w), a ratio which approximates that in surfactant. Use of DPPC-d62 permitted the FT-IR determination of the effect of protein on the thermotropic behavior of individual phospholipids in the binary mixture. High levels of SP-A induced an ordering of the phospholipids, as shown by an increase in the transition temperature of DPPC-d62 compared to the lipid model. In contrast, a mixture of the other surfactant proteins induced a progressive disordering of the phospholipids and disruption of the cooperativity of the melting event. Transition widths of about 3 degrees, 9 degrees, and 27 degrees were noted for protein:lipid ratios of 0, 1:1, and 2:1 (w/w), respectively. Possible roles for the various proteins in surfactant function are discussed in light of these data.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1989

Infrared spectroscopic investigations of pulmonary surfactant. Surface film transitions at the air-water interface and bulk phase thermotropism.

The molecular structure of the phospholipid component of intact pulmonary surfactant isolated from bovine lung lavage has been examined by Fourier transform infrared spectroscopy. Two different physical states of the surfactant were examined by means of different infrared spectroscopic sampling techniques. Transmission infrared experiments were used to study the surfactant in the bulk phase. In these experiments, the thermotropic behavior of the bulk surfactant was monitored by temperature-induced variations in the phospholipid acyl chain CH2 stretching frequencies. A broad phase transition (confirmed by differential scanning calorimetry) was noted with an onset temperature near 15 degrees C and a completion temperature near 42 degrees C. In addition to the bulk transmission experiments, external reflection infrared spectroscopy was used to examine surfactant films in situ at the air-water interface. As surface pressure was increased from 0 to 43 dyn/cm, a gradual and continuous decrease in the CH2 stretching frequency was noted for the surfactant. Thus, under surface pressures which correspond to large lung volumes in vivo, the surfactant acyl chains exist mostly in the ordered (trans) configuration. The frequency shift in the CH2 stretching mode is consistent with a continuous ordering of the acyl chains upon compression over the pressure range 0-43 dyn/cm, and implies that a weakly cooperative phase transition occurs in the hydrocarbon region of the surface film. The surface film transition is especially noted in the pressure-area curve of the surfactant and approximates in two dimensions the broad thermotropic phase transition of the bulk phase surfactant. Substantial differences were observed between the response to surface pressure changes of intact surfactant compared with the main surfactant phospholipid, 1,2-dipalmitoyl-sn--glycero-3-phosphocholine. The changes in response are attributed to the presence of additional surfactant components. The current work demonstrates the ability of infrared spectroscopy to obtain structural information on the surfactant in physical states that directly relate to those in vivo.

1,2-Dipalmitoylphosphatidylcholine↗