Bovine Lung Surfactant Extract Research Articles

Paradoxical Bactericidal Effects of Hydrophobic Lung Surfactant Proteins and Their Peptide Mimics Using Liposome Molecular Trojan.

Lung surfactant, besides alveolar stability, also provides defence against pathogens by surfactant proteins (SP), SP-A and SP-D. The hydrophobic proteins SP-B and SP-C enhance surface activity. An unusual and paradoxical effect of bovine LS and synthetic model LS with SP-B/-C was bactericidal to Staphylococcus aureus and Escherichia coli. Bacterial proliferation were investigated with bovine lung surfactant extract (BLES), dipalmitoylphosphatdylcholine, palmitooleylglycerol, in combination with SP-B/-C using standard microbiological colony forming unit (CFU) counts and structural imaging. BLES and other surfactant-SP-B/-C mixtures inhibit bacterial growth in the concentration range of 0 -7.5 mg/mL, at > 10 mg/mL paradoxical growth of both the bacterial species suggest antibiotic resistance. The lipid only LS have no effect on bacterial proliferation. Smaller peptide mimics of SP-B or SP-B1-25, were less efficient than SP-Cff. Ultra structural studies of the bacterial CFU using electron and atomic force microscopy suggest some membrane damage of S. aereus at inhibitory concentration of BLES, and some structural alteration of E. coli at dividing zones, suggesting utilization and incorporation of surfactant lipid species by both bacteria. The results depicted from in vitro studies are also in agreement with protein-protein interactions obtained from PatchDock, FireDock and ClasPro algorithm. The MD-simulation decipher a small range fluctuation of gyration radius of the LS proteins and their peptide mimics. The studies have alarming implications in the use of high dosages (100 mg/mL/kg body weight) of exogenous surfactant for treatment of respiratory distress syndrome, genetic knock-out abnormalities associated with these proteins, and the novel roles played by SP-B/C as bactericidal agents.

Orientation and depth of surfactant protein B C-terminal helix in lung surfactant bilayers

SP-BCTERM is a cationic amphipathic helical peptide and functional fragment composed of residues 63 to 78 of surfactant protein B (SP-B). Static oriented and magic angle spinning solid state NMR, along with molecular dynamics simulation was used to investigate its structure, orientation, and depth in lipid bilayers of several compositions, namely POPC, DPPC, DPPC/POPC/POPG, and bovine lung surfactant extract (BLES). In all lipid environments the peptide was oriented parallel to the membrane surface. While maintaining this approximately planar orientation, SP-BCTERM exhibited a flexible topology controlled by subtle variations in lipid composition. SP-BCTERM-induced lipid realignment and/or conformational changes at the level of the head group were observed using 31P solid-state NMR spectroscopy. Measurements of the depth of SP-BCTERM indicated the peptide center positions ~8Å more deeply than the phosphate headgroups, a topology that may allow the peptide to promote functional lipid structures without causing micellization upon compression.

Alterations In Phase And Morphology Of A Lung Surfactant Monolayer in contact with surfactant in the sub-phase induced by cholesterol and native surface active proteins

Although the presence of cholesterol, the major neutral lipid component, is well known in native surfactants (upto 10 % mass), its role in the surfactant remains uncertain. The most recently FDA approved clinical surfactant contain cholesterol, while two that have been used for 20 years have cholesterol carefully removed. However, they are all successful in treating neonatal respiratory distress syndrome (NRDS) resulting from a lack of surfactant. As a result the optimal concentration of cholesterol, if any at all, remains debated. Here we present indications for an optimal cholesterol concentration by presenting alterations to the phase and morphology of Survanta, a clinically used bovine lung surfactant extract, induced by both physiological and elevated concentrations of cholesterol when the monolayer is in contact with surfactant in the subphase. We find that low cholesterol concentrations (1-2 wt %) help to achieve a lower surface tension by enhancing surfactant material adsorption to the interface. However, increasing the cholesterol concentrations to higher values (≈ 20 wt %) significantly alters the normal surfactant isotherm. Alterations in a typical signature plateau for Survanta at ∼ 40 mN/m are noted suggesting a change in the solid phase fraction of the film. Fluorescence microscopic imaging reveals the coexistence of discrete monolayer along with “multilayer reservoir” adjacent to the air/water interface. Differences in the collapse structures of the monolayer are also noted indicating an alteration in the mechanical properties of the monolayer film. Alterations in these properties in the absence of surface active lung surfactant proteins were also observed indicating that interactions between the monolayer at the interface, the “multilayer reservoir” and the subphase are essential for the proper functioning of the lung surfactant.

Inhibitory effects of mycobacterial cell wall lipids on bovine lung surfactant extract: An in vitro study at the air–aqueous interface

Lung surfactant is a lipoprotein mixture synthesized by the type II alveolar cells in the lungs and lines the air–aqueous interface. It is essential for stabilizing the pulmonary alveoli. Deficiency or dysfunction of the surfactant due to chemicals or toxins results in alveolar collapse and if untreated leads to respiratory distress. In pulmonary tuberculosis, Mycobacterium tuberculosis bacteria reside in the alveoli, in close physical proximity with the alveolar surfactant. The surface active mycobacterial cell wall lipids may easily detach from bacterial cell wall and interact with surfactant components altering their surface activity. In this study, effects of mycolic acid and cord factor (the abundant surface lipids of mycobacterial cell wall) on the surface activity of modified bovine surfactant extract (Survanta™) were characterized in vitro using the Wilhelmy surface balance at physiological temperature (37 °C) and pH 7.4. Inhibitory effects of mycobacterial cell wall lipids on the terminal airway surfactant were evaluated using a capillary surfactometer. Addition of graded amounts of mycolic acid and cord factor significantly increased the minimum surface tension of Survanta™ monolayer from <1 mN/m to ∼11–20 mN/m. Presence of mycobacterial cell wall lipids decreased the rate and magnitude of Survanta™ adsorption and fluidized the surfactant monolayer. Capillary surfactometer study exhibited 100% capillary closure on addition of mycobacterial cell wall lipids to the modified bovine surfactant extract. AFM imaging revealed alteration of surface topography and presence of aggregates on addition of mycobacterial cell wall lipids to Survanta™ monolayer. This study suggests a biophysical inhibition of lung surfactant which may play a role in the pathogenesis of pulmonary tuberculosis.

Treatment of Premature Calves with Clinically Diagnosed Respiratory Distress Syndrome

Respiratory distress syndrome (RDS) has been reported previously in premature calves. However, there have been no published data on the effect of surfactant replacement therapy in the treatment of premature calves with RDS. Surfactant replacement therapy added to the standard treatment for premature calves clinically diagnosed with RDS would increase the viability of the calves. Twenty-seven premature calves with clinically diagnosed RDS. Twenty calves were instilled intratracheally with bovine lung surfactant extract and provided with standard treatment for RDS (surfactant group). Seven calves were given only standard care for RDS without surfactant therapy and placed in the control group. Standard treatment for newborn calves with RDS includes warming, administration of intranasal oxygen, fluid replacement, administration of antibiotics, and immunoglobulin solution. Arterial blood samples were collected from the calves at 3 observation points, the first just before treatment (hour 0) and at 2 hours (hour 2) and 24 hours (hour 24) after treatment was started to determine if ventilation was adequate, improving, or deteriorating. Blood gases, pH, bicarbonate, and lactate concentrations were measured. In the surfactant group, mean partial pressure of oxygen significantly increased at hours 2 and 24. Mean partial pressure of carbon dioxide decreased and mean arterial blood pH increased at hour 24 in the surfactant group compared with the control group (P < .05). Of the 20 calves in the surfactant group, 12 survived and 8 died. All 7 calves in the control group died. The results of this study suggest that surfactant replacement therapy may reduce neonatal deaths in premature calves with clinically diagnosed RDS.

Detection of tobacco smoke deposition by hyperpolarized krypton-83 MRI

Despite the importance of the tobacco smoke particulate matter in the lungs to the etiology of pulmonary disease in cigarette smokers, little is currently known about the spatial distribution of particle deposition or the persistence of the resulting deposits in humans, and no satisfactory technique currently exists to directly observe tobacco smoke condensate in airways. In this proof-of-principle work, hyperpolarized (hp) 83Kr MRI and NMR spectroscopy are introduced as probes for tobacco smoke deposition in porous media. A reduction in the hp-83Kr longitudinal (T1) relaxation of up to 95% under near-ambient humidity, pressure and temperature conditions was observed when the krypton gas was brought into contact with surfaces that had been exposed to cigarette smoke. This smoke-induced acceleration of the 83Kr self-relaxation was observed for model glass surfaces that, in some experiments, were coated with bovine lung surfactant extract. However, a similar effect was not observed with hp-(129)Xe indicating that the 83Kr sensitivity to smoke deposition was not caused by paramagnetic species but rather by quadrupolar relaxation due to high adsorption affinity for the smoke deposits. The 83Kr T1 differences between smoke-treated and untreated surfaces were sufficient to produce a strong contrast in variable flip angle FLASH hp-83Kr MRI, suggesting that hp-83Kr may be a promising contrast agent for in vivo pulmonary MRI.

Thermodynamic studies of bovine lung surfactant extract mixing with cholesterol and its palmitate derivative

Langmuir film behavior of bovine lipid extract surfactant (BLES), mixed with cholesterol (CHOL) and cholesterol palmitate (CHOLP), has been studied by surface pressure ( π)–area ( A) measurements. Associative interactions, observed for both systems, were less favored at lower BLES content. The presence of unsaturated phospholipids and surfactant proteins in BLES favored the association. Miscibility of BLES was better with CHOLP than with CHOL at all compositions, indicating more compact packing of the BLES–CHOLP than of the BLES–CHOL system. The most stable mixtures were found at 30–40 mol% CHOL and at low π and at 20–25 mol% CHOLP but at higher π. These results suggest that BLES–CHOL miscibility is better at low π and low CHOL concentrations, while BLES–CHOLP miscibility is better at high π and high CHOLP concentrations.

More Than a Monolayer: Relating Lung Surfactant Structure and Mechanics to Composition

Survanta, a clinically used bovine lung surfactant extract, in contact with surfactant in the subphase, shows a coexistence of discrete monolayer islands of solid phase coexisting with continuous multilayer “reservoirs” of fluid phase adjacent to the air-water interface. Exchange between the monolayer, the multilayer reservoir, and the subphase determines surfactant mechanical properties such as the monolayer collapse pressure and surface viscosity by regulating solid-fluid coexistence. Grazing incidence x-ray diffraction shows that the solid phase domains consist of two-dimensional crystals similar to those formed by mixtures of dipalmitoylphosphatidylcholine and palmitic acid. The condensed domains grow as the surface pressure is increased until they coalesce, trapping protrusions of liquid matrix. At ∼40 mN/m, a plateau exists in the isotherm at which the solid phase fraction increases from ∼60 to 90%, at which the surface viscosity diverges. The viscosity is driven by the percolation of the solid phase domains, which depends on the solid phase area fraction of the monolayer. The high viscosity may lead to high monolayer collapse pressures, help prevent atelectasis, and minimize the flow of lung surfactant out of the alveoli due to surface tension gradients.

Thermodynamic studies on mixed molecular langmuir films: Part 2. Mutual mixing of DPPC and bovine lung surfactant extract with long-chain fatty acids

Mixing of long-chain saturated fatty acids (FAs), from dodecanoic (lauric) to octadecanoic (stearic) acid with a disaturated phospholipid, dipalmitoylphosphatidylcholine (DPPC), and the lipid-extracted materials from bovine lung surfactant (BLES), a clinical pulmonary surfactant (PS) formulation, have been studied at the air–water interface using Langmuir surface balance. BLES is a natural mixture of both saturated (solid) and unsaturated (fluid) lipids having DPPC as its major constituent; besides it contains two hydrophobic proteins SP-B and -C that help in better surface activity. Surface pressure–area behavior of DPPC and BLES, mixed with different proportions of FA, were studied. While DPPC–FA mixtures showed positive deviations from ideality, BLES–FA mixtures showed negative deviations irrespective of the chain length of FAs. Minimum repulsion was observed for palmitic acid in DPPC–FA mixtures while maximum association occurred with palmitic acid for BLES–FA mixtures, especially at higher mole fractions of FAs, as revealed from the excess free energy or free energy of mixing. The miscibilities are discussed in terms of the chain lengths of the FAs, the size of lipid and the composition of BLES.

Thermodynamic studies on mixed molecular langmuir filmsPart 2. Mutual mixing of DPPC and bovine lung surfactant extract with long-chain fatty acids

Mixing of long-chain saturated fatty acids (FAs), from dodecanoic (lauric) to octadecanoic (stearic) acid with a disaturated phospholipid, dipalmitoylphosphatidylcholine (DPPC), and the lipid-extracted materials from bovine lung surfactant (BLES), a clinical pulmonary surfactant (PS) formulation, have been studied at the air–water interface using Langmuir surface balance. BLES is a natural mixture of both saturated (solid) and unsaturated (fluid) lipids having DPPC as its major constituent; besides it contains two hydrophobic proteins SP-B and -C that help in better surface activity. Surface pressure–area behavior of DPPC and BLES, mixed with different proportions of FA, were studied. While DPPC–FA mixtures showed positive deviations from ideality, BLES–FA mixtures showed negative deviations irrespective of the chain length of FAs. Minimum repulsion was observed for palmitic acid in DPPC–FA mixtures while maximum association occurred with palmitic acid for BLES–FA mixtures, especially at higher mole fractions of FAs, as revealed from the excess free energy or free energy of mixing. The miscibilities are discussed in terms of the chain lengths of the FAs, the size of lipid and the composition of BLES.

In vitro and in vivo intrapulmonary distribution of fluorescently labeled surfactant

To determine the distribution of endotracheally administered surfactant at the alveolar level in an animal model of acute respiratory distress syndrome. Prospective, randomized animal study. Research laboratory of a university hospital. Seventy-one male Sprague-Dawley rats, weighing 330-370 g. To measure surfactant distribution in vitro, a glass trough mimicking dichotomic lung anatomy was used to determine the spreading properties of bovine lung surfactant extract supplemented with fluorescent Bodipy-labeled surfactant protein B. To measure surfactant distribution in vivo, rats were anesthetized, and lipopolysaccharide was aerosolized (12 mg/kg body weight) to induce lung injury resembling acute respiratory distress syndrome; in control rats, buffered saline was aerosolized. Twenty-four hours later rats were anesthetized, tracheotomized, and mechanically ventilated (peak airway pressure = 20 mbar; positive end-expiratory pressure = 6 mbar; inspiration time = expiration time = 0.6 sec; Fio2 = 50%). Surfactant (bovine lung surfactant extract, supplemented with fluorescent Bodipy-labeled surfactant protein B; 50 mg/kg body weight) was applied as a bolus; in control rats, saline was administered as a bolus. Rats were ventilated for 5, 15, 30, or 60 mins (n = 8 or 9 for each group). Then, lungs were excised and sliced. Lung slices, divided into aerated (open), underinflated (dystelectatic), or collapsed (atelectatic) alveolar areas, were examined by both light and fluorescence microscopy. In vitro experiments revealed that surfactant spread independent of glass trough geometry and lowered the surface tension to equilibrium values (25 mN/m) within a few seconds. In vivo experiments showed that administered surfactant distributed preferentially into underinflated and aerated alveolar areas. Furthermore, surfactant distribution was not affected by length of mechanical ventilation. When conventional mechanical ventilation was used in lipopolysaccharide-induced lung injury, surfactant preferentially distributed into underinflated and aerated alveolar areas. Because surfactant rarely reached collapsed alveolar areas, methods aiding in alveolar recruitment (e.g., open lung concept or body positioning) should precede surfactant administration.

Surfactant (beractant) therapy for infants with congenital diaphragmatic hernia on ECMO: Evidence of persistent surfactant deficiency

Infants with congenital diaphragmatic hernia (CDH) on extracorporeal membrane oxygenation (ECMO) can have initial lung atelectasis which, in survivors, gradually improves over time. To test the hypothesis that these patients could benefit from surfactant therapy, infants with CDH (born at >34 weeks' gestation) on ECMO received either four doses of modified bovine lung surfactant extract (beractant) (surfactant group, n = 9) or an equal volume of air (control group, n = 8). Tracheal aspirate surfactant protein-A (SP-A) concentrations were initially low, and then increased over time in both CDH groups ( P = .0021); however, levels remained low when compared with those of infants on ECMO who had other diagnoses ( P = .04). Lung compliance (C L), time to extubation, time on oxygen, and total no. of hospital days were not different between the two groups. Infants with CDH had persistently elevated right ventricular pressure (RVP) at cessation of bypass when compared with non-CDH infants on ECMO (RVP = 53.25 mm Hg ± 19.52 in the CDH group, 32.90 ± 10.63 in the non-CDH group; P = .0121). The findings suggest that the postnatal surfactant deficiency may be more persistent in CDH infants than in non-CDH infants on ECMO. However, CDH remains a multifactorial condition, with delayed improvement, because of persistence of pulmonary hypertension, difficulties with vascular remodeling, degree of lung hypoplasia, or compromised respiratory mechanics.

Improved pulmonary outcome after exogenous surfactant therapy for respiratory failure in term infants requiring extracorporeal membrane oxygenation

A blinded, randomized, controlled study was designed to test whether multiple-dose surfactant therapy would improve pulmonary outcome in term infants with respiratory failure, resulting in a shortened period of extracorporeal membrane oxygenation (ECMO). Infants > or = 34 weeks of gestational age in severe respiratory failure and receiving ECMO were stratified by diagnosis and then randomly assigned to the treatment or the control group. Four doses of modified bovine lung surfactant extract (beractant) were administered to the surfactant group (n = 28), and an equal volume of air was administered to the control group (n = 28). Lung compliance was initially low in both groups; after treatment, values were higher with time in the surfactant group (F = 5.40, p = 0.026). The ECMO treatment period was significantly shorter in the surfactant group (mean +/- SD: 107 +/- 33 hours vs 139 +/- 54 hours for the control group; U = 232, p = 0.023). Tracheal aspirate concentrations of surfactant protein A were low in both groups, and then increased steadily to a higher level in the surfactant group (F = 2.58, p = 0.04). The overall incidence of complications after ECMO was decreased in the surfactant group (18% vs 46% for the control group; chi-square value = 5.004, p = 0.025). Radiographic scores, echocardiographic findings, incidence of intracranial or pulmonary hemorrhage and bronchopulmonary dysplasia, time to extubation, duration of oxygen therapy, and duration of hospitalization did not differ between the two groups. Beractant in this population improved pulmonary mechanics, increased surfactant protein A content in tracheal aspirate, decreased time on ECMO duration, and reduced disease complications.

Bovine Surfactant Replacement Therapy in Neonates of Less Than 30 Weeks' Gestation: a Randomized Controlled Trial of Prophylaxis Versus Treatment

The influence of the timing of surfactant replacement therapy for the treatment of neonatal respiratory distress syndrome was evaluated in a study of 182 neonates of less than 30 weeks' gestation who were randomly assigned prior to delivery to one of three study groups: control (dummy instillation of air given at birth), early surfactant (surfactant given at birth), or late surfactant (surfactant given at less than 6 hours of age). Subjects in the late surfactant group could avoid treatment if they had a clear chest roentgenogram and required no supplemental oxygen at a mean airway pressure of less than 7 cm of water. All treated neonates were eligible to receive up to three additional doses during the first 5 days of life. The three groups were comparable with respect to birth weight, gestational age, and other perinatal parameters with the exception of a lower cord arterial pH and 1-minute Apgar score in the early surfactant group. Of the 60 neonates randomly assigned to late treatment, 29 (48%) were deemed surfactant sufficient and thereby avoided treatment; the other 31 received their first dose at a mean age of 2.9 hours. There was a significant improvement in gas exchange during the first week of life in both surfactant groups compared with the control group, reflected by differences in fraction of inspired oxygen, arterial/alveolar PO2, and ventilation index (peak pressure x rate on the ventilator) (P less than .001). Surfactant therapy also resulted in a lower incidence of pulmonary air leak and severe chronic lung disease (defined as requirement for respiratory support beyond 36 weeks post-conceptional age). There were no differences between early and late surfactant groups in any of these parameters. The only statistically significant difference between the surfactant groups was that the early group had a higher incidence of mild chronic lung disease (respiratory support beyond 28 days of age) than the late treatment group (P less than .005). Neonates in the late treatment group were extubated earlier and had a shorter neonatal intensive care unit stay than control neonates (P less than .05), whereas those in the early group were not significantly different from control neonates in these parameters. It is concluded that replacement therapy with bovine lung surfactant extract in neonates of less than 30 weeks' gestation results in decreased oxygen and ventilatory requirements during the first week of life and a lower incidence of pulmonary air leak and severe chronic lung disease.(ABSTRACT TRUNCATED AT 400 WORDS)