Synthetic surfactant with a recombinant surfactant protein C analogue improves lung function and attenuates inflammation in a model of acute respiratory distress syndrome in adult rabbits.

AIM: In acute respiratory distress syndrome (ARDS) damaged alveolar epithelium, leakage of plasma proteins into the alveolar space and inactivation of pulmonary surfactant lead to respiratory dysfunction. Lung function could potentially be restored with exogenous surfactant therapy, but clinical trials have so far been disappointing. These negative results may be explained by inactivation and/or too low doses of the administered surfactant. Surfactant based on a recombinant surfactant protein C analogue (rSP-C33Leu) is easy to produce and in this study we compared its effects on lung function and inflammation with a commercial surfactant preparation in an adult rabbit model of ARDS. METHODS: ARDS was induced in adult New Zealand rabbits by mild lung-lavages followed by injurious ventilation (VT 20 m/kg body weight) until P/F ratio < 26.7 kPa. The animals were treated with two intratracheal boluses of 2.5 mL/kg of 2% rSP-C33Leu in DPPC/egg PC/POPG, 50:40:10 or poractant alfa (Curosurf®), both surfactants containing 80 mg phospholipids/mL, or air as control. The animals were subsequently ventilated (VT 8-9 m/kg body weight) for an additional 3 h and lung function parameters were recorded. Histological appearance of the lungs, degree of lung oedema and levels of the cytokines TNFα IL-6 and IL-8 in lung homogenates were evaluated. RESULTS: Both surfactant preparations improved lung function vs. the control group and also reduced inflammation scores, production of pro-inflammatory cytokines, and formation of lung oedema to similar degrees. Poractant alfa improved compliance at 1 h, P/F ratio and PaO2 at 1.5 h compared to rSP-C33Leu surfactant. CONCLUSION: This study indicates that treatment of experimental ARDS with synthetic lung surfactant based on rSP-C33Leu improves lung function and attenuates inflammation.

Synthetic surfactants with SP-B and SP-C analogues to enable worldwide treatment of neonatal respiratory distress syndrome and other lung diseases.

Treatment of neonatal respiratory distress syndrome (RDS) using animal-derived lung surfactant preparations has reduced the mortality of handling premature infants with RDS to a 50th of that in the 1960s. The supply of animal-derived lung surfactants is limited and only a part of the preterm babies is treated. Thus, there is a need to develop well-defined synthetic replicas based on key components of natural surfactant. A synthetic product that equals natural-derived surfactants would enable cost-efficient production and could also facilitate the development of the treatments of other lung diseases than neonatal RDS. Recently the first synthetic surfactant that contains analogues of the two hydrophobic surfactant proteins B (SP-B) and SP-C entered clinical trials for the treatment of neonatal RDS. The development of functional synthetic analogues of SP-B and SP-C, however, is considerably more challenging than anticipated 30 years ago when the first structural information of the native proteins became available. For SP-B, a complex three-dimensional dimeric structure stabilized by several disulphides has necessitated the design of miniaturized analogues. The main challenge for SP-C has been the pronounced amyloid aggregation propensity of its transmembrane region. The development of a functional non-aggregating SP-C analogue that can be produced synthetically was achieved by designing the amyloidogenic native sequence so that it spontaneously forms a stable transmembrane α-helix.

Therapeutic effects of exogenous surfactant enriched with dextran in newborn rabbits with respiratory failure induced by airway instillation of albumin.

The aim of the study was to investigate lung function and morphology in newborn rabbits with acute respiratory failure induced by airway instillation of albumin and to test whether the therapeutic effect of exogenous modified natural surfactant could be enhanced by addition of dextran. Rabbits (gestational age 29 days) were ventilated with 100% O2 and tidal volume 8-10 ml/kg. Respiratory failure was induced by tracheal instillation of albumin (50 or 100mg/ml; 2 ml/kg) and the instillation was repeated 15 min later. Surfactant (Curosurf, 200mg/kg) with or without addition of dextran (30 mg/ml) was administered 15 min after the second dose of albumin and animals were then ventilated for another 60 min. In one series of experiments, instillation of albumin was followed by administration of diluted Curosurf (40 mg/kg) with or without dextran (30 mg/ml). Animals with initial lung-thorax compliance (CLT)<0.6 ml/kg cm H2O were directly treated with surfactant without previous instillation of albumin. Animals treated with a larger dose of Curosurf (200mg/kg) showed a significant increase in CLT at 60 min with no difference between groups receiving high or low dose of albumin. Addition of dextran to Curosurf at high dose (200mg/kg) reduced the therapeutic response (P<0.05 vs. Curosurf without dextran). After treatment with low-dose Curosurf (40 mg/kg) CLT increased more prominently after Curosurf+dextran than after non-enriched surfactant (P<0.05). Dextran potentiated the treatment effect of Curosurf at suboptimal dose (40 mg/kg) but not at high dose (200mg/kg) in animals with low-grade surfactant deficiency or respiratory failure induced by airway instillation of albumin.

Biophysical and physiological properties of porcine surfactant enriched with polymyxin B.

OBJECTIVE: We examined whether the biophysical and physiological properties of Curosurf were improved by the cyclic amphipathic decapeptide polymyxin B (PxB). METHODS: Curosurf was diluted to 1-5 mg/ml with PxB added at 1, 2 or 3% (w/w). Albumin was added at 40 mg/ml. Minimum surface tension (gammamin) during surface compression was determined for each mixture with pulsating bubble. Immature newborn rabbits were treated with 2.5 ml/kg of Curosurf 80 mg/ml, or Curosurf 32 mg/ml with or without 2% PxB and ventilated for up to 5 h. RESULTS: At surfactant concentration 2 mg/ml, gammamin was high (17 +/- 8.9 mN/m) but remained low (2.7 +/- 0.8 mN/m) when PxB was added. Albumin inactivated Curosurf at both 2 and 3.5 mg/ml; this inactivation was prevented by 2% PxB. Treatment of newborn rabbits with Curosurf 80 mg/kg + 2% PxB significantly decreased incidence of pneumothorax in comparison with controls but had no significant effect on lung-thorax compliance or alveolar expansion. CONCLUSION: Addition of 2% PxB improves surface activity of Curosurf at low concentration, increases its resistance to inactivation by albumin, and reduces the incidence of pneumothorax in immature newborn rabbits undergoing prolonged ventilation.

A lung recruitment maneuver immediately before rescue surfactant therapy does not affect the lung mechanical response in immature lambs with respiratory distress syndrome.

BACKGROUND: In animals with acquired surfactant-deficiency, a recruitment maneuver by increased tidal volumes enhances the effect of exogenous surfactant. In contrast, in the preterm lamb model, hyperinflation early after birth impairs the effect of surfactant prophylaxis. Here we examined whether a lung recruitment maneuver just before surfactant would affect the response to rescue treatment in immature lambs with established respiratory distress syndrome (RDS). METHODS: Five pairs of preterm twin lambs with gestational age 127 days were delivered by cesarean section and supported by pressure-limited mechanical ventilation for 4 h. At 30 min of age, when all the lambs were in severe respiratory failure, they were treated with porcine surfactant, 200 mg x kg-1. One lamb in each pair was subjected to a lung recruitment maneuver consisting of five sustained inflations of 20 ml x kg-1 just before surfactant instillation. RESULTS: At 10 min after surfactant treatment, all the lambs showed a large improvement in oxygenation and an increase in inspiratory capacity and static compliance. Except for a transiently better oxygenation after surfactant therapy in the recruitment group (P < 0.05), there was no significant between-group differences in gas exchange or lung mechanics at any time point during the study. There was no difference in post mortem intrapulmonary air volume or alveolar expansion in histologic lung sections between groups. CONCLUSION: This small study does not show any positive or negative effect of a lung recruitment maneuver on the response to rescue surfactant therapy in immature animals with RDS.

Artificial surfactants based on analogues of SP-B and SP-C.

The hydrophobic proteins SP-B and SP-C are important components of natural surfactant preparations currently used in clinical practice, and physiologically active surfactants can be made from isolated SP-B and/or SP-C reconstituted with synthetic lipids. Efforts have been made to produce these polypeptides, or analogues with similarfunction, by organic synthesis or expression in heterologous systems. It is important to obtain proper folding of the synthetic peptides, as required for optimal interaction with the surfactant lipids. Another issue is to avoid loss of SP-C activity due to alpha-helix to beta-sheet transition. This latter problem can be circumvented by replacing the polyvaline stretch of SP-C with a polyleucine stretch containing a few lysines. Palmitoylation of cysteines or serines at positions 5 and 6 also seems important for the properties of SP-C. SP-B, which is too big a molecule to be easily produced by organic synthesis. apparently can be replaced in an artificial surfactant by a peptide capable of cross-linking phospholipid bilayers. The development of synthetic analogues of the surfacant proteins might make it possible to tailor artificial surfactants for specific therapeutic missions, for instance by enhancing resistance to inactivation by meconium, plasma proteins, or oxygen radicals or maximizing bacteriostatic effects.

Lung recruitment at birth does not improve lung function in immature lambs receiving surfactant.

BACKGROUND: In mature animals with surfactant deficiency induced by lung lavage, the therapeutic effect of exogenous surfactant is enhanced by a lung recruitment maneuver. We then tested whether a lung recruitment maneuver at birth immediately before surfactant treatment would improve lung function also in preterm lambs with surfactant deficiency due to immaturity. METHODS: Ten newborn lambs with a gestational age of 127 days were randomized to receive surfactant either before the first breath or immediately after a lung recruitment maneuver consisting of five sustained inflations of 8, 16 or 32 ml/kg. Functional residual capacity was measured by sulfur hexafluoride washout, and inspiratory capacity as well as maximal compliance were obtained from a static expiratory pressure-volume curve after the lungs had been inflated to 35 cm H2O. In addition, blood gases were obtained. Measurements were made at 15, 45, 175, 135, 170 and 230 min after birth. Post mortem histological examinations of the lungs were performed in a blinded fashion. RESULTS: The lung recruitment maneuvers did not improve oxygenation. Inspiratory capacity, static compliance and functional residual capacity at 4 h, as well as post mortem intrapulmonary air volume, had an inverse relation to the size of inflations given at birth. There was also a negative correlation between size of inflations at birth and response to surfactant therapy, as assessed by lung microscopy. CONCLUSION: Lung recruitment at birth does not improve the response to surfactant in immature lambs, but may instead have an adverse effect on lung function and morphology.

Surface activity and film formation from the surface associated material of artificial surfactant preparations.

Surfactant proteins B and C (SP-B and SP-C) are present in natural derived surfactant preparations used for treatment of respiratory distress syndrome. Herein the surface activity of an SP-C analogue (SP-C(LKS)), a hybrid peptide between SP-C and bacteriorhodopsin (SP-C/BR) and a model peptide (KL(4)) was studied with a captive bubble surfactometer (CBS). The peptides were mixed with either 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC)/phosphatidylglycerol (PG) (7:3, by weight) or DPPC/PG/palmitic acid (68:22:9, by weight) at a concentration of 1 mg/ml in HEPES buffer, pH 6.9 and a polypeptide/lipid weight ratio of 0.02--0.03. In some lipid/peptide preparations also 2% of SP-B was included. Adsorption, monitored as surface tension vs. time for 10 min after bubble formation did not show discernible differences for the whole set of preparations. Equilibrium surface tensions of approximately 25 mN/m were reached after 5--10 min for all preparations, although those with SP-C/BR appeared not to reach end point of adsorption within 10 min. Area compression needed to reach minimum surface tension of 0.5--2.0 mN/m was least for the KL(4) preparation, about 13% in the first cycle. 3% SP-C(LKS) in DPPC:PG (7:3, by weight) reached minimum surface tension upon 27% compression in the first cycle. If DPPC:PG:PA (68:22:9, by weight) was used instead only 16% area compression was needed and 14% if also 2% SP-B was included. 3% SP-C(LKS) in DPPC:PG (7:3, by weight)+2% SP-B needed 34% compression to reach minimum surface tension. The replenishment of material from a surface associated surfactant reservoir was estimated with subphase depletion experiments. With the 2% KL(4) preparation incorporation of excess material took place at a surface tension of 25--35 mN/m during stepwise bubble expansion and excess material equivalent to 4.3 monolayers was found. When 2% SP-B was added to 3% SP-C(LKS) in DPPC:PG (7:3, by weight) the number of excess monolayers increased from 1.5 to 3.6 and the incorporation took place at 30--40 mN/m. When SP-B was added to 3% SP-C(LKS) in DPPC:PG:PA (68:22:9, by weight) the number of excess monolayers increased from 0.5 to 3.4 and incorporation took place at 40--50 mN/m. With 2% SP-C/BR incorporation took place at 40--45 mN/m, frequent instability clicks were observed and excess material of approximately 1.1 monolayer was estimated.

Pilot study of nebulized surfactant therapy for neonatal respiratory distress syndrome.

Thirty-four spontaneously breathing newborns with respiratory distress syndrome (RDS) requiring nasal continuous positive airway pressure (CPAP) and an arterial-to-alveolar oxygen tension ratio (a/A PO2) of 0.15-0.22 were randomized to treatment with nebulized surfactant (Curosurf) or to serve as controls. All children were first supported by nasal CPAP according to normal clinical routines. Surfactant was administered using a modified Aiolos nebulizer, and a total of 480 mg was aerosolized in each case. The control group received no nebulized material, but had the same CPAP support. Acid-base status and a/A PO2 were determined at regular intervals before, during and after surfactant administration. Both groups included in the study were similar with regard to gestational age, birthweight, steroids given before birth, sex and Apgar scores as well as a/A PO2 when entering the study. There were no significant differences between the groups in a/A PO2 1-12 h after randomization, number of infants needing mechanical ventilation, time on ventilator or CPAP. Two children in the treated group developed bronchopulmonary dysplasia. No side effects of the surfactant therapy were noted. No beneficial effects of aerosolized surfactant were demonstrated in our trial, contrary to data from animal experiments. This finding probably reflects differences in administration techniques. Our findings do not justify large clinical trials with the same protocol. Further work is needed to optimize delivery of aerosolized surfactant to the neonatal lung in clinical practice.

Palmitoylation of a pulmonary surfactant protein C analogue affects the surface associated lipid reservoir and film stability.

Surfactant protein C (SP-C) is a lipopeptide that contains two thioester-linked palmitoyl groups and is considered to be important for formation of the alveolar surface active lipid film. Here, a non- or dipalmitoylated SP-C analogue (SP-C(Leu)), in which all helical Val residues were replaced with Leu and Cys-5 and Cys-6 were replaced with Ser, was tested for surface activity in a captive bubble system (CBS). SP-C(Leu), either palmitoylated at Ser-5 and Ser-6 or non-palmitoylated, was added to mixtures of 1, 2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC)/phosphatidyl glycerol (PG)/palmitic acid (PA), 68:22:9, (by mass) at a concentration of 2 and 5%. With 2% peptide, surface film formation was rapid, reaching a surface tension below 25 mN/m within 5 s, but the samples with 5% SP-C(Leu) required more than 20 s to reach values below 25 mN/m. Minimum surface tension for the samples with dipalmitoylated SP-C(Leu) was below 1.5 mN/m and very stable, as the surface tension increased by less than 0.5 mN/m within 10 min at constant bubble volume. Minimum surface tension for the non-palmitoylated SP-C(Leu) was approximately 2 and 5 mN/m for 2 and 5% peptide, respectively, but the films were less stable as seen by frequent bubble clicking at low surface tensions. Films with dipalmitoylated SP-C(Leu) that were dynamically cycled at 20-30 cycles/min were substantially less compressible at a surface tension of 20 mN/m (0.007 m/mN) than those that contained the non-palmitoylated peptide (0.02 m/mN). After subphase depletion, the incorporation of lipids into the surface active film during initial bubble expansion occurred at a relatively low surface tension (about 35 mN/m) for the samples with dipalmitoylated SP-C(Leu) compared to approximately 45 mN/m for those containing the non-palmitoylated peptide. Furthermore, for samples that contained non-palmitoylated SP-C(Leu), the ability to reach near zero stable surface tension was lost after a few adsorption steps, whereas with the dipalmitoylated peptide the film quality did not deteriorate even after more than 10 expansion steps and the incorporation of reservoir material equivalent to more than two monolayers. It appears that the covalently linked palmitoyl groups of the SP-C analogue studied are important for the mechanical stability of the lipid film, for the capacity to incorporate material from the reservoir into the surface active film upon area expansion, and for the low film compressibility of dynamically cycled films.

Pulmonary surfactant protein B: a structural model and a functional analogue.

Surfactant proteins B and C (SP-B and SP-C), together with phospholipids, are important constituents of pulmonary surfactant and of preparations used for treatment of respiratory distress syndrome (RDS). SP-B belongs to the saposin family of homologous proteins, which include other lipid-interacting proteins, like the membranolytic NK-lysin. SP-B, in contrast to other saposins, is hydrophobic and a disulfide-linked dimer, and its mechanism of action is not known. A model of the three-dimensional structure of one SP-B subunit was generated from the structure of monomeric NK-lysin determined by nuclear magnetic resonance, and the SP-B dimer was formed by joining two subunits via the intersubunit disulfide bond Cys48-Cys48'. After energy minimization, intersubunit hydrogen bonds/ion pairs were formed between the strictly conserved residues Glu51 and Arg52, which creates a central non-polar region located in between two clusters of positively charged residues. The structural features support a function of SP-B in cross-linking of lipid membranes. Mixtures of phospholipids, an SP-C analogue and polymyxin B (which cross-links lipid vesicles but is structurally unrelated to SP-B) exhibit in vitro surface activity which is indistinguishable from that of analogous mixtures containing SP-B instead of polymyxin B. This suggests an avenue for identification of SP-B analogues that can be used in synthetic surfactants for treatment of RDS.

Aerosolized surfactant in lung-lavaged adult rats: factors influencing the therapeutic response.

OBJECTIVE: To evaluate the effect of aerosolized modified natural surfactant in adult rats with respiratory failure. METHODS: Lung-lavaged adult rats were treated with aerosolized surfactant, aerosolized saline or a bolus of surfactant. Surfactant was labelled with dimyristoylphosphatidylcholine (DMPC) and human serum albumin was given intravenously for evaluation of lung protein leakage. Blood gases and dynamic compliance were measured intermittently. At the end of ventilation, the lungs were either fixed by vascular perfusion for histological examination or washed for determination of total phospholipids, DMPC and human albumin in the lavage fluid. RESULTS: Treatment with bolus surfactant led to a quick and sustained restoration of pre-lavage blood gas values in most animals. The response to aerosolized surfactant varied considerably, with an overall moderate improvement of gas exchange. The saline-treated group failed to show any significant recovery of lung function. No histopathological differences were found between any of the groups. On average 0.46% of total administered aerosolized surfactant could be recovered. Vascular-to-alveolar leakage of human albumin averaged 11%, with no significant differences between the groups. Final values for PaO2 were significantly correlated with total phospholipids in the lavage fluid, and inversely related to the vascular-to-alveolar leakage of albumin. CONCLUSION: Neither bolus nor aerosolized surfactant influenced lung morphology. Nebulized surfactant improved lung function but the effect was inferior to that obtained with bolus surfactant, and the outcome depended on the balance between the combined pool size of exogenous and endogenous surfactant and the vascular-to-alveolar leakage of serum protein.

Synthetic surfactants to treat neonatal lung disease.

Pulmonary surfactant is a complex of surface-active lipids mixed with specific proteins. Two of these, SP-B and SP-C, are essential for adsorption of surfactant lipids to the air-liquid interfaces of the lungs and, hence, are also essential for alveolar stability and effective gas exchange. Surfactant substitutes must contain at least one of these proteins (or analogues of them) to be optimally effective when administered into the airways of babies with surfactant deficiency or dysfunction. This review describes how an increased understanding of the properties of surfactant proteins has led to the development of improved synthetic surfactants with the potential to treat a wide range of respiratory disorders.

Prospects for a new synthetic surfactant.

Physiologically active artificial surfactants can be made from lipids and analogs of SP-C and SP-B, the latter serving as cross-linking peptides promoting the formation of a surface-associated surfactant reservoir. Artificial surfactants can probably be tailored to maximize resistance to inactivation by meconium and plasma proteins by modifying the amino acid sequence of the protein analogs or adding non-ionic polymers to the final product. Comparative clinical trials are required to evaluate the efficacy of these new artificial surfactants in various forms of neonatal lung disease.

Porcine pulmonary surfactant preparations contain the antibacterial peptide prophenin and a C-terminal 18-residue fragment thereof.

Surfactant preparations obtained from porcine lungs by extraction with chloroform/methanol followed by chromatography over Lipidex-5000 are used for treatment of respiratory distress syndrome in preterm infants. These preparations contain about 98% phospholipids and 1-2% of the hydrophobic pulmonary surfactant-associated proteins B and C (SP-B and SP-C). Separation of the proteins in the surfactant preparation by reversed-phase high performance liquid chromatography revealed, in addition to SP-B and SP-C, the presence of three peptides derived from the cathelicidin family of antibacterial peptides. The 79-residue proline-rich peptide prophenin (identical to that isolated from leukocytes), an 80-residue prophenin with an N-terminal pyroglutamic acid residue, and a C-terminal 18-residue fragment of prophenin were found in approximate molar ratios of 1:20:5. A synthetic version of the C-terminal 18-residue peptide exhibits salt-dependent antibacterial activity (higher activity in the absence of salt) against the Gram-positive bacterium Bacillus megaterium Bm11 and, to a lesser extent, against Gram-negative Escherichia coli D21 cells. It appears possible that the presence of prophenin peptides may contribute to the antibacterial properties of surfactant preparations.

Computerized image analysis of lung expansion patterns in surfactant treated immature newborn rabbits.

This study evaluates the alveolar stereological profile of immature rabbit lungs after treatment with various surfactant preparations, deliberately modified to represent different rates of film adsorption, minimum surface tension and compressibility. Surfactant was isolated from porcine or bovine lungs, and some of these preparations were enriched with dipalmitoylphosphatidylcholine and other synthetic lipids. Alveolar stereological parameters were evaluated in histological lung sections by computerized interactive image analysis. Surfactant treatment enhanced lung expansion, with a significant correlation coefficient between dynamic compliance and alveolar volume density in all groups of animals. Surfactant treatment also significantly increased alveolar image area. Enrichment of surfactant with synthetic lipids did not improve stereological parameters, but caused a significant increase in the coefficient of variation for alveolar perimeter and image area, suggesting more heterogeneous expansion pattern. Experimental evaluation of exogenous surfactants in immature newborn animals should include assessment of alveolar stereological parameters to detect deviations from the uniform expansion pattern seen after treatment with an optimal surfactant substitute.

Lysophosphatidylcholine abrogates the CR1 preserving effect of surfactant on quartz-exposed human granulocytes.

The effects of pulmonary surfactant on granulocytes were studied by flow cytofluorometry. Cells from hemolyzed blood were first activated by N-formylmethionyl-leucyl-phenylalanine (fMLP) which mobilizes complement receptor 1 (CR1) from the intracellular pool to the cell surface. The reduced CR1 expression observed after quartz incubation was abolished by a porcine surfactant preparation containing phospholipids and the hydrophobic surfactant proteins. Phospholipids alone had no preservative capacity. However, addition of the surfactant proteins to the phospholipids did not restore the CR1 values to that of intact surfactant, probably due to changed protein structure during the purification procedure. Heating of surfactant at 37 degrees C up to 72 h reduced the preservative effect of surfactant on CR1 expression. Congruent results of CR1 expression were achieved when 1-10% lysophosphatidylcholine was added to the surfactant preparations. Our results imply that lysophosphatidylcholine formed during storage of surfactant at elevated temperatures influences CR1 expression on granulocytes.

Biophysical activity of an artificial surfactant containing an analogue of surfactant protein (SP)-C and native SP-B.

Natural surfactant preparations containing phospholipids and the hydrophobic surfactant proteins B and C (SP-B and SP-C) are effective in the treatment of respiratory distress syndrome in premature infants. The limited supply, and the risk of infectious agents and immunological reactions have promoted the evaluation of synthetic peptides in surfactant preparations. However, the folding of synthetic SP-C into an alpha-helix is inefficient and alpha-helical SP-C analogues with Val-->Leu substitutions form oligomers. In order to circumvent these problems we have synthesized an SP-C analogue, named SP-C(LKS), which differs from SP-C mainly by the exchange of most of the Val residues in positions 16-28 with Leu residues to promote an alpha-helical conformation, and by the introduction of Lys residues at positions 17, 22 and 27 in order to locate positive charges around the helical circumference and thereby avoid self polymerization. CD spectroscopy showed a spectrum typical for alpha-helical peptides and SDS/PAGE disclosed a single band. The biophysical activity of artificial surfactant preparations containing SP-C(LKS) and phospholipids, with and without native SP-B, was measured using a Wilhelmy balance and a pulsating bubble surfactometer. SP-C(LKS) (3%, w/w) in a mixture of 1, 2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC)/phosphatidylglycerol/palmitic acid (68:22:9, by wt.) suspended in 150 mM NaCl, showed rapid spreading at the air-liquid interface and produced a surface tension of <1 mN/m at minimum bubble size (gammamin) and 42 mN/m at maximum bubble size (gammamax) in the pulsating bubble surfactometer. The addition of 2% (w/w) SP-B to the preparation reduced the maximum surface tension to 33-35 mN/m, i.e. both gammamin and gammamax values were similar to those of natural surfactant preparations. Optimal in vitro characteristics were also obtained from a preparation containing SP-C(LKS), SP-B, DPPC and phosphatidylglycerol, i.e. when palmitic acid was omitted from the lipid mixture. SP-B containing surfactant preparations made up in Hepes buffer at pH 6.9, instead of in 150 mM NaCl, had similar biophysical activity provided that palmitic acid was omitted, but decreased activity in the presence of palmitic acid.

Experimental neonatal respiratory failure induced by lysophosphatidylcholine: effect of surfactant treatment.

The purpose of this study was to characterize the toxic effects of lysophosphatidylcholine (lyso-PC) on neonatal lung function. Various doses of lyso-PC (from 0 to 40 mg/kg) were administered to near-term newborn rabbits. Lung-thorax compliance during mechanical ventilation was significantly decreased by doses >/=10 mg/kg, and static lung volumes during deflation were decreased by doses >/=20 mg/kg. Using the same experimental model, we investigated the effects of modified porcine surfactant (Curosurf, 200 mg/kg). Animals exposed to lyso-PC at birth and treated simultaneously with surfactant showed a satisfactory therapeutic response, whereas those treated after 30 min failed to respond. These animals also had a much larger leak of albumin into the air spaces and an elevated minimum surface tension of the lavage fluid in a pulsating bubble surfactometer, suggesting inactivation of the exogenous surfactant. Timing of surfactant administration may thus be essential for the therapeutic effect in this experimental model of acute lung injury.

Combined effects of high-frequency ventilation and surfactant treatment in experimental meconium aspiration syndrome.

BACKGROUND: Deterioration of lung function in meconium aspiration syndrome may in part be due to inactivation of endogenous surfactant. We evaluated the efficacy of high-frequency ventilation (HFV) and the combination of HFV and surfactant therapy in the management of respiratory failure induced by experimental meconium aspiration in adult rats. METHODS: Animals were anesthetized and tracheotomized, and received via the tracheal cannula a suspension of human meconium (25 mg/ml, dose 4 or 5 ml/kg). After 30 min of conventional ventilation (CV) with 100% oxygen, animals were in respiratory failure as indicated by a decrease in lung-thorax compliance of > or = 30% and PaO2 < 10 kPa. They were then ventilated for an additional 3 h with either CV (frequency 40/min, inspiration time 50%) or HFV (frequency 15 Hz, inspiration time 50%) using comparable mean airway pressures (about 20 cmH2O) and variable FiO2. Subgroups of animals were treated with Curosurf (80 mg/ml, dose 200 mg/kg) 30 min after meconium aspiration; no additional material was instilled in controls. RESULTS: From 30 min onwards, values for PaO2/FiO2 were significantly higher in animals ventilated with HFV without receiving surfactant than in control animals subjected to CV alone (at 120 min; 18 +/- 7.3 vs. 7.4 +/- 0.8 kPa, P < 0.05). Additional improvement in oxygenation was seen in HFV-treated animals receiving Curosurf (at 120 min: 39 +/- 16 kPa: P vs. HFV alone < 0.01). Relative lung volumes at a deflation pressure of 10 cmH2O, expressed as percent of maximum volume, were larger in animals ventilated with HFV than in those undergoing CV (41 +/- 12 vs. 33 +/- 10%; P < 0.05), and were further increased in the groups of animals treated with surfactant and ventilated with HFV (55 +/- 9.9%; P vs. animals ventilated with HFV not receiving surfactant < 0.01) or CV (49 +/- 9.6%; P vs. animals ventilated with CV without receiving surfactant < 0.05). Hyaline membranes, granulocytes in cytospin preparations from lung lavage fluid, and vascular-to-alveolar leak of albumin were less prominent in the HFV than in the CV group, particularly in animals treated with surfactant. CONCLUSION: Our data indicate that HFV, especially in combination with surfactant therapy, may be superior to CV for treatment of respiratory failure in this animal model of meconium aspiration syndrome.