A 20-Mer Peptide Derived from the Lectin Domain of SP-A2 Decreases Tumor Necrosis Factor Alpha Production during Mycoplasma pneumoniae Infection.

Human surfactant protein-A2 (hSP-A2) is a component of pulmonary surfactant that plays an important role in the lung's immune system by interacting with viruses, bacteria, and fungi to facilitate pathogen clearance and by downregulating inflammatory responses after an allergic challenge. Genetic variation in SP-A2 at position Gln223Lys is present in up to ∼30% of the population and has been associated with several lung diseases, such as asthma, pulmonary fibrosis, and lung cancer (M. M. Pettigrew, J. F. Gent, Y. Zhu, E. W. Triche, et al., BMC Med Genet 8:15, 2007, https://bmcmedgenet.biomedcentral.com/articles/10.1186/1471-2350-8-15; Y. Wang, P. J. Kuan, C. Zing, J. T. Cronkhite, et al., Am J Hum Genet 84:52-59, 2009, https://www.cell.com/ajhg/fulltext/S0002-9297(08)00595-8). Previous work performed by our group showed differences in levels of SP-A binding to non-live mycoplasma membrane fractions that were dependent on the presence of a lysine (K) or a glutamine (Q) at amino acid position 223 in the carbohydrate region of SP-A2. On the basis of these differences, we have derived 20-amino-acid peptides flanking this region of interest in order to test the ability of each to regulate various immune responses to live Mycoplasma pneumoniae in SP-A knockout mice and RAW 264.7 cells. In both models, the 20-mer containing 223Q significantly decreased both tumor necrosis factor alpha (TNF-α) mRNA levels and protein levels in comparison to the 20-mer containing 223K during M. pneumoniae infection. While neither of the 20-mer peptides (223Q and 223K) had an effect on p38 phosphorylation during M. pneumoniae infection, the 223Q-20mer peptide significantly reduced NF-κB p65 phosphorylation in both models. Taken together, our data suggest that small peptides derived from the lectin domain of SP-A2 that contain the major allelic variant (223Q) maintain activity in reducing TNF-α induction during M. pneumoniae infection.

Structural and Functional Determinants of Rodent and Human Surfactant Protein A: A Synthesis of Binding and Computational Data.

Surfactant protein A (SP-A) provides surfactant stability, first line host defense, and lung homeostasis by binding surfactant phospholipids, pathogens, alveolar macrophages (AMs), and epithelial cells. Non-primates express one SP-A protein whereas humans express two: SP-A1 and SP-A2 with core intra- and inter-species differences in the collagen-like domain. Here, we used macrophages and solid phase binding assays to discern structural correlates of rat (r) and human (h) SP-A function. Binding assays using recombinant rSP-A expressed in insect cells showed that lack of proline hydroxylation, truncations of amino-terminal oligomerization domains, and site-directed serine (S) or alanine (A) mutagenesis of cysteine 6 (C6S), glutamate 195 (E195A), and glutamate 171 (E171A) in the carbohydrate recognition domain (CRD) all impaired SP-A binding. Replacement of arginine 197 with alanine found in hSP-A (R197A), however, restored the binding of hydroxyproline-deficient rSP-A to the SP-A receptor SP-R210 similar to native rat and human SP-A. In silico calculation of Ca++ coordination bond length and solvent accessibility surface area revealed that the "humanized" R197A substitution alters topology and solvent accessibility of the Ca++ coordination residues of the CRD domain. Binding assays in mouse AMs that were exposed to either endogenous SP-A or hSP-A1 (6A2) and hSP-A2 (1A0) isoforms in vivo revealed that mouse SP-A is a functional hybrid of hSP-A1 and hSP-A2 in regulating SP-A receptor occupancy and binding affinity. Binding assays using neonatal and adult human AMs indicates that the interaction of SP-A1 and SP-A2 with AMs is developmentally regulated. Furthermore, our data indicate that the auxiliary ion coordination loop encompassing the conserved E171 residue may comprise a conserved site of interaction with macrophages, and SP-R210 specifically, that merits further investigation to discern conserved and divergent SP-A functions between species. In summary, our findings support the notion that complex structural adaptation of SP-A regulate conserved and species specific AM functions in vertebrates.

Epigallocatechin-3-gallate (EGCG) inhibits aggregation of pulmonary fibrosis associated mutant surfactant protein A2 via a proteasomal degradation pathway.

BACKGROUND/AIMS: Epigallocatechin-3-gallate (EGCG), a major catechin found in green tea, plays an important anti-tumor role and is involved in various other biological processes, such as, neuroprotection by prevention of aggregation of misfolded proteins generated because of genetic defects. Surfactant protein A2 mutations (G231V and F198S) have been identified to be associated with pulmonary fibrosis and lung cancer, and these mutations cause protein aggregation, instability as well as secretion deficiency. The present study focused on investigating the inhibitory effects of EGCG on aggregation of mutant SP-A2 and elucidating the potential mechanisms underlying this action. METHODS: Wild-type and mutant SP-A2 were transiently expressed in CHO-K1 cells. The aggregated and soluble proteins were separated into NP-40-insoluble and NP-40-soluble fractions. Protein stability was validated by chymotrypsin limited proteolysis assay. Western blot and RT-PCR were used to determine the protein and mRNA expression level, respectively. RESULTS: Mutant SP-A2 alone or wild-type SP-A2 co-expressed with G231V formed NP-40-insoluble aggregates in CHO-K1 cells. EGCG significantly suppressed this aggregation and alleviated mutant SP-A2 accumulation in the ER. When combined with 4-PBA, EGCG treatment completely blocked mutant SP-A2 aggregate formation. Though secretion of mutant protein was not affected, EGCG facilitated protein instability in both wild-type and mutant protein. Importantly, MG132, a proteasome inhibitor, reversed EGCG-induced aggregate reduction. CONCLUSIONS: EGCG inhibits aggregation of misfolded SP-A2 via induction of protein instability and activation of proteasomal pathway for aggregate degradation.

Surfactant Proteins A and D: Trimerized Innate Immunity Proteins with an Affinity for Viral Fusion Proteins.

Innate recognition of viruses is an essential part of the immune response to viral pathogens. This is integral to the maintenance of healthy lungs, which are free from infection and efficient at gaseous exchange. An important component of innate immunity for identifying viruses is the family of C-type collagen-containing lectins, also known as collectins. These secreted, soluble proteins are pattern recognition receptors (PRRs) which recognise pathogen-associated molecular patterns (PAMPs), including viral glycoproteins. These innate immune proteins are composed of trimerized units which oligomerise into higher-order structures and facilitate the clearance of viral pathogens through multiple mechanisms. Similarly, many viral surface proteins form trimeric configurations, despite not showing primary protein sequence similarities across the virus classes and families to which they belong. In this review, we discuss the role of the lung collectins, i.e., surfactant proteins A and D (SP-A and SP-D) in viral recognition. We focus particularly on the structural similarity and complementarity of these trimeric collectins with the trimeric viral fusion proteins with which, we hypothesise, they have elegantly co-evolved. Recombinant versions of these innate immune proteins may have therapeutic potential in a range of infectious and inflammatory lung diseases including anti-viral therapeutics.

Surfactant protein-A nanobody-conjugated liposomes loaded with methylprednisolone increase lung-targeting specificity and therapeutic effect for acute lung injury.

The advent of nanomedicine requires novel delivery vehicles to actively target their site of action. Here, we demonstrate the development of lung-targeting drug-loaded liposomes and their efficacy, specificity and safety. Our study focuses on glucocorticoids methylprednisolone (MPS), a commonly used drug to treat lung injuries. The steroidal molecule was loaded into functionalized nano-sterically stabilized unilamellar liposomes (NSSLs). Targeting functionality was performed through conjugation of surfactant protein A (SPANb) nanobodies to form MPS-NSSLs-SPANb. MPS-NSSLs-SPANb exhibited good size distribution, morphology, and encapsulation efficiency. Animal experiments demonstrated the high specificity of MPS-NSSLs-SPANb to the lung. Treatment with MPS-NSSLs-SPANb reduced the levels of TNF-α, IL-8, and TGF-ß1 in rat bronchoalveolar lavage fluid and the expression of NK-κB in the lung tissues, thereby alleviating lung injuries and increasing rat survival. The nanobody functionalized nanoparticles demonstrate superior performance to treat lung injury when compared to that of antibody functionalized systems.

Disruption of the structural and functional features of surfactant protein A by acrolein in cigarette smoke.

The extent to which defective innate immune responses contribute to chronic obstructive pulmonary disease (COPD) is not fully understood. Pulmonary surfactant protein A (SP-A) plays an important role in regulating innate immunity in the lungs. In this study, we hypothesised that cigarette smoke (CS) and its component acrolein might influence pulmonary innate immunity by affecting the function of SP-A. Indeed, acrolein-modified SP-A was detected in the lungs of mice exposed to CS for 1 week. To further confirm this finding, recombinant human SP-A (hSP-A) was incubated with CS extract (CSE) or acrolein and then analysed by western blotting and nanoscale liquid chromatography-matrix-assisted laser desorption/ionisation time-of-flight tandem mass spectrometry. These analyses revealed that CSE and acrolein induced hSP-A oligomerisation and that acrolein induced the modification of six residues in hSP-A: His39, His116, Cys155, Lys180, Lys221, and Cys224. These modifications had significant effects on the innate immune functions of hSP-A. CSE- or acrolein-induced modification of hSP-A significantly decreased hSP-A's ability to inhibit bacterial growth and to enhance macrophage phagocytosis. These findings suggest that CS-induced structural and functional defects in SP-A contribute to the dysfunctional innate immune responses observed in the lung during cigarette smoking.

Surfactant protein A (SP-A) and SP-A-derived peptide attenuate chemotaxis of mast cells induced by human ß-defensin 3.

Human ß-defensin 3 (hBD3) is known to be involved in mast cell activation. However, molecular mechanisms underlying the regulation of hBD3-induced mast cell activation have been poorly understood. We previously reported that SP-A and SP-A-derived peptide 01 (SAP01) regulate the function of hBD3. In this study, we focused on the effects of SP-A and SAP01 on the activation of mast cells induced by hBD3. SAP01 directly bound to hBD3. Mast cell-mediated vascular permeability and edema in hBD3 administered rat ears were decreased when injected with SP-A or SAP01. Compatible with the results in rat ear model, both SP-A and SAP01 inhibited hBD3-induced chemotaxis of mast cells in vitro. Direct interaction between SP-A or SAP01 and hBD3 seemed to be responsible for the inhibitory effects on chemotaxis. Furthermore, SAP01 attenuated hBD3-induced accumulation of mast cells and eosinophils in tracheas of the OVA-sensitized inflammatory model. SP-A might contribute to the regulation of inflammatory responses mediated by mast cells during infection.

Malondialdehyde-Acetaldehyde-Adducted Surfactant Protein Alters Macrophage Functions Through Scavenger Receptor A.

BACKGROUND: Reactive aldehydes such as acetaldehyde and malondialdehyde generated as a result of alcohol metabolism and cigarette smoke exposure lead to the formation of malondialdehyde-acetaldehyde-adducted proteins (MAA adducts). These aldehydes can adduct to different proteins such as bovine serum albumin and surfactant protein A or surfactant protein D (SPD). Macrophages play an important role in innate immunity, but the effect of MAA adducts on macrophage function has not yet been examined. Because macrophage scavenger receptor A (SRA; CD204) mediates the uptake of modified proteins, we hypothesized that the effects of MAA-modified proteins on macrophage function are primarily mediated through SRA. METHODS: We tested this hypothesis by exposing SPD-MAA to macrophages and measuring functions. SPD-MAA treatment significantly stimulated pro-inflammatory cytokine tumor necrosis factor-alpha (TNF-α) release in the macrophage cell line, RAW 264.7. RESULTS: A significant reduction in phagocytosis of zymosan particles was also observed. SPD-MAA stimulated a significant dose-dependent increase in TNF-α and interleukin (IL)-6 release from peritoneal macrophages (PMs) of wild-type (WT) mice. But significantly less TNF-α and IL-6 were released from PMs of SRA-/- mice. We observed a significant reduction in phagocytosis of zymosan particles in PMs from WT mice treated with SPD-MAA. No further SPD-MAA-induced reduction was seen in PMs from SRA-/- mice. SPD-MAA treatment significantly increased SRA mRNA expression, but had no effect on surface receptor protein expression. Protein kinase C alpha inhibitor and NF-κB inhibitor significantly reduced pro-inflammatory cytokine release in response to SPD-MAA. CONCLUSIONS: In conclusion, our data demonstrate that SRA is important for MAA-adducted protein-mediated effect on macrophage functions.

Identification and Quantitation of Coding Variants and Isoforms of Pulmonary Surfactant Protein A.

Pulmonary surfactant protein A (SP-A), a heterooligomer of SP-A1 and SP-A2, is an important regulator of innate immunity of the lung. Nonsynonymous single nucleotide variants of SP-A have been linked to respiratory diseases, but the expressed repertoire of SP-A protein in human airway has not been investigated. Here, we used parallel trypsin and Glu-C digestion, followed by LC-MS/MS, to obtain sequence coverage of common SP-A variants and isoform-determining peptides. We further developed a SDS-PAGE-based, multiple reaction monitoring (GeLC-MRM) assay for enrichment and targeted quantitation of total SP-A, the SP-A2 isoform, and the Gln223 and Lys223 variants of SP-A, from as little as one milliliter of bronchoalveolar lavage fluid. This assay identified individuals with the three genotypes at the 223 position of SP-A2: homozygous major (Gln223/Gln223), homozygous minor (Lys223/Lys223), or heterozygous (Gln223/Lys223). More generally, our studies demonstrate the challenges inherent in distinguishing highly homologous, copurifying protein isoforms by MS and show the applicability of MRM mass spectrometry for identification and quantitation of nonsynonymous single nucleotide variants and other proteoforms in airway lining fluid.

Surfactant protein A binds TGF-ß1 with high affinity and stimulates the TGF-ß pathway.

We were able to demonstrate reversible, specific and high-affinity binding of radioactively-labelled TGF-ß1 ((125)I-TGF-ß1) to immobilized surfactant protein A (SP-A), with an apparent dissociation constant of 53 picomolar at ∼21. Addition of a 200-fold molar excess of the latency associated peptide (LAP) prevented and dissociated the binding of (125)I-TGF-ß1 to SP-A, whereas latent TGF-ß1 had no effect. Using a bioassay for TGF-ß1 activity--a luciferase reporter assay--we were able to show that SP-A in the presence of TGF-ß1 stimulated the TGF-ß1 pathway, whereas SP-A alone had no effect. Studies with structural analogues of the distinct SP-A tail domain and head domain indicated that stimulatory activity of SP-A resided in the head domain. No activation of latent TGF-ß1 by SP-A was observed. In addition, we observed that SP-A inhibited TGF-ß1 inactivation by LAP. These results indicate that SP-A may have a regulatory role in the TGF-ß1-mediated processes in the lung.

Transient exposure of pulmonary surfactant to hyaluronan promotes structural and compositional transformations into a highly active state.

Pulmonary surfactant is a lipid-protein complex that lowers surface tension at the respiratory air-liquid interface, stabilizing the lungs against physical forces tending to collapse alveoli. Dysfunction of surfactant is associated with respiratory pathologies such as acute respiratory distress syndrome or meconium aspiration syndrome where naturally occurring surfactant-inhibitory agents such as serum, meconium, or cholesterol reach the lung. We analyzed the effect of hyaluronan (HA) on the structure and surface behavior of pulmonary surfactant to understand the mechanism for HA-promoted surfactant protection in the presence of inhibitory agents. In particular, we found that HA affects structural properties such as the aggregation state of surfactant membranes and the size, distribution, and order/packing of phase-segregated lipid domains. These effects do not require a direct interaction between surfactant complexes and HA and are accompanied by a compositional reorganization of large surfactant complexes that become enriched with saturated phospholipid species. HA-exposed surfactant reaches very high efficiency in terms of rapid and spontaneous adsorption of surfactant phospholipids at the air-liquid interface and shows significantly improved resistance to inactivation by serum or cholesterol. We propose that physical effects pertaining to the formation of a meshwork of interpenetrating HA polymer chains are responsible for the changes in surfactant structure and composition that enhance surfactant function and, thus, resistance to inactivation. The higher resistance of HA-exposed surfactant to inactivation persists even after removal of the polymer, suggesting that transient exposure of surfactant to polymers like HA could be a promising strategy for the production of more efficient therapeutic surfactant preparations.

Specific cleavage of the lung surfactant protein A by human cathepsin S may impair its antibacterial properties.

Human cysteine cathepsins (Cats) are implicated in lung injuries and tissue remodeling and have recently emerged as important players in pulmonary inflammations. The proteolytic activities of Cat B, L, K, S and H are dramatically increased in the sputum of patients with cystic fibrosis (CF), suggesting a possible involvement in the CF pathophysiology. We found that pulmonary surfactant protein A (SP-A) that participates to innate host defense is extensively degraded in CF expectorations. Breakdown of SP-A was markedly decreased in CF sputum by E-64 and Mu-Leu-Hph-VSPh, a Cat S inhibitor. Cat S cleaved efficiently and specifically SP-A within critical residues of the solvent-exposed loop of its carbohydrate recognition (C-type lectin) domain that allows binding to pathogens. Cat S decreased aggregation properties of SP-A (self-aggregation, aggregation of phospholipid vesicles and rough LPS). Moreover cleavage of SP-A by Cat S reduced binding to yeast mannan and impaired agglutination of Escherichia coli and Pseudomonas aeruginosa, a foremost detrimental pathogen colonizing the lungs of CF patients. Besides human neutrophil serine proteases and bacterial proteases, we propose that Cat S may participate in the pathophysiology of CF by weakening the antibacterial activity of SP-A. More broadly, present results provide further indication that Cat S, along with Cats B and L, could display immuno-modulatory functions by inactivating key proteins involved in the innate immunity defense.

Surfactant protein A influences reepithelialization in an alveolocapillary model system.

PURPOSE: Restoring the barrier integrity of the alveolar epithelium after injury is pivotal. In the current study, we evaluated the effects of surfactant, surfactant protein A (SP-A), transforming growth factor ß (TGF-ß), and analogues of SP-A on alveolar epithelial repair. Additionally, we assessed the influence of microvascular endothelial cells on reepithelialization. METHODS: Repair was studied in an in vitro model system consisting of a bilayer coculture of A549 and human pulmonary microvascular endothelial cells (HPMECs), which stably expressing fluorescent proteins. The epithelial repair was assessed in a scratch assay using vital fluorescence microscopy and compared with a monolayer of A549 cells. RESULTS: HMPEC cells differentially modulated the response of the A549 cells. Surfactant and SP-A augmented the reepithelialization in the presence of HPMECs, whereas in the absence of HPMECs, surfactant inhibited wound healing and SP-A failed to alter the response. Like SP-A, a structural analogue of its collagenous tail domain augmented the reepithelialization in the model system, whereas an analogue of its head domain did not alter the response. Additionally, we demonstrated that TGF-ß associated with SP-A was able to initiate the Smad-dependent TGF-ß pathway and that both TGF-ß and TGF-ß free SP-A were able to stimulate wound healing in the bilayer model. CONCLUSIONS: These data show that surfactant, SP-A and TGF-ß, influence epithelial repair in vitro and that the microvascular endothelial cells can modulate the response. This indicates that surfactant and SP-A could play a role in alveolar epithelial repair and that the microvascular endothelium may be involved in these processes.

SFTA2--a novel secretory peptide highly expressed in the lung--is modulated by lipopolysaccharide but not hyperoxia.

Tissue-specific transcripts are likely to be of importance for the corresponding organ. While attempting to define the specific transcriptome of the human lung, we identified the transcript of a yet uncharacterized protein, SFTA2. In silico analyses, biochemical methods, fluorescence imaging and animal challenge experiments were employed to characterize SFTA2. Human SFTA2 is located on Chr. 6p21.33, a disease-susceptibility locus for diffuse panbronchiolitis. RT-PCR verified the abundance of SFTA2-specific transcripts in human and mouse lung. SFTA2 is synthesized as a hydrophilic precursor releasing a 59 amino acid mature peptide after cleavage of an N-terminal secretory signal. SFTA2 has no recognizable homology to other proteins while orthologues are present in all mammals. SFTA2 is a glycosylated protein and specifically expressed in nonciliated bronchiolar epithelium and type II pneumocytes. In accordance with other hydrophilic surfactant proteins, SFTA2 did not colocalize with lamellar bodies but colocalized with golgin97 and clathrin-labelled vesicles, suggesting a classical secretory pathway for its expression and secretion. In the mouse lung, Sfta2 was significantly downregulated after induction of an inflammatory reaction by intratracheal lipopolysaccharides paralleling surfactant proteins B and C but not D. Hyperoxia, however, did not alter SFTA2 mRNA levels. We have characterized SFTA2 and present it as a novel unique secretory peptide highly expressed in the lung.

The untranslated exon B of human surfactant protein A2 mRNAs is an enhancer for transcription and translation.

Two human genes, SFTPA1 (SP-A1) and SFTPA2 (SP-A2), encode surfactant protein A, a molecule of innate immunity and surfactant-related functions. Several genetic variants have been identified for both genes. These include nucleotide (nt) polymorphisms, as well as alternative splicing patterns at the 5' untranslated region (5'UTR). Exon B (eB) is included in the 5'UTR of most SP-A2, but not SP-A1 splice variants. We investigated the role of eB in the regulation of gene expression and translation efficiency. A luciferase (Luc) reporter gene was cloned downstream of the entire (AeBD) or eB deletion mutants (del_mut) of the SP-A2 5'UTR, or heterologous 5'UTRs containing the eB sequence, or a random sequence of equal length. The del_mut constructs consisted in consecutive deletions of five nucleotides (n = 8) within eB and the exon-exon junctions in the AeBD 5'UTR. Luc activities and mRNA levels were compared after transfection of NCI-H441 cells. We found that 1) eB increased Luc mRNA levels when placed upstream of heterologous 5'UTR sequences or the promoter region, regardless of its position and orientation; 2) translation efficiency of in vitro-generated mRNAs containing eB was higher than that of mRNAs without eB; and 3) the integrity of eB sequence is crucial for transcription and translation of the reporter gene. Thus eB 1) is a transcription enhancer, because it increases mRNA content regardless of position and orientation, 2) enhances translation when placed in either orientation within its natural 5'UTR sequence and in heterologous 5'UTRs, and 3) contains potential regulatory elements for both transcription and translation. We conclude that eB sequence and length are determinants of transcription and translation efficiency.

Surfactant protein A2 mutations associated with pulmonary fibrosis lead to protein instability and endoplasmic reticulum stress.

Rare heterozygous mutations in the gene encoding surfactant protein A2 (SP-A2, SFTPA2) are associated with adult-onset pulmonary fibrosis and adenocarcinoma of the lung. We have previously shown that two recombinant SP-A2 mutant proteins (G231V and F198S) remain within the endoplasmic reticulum (ER) of A549 cells and are not secreted into the culture medium. The pathogenic mechanism of the mutant proteins is unknown. Here we analyze all common and rare variants of the surfactant protein A2, SP-A2, in both A549 cells and in primary type II alveolar epithelial cells. We show that, in contrast with all other SP-A2 variants, the mutant proteins are not secreted into the medium with wild-type SP-A isoforms, form fewer intracellular dimer and trimer oligomers, are partially insoluble in 0.5% Nonidet P-40 lysates of transfected A549 cells, and demonstrate greater protein instability in chymotrypsin proteolytic digestions. Both the G231V and F198S mutant SP-A2 proteins are destroyed via the ER-association degradation pathway. Expression of the mutant proteins increases the transcription of a BiP-reporter construct, expression of BiP protein, and production of an ER stress-induced XBP-1 spliced product. Human bronchoalveolar wash samples from individuals who are heterozygous for the G231V mutation have similar levels of total SP-A as normal family members, which suggests that the mechanism of disease does not involve an overt lack of secreted SP-A but instead involves an increase in ER stress of resident type II alveolar epithelial cells.

Lung surfactant proteins A and D as pattern recognition proteins.

Lung surfactant proteins A and D belong to a group of soluble humoral pattern recognition receptors, called collectins, which modulate the immune response to microorganisms. They bind essential carbohydrate and lipid antigens found on the surface of microorganisms via low affinity C-type lectin domains and regulate the host's response by binding to immune cell surface receptors. They form multimeric structures that bind, agglutinate, opsonise and neutralize many different pathogenic microorganisms including bacteria, yeast, fungi and viruses. They modulate the uptake of these microorganisms by phagocytic cells as well as both the inflammatory and the adaptive immune responses. Recent data have also highlighted their involvement in clearance of apoptotic cells, hypersensitivity and a number of lung diseases.

Environmental tobacco smoke effects on lung surfactant film organization.

Adsorption of the clinical lung surfactants (LS) Curosurf or Survanta from aqueous suspension to the air-water interface progresses from multi-bilayer aggregates through multilayer films to a coexistence between multilayer and monolayer domains. Exposure to environmental tobacco smoke (ETS) alters this progression as shown by Langmuir isotherms, fluorescence microscopy and atomic force microscopy (AFM). After 12 h of LS exposure to ETS, AFM images of Langmuir-Blodgett deposited films show that ETS reduces the amount of material near the interface and alters how surfactant is removed from the interface during compression. For Curosurf, ETS prevents refining of the film composition during cycling; this leads to higher minimum surface tensions. ETS also changes the morphology of the Curosurf film by reducing the size of condensed phase domains from 8-12 microm to approximately 2 microm, suggesting a decrease in the line tension between the domains. The minimum surface tension and morphology of the Survanta film are less impacted by ETS exposure, although the amount of material associated with the film is reduced in a similar way to Curosurf. Fluorescence and mass spectra of Survanta dispersions containing native bovine SP-B treated with ETS indicate the oxidative degradation of protein aromatic amino acid residue side chains. Native bovine SP-C isolated from ETS exposed Survanta had changes in molecular mass consistent with deacylation of the lipoprotein. Fourier Transform Infrared Spectroscopy (FTIR) characterization of the hydrophobic proteins from ETS treated Survanta dispersions show significant changes in the conformation of SP-B and SP-C that correlate with the altered surface activity and morphology of the lipid-protein film.

Effect of maternal dexamethasone treatment on the type II pneumocytes in hypoplastic lung by oligohydramnios: an ultrastructural study.

A previous study documented the effects of maternal corticosteroid treatment on structural growth and functional development in fetal lungs associated with pathogenic conditions such as oligohydramnios using immunohistochemical and morphometric analyses. The purpose of the present study was to examine the effect of maternal dexamethasone treatment the expression of lamellar body in type II pneumocytes of the fetal rabbit lungs with hypoplasia induced by oligohydramnios using electron microscopy. Using an amniotic shunting rabbit model, pregnant rabbits were injected intravenously with either 0.1 ml of saline or 0.25 mg/kg/day of dexamethasone in 0.1 ml of saline 48 and 24 h before the delivery of fetuses, at day 30 of gestation. The number of lamellar bodies per type II pneumocyte was counted in each group using electron micrographs. The number of lamellar bodies per type II pneumocyte in the lungs of the shunted group that received saline was consistently and significantly less than that of the other three groups (5.49 +/- 2.07 vs. 7.34 +/- 2.27: shunted group that received dexamethasone, 7.58 +/- 2.08: non-shunted group that received saline, 7.79 +/- 1.90: non-shunted group that received dexamethasone; P < 0.01). These results suggest that maternal dexamethasone treatment accelerates the maturation of the surfactant system, especially the expression of lamellar bodies in type II pneumocytes, even in hypoplastic lungs induced by oligohydramnios.