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The hallmarks of pediatric acute respiratory failure (ARF) are dysregulated inflammation and surfactant dysfunction. The objective is to study association of surfactant protein (SP) genes' single nucleotide polymorphisms (SNPs) with ARF and its morbidity: pulmonary dysfunction at discharge (PDAD), employing a single-, two-, and three-SNP interaction model. We enrolled 468 newborn controls and 248 children aged ≤ 24 months with ARF; 86 developed PDAD. Using quantitative genetic principles, we tested the association of SP genes SNPs with ARF and PDAD. We observed a dominant effect of rs4715 of the SFTPC on ARF risk. In a three-SNP model, we found (a) 34 significant interactions among SNPs of SFTPA1, SFTPA2, and SFTPC associated with ARF (p = 0.000000002-0.05); 15 and 19 of those interactions were associated with increased and decreased risk for ARF, respectively; (b) intergenic SNP-SNP interactions of both hydrophobic and hydrophilic SP genes associated with PDAD (p = 0.00002-0.03). The majority of intra- and intergenic interactions associated with ARF involve the SFTPA2 SNPs, whereas most of the intra- and intergenic interactions associated with PDAD are of SFTPA1 SNPs. We also observed a dominant effect of haplotypes GG of SFTPA1 associated with increased and AA of SFTPC associated with decreased ARF risk (p = 0.02). To the best of our knowledge, this is the first study showing an association of complex interactions of SP genes with ARF and PDAD. Our data indicate that SP genes polymorphisms may contribute to ARF pathogenesis and subsequent PDAD and/or may serve as markers for disease susceptibility in healthy children.

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PURPOSE: Gene polymorphisms of surfactant proteins, key players in lung innate immunity, have been associated with various lung diseases. The aim of this study was to investigate the potential association between variations within the surfactant protein (SP)-A gene of the donor lung allograft and recipient post-transplant outcome. METHODS: Lung-transplant patients (n=192) were prospectively followed-up with pulmonary function tests, bronchoscopies with bronchoalveolar lavage and biopsies. Donor lungs were assayed for SP-A1 (6An) and SP-A2 (1An) gene polymorphism using the pyrosequencing method. Unadjusted and adjusted stratified Cox survival models are reported. RESULTS: SP-A1 and SP-A2 genotype frequency and lung transplant recipient and donor characteristics as well as cause of death are noted. Recipients were grouped per donor SP-A2 variants. Individuals that received lungs from donors with the SP-A2 1A0 (n=102) versus 1A1 variant (n=68) or SP-A2 genotype 1A01A0 (n=54) versus 1A0A1 (n=38) had greater survival at 1 year (log-rank p<0.025). No significant association was noted for SP-A1 variants. Stratified adjusted survival models for 1-year survival and diagnosis showed a reduced survival for 1A1 variant and the 1A01A1 genotype. Furthermore, when survival was conditional on 1-year survival no significance was observed, indicating that the survival difference was due to the first year's outcome associated with the 1A1 variant. CONCLUSION: Donor lung SP-A gene polymorphisms are associated with post-transplant clinical outcome. Lungs from donors with the SP-A2 variant 1A1 had a reduced survival at 1 year. The observed donor genetic differences, via innate immunity relate to the post-transplant clinical outcome.

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OBJECTIVE: Surfactant proteins A and D are important molecules involved in lung allograft innate immunity. Genetic polymorphisms of surfactant proteins A and D are associated with various lung diseases. In this study, surfactant protein A and D expression responses were investigated during pharmacogenetics upon methylprednisolone treatment as observed during lung transplantation. METHODS: A human cell line (NCI-H441) and precision-cut lung slices from 16 human donors were incubated with methylprednisolone, and surfactant protein A1, surfactant protein A2, and surfactant protein D messenger RNA and surfactant protein A protein expression were assayed. Surfactant protein A1, A2, and D polymorphisms and surfactant protein A gene and protein expressions were determined. RESULTS: In NCI-H441 cells, methylprednisolone treatment at 10-5 M and 10-6 M reduced surfactant protein A1 and surfactant protein A2 messenger RNA and surfactant protein A protein expression (P < .05). A pharmacogenetic relationship was observed in human donor precision-cut lung slices between the surfactant protein A2 (1Ax) variants: Surfactant protein A1, A2, and D messenger RNA expression were greater for 1A0 versus 1A1 (P < .05); surfactant protein A1/surfactant protein A2 genotype 6A26A2/1A01A0 (n = 5) showed greater surfactant protein A1, A2, and D messenger RNA expression and surfactant protein A protein expression compared with the other surfactant protein A1/surfactant protein A2 genotypes (n = 11) (P < .05). CONCLUSIONS: The surfactant protein A genotype and methylprednisolone stimuli influence donor lung surfactant protein A and D expression. Lungs carrying the surfactant protein A2 variant 1A0 have a greater expression of surfactant protein A when treated with methylprednisolone. Surfactant protein A polymorphisms could be used to personalize immunosuppressive regimens.

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Surfactant proteins (SP) are involved in surfactant function and innate immunity in the human lung. Both lung function and innate immunity are altered in CF, and altered SP levels and genetic association are observed in Cystic Fibrosis (CF). We hypothesized that single nucleotide polymorphisms (SNPs) within the SP genes associate with CF or severity subgroups, either through single SNP or via SNP-SNP interactions between two SNPs of a given gene (intragenic) and/or between two genes (intergenic). We genotyped a total of 17 SP SNPs from 72 case-trio pedigree (SFTPA1 (5), SFTPA2 (4), SFTPB (4), SFTPC (2), and SFTPD (2)), and identified SP SNP associations by applying quantitative genetic principles. The results showed (a) Two SNPs, SFTPB rs7316 (p = 0.0083) and SFTPC rs1124 (p = 0.0154), each associated with CF. (b) Three intragenic SNP-SNP interactions, SFTPB (rs2077079, rs3024798), and SFTPA1 (rs1136451, rs1059057 and rs4253527), associated with CF. (c) A total of 34 intergenic SNP-SNP interactions among the 4 SP genes to be associated with CF. (d) No SNP-SNP interaction was observed between SFTPA1 or SFTPA2 and SFTPD. (e) Equal number of SNP-SNP interactions were observed between SFTPB and SFTPA1/SFTPA2 (n = 7) and SP-B and SFTPD (n = 7). (f) SFTPC exhibited significant SNP-SNP interactions with SFTPA1/SFTPA2 (n = 11), SFTPB (n = 4) and SFTPD (n = 3). (g) A single SFTPB SNP was associated with mild CF after Bonferroni correction, and several intergenic interactions that are associated (p < 0.01) with either mild or moderate/severe CF were observed. These collectively indicate that complex SNP-SNP interactions of the SP genes may contribute to the pulmonary disease in CF patients. We speculate that SPs may serve as modifiers for the varied progression of pulmonary disease in CF and/or its severity.

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New influenza A viruses that emerge frequently elicit composite inflammatory responses to both infection and structural damage of alveolar-capillary barrier cells that hinders regeneration of respiratory function. The host factors that relinquish restoration of lung health to enduring lung injury are insufficiently understood. Here, we investigated the role of endophilin B2 (B2) in susceptibility to severe influenza infection. WT and B2-deficient mice were infected with H1N1 PR8 by intranasal administration and course of influenza pneumonia, inflammatory, and tissue responses were monitored over time. Disruption of B2 enhanced recovery from severe influenza infection as indicated by swift body weight recovery and significantly better survival of endophilin B2-deficient mice compared to WT mice. Compared to WT mice, the B2-deficient lungs exhibited induction of genes that express surfactant proteins, ABCA3, GM-CSF, podoplanin, and caveolin mRNA after 7 days, temporal induction of CCAAT/enhancer binding protein CEBPα, ß, and δ mRNAs 3-14 days after infection, and differences in alveolar extracellular matrix integrity and respiratory mechanics. Flow cytometry and gene expression studies demonstrated robust recovery of alveolar macrophages and recruitment of CD4+ lymphocytes in B2-deficient lungs. Targeting of endophilin B2 alleviates adverse effects of IAV infection on respiratory and immune cells enabling restoration of alveolar homeostasis.

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BackgroundMaternal breast milk (MBM) is enriched in microRNAs, factors that regulate protein translation throughout the human body. MBM from mothers of term and preterm infants differs in nutrient, hormone, and bioactive-factor composition, but the microRNA differences between these groups have not been compared. We hypothesized that gestational age at delivery influences microRNA in MBM, particularly microRNAs involved in immunologic and metabolic regulation.MethodsMBM from mothers of premature infants (pMBM) obtained 3-4 weeks post delivery was compared with MBM from mothers of term infants obtained at birth (tColostrum) and 3-4 weeks post delivery (tMBM). The microRNA profile in lipid and skim fractions of each sample was evaluated with high-throughput sequencing.ResultsThe expression profiles of nine microRNAs in lipid and skim pMBM differed from those in tMBM. Gene targets of these microRNAs were functionally related to elemental metabolism and lipid biosynthesis. The microRNA profile of tColostrum was also distinct from that of pMBM, but it clustered closely with tMBM. Twenty-one microRNAs correlated with gestational age demonstrated limited relationships with method of delivery, but not other maternal-infant factors.ConclusionPremature delivery results in a unique MBM microRNA profile with metabolic targets. This suggests that preterm milk may have adaptive functions for growth in premature infants.

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BACKGROUND: Acute ozone (O3) exposure has known deleterious effects on the respiratory system and has been linked with respiratory disease and infection. Inflammatory lung disease induced by air pollution has demonstrated greater severity and poorer prognosis in women vs. men. Both severe damage to the bronchial-alveolar epithelium and malfunctioning of bronchial-blood barrier have been largely attributed to the pathobiology of O3-induced inflammatory response, but the associated mechanisms in the male and female lung remain unknown. METHODS: Here, we investigated sex-based differential regulation of lung interleukin-6 (IL-6) and its downstream signaling pathways JAK2/STAT3 and AKT1/NF-κB in response to O3 exposure in a mouse model. We exposed male and female mice (in different stages of the estrous cycle) to 2 ppm of O3 or filtered air (FA) for 3 h, and we harvested lung tissue for protein expression analysis by Western blot. RESULTS: We found significant up-regulation of IL-6 and IL-6R in females and IL-6 in males in response to O3 vs. FA. Ozone exposure induced a significant increase in STAT3-Y705 phosphorylation in both females and males. Males exposed to O3 had decreased levels of JAK2, but increased JAK2 (Y1007+Y1008) phosphorylation, while females exposed to O3 showed significant up-regulation of both proteins. Both NF-κB (p105/p50) and AKT1 protein levels were significantly increased only in females exposed to O3. In addition, females exposed to O3 during proestrus displayed increased expression of selected genes when compared to females exposed to O3 in other estrous cycle stages. CONCLUSIONS: Together, our observations indicate a sex-based and estrous cycle-dependent differential lung inflammatory response to O3 and involvement of two converging JAK2/STAT3 and AKT1/NF-κB pathways. To our knowledge, this is the first study specifically addressing the impact of the estrous cycle in O3-associated lung inflammatory pathways.

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Sex differences in the incidence of respiratory diseases have been reported. Women are more susceptible to inflammatory lung disease induced by air pollution and show worse adverse pulmonary health outcomes than men. However, the mechanisms underlying these differences remain unknown. In the present study, we hypothesized that sex differences in the expression of lung inflammatory mediators affect sex-specific immune responses to environmental toxicants. We focused on the effects of ground-level ozone, a major air pollutant, in the expression and regulation of lung immunity genes. We exposed adult male and female mice to 2 ppm of ozone or filtered air (control) for 3 h. We compared mRNA levels of 84 inflammatory genes in lungs harvested 4 h postexposure using a PCR array. We also evaluated changes in lung histology and bronchoalveolar lavage fluid cell counts and protein content at 24 and 72 h postexposure. Our results revealed sex differences in lung inflammation triggered by ozone exposure and in the expression of genes involved in acute phase and inflammatory responses. Major sex differences were found in the expression of neutrophil-attracting chemokines (Ccl20, Cxcl5, and Cxcl2), the proinflammatory cytokine interleukin-6, and oxidative stress-related enzymes (Ptgs2, Nos2). In addition, the phosphorylation of STAT3, known to mediate IL-6-related immune responses, was significantly higher in ozone-exposed mice. Together, our observations suggest that a differential regulation of the lung immune response could be implicated in the observed increased susceptibility to adverse health effects from ozone observed in women vs. men.

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The surfactant protein (SP-A) receptor SP-R210 has been shown to increase phagocytosis of SP-A-bound pathogens and to modulate cytokine secretion by immune cells. SP-A plays an important role in pulmonary immunity by enhancing opsonization and clearance of pathogens and by modulating macrophage inflammatory responses. Alternative splicing of the Myo18A gene results in two isoforms: SP-R210S and SP-R210L, with the latter predominantly expressed in alveolar macrophages. In this study we show that SP-A is required for optimal expression of SP-R210L on alveolar macrophages. Interestingly, pre-treatment with SP-A prepared by different methods either enhances or suppresses responsiveness to LPS, possibly due to differential co-isolation of SP-B or other proteins. We also report that dominant negative disruption of SP-R210L augments expression of receptors including SR-A, CD14, and CD36, and enhances macrophages' inflammatory response to TLR stimulation. Finally, because SP-A is known to modulate CD14, we used a variety of techniques to investigate how SP-R210 mediates the effect of SP-A on CD14. These studies revealed a novel physical association between SP-R210S, CD14, and SR-A leading to an enhanced response to LPS, and found that SP-R210L and SP-R210S regulate internalization of CD14 via distinct macropinocytosis-like mechanisms. Together, our findings support a model in which SP-R210 isoforms differentially regulate trafficking, expression, and activation of innate immune receptors on macrophages.

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Knowledge of the methylation profile of genes allow for the identification of biomarkers that may guide diagnosis and effective treatment of disease. Human surfactant protein A (SP-A) plays an important role in lung homeostasis and immunity, and is encoded by two genes (SFTPA1 and SFTPA2). The goal of this study was to identify differentially methylated CpG sites in the promoter region of the SFTPA2 gene in lung cancer tissue, and to determine the correlation between the promoter's methylation profile and gene expression. For this, we collected 28 pairs of cancerous human lung tissue and adjacent noncancerous (NC) lung tissue: 17 adenocarcinoma (AC), 9 squamous cell carcinoma (SCC), and 2 AC with SCC features, and we evaluated DNA methylation of the SFTPA2 promoter region by bisulfite conversion. Our results identified a higher methylation ratio in one CpG site of the SFTPA2 gene in cancerous tissue versus NC tissue (0.36 versus 0.11, p = 0.001). When assessing AC samples, we also found cancerous tissues associated with a higher methylation ratio (0.43 versus 0.10, p = 0.02). In the SCC group, although cancerous tissue showed a higher methylation ratio (0.22 versus 0.11), this difference was not statistically significant (p = 0.35). Expression of SFTPA2 mRNA and total SP-A protein was significantly lower in cancer tissue when compared to adjacent NC tissue (p < 0.001), and correlated with the hypermethylated status of an SFTPA2 CpG site in AC samples. The findings of this pilot study may hold promise for future use of SFTPA2 as a biomarker for the diagnosis of lung cancer.

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Human surfactant protein A (SP-A) plays an important role in surfactant metabolism and lung innate immunity. SP-A is synthesized and secreted by alveolar type II (ATII) cells, one of the two cell types of the distal lung epithelium (ATII and ATI). We have shown that miRNA interactions with sequence polymorphisms on the SP-A mRNA 3'UTRs mediate differential expression of SP-A1 and SP-A2 gene variants in vitro. In the present study, we describe a physiologically relevant model to study miRNA regulation of SP-A in human ATII. For these studies, we purified and cultured human ATII on an air-liquid interface matrix (A/L) or plastic wells without matrix (P). Gene expression analyses confirmed that cells cultured in A/L maintained the ATII phenotype for over 5 days, whereas P-cultured cells differentiated to ATI. When we transfected ATII with siRNAs to inhibit the expression of Drosha, a critical effector of miRNA maturation, the levels of SP-A mRNA and protein increased in a time dependent manner. We next characterized cultured ATII and ATI by studying expression of 1,066 human miRNAs using miRNA PCR arrays. We detected expression of >300 miRNAs with 24 miRNAs differentially expressed in ATII versus ATI, 12 of which predicted to bind SP-A 3'UTRs, indicating that these may be implicated in SP-A downregulation in ATI. Thus, miRNAs not only affect SP-A expression, but also may contribute to the maintenance of the ATII cell phenotype and/or the trans-differentiation of ATII to ATI cells, and may represent new molecular markers that distinguish ATII and ATI.

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Surfactant protein A (SP-A) plays a vital role in maintaining normal lung function and in host defense. Two genes encode SP-A in humans (SFTPA1, SFTPA2), and several gene variants have been identified for these. We have previously shown that sequence elements of SFTPA1 and SFTPA2 3' untranslated regions (UTRs) differentially affect translation efficiency in vitro. Polymorphisms at the 3'UTRs of mRNA variants may account for differential binding of miRNAs, a class of small noncoding RNAs that regulate gene expression. In this work, we generated 3'UTR reporter constructs of the SFTPA1 and SFTPA2 variants most frequently found in the population, as well as mutants of a previously described 11-nt indel element (refSNP rs368700152). Reporter constructs were transfected in NCI-H441 cells in the presence or absence of miRNA mimics, and reporter gene expression was analyzed. We found that human miRNA mir-767 negatively affected expression of constructs containing SFTPA1 and SFTPA2 variants, whereas mir-4507 affected only constructs with 3'UTRs of SFTPA1 variants 6A, 6A(3), and 6A(4) (not containing the 11-nt element). Three miRNAs (mir-183, mir-449b, and mir-612) inhibited expression of recombinants of SFTPA2 variants and the SFTPA1 variant 6A(2), all containing the 11-nt element. Similar results were obtained for SP-A expression when these miRNAs were transfected in Chinese hamster ovary cells expressing SFTPA1 or SFTPA2 variants or in NCI-H441 cells (genotype 1A(5)/1A(5)-6A(4)/6A(4)). Moreover, transfection with a specific antagomir (antagomir-183) reversed the effects of mir-183 on SP-A mRNA levels. Our results indicate that sequence variability at the 3'UTR of SP-A variants differentially affects miRNA regulation of gene expression.

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The first half of the surfactant protein B (SP-B) gene intron 4 is a CA-repeat-rich region that contains 11 motifs. To study the role of this region on SP-B mRNA splicing, minigenes were generated by systematic removal of motifs from either the 5' or 3' end. These were transfected in CHO cells to study their splicing efficiency. The latter was determined as the ratio of completely to incompletely spliced SP-B RNA. Our results indicate that SP-B intron 4 motifs differentially affect splicing. Motifs 8 and 9 significantly enhanced and reduced splicing of intron 4, respectively. RNA mobility shift assays performed with a Motif 8 sequence that contains a CAUC cis-element and cell extracts resulted in a RNA:protein shift that was lost upon mutation of the element. Furthermore, in silico analysis of mRNA secondary structure stability for minigenes with and without motif 8 indicated a correlation between mRNA stability and splicing ratio. We conclude that differential loss of specific intron 4 motifs results in one or more of the following: a) altered splicing, b) differences in RNA stability and c) changes in secondary structure. These, in turn, may affect SP-B content in lung health or disease.

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INTRODUCTION: The genetic contribution to the development of bronchopulmonary dysplasia (BPD) in prematurely born infants is substantial, but information related to the specific genes involved is lacking. RESULTS: Genotype analysis revealed, after multiple comparisons correction, two significant single-nucleotide polymorphism (SNPs), rs3771150 (IL-18RAP) and rs3771171 (IL-18R1), in African Americans (AAs) with BPD (vs. AAs without BPD; q < 0.05). No associations with Caucasian (CA) BPD, AA or CA respiratory distress syndrome (RDS), or prematurity in either AAs or CAs were identified with these SNPs. Respective frequencies were 0.098 and 0.093 in infants without BPD and 0.38 for each SNP in infants with BPD. In the replication set (82 cases; 102 controls), the P values were 0.012 for rs3771150 and 0.07 for rs3771171. Combining P values using Fisher's method, overall P values were 8.31 × 10(-7) for rs3771150 and 6.33 × 10(-6) for rs3771171. DISCUSSION: We conclude that IL-18RAP and IL-18R1 SNPs identify AA infants at risk for BPD. These genes may contribute to AA BPD pathogenesis via inflammatory-mediated processes and require further study. METHODS: We conducted a case-control SNP association study of candidate genes (n = 601) or 6,324 SNPs in 1,091 prematurely born infants with gestational age <35 weeks, with or without neonatal lung disease including BPD. BPD was defined as a need for oxygen at 28 days.

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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.

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Surfactant protein A (SP-A) plays a role in lung innate immunity and surfactant-related functions. Two functional genes, SP-A1 (SFTPA1) and SP-A2 (SFTPA2), are present in humans and primates (rodents have one gene). Single gene SP-A1 or SP-A2 proteins expressed in vitro are functional. To study their role in vivo, we generated humanized transgenic (hTG) C57BL/6 mice, SP-A1(6A(4)) and SP-A2(1A(3)). The SP-A cDNA in experimental constructs was driven by the 3.7-kb SP-C promoter. Positive hTG mice were bred with SP-A knock-out mice to generate F8 offspring for study. Epithelial alveolar type II cells were SP-A-positive, and Clara cells were negative by immunohistochemistry in hTG mice. The levels of SP-A in lungs of two hTG lines used were comparable with those in human lungs. Southern blot analysis indicated that two cDNA copies of either SP-A1(6A(4)) or SP-A2(1A(3)) were integrated as a concatemer into the genome of each of the two hTG lines. Electron microscopy analysis revealed that hTG mice with a single SP-A1(6A(4)) or SP-A2(1A(3)) gene product lacked tubular myelin (TM), but hTG mice carrying both had TM. Furthermore, TM was observed in human bronchoalveolar lavage fluid only if both SP-A1 and SP-A2 gene products were present and not in those containing primarily (>99.7%) either SP-A1 or SP-A2 gene products. In vivo rescue study confirmed that TM can only be restored after administering exogenous SP-A containing both SP-A1 and SP-A2 into the lungs of SP-A knock-out mice. These observations indicate that SP-A1 and SP-A2 diverged functionally at least in terms of TM formation.

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Severity of lung injury with respiratory syncytial virus (RSV) infection is variable and may be related to genetic variations. This preliminary report describes a prospective, family-based association study of children hospitalized secondary to RSV, aimed to determine whether intragenic and other haplotypes of surfactant proteins (SP)-A and SP-D are transmitted disproportionately from parents to offspring with RSV disease. Genomic DNA was genotyped for several SP-A and SP-D single nucleotide polymorphisms (SNPs). Transmission disequilibrium test analysis was used to determine transmission of variants and haplotypes from parents to affected offspring. Three hundred seventy-five individuals were studied, including 148 children with active RSV disease and one or both parents. The SP-A2 intragenic haplotype 1A was found to be protective (p = 0.013). The SP-D SNP DA160_A may possibly be an "at-risk" marker (p = 0.0058). Additional two- and three-marker haplotypes were associated with severe RSV disease, with two being protective (DA11_T/DA160_G and DA160_G/SP-A2 1A/SP-A1 6A). We conclude that there may be associations between SP-A and SP-D and RSV disease. Further study is required to determine whether these variants can be used to target a high-risk patient population in clinical trials aimed at reducing either the symptoms of acute infection or long-term pulmonary sequelae.

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AIMS: Polymorphisms of genes are transmitted together in haplotypes, which can be used in the study of the development of complex diseases such as respiratory distress syndrome (RDS). The surfactant proteins (SPs) play important roles in lung function, and genetic variants of these proteins have been linked with lung diseases, including RDS. To determine whether haplotypes of SP-A and SP-D are transmitted disproportionately from parents to offspring with RDS, we hypothesized that previously unstudied genetic haplotypes of these SP genes are associated with the development of RDS. METHODS: DNA was collected from 132 families of neonates with RDS. Genotyping was performed, and haplotype transmission from parent to offspring was determined by transmission disequilibrium test. RESULTS: The two-marker SP-D/SP-A haplotype DA160_A/SP-A2 1A(1) is protective against the development of RDS (p = 0.035). Four three- and four-marker haplotypes containing one or both loci from the significant two-marker haplotype are also protective against the development of RDS. CONCLUSIONS: These data identify protective haplotypes against RDS and support findings related to SP genetic differences in children who develop RDS. Study of haplotypes in complex diseases with both genetic and environmental risk factors may lead to better understanding of these types of diseases.

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Genetic variants of surfactant protein B (SP-B) have been associated with respiratory distress syndrome (RDS) in the prematurely born infant. We wished to determine linkage between RDS and SP-B single nucleotide polymorphisms (SNPs) [-18 (A/C), 1013 (A/C), 1580 (C/T), and 9306 (A/G)] or SP-B-linked microsatellite [(D2S388, D2S2232, (AAGG)n, and GATA41E01 (or D2S1331)] loci and identify susceptibility or protective alleles and haplotypes. We genotyped 132 families consisting of one or two parents and at least one child affected with RDS and performed biallelic and multiallelic family-based association test (FBAT) analysis, and extended transmission disequilibrium test (ETDT). ETDT analysis identified the microsatellite SP-B-linked loci (except D2S2232) to be linked to RDS. One allele from each of these three marker loci contributes to the risk of RDS. Multiallelic FBAT analysis detected a signal of linkage for the region of the four SNP loci. Three haplotypes within this region contribute to RDS risk. Although no other region showed significant linkage as judged by multiallelic FBAT, biallelic FBAT analysis revealed three potential susceptibility haplotypes formed by two to four loci within the SP-B and SP-B-linked microsatellite region. Each haplotype included GATA41E01, which was identified by ETDT analysis to be linked to RDS. We conclude that SP-B or SP-B-linked loci are linked to RDS and certain alleles or haplotypes are susceptibility or protective factors for the development of RDS in infants born prematurely.

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Derangement in pulmonary surfactant or its components and alveolar collapse are common findings in idiopathic pulmonary fibrosis (IPF). Surfactant proteins play important roles in innate host defense and normal function of the lung. We examined associations between IPF and genetic polymorphic variants of surfactant proteins, SP-A1, SP-A2, SP-B, SP-C, and SP-D. One SP-A1 (6A(4)) allele and single nucleotide polymorphisms (SNPs) that characterize the 6A(4) allele, and one SP-B (B1580_C) were found with higher frequency ( P

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