Surfactant plus budesonide decreases lung and systemic responses to injurious ventilation in preterm sheep
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PROVIDER: PRJNA542880 | ENA |
REPOSITORIES: ENA
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Project description:Background: Intratracheal budesonide mixed with surfactant has been investigated as an alternative to decrease lung injury and prevent bronchopulmonary dysplasia in preterm infants. However, possible systemic effects are not known. Our goal was identify systemic effects of intra-tracheal instilled budesonide by transcriptome analysis of the brain and lung in a preterm lamb model of mechanical ventilation. Methods: We performed RNA-sequencing of the fetal periventricular white matter and liver from preterm sheep after treatment with intratracheal budesonide mixed with surfactant or surfactant alone followed by mechanical ventilation. Unventilated animals were used as controls. mRNA profiles were generated from polyA selected RNA using Illumina platform. Read sequences were aligned to the sheep genome (Oar 4.0) using Bowtie2 followed read counts using featureCounts. We performed differential expression analysis using EdgeR. Results: We identified large and significant gene expression genes both in the liver and in the priventricular white matter in animals treated with budesonide+surfactant compared to surfactant alone and unventilated animals. There were complexes synergies and antagonism of effect of ventilation and budesonide on gene expression. Ventilation induced inflammatory signaling in the periventricular white matter and liver which was partially antagonized by budesonide. Budesonide also suppressed developmental pathways in the periventricular white matter. Conclusions: Intratracheal budesonide had systemic effects including suppression of developmental pathways in the brain. These effects could have clinical implication in affecting the neurodevelopmental outcomes of preterm newborns.
2019-11-26 | GSE131195 | GEO
Project description: Not available
| S-EPMC6985873 | biostudies-literature
Project description:This study uses microarray analyses to examine gene expression profiles for Mycobacterium tuberculosis (Mtb) induced by exposure in vitro to bovine lung surfactant preparations that vary in apoprotein content: (i) whole lung surfactant (WLS) containing the complete mix of endogenous lipids and surfactant proteins (SP)-A, -B, -C, and -D; (ii) extracted lung surfactant (CLSE) containing lipids plus SP-B and -C; (iii) column-purified surfactant lipids (PPL) containing no apoproteins, and (iv) purified human SP-A. Exposure to WLS evoked a multitude of transcriptional responses in Mtb, with 52 genes up-regulated and 23 genes down-regulated at 30 min exposure, plus 146 genes up-regulated and 27 genes down-regulated at 2 h. Notably, WLS rapidly induced several membrane-associated lipases that presumptively act on surfactant lipids as substrates, and a large number of genes involved in the synthesis of phthiocerol dimycocerosate (PDIM), a cell wall component known to be important in macrophage interactions and Mtb virulence. Exposure of Mtb to CLSE, PPL, or purified SP-A caused a substantially weaker transcriptional response (≤ 20 genes were induced) suggesting that interactions among multiple lipid-protein components of WLS may contribute to its effects on Mtb transcription.
2009-06-01 | GSE14005 | GEO
Project description:Purpose: The goal of this study was to measure by RNA-seq the impact of preterm birth and invasive mechanical ventilation verses noninvasive respiratory support on the lung transcriptome Methods: Lung mRNA profiles were generated for: 1) Unventilated preterm lamb, 2) preterm lambs delivered at gd131 and intubated and mechanically ventilated for 3 days, 3) preterm lambs delivered at gd131 and not intubated and resuscitated by placing a face mask over the nose and mouth and controlling O2 delivery via a computer-controlled electronic blower device, 4) unventilated naturally delivered full-term lambs. Results: Using an optimized data analysis workflow, we mapped between 61 and 81 million sequence reads per sample to the sheep genome Oar_v3.1/oviAri3 Conclusions: Our study represents the first detailed analysis of ventilated preterm lung transcriptomes, with biologic replicates, generated by RNA-seq
2021-06-30 | GSE163806 | GEO
Project description:Sash1 acts as a scaffold in TLR4 signaling. We generated Sash1-/- mice, which die in the perinatal period due to respiratory distress. Constitutive or endothelial-restricted Sash1 loss leads to a reduction of surfactant-associated protein synthesis. We show that Sash1 interacts with β-arrestin 1 downstream of the TLR4 pathway to activate Akt and eNOS in microvascular endothelial cells. Generation of nitric oxide downstream of Sash1 in endothelial cells activated cGMP in adjacent alveolar type 2 cells to induce transcription of surfactant genes. Thus we identify a critical cell nonautonomous function for Sash1 in embryonic development in which endothelial Sash1 affects alveolar type 2 cells and promotes pulmonary surfactant production through nitric oxide signaling. Lack of pulmonary surfactant is a major cause of respiratory distress and mortality in preterm infants, and these findings identify the endothelium as a potential target for therapy.
2019-05-01 | GSE123889 | GEO
Project description:Purpose: ATAC-seq was performed on preterm lambs that were ventilated by invasive mechanical ventilation or noninvasive respiratory support utilizing a mask and compared to gestation-age-matched preterm lambs that were not ventilated and naturally delivered term lambs. Methods: Lung chromatin access profiles were generated for: 1) Unventilated preterm lamb, 2) preterm lambs delivered at gd131 and intubated and mechanically ventilated for 3 days, 3) preterm lambs delivered at gd131 and not intubated and resuscitated by placing a face mask over the nose and mouth and controlling O2 delivery via a computer-controlled electronic blower device, 4) unventilated naturally delivered full-term lambs. Results: Using an optimized data analysis workflow, we mapped between 76 and 96 million sequence reads per sample to the sheep genome Conclusions: Our study represents the first detailed analysis of ventilated preterm lung chromatin access, with biologic replicates, generated by ATAC-seq
2021-06-30 | GSE163807 | GEO
Project description:To understand the molecular mechanisms of human lung macrophage development, function, and role in BPD pathogenesis, we conducted a clinical study using isolated tracheal aspirate macrophages from intubated preterm infants born before 30 wk gestation. One hundred twenty-eight patients intubated for respiratory distress syndrome and surfactant administration were consented for the study.
2020-05-28 | GSE149490 | GEO
Project description:This study uses microarray analyses to examine gene expression profiles for Mycobacterium tuberculosis (Mtb) induced by exposure in vitro to bovine lung surfactant preparations that vary in apoprotein content: (i) whole lung surfactant (WLS) containing the complete mix of endogenous lipids and surfactant proteins (SP)-A, -B, -C, and -D; (ii) extracted lung surfactant (CLSE) containing lipids plus SP-B and -C; (iii) column-purified surfactant lipids (PPL) containing no apoproteins, and (iv) purified human SP-A. Exposure to WLS evoked a multitude of transcriptional responses in Mtb, with 52 genes up-regulated and 23 genes down-regulated at 30 min exposure, plus 146 genes up-regulated and 27 genes down-regulated at 2 h. Notably, WLS rapidly induced several membrane-associated lipases that presumptively act on surfactant lipids as substrates, and a large number of genes involved in the synthesis of phthiocerol dimycocerosate (PDIM), a cell wall component known to be important in macrophage interactions and Mtb virulence. Exposure of Mtb to CLSE, PPL, or purified SP-A caused a substantially weaker transcriptional response (⤠20 genes were induced) suggesting that interactions among multiple lipid-protein components of WLS may contribute to its effects on Mtb transcription. Whole lung surfactant (WLS) was obtained by centrifugation of lavage fluid (0.15M NaCl) from the intact lungs of freshly killed calves (ONY, Inc, Amherst, NY). Centrifugation was done at 12,000 x g at room temperature to pellet the active large aggregate surfactant fraction. CLSE was prepared by extraction of WLS into chloroform-methanol by the method of Bligh and Dyer (1959), as described in our prior work (Notter et al. 2002). CLSE is conceptually identical to the substance of the clinical surfactant Infasurf®. The composition of CLSE by weight was approximately 94% phospholipid, 4.5% cholesterol, and 1.5% protein (a mixture of SP-B and SP-C) as defined previously (Notter et al. 2002). Mixed lung surfactant phospholipids (PPL) were isolated from CLSE by column chromatography with an elution solvent of 1:1 (v/v) chloroform-methanol plus 5% 0.1 N HCl. Two passes through a Sephadex LH-20 column (Pharmacia-LKB Biotechnology, Piscataway, NJ) separated phospholipids away from hydrophobic surfactant apoproteins and neutral lipids. Acid remaining in PPL fractions was removed by a second chloroform-methanol extraction. Final PPL had a protein content below the limits of detection of both the Folin-phenol assay of Lowry et al. (1951) and the amido black assay of Kaplin and Pedersen (1989) modified by the addition of 15% sodium dodecyl sulfate (SDS) to allow accurate quantitation in the presence of lipid. Human SP-A was a gift from Dr. J. R. Wright at Duke University, and was isolated from bronchoalveolar lavage from patients with alveolar proteinosis as described previously (Wright et al. 1987). For exposure studies with Mtb, amounts of WLS, CLSE, and PPL used per experiment were 3.3 mg in a total volume of 4 ml. In preliminary studies, this concentration was shown to induce a transcriptional response in Mtb without affecting viability after a 2 h treatment as determined by serial dilutions compared to untreated Mtb. SP-A was studied at a level of 0.165 mg based on its putative concentration of approximately 5% in endogenous surfactant. To determine the effects of lung surfactant on the transcriptional response of Mtb, preparations of WLS, CLSE, or PPL (in 650 µl PBS) were initially coated onto the bottom surface of a 25 cm2 cell culture flask for 30 min at 37oC to mimic a surfactant layer. A minimum of two independent biological replicates were studied for each surfactant preparation. Purified human SP-A was also examined for its effects on Mtb transcription, and phosphate buffered saline (PBS) was used as a âno treatmentâ control. In experiments with SP-A (in 650 µl PBS), the pure apoprotein was either pre-coated on the flask as done with the surfactant preparations, or was directly added and incubated with Mtb during study. Mtb was added to the flask in 3.35 ml of DMEM (supplemented with 2% FCS and 5 mM HEPES) and exposed to coated surfactant/SP-A or PBS control for 30 min or 2h at 37oC, followed by RNA isolation as detailed by Rohde et al. (2008). GTC lysis/stabilization buffer was added, and pelleted Mtb were washed and then digested with lysozyme. TRIzol (Invitrogen) was added, and organisms were disrupted using glass beads in a Mini-Bead Beater (BioSpec Products, Inc. Bartlesville, OK). After extraction of Mtb lysates with chloroform, ethanol was added to the aqueous phase before loading samples onto RNeasy mini columns (Qiagen, Valencia, CA). Subsequent steps were then performed according to the manufacturerâs instructions. Following elution with RNase-free water, the RNA samples were treated with DNase (Turbo DNAfree, Applied Biosystems, Foster City, CA), and stored at -70oC until analysis. The MessageAmpTMII-Bacteria system (Applied Biosystems) was used to amplify mycobacterial RNA for all samples [16]. Total RNA was reverse transcribed to yield double-stranded amino-allyl modified aRNA, which was labeled with Alexa Fluor 555 and Alexa Fluor 647 (Invitrogen). Excess reactive dyes were removed with a Mega Clear purification kit (Applied Biosystems). Next, Alexa-labeled aRNA from paired samples was dried using a Speedvac, and re-suspended in hybridization buffer (5x SSC, 25% formamide, 0.15% SDS, and 25 μg salmon sperm DNA). The mixed samples were denatured by being heated to 95oC for and then cooled to 50oC prior to loading them onto microarrays. The arrays were hybridized for approximately 15 h at 45oC, washed and then dried by centrifugation. Fluorescence intensity data from each array were collected with a GenePix 4000B scanner (Axon Instruments, Union City, CA) using GenePixPro3.0 software (Axon Instruments). Image analysis, i.e. spot identification, background measurements, quality assessment and flagging were conducted with Imagene software (Version 6.0, BioDiscovery, Inc., El Segundo, CA). Poor quality spots were filtered and removed from the analysis. Subsequent normalization, statistical analysis (i.e. t-test), and visualization of array data was conducted using Genespring (Version 7, Agilent Technologies, Palo Alto, CA).
2010-05-18 | E-GEOD-14005 | biostudies-arrayexpress
Project description:This project focuses both on the separate and combined effects of large tidal volume ventilation and hyperoxia on gene expression in the lungs of anesthetized rats. Rats were either mechanically or spontaneously ventilated at either 21 or 100% oxygen and expression levels were evaluated on RG_U34 GeneChips. Keywords: other
2003-07-16 | GSE489 | GEO
Project description:This study aimed to use a lamb model of preterm birth to investigate the impact of different tidal volume strategies, applied during positive pressure ventilation, on lung injury development. Preterm lambs were ventilated for a total of 15 minutes, followed by a 30 minute ‘rest’ period where lambs were supported via the intact placenta. A group of lambs was also euthanised at birth to act as an unventilated control group. At post-mortem lung tissue was sampled for proteomic analysis. Lung tissue samples were analysed by mass spectrometry-based proteomics using a TMT-11plex labelling approach, followed by nano-liquid chromatography coupled to an Q Exactive HF-X benchtop Orbitrap mass spectrometer in a data-dependent acquisition mode.
2023-10-09 | PXD041917 | Pride