Murine bronco alveolar lavage fluid from wild type and mannose receptor knock out mice, unchallenged and LPS challenged
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ABSTRACT:
INSTRUMENT(S):
ORGANISM(S): Mus Musculus (mouse)
TISSUE(S): Lung
SUBMITTER: 
Kirstine Nørregaard
LAB HEAD: Niels Behrendt
PROVIDER: PXD049201 | Pride | 2024-04-17
REPOSITORIES: Pride
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Project description:Rationale: Lipopolysaccharide (LPS) is ubiquitous in the environment. Inhalation of LPS has been implicated in the pathogenesis and/or severity of several lung diseases, including pneumonia, chronic obstructive pulmonary disease and asthma. Alveolar macrophages are the main resident leukocytes exposed to inhaled antigens. Objectives: To obtain insight into which innate immune pathways become activated within human alveolar macrophages upon exposure to LPS in vivo. In seven healthy humans sterile saline was instilled into a lung segment by bronchoscope, followed by instillation of LPS into the contralateral lung. Six hours later a bilateral bronchoalveolar lavage was performed and whole-genome transcriptional profiling was done (Affymetrix HG-U133 Plus 2.0) on purified alveolar macrophages, comparing cells exposed to saline or LPS from the same individuals.
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Project description:Thirty female BALB/c mice of 6-8 weeks of age were purchased from Charles River (Montreal, QC, Canada) and divided randomly into control (n=15) and treated (n=15) groups. Mice were maintained on an egg free diet in the pathogen-free conditions at the University of Guelph Central Animal Facility. Animals were maintained between 2224°C and at a relative humidity of 4050%. The light-to-dark cycle was maintained at 12 h from 7 AM to 7 PM. All of the animal experiments were approved by the University of Guelph Animal Care Committee according the Canadian Council on Animal Care (CCAC) guidelines.Mice were exposed to intra-gastric gavage (i.g.) twice every week for 5 weeks with OVM (1 mg/100 μl) and 10 μg of cholera toxin adjuvant. An amino acid (Sigma, St. Louis, MO) solution of 10 μg/μl (100 μl per mouse with 10 μg of cholera toxin) was used as a negative control. The amino acids were mixed accordingly to match the OVM amino acid composition. Five weeks after the first sensitization, mice were challenged by i.p. injection with 1 mg of OVM in 50 μl of sterile water and aluminum hydroxide (50 μl, 2% Alhydrogel) adjuvant (Superfos Biosector, Denmark), to induce an immediate type-1 hypersensitivity reaction. The anaphylactic responses were visualized and scored depending on the response of the individual mice in each of the treated and control groups. After 40 min, mice were euthanized with CO2 and partial spleen tissue samples were collected in RNAlater solution for microarray experiment (Ambion Inc., TX, USA). Spleen samples in RNAlater solution were kept at 4oC overnight prior to long term storage at -80oC.
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Project description:Protein expression is regulated by production and degradation of mRNAs and proteins, but their specific relationships remain unknown. We combine measurements of protein production and degradation and mRNA dynamics to build a quantitative genomic model of the differential regulation of gene expression in LPS stimulated mouse dendritic cells. Changes in mRNA abundance play a dominant role in determining most dynamic fold changes in protein levels. Conversely, the preexisting proteome of proteins performing basic cellular functions is remodeled primarily through changes in protein production or degradation, accounting for over half of the absolute change in protein molecules in the cell. Thus, the proteome is regulated by transcriptional induction of novel cellular functions and remodeling of preexisting functions through the protein life cycle. Mouse primary dendritic cells were treated with LPS or mock stimulus and profiled over a 12-hour time course. Cells were grown in M-labeled SILAC media, which was replaced with H-labeled SILAC media at time 0. Aliquots were taken at 0, 0.5, 1, 2, 3, 4, 5, 6, 9, and 12 hours post-stimulation and added to equal volumes of a master mix of unlabeled (L) cells for the purpose of normalization. RNA-Seq was performed at 0, 1, 2, 4, 6, 9, and 12 hours post-stimulation.
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2018-10-22 | PXD006770 | Pride
Project description:Background: It has been shown previously that administration of Francisella tularensis (Ft) LVS lipopolysaccharide (LPS) protects mice against subsequent challenge with Ft LVS and blunts the pro-inflammatory cytokine response. Methods: To investigate further the molecular mechanisms that underlie Ft LVS LPS-mediated protection, we profiled global hepatic gene expression following Ft LVS LPS or saline pretreatment and subsequent Ft LVS challenge using Affymetrix arrays. Results: A large number of genes (> 3000) were differentially expressed at 48 hours post-infection. The degree of modulation of inflammatory genes by infection was clearly attenuated by LPS-pretreatment in the surviving mice. However, LPS alone had a subtle but significant effect on the gene expression profile of the uninfected mice. By employing gene set enrichment analysis, we discovered significant up-regulation of peroxisome proliferator activated receptors (PPARs) and their target genes in LPS-treated liver. Conclusions: We hypothesize that the LPS-induced blunting of pro-inflammatory response in mouse is, in part, mediated by PPARs (alpha and gamma). Experiment Overall Design: Two groups of 9 mice each were used for this experiment. 48 hours prior to Ft LVS challenge, mice were injected i.p. with either 100 ng of Ft LVS LPS or an equivalent volume of saline. On the day of challenge, 3 saline- and 3 Ft LVS LPS-pretreated animals were sacrificed (uninfected controls), while all remaining mice were challenged i.p. with ~4-5 X 105 Ft LVS. Ft LVS-challenged mice were sacrificed (in groups of 3) at 24 and 48 hours post-infection.
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Project description:Abstract Isorhynchophylline, a tetracyclic indole alkaloid, has anti-inflammatory and antioxidant activities against cardiovascular diseases and central nervous system disorders. Acute lung injury (ALI) is a manifestation of inflammation concentrated in the lungs and has a high incidence rate and mortality. Here, we established a mouse model of ALI and observed the effects of isorhynchophylline. Proteomic results showed that 5727 proteins were detected in mouse lung tissues, and 16 proteins were screened out. Isorhynchophylline could reverse the trend of these differential proteins. In addition, isorhynchophylline can act on integrin alpha M to reduce neutrophil recruitment and thereby produce anti-inflammatory effects and may suppress neutrophil migration through the leukocyte transendothelial migration pathway. TUNEL and RT-PCR experiments revealed that LPS-induced ALI in mice increases the apoptosis of lung tissues, damage to alveolar epithelial cells and levels of inflammatory factors. Treatment with isorhynchophylline can repair tissues, improve lung tissue pathology and reduce lung inflammation.
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