Microarray analysis on RNA from lung eosinophil subpopulation from naive or HDM-treated mice using Affymetrix Mouse Gene 2.0 ST
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ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Dimitri Pirottin
PROVIDER: E-MTAB-4084 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Innate immune cells control acute eosinophilic lung inflammation induced by cystein proteases. Here we characterize the dynamic change of gene expression profile in basophils, natural helper cells and eosinophils during lung inflammation via cystein protease Examination of mRNA levels in individual cell populations, basophils, natural helper cells and eosinophils of the lung from naïve mice and papain treated mice.
2014-03-28 | E-GEOD-56292 | biostudies-arrayexpress
Project description:Levels of asymmetric dimethylarginine (ADMA), an endogenous inhibitor of nitric oxide synthase, are increased in lung, sputum, exhaled breath condensate and plasma samples from asthma patients. ADMA is metabolized primarily by dimethylarginine dimethylaminohydrolase 1 (DDAH1) and DDAH2. We determined the effect of DDAH1 overexpression on development of allergic inflammation in mouse models of asthma. Wild type and DDAH1-transgenic mice were challenged with PBS or house dust mite (HDM). Airway inflammation was assessed by bronchoalveolar lavage (BAL) total and differential cell counts. Gene expression in lungs was determined by RNA-Seq and RT-quantitative PCR (qPCR). The expression of DDAH1 and DDAH2 was decreased in the lungs of mice following HDM exposure. Transgenic overexpression of DDAH1 resulted in decreased BAL total cell and eosinophil numbers following HDM exposure. Total IgE levels in serum and BAL fluid were decreased in HDM-exposed DDAH1-transgenic mice compared to HDM-exposed wild type mice. RNA-Seq results showed downregulation of genes in inducible nitric oxide synthase (iNOS) signaling pathway in PBS-treated DDAH1 transgenic mice versus PBS-treated wild type mice and downregulation of genes in IL-13/FOXA2 signaling pathway in HDM-treated DDAH1 transgenic mice versus HDM-treated wild type mice. Our findings suggest that decreased expression of DDAH1 in airway epithelial cells may contribute to allergic asthma and overexpression of DDAH1 attenuates allergen-induced airway inflammation through modulation of Th2 responses. mRNA profiles of WT and DDAH1-transgenic mice treated with PBS or house dust mite (HDM).
2013-12-03 | E-GEOD-49047 | biostudies-arrayexpress
Project description:Eosinophils are major effector cells in type 2 inflammatory responses and become activated in response to IL-4 and IL-33, yet the molecular mechanism remains unclear. We examined the direct effect of these cytokines on eosinophils and demonstrated that murine eosinophils respond to IL-4 and IL-33 by phosphorylation of STAT-6 and NFkB, respectively. RNA sequencing analysis of murine eosinophils indicated that IL-33 regulates 519 genes, whereas IL-4 regulates only 28 genes, including 19 IL-33-regulated genes. Interestingly, IL-33 induced eosinophil activation via two distinct mechanisms, IL-4 independent and IL-4 secretion/auto-stimulation dependent. Anti-IL-4 or anti-IL-4Ra antibody-treated eosinophils, as well as Il4- or Stat6-deficient eosinophils, had attenuated protein secretion of a subset of IL-33-induced genes, including Retnla and Ccl17. However, the induction of most IL-33-regulated transcripts (e.g. Il6 and Il13) was IL-4 independent and blocked by NFkB inhibition. Indeed, IL-33 induced the rapid release of pre-formed IL-4 protein from eosinophils by an NFkB-dependent mechanism. Thus, we have identified a novel activation pathway in murine eosinophils that is induced by IL-33 and differentially dependent upon IL-4. These data suggest that IL-4 plays a critical role in auto-amplification of IL-33-induced eosinophil activation and could be a potential target for therapeutic approaches in IL-33-related eosinophil-associated diseases. Low density bone marrow derived murine eosinophils were generated in culture over the period of 14 days. Eosinophils were activated by either IL-33 or IL-4 at 10 ng/ml for 1hr and 4hr. RNA was collected and subjected to next generation sequencing.
2013-08-01 | E-GEOD-43660 | biostudies-arrayexpress
Project description:Purpose: we aimed to characterized eosinophils in the murine model of asthma Methods: Eosinophils were sorted from the lung of naïve and and HDM-induced asthma mice. There after RNA was extracted from the sorted eosinophils and subjected to RNAseq. Results:we demonstrated that resident lung eosinophils require IL-5 for their survival and that the expression of Siglec-F, is regulated by IL-5 . Conclusion: Our data suggest that the distinct eosinophil populations in the asthmatic lung represent a continuum of activation states rather than distinct eosinophil subsets.
2023-04-17 | GSE203225 | GEO
Project description:Activated eosinophils contribute to airway dysfunction and tissue remodeling in asthma, and thus are considered an important factor in asthma pathology. We report here comparative proteomic and phosphoproteomic changes upon activation of eosinophils using eight cytokines individually and in selected cytokine combinations, in time-course reactions. Differential protein and phosphoprotein expressions were determined by mass spectrometry after 2-dimensional gel electrophoresis (2DGE) followed by protein identification by MS, and by label-free LC-MS/MS. We found that each cytokine-stimulation produced significantly different changes in the eosinophil proteome and phosphoproteome, with phosphoproteomic changes being more pronounced and having an earlier onset. Furthermore, we observed that IL-5, GM-CSF, and IL 3 showed the greatest change in protein expression and phosphorylation, and this expression differed markedly from those of the other five cytokines evaluated. Comprehensive univariate and multivariate statistical analyses were employed to evaluate the comparative results. We also monitored eosinophil activation using flow cytometry (FC) analysis of CD69 expression. In agreement with our proteomic studies, FC indicated that IL-5, GM-CSF, and IL-3 were more effective than the other five cytokines studied in stimulating a cell surface CD69 increase indicative of eosinophil activation. Moreover, selected combinations of cytokines revealed proteomic patterns with many proteins in common with single cytokine expression patterns but also showed a greater than additive effect of the two cytokines employed, indicating a more complex signaling pathway that was reflective of a more typical inflammatory pathology.
2017-12-19 | PXD004043 | Pride
Project description:Global expression pattern in eosinophils in various conditions such as eosinophilic asthma, hypereosinophilic dermatological diseases, parasitosis and pulmonary aspergillosis. Eosinophil were isolated from peripheral blood by FACS from 4 hypereosinophilic conditions: eosinophilic asthma (4 replicates), hypereosinophilic dermatological diseases (3 replicates), parasitosis (3 replicates) and pulmonary aspergillosis (4 replicates).
2015-12-10 | E-GEOD-65239 | biostudies-arrayexpress
Project description:Segmental allergen challenge increases the percentage of eosinophils in bronchoalveolar lavage (BAL) cells. Mepolizumab, an anti-IL-5 therapeutic antibody, decreases the number of eosinophils in bronchoalveolar lavages (BAL). The use of both procedures allows to define genes that are either expressed by eosinophils or dependent on eosinophil presence in the airways. Cells from Bronchoalveolar lavages (BAL) are obtained by bronchoscopy before (V5) and 48 h after a segmental allergen challenge (V6) in atopic and mild astmatics. This procedure was repeated in the same two subjects 2 months later and 1 month after an injection of mepolizumab (V22 is before challenge and V23 after challenge). Cells were immediately lysed and isolated total RNAs were analyzed using the Human Genome 1.0 ST GeneChip arrays (Affymetrix, Santa Clara, CA).
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Project description:Objectives: Eosinophils cultured with all-trans RA (ATRA) and 9-cis RA showed dramatically induced cell survival, and the efficacy of RAs (10-6 M) was similar to that of IL-5 (1 ng/ml), the most critical cytokine for eosinophil activation. Using a gene microarray, we studied mRNA profile regulated by RAs. Methods: Peripheral venous blood was obtained from subjects with mild eosinophilia. Eosinophils were isolated by sedimentation with 6% dextran followed by centrifugation on 1.088 Percoll (Pharmacia, Uppsala, Sweden) density gradients. The cells were further purified by negative selection using anti-CD16 immunomagnetic beads and a MACS system (Miltenyi Biotec, Bergisch Gladbach, Germany). Purified eosinophils were resuspended in RPMI 1640 medium (Gibco, Grand Island, NY) with 10% FCS, and incubated with vehicle, 10-6 M of 9-cis RA, or ATRA in 1% human serum albumin-coated plates, each 4 Ã 106 cells, for 4 h. Then cells were lysed, and total RNA was isolated using Isogen (Nippon Gene, Tokyo, Japan) as per the manufacturerâs instructions. RNA was repurified with phenol-chloroform extraction and ethanol precipitation. For each cell culture condition, the total RNA from three donors was pooled, enabling us to acquire adequate amounts of RNA for microarray analysis. Results: There was little variation between the samples treated by 9-cisRA and ATRA. The comparison between vehicle control and 9-cisRA, or vehicle control and ATRA, identified an increase (greater than twofold change over control) in 19 and 11 transcripts, respectively. The number of decreased transcripts (less than 0.5-fold) resulting fromby 9-cisRA or ATRA exposure wasere 19 and 8 transcripts, respectively. In these transcripts, the expression changes relative to vehicle control were smaller in withthe ATRA compared with 9-cis RA, suggesting that 9-cis RA induced gene transcription more effectively rather than ATRA. Of note, among the ten genes with the biggestmost decrease relative tofrom vehicle control, we found an apoptosis- related gene, caspase-3. Conclusions: We screened several genes involved in RA-induced prolonged eosinophil survival. Purified peripheral blood eosinophils were incubated with vehicle, 10-6 M of 9-cis RA, or ATRA in 1% human serum albumin-coated plates, each 4 Ã 106 cells, for 4 h. Then cells were lysed, and total RNA was isolated. RNA was repurified with phenol-chloroform extraction and ethanol precipitation. For each cell culture condition, the total RNA from three donors was pooled, enabling us to acquire adequate amounts of RNA for microarray analysis.
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