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Microarray analysis to identify novel genes regulated by Cholera toxin in bone marrow-derived dendritic cells (BMDCs)

ABSTRACT: The mechanisms by which dendritic cells (DCs) induce differentiation of naïve CD4+ T cells along the Th2 lineage is not well understood given that DCs themselves do not produce IL-4. In the present study, we undertook a microarray approach to identify genes involved in the induction of Th2 differentiation by DCs treated with a Th2-skewing adjuvant cholera toxin (CT). In the microarray analysis, murine bone marrow derived immature DCs were treated with CT. Of particular interest was tthe significant upregulation of the expression of c-kit. The upregulation of c-kit on DCs is critical for the induction of a Th2 response. Keywords: cholera toxin, bone marrow derived dendritic cells, gene expression array-based (RNA / spotted DNA/cDNA)

ORGANISM(S): Mus musculus

PROVIDER: GSE10815 | GEO | 2008/04/14

SECONDARY ACCESSION(S): PRJNA107411

REPOSITORIES: GEO

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Microarray analysis to identify novel genes regulated by Cholera toxin in bone marrow-derived dendritic cells (BMDCs)

Project description:The mechanisms by which dendritic cells (DCs) induce differentiation of naÃ¯ve CD4+ T cells along the Th2 lineage is not well understood given that DCs themselves do not produce IL-4. In the present study, we undertook a microarray approach to identify genes involved in the induction of Th2 differentiation by DCs treated with a Th2-skewing adjuvant cholera toxin (CT). In the microarray analysis, murine bone marrow derived immature DCs were treated with CT. Of particular interest was tthe significant upregulation of the expression of c-kit. The upregulation of c-kit on DCs is critical for the induction of a Th2 response. Keywords: cholera toxin, bone marrow derived dendritic cells, gene expression array-based (RNA / spotted DNA/cDNA) Murine bone marrow cells were cultured in the presence of GM-CSF (10 ng/ml) for 6 days . On day 6 the cells were harvested and purified using magnetically labeled anti mouse CD11c+ beads . The DCs were stimulated with CT (1 ug/ml) for 24 hours and the RNA was isolated.

2008-04-14 | E-GEOD-10815 | biostudies-arrayexpress

Effect of cholera toxin or forskolin on mouse bone marrow-derived dendritic cells

Project description:Bone marrow-derived dendritic cells from C57BL/6 mice were treated with 1 ug/ml cholera toxin, 10 uM forskolin or control medium for 2 h.

2010-05-07 | GSE21711 | GEO

Cholera toxin mediates cAMP-dependent CTLA-2 secretion by dendritic cells to induce regulatory T cells in the EAE model

Project description:Therapy of autoimmune diseases by tolerogenic dendritic cells (DCs) represents an interesting option for clinical application. Mostly, immature or semi-mature stages of DCs have been shown to convert naïve T cells into FoxP3+ induced regulatory T cells (iTreg). Here we found that murine bone marrow-derived DCs (BM-DCs) matured with cholera toxin (CT) produced cytotoxic T lymphocyte antigen 2 (CTLA-2) via cAMP signaling. CTLA-2 is a cathepsin L inhibitor, however, it exerted cathepsin L-independent tolerogenic functions as shown with cathepsin L knock-out BM-DCs. High or low dose CT (CThi, CTlo) or cAMP treatment of BM-DCs, but not other stimuli such as LPS or TNF, induced mRNA of both isoforms of CTLA-2α and CTLA-2β. CThi-, CTlo- or cAMP-DCs display a mature phenotype (MHC IIhi, costimulationhi). Importantly, only CThi DCs secreted IL-1β, IL-6 and IL-23. Consequently, CThi DCs instructed Th17 cell polarization while, CTlo- or cAMP-DCs induced TGF-β-dependent FoxP3+ iTreg conversion but little or no Th17 cells. iTreg conversion could be reduced by blocking of CTLA-2 with siRNAs and in accordance, addition of recombinant CTLA-2α increased the conversion towards iTreg in vitro. Injection of CTlo- or cAMP-DCs exerted MOG peptide-specific protective effects in experimental autoimmune encephalomyelitis (EAE). Together, we identified CTlo- or cAMP-induced CTLA-2 production by BM-DCs as a tool to enhance their iTreg induction in vitro and in vivo. These findings point to CTLA-2 as a novel mediator of immune evasion by cholera bacteria and its potential use to treat autoimmune diseases.

2017-03-15 | GSE79274 | GEO

Effect of cholera toxin or forskolin on mouse bone marrow-derived dendritic cells

Project description:Bone marrow-derived dendritic cells from C57BL/6 mice were treated with 1 ug/ml cholera toxin, 10 uM forskolin or control medium for 2 h. drug treatment groups

2010-05-27 | E-GEOD-21711 | biostudies-arrayexpress

Transcription profiling of pig IPEC-J2 cells treated with cholera toxin for 8h

Project description:The molecular mechanisms involved in host pathogen interactions at mucosal surfaces needs to be better understood in order to develop immune-mediated methods of protection against pathogens. Cholera toxin (CT) has the rare ability for a protein of inducing robust mucosal immunity in the gut and is therefore an excellent model with which to determine mechanisms of adjuvanticity and immunogenicity at intestinal mucosal surfaces. Jejunal epithelial cells are one of the first sites of antigen encounter. Therefore a porcine intestinal epithelial cell line, IPEC-J2, was cultured in 6-well transwell plates in the presence or absence of 50 ng/ml cholera toxin for up to 8 hours and the cell layer was harvested for gene expression analysis using the Affymetrix porcine genome array and real-time PCR analysis. Affymetrix analysis identified, and real-time PCR analysis of 15 genes confirmed, an increase in gene expression for 59 genes and a decrease in gene expression for 14 genes under CT treatment. An 8 hour time course of expression revealed that by 2-4 hours after CT treatment, all 10 upregulated genes were differentially expressed and by 4-6 hours after CT treatment 3 of the 5 downregulated genes were differentially expressed. These data suggest that the potent mucosal adjuvanticity and immunogenicity of CT derives from rapid alterations in gene expression at the site of first antigen encounter with the immune system. Characterization of early immune gene expression may elucidate potential biological mechanisms for mucosal immune induction leading to the development of effective vaccines against enteric pathogens. Experiment Overall Design: Confluent IPEC-J2 cell monolayers were treated with or without 50 ng/ml cholera toxin in transwell dishes for 8h. The experiment was repeated 3 times for a total of 6 chips, 3 cholera toxin treated and 3 untreated. Gene expression was compared between cholera toxin treated and untreated cells.

2008-06-11 | E-GEOD-10314 | biostudies-arrayexpress

IPEC-J2 cells treated with cholera toxin for 8h

Project description:The molecular mechanisms involved in host pathogen interactions at mucosal surfaces needs to be better understood in order to develop immune-mediated methods of protection against pathogens. Cholera toxin (CT) has the rare ability for a protein of inducing robust mucosal immunity in the gut and is therefore an excellent model with which to determine mechanisms of adjuvanticity and immunogenicity at intestinal mucosal surfaces. Jejunal epithelial cells are one of the first sites of antigen encounter. Therefore a porcine intestinal epithelial cell line, IPEC-J2, was cultured in 6-well transwell plates in the presence or absence of 50 ng/ml cholera toxin for up to 8 hours and the cell layer was harvested for gene expression analysis using the Affymetrix porcine genome array and real-time PCR analysis. Affymetrix analysis identified, and real-time PCR analysis of 15 genes confirmed, an increase in gene expression for 59 genes and a decrease in gene expression for 14 genes under CT treatment. An 8 hour time course of expression revealed that by 2-4 hours after CT treatment, all 10 upregulated genes were differentially expressed and by 4-6 hours after CT treatment 3 of the 5 downregulated genes were differentially expressed. These data suggest that the potent mucosal adjuvanticity and immunogenicity of CT derives from rapid alterations in gene expression at the site of first antigen encounter with the immune system. Characterization of early immune gene expression may elucidate potential biological mechanisms for mucosal immune induction leading to the development of effective vaccines against enteric pathogens. Keywords: Treatment comparison, cholera toxin vs untreated at 8h time point

2008-04-01 | GSE10314 | GEO

Genome-wide effect of cholera toxin and nitrate treatment in etiolated rice plants

2010-12-01 | GSE20924 | GEO

Total and cholera toxin B-tetramer specific T cells from Peyer's patches and mesenteric lymph nodes

Project description:T cells were separately isolated from MLN and PP from two naive mice, and T cells binding to a cholera toxin B subunit specific MHC II tetramer from mice orally immunized with cholera toxin 7 days or boosted 5 days with a second dose of cholera toxin. Cells were purified using flow cytometric cell sorting and 3´ (n=5) or 5´ (n=5) libraries were constructed using the 10X workflow. From 5´libraries TCR libraries were also constructed and sequenced.

2022-06-22 | E-MTAB-11805 | biostudies-arrayexpress

Genome-wide effect of cholera toxin and nitrate treatment in etiolated rice plants

Project description:The present study used microarray approach to identify the genomewide response to cholera toxin in the presence of nitrate. Considering that fact that the possibility of the existence of multiple GM-NM-1 genes/proteins in plants has not been conclusively ruled out, analysis of the genomewide impact of RGA1 mutation in rice and GPA1 mutation in Arabidopsis reveal only those genes that are under their direct control. On the other hand, assuming that all those different GM-NM-1 subunits in any given plant are regulated by cholera toxin, analysis of the genomwide response to cholera toxin could capture the entire G-protein responsive transcriptome, beyond what can be revealed by the mutant approach. This could reveal even those genes that respond to other, as yet unidentified GM-NM-1 subunits, as well as reveal some genes that are non-specifically regulated by cholera toxin, independent of any G-proteins. Total RNA was isolated from 10 day old excised rice leaves treated with or without cholera toxin in the presence of nitrate.Total RNA was first converted into cDNA and then into cRNA labeled with Cy3 and Cy5 dyes. These were hybridized on the 44K 60-mer whole genome arrays on glass slides. The slides were washed and scanned on an Agilent scanner (G2565B) at 100 % laser power. Data extraction was carried out with Agilent Feature Exiraction software (version 9.1). The experiment was repeated with a biological-cum-technical replicate where total RNA samples were isolated from a fresh batch of rice plants and the cRNAs were labeled with opposite dyes (dye-swap).

2010-12-01 | E-GEOD-20924 | biostudies-arrayexpress

single cell RNA sequencing of migratory dendritic cells after intranasal adjuvant/tolerogen administration

Project description:We compared migratory DCs in draining mediastinal lymph node after intranasal administration of different vaccine adjuvants based on CTA1 subunit of cholera toxin. The goal was to determine different characteristics of dendritic cells subsets after pro-inflammatory vs tolerogenic adjuvant. We had 3 groups: CTA1-DD (pro- inflammatory), CTA1(R7K)-DD (tolerogenic) and PBS to define DC which migrate to LN. All vaccine constructs included 3Ea peptide to allow for sorting of DC which took up the vaccine.

2023-05-15 | E-MTAB-12942 | biostudies-arrayexpress

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