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Transcription profiling of wild type and aire -/- murine meduallary thymic epithelial cells to study the immunological roles of aire protein in thymus.

ABSTRACT: Mice used were B6/129 F2's, 3-5 weeks of age, either wild type or with both copies of the autoimmune regulator gene (aire, GenBank #AF079536) disrupted. Thymi from five of these mice of both sexes were removed and pooled. After collagenase/dispase digestion, density gradient fractionation, and fluorescent antibody staining, cells with the phenotype CD45-, G8.8+, CDR1int and B7.1hi were FACS-sorted and total RNA was made from them. RNA was twice-amplified using a T7 polymerase-based method.

ORGANISM(S): Mus musculus

SUBMITTER: CBDM Lab

PROVIDER: E-GEOD-85 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Projection of an immunological self shadow within the thymus by the aire protein.

Anderson Mark S MS Venanzi Emily S ES Klein Ludger L Chen Zhibin Z Berzins Stuart P SP Turley Shannon J SJ von Boehmer Harald H Bronson Roderick R Dierich Andrée A Benoist Christophe C Mathis Diane D

Science (New York, N.Y.) 20021010 5597

Humans expressing a defective form of the transcription factor AIRE (autoimmune regulator) develop multiorgan autoimmune disease. We used aire- deficient mice to test the hypothesis that this transcription factor regulates autoimmunity by promoting the ectopic expression of peripheral tissue- restricted antigens in medullary epithelial cells of the thymus. This hypothesis proved correct. The mutant animals exhibited a defined profile of autoimmune diseases that depended on the absence of aire in ...[more]

PMID: 12376594

Similar Datasets

wild type and aire -/- murine meduallary thymic epithelial cells

Project description:Mice used were B6/129 F2's, 3-5 weeks of age, either wild type or with both copies of the autoimmune regulator gene (aire, GenBank #AF079536) disrupted. Thymi from five of these mice of both sexes were removed and pooled. After collagenase/dispase digestion, density gradient fractionation, and fluorescent antibody staining, cells with the phenotype CD45-, G8.8+, CDR1int and B7.1hi were FACS-sorted and total RNA was made from them. RNA was twice-amplified using a T7 polymerase-based method. Keywords: ordered

2002-09-19 | GSE85 | GEO

Transcription profiling of mouse lymphtoxin beta receptor-deficient thymic medullary epithelial cells vs. wild-type and Aire-deficient vs wild-type littermates

Project description:We compared gene expression profiles of lymphotoxin alpha- and lymphtoxin beta receptor-deficient thymic medullary epithelial cells with their wild-type littermates, as well as with Aire-deficient and wild-type littermates. This was done in order to determine whether there was overlap in the effects of lymphotoxin and aire. Experiment Overall Design: Individual thymi were digested with collagenase, dispase and DNAse, and meduallary epithelial cells with the phenotype CD45-G8.8+Ly51intMHCclasssIIhi were isolated by FACS.

2008-06-16 | E-GEOD-8563 | biostudies-arrayexpress

Transcription profiling of mouse ire-deficient and wild-type littermate thymic medullary epithelial cells

Project description:We compared gene expression profiles of aire-deficient and wild-type littermate thymic medullary epithelial cells. This was done in order to determine whether Aire's effects differed among strains, and also among individuals of the same strain. Experiment Overall Design: Individual thymi were digested with collagenase, dispase and DNAse, and meduallary epithelial cells with the phenotype CD45-G8.8+Ly51intMHCclasssIIhi were isolated by FACS.

2008-10-18 | E-GEOD-8564 | biostudies-arrayexpress

Transcription profiling of mouse thymic stromal cells from 6-12 week old wild type (C57BL/6) and Aire knock-outs (mixed background)

Project description:Gene expression in different thymic stromal cells and subsets thereof was analyzed in 6-12 week old wild type (C57BL/6) and Aire knock-out (mixed background) mice. Thymic stromal cells were purified by sequential enzymatic digestion (collagenase, collagenase/dispase and trypsin) followed by gradient centrifugation and FACS sorting. Sort criteria were as follows: dendritic cells (CD11c+, F4/80 -), macrophages (F4/80+, CD11c-), cTECs (CD45â/lo, CDR1/Ly51+, Ep-CAM+) and mTECs (CD45â/lo, CDR1/Ly51â, Ep-CAM+). mTECs of wild-type and Aire knock-out mice were further subdivided according to CD80 expression levels.,For microarray analysis total RNA from thymic stromal cell samples of two independent experiments was pre-amplified and biotinylated by two rounds of cDNA synthesis and in vitro transcription. Fluorescence readings were evaluated by using Microarray Suite 5.0 software.

2007-12-15 | E-GEOD-2585 | biostudies-arrayexpress

promiscuous gene expression in the mouse thymus

Project description:Gene expression in different thymic stromal cells and subsets thereof was analyzed in 6-12 week old wild type (C57BL/6) and Aire knock-out (mixed background) mice. Thymic stromal cells were purified by sequential enzymatic digestion (collagenase, collagenase/dispase and trypsin) followed by gradient centrifugation and FACS sorting. Sort criteria were as follows: dendritic cells (CD11c+, F4/80 -), macrophages (F4/80+, CD11c-), cTECs (CD45–/lo, CDR1/Ly51+, Ep-CAM+) and mTECs (CD45–/lo, CDR1/Ly51–, Ep-CAM+). mTECs of wild-type and Aire knock-out mice were further subdivided according to CD80 expression levels. For microarray analysis total RNA from thymic stromal cell samples of two independent experiments was pre-amplified and biotinylated by two rounds of cDNA synthesis and in vitro transcription. Fluorescence readings were evaluated by using Microarray Suite 5.0 software. Keywords = thymus Keywords = promiscuous gene expression Keywords = AIRE Keywords: ordered

2005-04-28 | GSE2585 | GEO

RNA-seq of medullary thymic epithelial cell (mTEC) subsets in inducible Aire-lineage tracing mice

Project description:The goal of the study was to sequence mRNA expression from sorted medullary thymic epithelial cell (mTEC) subsets in inducible Aire-CreERT2.R26-Stopfl-tdTomato lineage tracing mice after a pulse chase. Four cell subsets were sorted 7 days after a single 2mg pulse of tamoxifen administered by oral gavage. 4 biological replicates (1,2,3,4) were collected derived from 12 pooled thymi per replicate. From the DAPI-;CD45-;EpCAM+ TEC pool, cells were sorted as: pre-Aire (MHCIIlo;RFP-), early-Aire (MHCIIhi;RFP-), late-Aire (MHCIIhi;RFP+), and post-Aire (MHCIIlo;RFP+). The data were used to identify differentially expressed genes across the four mTEC subsets to examine mTEC heterogeneity and identify novel mTEC subpopulations.

2018-05-19 | GSE114651 | GEO

Gene expression analysis of a human thymic cell line model of AIRE molecular function

Project description:Microarray analysis of AIRE regulated gene expression has been performed extensively in studies using the thymi of different mouse strains, but to date, has only been rarely performed using human model systems, and critically, not yet with a human thymic model. We have generated a Flp-In host cell line variant of the TEC 1A3 human thymic epithelial cell line and subsequently used site-specific recombination to generate a variant that stably expresses recombinant AIRE. We used microarray analysis to determine the transcription expression profiles of these two variants and determine the repertoire of AIRE regulated gene expression in a human thymic model system. The aim of the present study is to use microarray analysis to identify the repertoire of AIRE regulated genes within a human model system, and to validate this model as an appropriate human model system for the study of the molecular function of AIRE

2018-06-01 | GSE35244 | GEO

Messenger RNA expression profile of medullary thymic epithelial-mTEC-cells by in vivo thymus Aire knockdown-BALB-c mice

Project description:Small interfering RNA (siRNA) was used to knockdown Autoimmune regulator(Aire) gene by in vivo electroporation of the thymus of BALB-c mice. In This set of data we include control and Aire-silenced mTEC cells isolated from thymus of BALB-c mice.

2012-08-11 | E-MEXP-3343 | biostudies-arrayexpress

MicroRNA profiling by array of control and Aire-knockdown medullary thymic epithelial cells isolated from thymus of BALB-c mice.

Project description:Small interfering RNA ( siRNA) was used to knockdown Autoimmune regulator (Aire) gene by in vivo electroporation of the thymus of BALB-c mice. In this set of data we include control and Aire-knockdown mTEC cells isolated from thymus of BALB-c mice.

2013-06-28 | E-MEXP-3665 | biostudies-arrayexpress

Transcriptional regulation by Aire in medullary thymic epithelial cells revisited

Project description:The deficiency of Aire, a transcriptional regulator whose defect results in the development of autoimmunity, is associated with reduced expression of tissue-restricted self-Ags (TRAs) in medullary thymic epithelial cells (mTECs). Although the mechanisms underlying Aire-dependent expression of TRAs need to be explored, the physical identification of the target(s) of Aire has been hampered by the low and promiscuous expression of TRAs. We have tackled this issue by engineering mice with augmented Aire expression. Integration of the transcriptomic data from Aire-augmented and Aire-deficient mTECs revealed that a large proportion of so-called Aire-dependent genes, including those of TRAs, may not be direct transcriptional targets downstream of Aire. Rather, Aire induces TRA expression indirectly through controlling the heterogeneity of mTECs, as revealed by single-cell analyses. In contrast, Ccl25 emerged as a canonical target of Aire, and we verified this both in vitro and in vivo. Our approach has illuminated the Aire?s primary targets while distinguishing them from the secondary targets.

2021-10-13 | GSE155331 | GEO

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