Expression data from subsets of mouse spleen dendritic cells
Ontology highlight
ABSTRACT: Mouse splenic dendritic cells are divided into different subsets based on their phynotype. CD8α+ and CD8α- dendritic cells play different roles against pathogens. Cross-presentation is essential for immune defense against viruses, tumors and intracellular bacteria and CD8α+ DCs are more potent in cross-presentation compared to CD8α- DCs. We used microarrays to detail the global gene expression to analyze the underlying mechanism in CD8α+ dendritic cells cross-presentation Splenic CD8α+ and CD8α- dendritic cells were sorted by flow-cytometer for RNA extraction and hybridization on Affymetrix microarrays. Thirty C57BL/6J mice were sacrified for the splenic cells, splenic DCs were enriched by negtive selection, then CD8α+ and CD8α- dendritic cells were seprated based on CD11c and CD8α markers.
Project description:Despite the significance of nucleus genes in plant growth and development, little is known of the molecular bases of regulation of photosynthesis in woody plants.Hence discovering the genetic basis for photosynthesis related phenotypic variation and identifying the major genes controlling these complex traits in trees is essential. Combining the microarray technique and bulked segregant analysis strategy, we used poplar as a model to detect the nucleus genes differentially expressed in segregation population of photosynthesis traits. We measured the F1 interspecific population and segregated the stable extrem samples into two groups including three pools containing five incividuals.Use the Affymetrix poplar gene chip to decrypt the gene functions and mechanisms in different photosynthetic rate. Leaves were taken from high and low photosynthetic rate individuals for RNA extraction and hybridization on Affymetrix microarrays. H_A, H_B, H_C are from the high photosynthetic rate individuals. L_D,L_E, L_F are from the low photosynthetic rate incividuals.
Project description:Insulin signalling activity is well known to mediate transcriptional changes in response to nutrients and it is changed in the spargel mutant. We performed a genome-wide microarray study to investigate the role of Spargel as a transcriptional coregulator downstream of insulin receptor signalling. Experiment Overall Design: To investigate the effect of Spargel on transcriptional changes mediated by the insulin-signalling pathway we analyzed 12 samples in total: three biological replicates of the control with or without UAS-Inr (Inr_wt and ctrl_wt) and three biological replicates of the spargel mutant with or without UAS-Inr (Inr_KGand ctrl_KG).
Project description:This experiment compares the transciptional changes in antigen specific murine CD8 T cells (P14 T cells) after exposure in vivo to dendritic cells (DC) pulsed with low dose cognate peptide (1uM KAVYNFATC), high dose cognate peptide (100uM KAVYNFATC) or no antigen. Splenic dendritic cells were freshly isolated, peptide pulsed, washed and then adoptively transferred s.c. to the right footpad of C57BL/6 hosts. After 18h, freshly isolated P14 CD8 T cells were labelled with CMFDA and adoptively transferred iv. Two hours after T cell transfer, anti-L selectin antibody was given iv. At 12 and 24 hours, recipients were sacrificed and The right popliteal LN was harvested at 12 or 24h post T cell transfer and a single cell suspension was created and stained with PE CD4, B220 and CD19 (dump channel). Cells were then sorted on a FacsARIA for being non-doublets, CMFDA positive and dump channel negative. This experiment compares the transciptional changes in antigen specific murine CD8 T cells (P14 T cells) after exposure in vivo to dendritic cells (DC) pulsed with low dose cognate peptide (1uM KAVYNFATC), high dose cognate peptide (100uM KAVYNFATC) or no antigen. Splenic dendritic cells were freshly isolated, peptide pulsed, washed and then adoptively transferred s.c. to the right footpad of C57BL/6 hosts. After 18h, freshly isolated P14 CD8 T cells were labelled with CMFDA and adoptively transferred iv. Two hours after T cell transfer, anti-L selectin antibody was given iv. At 12 and 24 hours, recipients were sacrificed and The right popliteal LN was harvested at 12 or 24h post T cell transfer and a single cell suspension was created and stained with PE CD4, B220 and CD19 (dump channel). Cells were then sorted on a FacsARIA for being non-doublets, CMFDA positive and dump channel negative. The experiment was conducted for 39 samples out of which 35 passsed transcriptional quality control tests. The phenotypic distribution for the 35 samples includes: (1) high dose (100uM KAVYNFATC ) cognate peptide pulsed samples harvested at 12h post T cell transfer: 6 biological replicates (2) high dose (100uM KAVYNFATC ) cognate peptide pulsed samples harvested at 24h post T cell transfer: 7 biological replicates (3) low dose (1uM KAVYNFATC) cognate peptide pulsed samples harvested at 12h post T cell transfer: 6 biological replicates (4) low dose (1uM KAVYNFATC) cognate peptide pulsed samples harvested at 24h post T cell transfer: 9 biological replicates (5) no antigen pulsed samples harvested at 12h post T cell transfer: 3 biological replicates (6) no antigen pulsed samples harvested at 24h post T cell transfer: 4 biological replicates.
Project description:Deletion of the gene encoding Foxa2, a winged helix transcription factor selectively expressed in respiratory epithelial cells, caused spontaneous pulmonary eosinophilic inflammation and goblet cell metaplasia. Loss of Foxa2 induced the recruitment and activation of myeloid dendritic cells (mDCs) and Th2 cells in the lung, and was associated with the increased production of T helper 2 (Th2) cytokines and chemokines. mRNA microarray analysis demonstrated that deletion of Foxa2 induced the expression of a number of mRNAs regulating pulmonary dendritic cell activation, Th2 mediated inflammation, and goblet cell differentiation. The spontaneous pulmonary inflammation and goblet cell metaplasia caused by loss of Foxa2 was inhibited by treatment of newborn Foxa2â??/â?? mice with monoclonal IL-4Ralpha antibody. Expression of Foxa2 in non-ciliated secretory cells (Clara cells) in vivo inhibited goblet cell differentiation induced by pulmonary allergen exposure. The respiratory epithelium plays a central role in the regulation of Th2-mediated inflammation and innate immunity in the developing lung in a process regulated by Foxa2. To investigate the role of Foxa2 and its downstream targets associated with the Th2 inflammation and goblet cell hyperplasia, RNAs were isolated from the lungs of Foxa2-/- and control littermates at PN15. Lung cRNA was hybridized to the murine genome MOE430 V2 chips.
Project description:Mouse splenic dendritic cells are divided into different subsets based on their phynotype. CD8α+ and CD8α- dendritic cells play different roles against pathogens. Cross-presentation is essential for immune defense against viruses, tumors and intracellular bacteria and CD8α+ DCs are more potent in cross-presentation compared to CD8α- DCs. We used microarrays to detail the global gene expression to analyze the underlying mechanism in CD8α+ dendritic cells cross-presentation
Project description:Zygotic genome activation (ZGA), which is according to the midblastula transition in zebrafish, is an important event during the maternal-zygotic transition in animals. Our preliminary study and other groupâs works indicate that epigenetic regulations play an essential role in ZGA. Morpholino was employed to knockdown PRMT6. We used microarrays to analyze the global gene expression in prmt6 morphants. prmt6 MO (0.3mM) was injected into the one-two cell zebrafish, prmt6 cMO (0.3mM) injection as a control. At 6 hpf, embryos were classified into three subtypes (normal, mild and severe) and prepared for global gene expression analysis with Affymetrix Zebrafish Genome Arrays. The severe subtype and the control were repeated three times.
Project description:Four days old rice calli were selected to grow 324h under spaceflight controls, 1g-flight controls and 1g-ground controls. We used microarrays to detail the global programme of gene expression underlying cellularisation and identified distinct classes of up-regulated genes during this process. Rice calli were selected at different treatment for RNA extraction and hybridization on Affymetrix microarrays. We sought to obtain the diff-genes that caused by the microgravity.
Project description:Gastric cancer (GC) is one of the most common cancer worldwide. Specific and reliable molecular markers are limited; it is critical to identify new biomarkers for GC to aid in early diagnosis, treatment strategy, and prognosis evaluation. Microarray technology makes it possible to measure the expression levels of thousands of genes, and identifying meaningful and useful molecular targets from these large data. Total RNA was extracted from 10 pairs of GC tissue and adjacent non-tumor mucosa using TRIzol® reagent (Invitrogen, Carlsbad, CA, US) following the manufacturer's instructions. The RNA integrity number (RIN), detected by an Agilent Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA, US), was used to determine the RNA integrity. Total RNA meeting the specified quality criteria (RIN ⥠7.0 and 28S/18S ⥠0.7) were further purified using an RNase-Free DNase Set (Qiagen, Hilden, Germany) and an RNeasy® Micro Kit (Qiagen, Hilden, Germany) according to the manufacturer's instructions. Purified total RNA was then used for obtaining biotin-labeled cRNA by applying the GeneChip® 3'IVT Express Kit (Affymetrix, Santa Clara, CA, US), according to the manufacturer's instructions. Then, the GeneChip® Hybridization, Wash, and Stain Kit (Affymetrix, Santa Clara, CA, US) was used for performing the array hybridization and wash, with the use of a GeneChip® Hybridization Oven 645 (Affymetrix, Santa Clara, CA, US) and a GeneChip® Fluidics Station 450 (Affymetrix, Santa Clara, CA, US). All arrays were scanned using a GeneChip® Scanner 3000 (Affymetrix, Santa Clara, CA, US) and Command Console Software 3.1 (Affymetrix, Santa Clara, CA, US) with the default settings. Raw data were normalized by the MAS 5.0 algorithm in GeneSpring Software 11.0 (Agilent Technologies, Santa Clara, CA, US). The significance analysis of microarray (SAM) was used to identify genes that were differentially expressed between the GC and normal tissues. Genes were considered to be differentially expressed when the Tumor versus Normal signal log ratio values were â¥2 or â¤0.5.
Project description:Chromatin modifying activities for construction of appropriate epigenetic landscapes by polycomb repressive complex 2 (PRC2) play an essential role in development and tumorigenesis. However, the spatiotemporal mechanisms by which PRC2 achieves diverse epigenomes for specific tissue or cellular contexts remain poorly understood. Here, we discovered that LATS2 knockout causes dysregulation of PRC2 and subsequent transcriptome changes for differentiation in both mouse and human cells. LATS2 depletion dependent dysregulation of PRC2 also effects H3K4me3 and forms negative feedback loop for maintenance of PRC2. Further analyses reveal that LATS2 on chromatin binds to EZH2 and LATS2 has ability to phosphorylate PRC2 in vitro. These LATS2 dependent H3K27me3 targets are highly induced during neurogenesis, and statistical analysis of glioblastoma multiforme reveals that LATS2-high cases show more dedifferentiated transcriptome and poor prognosis with silencing of H3K27me3 targets. These observations suggest that LATS2-mediated epigenome coordination is pivotal for development and disease, including cancer. The rabeled cRNAs from Lats2-/- mouse embryonic fibroblasts (MEFs) and litter wild type MEFs were subjected to Agilent Mouse miRNA Microarray 8 x 15K. Each sample was run in biological dupulicate (different PDL).
Project description:The mechanism of egress of mature regulatory T cells (Tregs) from the thymus to the periphery remains enigmatic, as does the nature of those factors expressed in the thymic environment. Here, we examined the fate of thymic Tregs in TNFα/RelA double-knockout (TA-KO) mice, because TA-KO mice retain a Treg population in the thymus but have only a small Treg population at the periphery. Transplantation of whole TA-KO thymus to under the kidney capsule of Rag1 null mice failed to induce the production of donor-derived splenic Tregs expressing neuropilin-1 (Nrp1), which was reported to be a marker of naturally occurring Tregs, indicating that TA-KO thymic Tregs either do not leave the thymus or are lost at the periphery. We next transplanted enriched TA-KO thymic Tregs to the peripheries of TA-KO mice and traced mouse survival. Transplantation of TA-KO thymic Tregs rescued the lethality in TA-KO mice, demonstrating that TA-KO thymic Tregs remain functional at the periphery. The TA-KO thymic Treg population had highly demethylated CpG motifs in the foxp3 locus, indicating that the cells were arrested at a late-mature stage. Also, the population included a large subpopulation of Tregs expressing IL-7Rα, which is a possible marker of late-mature Tregs. Finally, TA-KO fetal liver chimeric mice developed an Nrp1+ splenic Treg population from TA-KO cells, suggesting that Treg arrest is caused by a lack of RelA in the thymic environment. Together, these results suggest that egress of mature Tregs from the thymus depends on RelA in the thymic environment. For the isolation of thymic Tregs, CD4+CD8α-CD25hi thymocytes were isolated from five 1.5- to 2-week-old TNFα-KO or TA-KO mice by using a FACSAria cell sorter. For the isolation of thymic stromal cells, 10 thymi from 1.5- to 2-week-old TNFα-KO or TA-KO mice were minced with scissors and treated with RPMI 1640 supplemented with 2% FCS, 0.2 mg/ml collagenase (Roche, Basel, Switzerland), 0.2 mg/ml dispase I (Roche), and 100 U/ml DNase I (Life Technologies) for 30 min with stirring. Digested thymi were centrifuged in a Percoll (GE Healthcare Bio-Sciences, Piscataway, NJ, USA) gradient (density, 1.115, 1.065, and PBS) at 1400g for 30 min. Cells in the upper layer were collected, and the CD45-EpCAM+ (thymic epithelial cells) and CD45+EpCAM- populations (enriched thymic stromal cells containing macrophages or dendritic cells) were sorted.