In order to define the transcriptional network functionally regulated by Pax8 as well as infer its direct targets, we performed RNAi to knock-down Pax8 gene in FRTL-5 thyroid cells. Expression data from three independent silencing experiments were analyzed by microarray technology unraveling 2815 genes differentially expressed between silenced cells and controls. Of these, 1421 genes were down-regulated and 1394 genes were up-regulated 72hrs after Pax8 silencing.
Identification of novel Pax8 targets in FRTL-5 thyroid cells by gene silencing and expression microarray analysis.
Cell line
View SamplesIn contrast to the migration of leukocytes from blood vessels into tissues, and the involvement of adhesion molecules and chemokines in this process, the migration of leukocytes from the tissue into lymphatic vessels is much less well understood. This can, in part be explained by the fact that murine lymphatic endothelial cells (LECs) have proven particularly hard to isolate and propagate in culture. Hence, it has been difficult to establish suitable models to study this process in vitro. Combining magnetic bead-based purification and fluorescence-activated cell sorting (FACS), we have isolated LECs (immorto-LECs) from the skin of mice which express a temperature-sensitive SV40 large T antigen (H-2Kb-tsA58 mice; ImmortoMice) in all cell types under the control of the MHC-class-I-promotor, H-2Kb. The isolated cells are viable for more than 30 passages when cultured at 33 C, the temperature at which the large T antigen is stably expressed. Furthermore, immorto-LECs tolerate several days of culture at 37 C, but become senescent if continuously cultured at this temperature. All cells stably express endothelial and lymphatic markers like CD31, podoplanin, Prox-1 and VEGFR-3 up to passage 30. When cultured in presence of tumor necrosis factor-alpha (TNF-a), immorto-LECs upregulate adhesion molecules, such as ICAM-1, VCAM-1 and E-selectin, similarly to what has been reported to occur under inflammatory conditions in vivo. Overall, our findings establish immorto-LECs as a useful and handy tool for the in vitro investigation of immune cell transmigration across lymphatic endothelium.
Tissue inflammation modulates gene expression of lymphatic endothelial cells and dendritic cell migration in a stimulus-dependent manner.
Specimen part
View SamplesThe AIL transcription factor BABY BOOM (BBM) is required together with the related PLETHORA proteins for embryo and root meristem development and its expression is sufficient to confer pluripotency and totipotency to somatic tissues. We show that BBM and other AIL proteins interact with multiple members of the L1/epidermal-expressed HD-ZIP class IV / HOMEODOMAIN GLABROUS (HDG) transcription factor family. Ectopic overexpression of HDG1, HDG11 and HDG12 genes induces a reduced growth phenotype, and analysis of HDG1 overexpression lines shows that this growth reduction is due to both root and shoot meristem arrest. To understand how HDG1 controls cell proliferation, as well as its functional relationship with BBM, we performed microarray experiments to identify candidate genes that are directly regulated by HDG1, and compared these to the set of genes that are directly regulated by BBM expression.
AIL and HDG proteins act antagonistically to control cell proliferation.
Specimen part, Treatment
View SamplesTo analyze the gene expression alteration after stroke, we used Middle Cerebral Artery Occlusion model of rats. By comparing with Sham operated rats, we extracted the mRNAs whose expressions are alterated by stroke.
Gene Expression Analysis of the Effect of Ischemic Infarction in Whole Blood.
Sex, Age, Specimen part
View SamplesTranscriptional profiling revealed that murine VH11 and non-VH11 CLL differed in the upregulation of specific pathways implicated in cell signaling and metabolism. We identified a gene expression signature (including Ccdc88a, Clip3, Zcchc18, Chd3 and Itm2a) that was significantly upregulated in T cell-dependent non-VH11 CLL compared with T cell-independent VH11/Vk14 or mutated IgH.TEµ CLL. Overall design: biological replicate (n=3-4) RNA-Seq experiments Please note that the ''countTable_exons_def_norm_rpkm_all.txt'' contains the ''FPKM'' column headers (as a default output setting for the HOMER software package). However, the .txt file contains RPKM value as described in the sample data processing field.
Identification of Distinct Unmutated Chronic Lymphocytic Leukemia Subsets in Mice Based on Their T Cell Dependency.
Specimen part, Cell line, Subject
View SamplesWe have studied the genes activated in human liver transplantation to identify potential target genes for the prevention or treatment of related injuries.
Wide gene expression profiling of ischemia-reperfusion injury in human liver transplantation.
Sex, Age, Specimen part, Subject
View SamplesThe innate immune system is the organisms first line of defense against pathogens. Pattern recognition receptors (PRRs) are responsible for sensing the presence of pathogen-associated molecules. The prototypic PRRs, the membrane-bound receptors of the Toll-like receptor (TLR) family, recognize pathogen-associated molecular patterns (PAMPs) and initiate an innate immune response through signaling pathways that depend on the adaptor molecules MyD88 and TRIF. Deciphering the differences in the complex signaling events that lead to pathogen recognition and initiation of the correct response remains challenging. Here we report the discovery of temporal changes in the protein signaling components involved in innate immunity. Using an integrated strategy combining unbiased proteomics, transcriptomics and macrophage stimulations with three different PAMPs, we identified differences in signaling between individual TLRs and revealed specifics of pathway regulation at the protein level. In addition to forming macrophages and dendritic cells, monocytes in adult peripheral blood retain the ability to develop into osteoclasts, mature bone-resorbing cells. The extensive morphological and functional transformations that occur during osteoclast differentiation require substantial reprogramming of gene and protein expression. Here we employ -omic-scale technologies to examine in detail the molecular changes at discrete developmental stages in this process (precursor cells, intermediate osteoclasts, and multinuclear osteoclasts), quantitatively comparing their transcriptomes and proteomes.
Characterization of functional reprogramming during osteoclast development using quantitative proteomics and mRNA profiling.
Specimen part, Cell line
View SamplesTriggering of B cell receptors (BCR) induces a massive synthesis of NFATc1 in splenic B cells. By inactivating the Nfatc1 gene and re-expressing NFATc1 we show that NFATc1 levels are critical for the survival of splenic B cells upon BCR stimulation. NFATc1 ablation led to decreased BCR-induced Ca++ flux and proliferation of splenic B cells, increased apoptosis and suppressed germinal centre formation and immunoglobulin class switch by T cell-independent antigens. By controlling IL-10 synthesis in B cells, NFATc1 supported the proliferation and IL-2 synthesis of T cells in vitro and appeared to contribute to the mild clinical course of Experimental Autoimmune Encephalomyelitis in mice bearing NFATc1-/- B cells. These data indicate NFATc1 as a key factor controlling B cell function.
NFATc1 affects mouse splenic B cell function by controlling the calcineurin--NFAT signaling network.
Specimen part
View SamplesClassical dendritic cells (DCs) are key players at the interface between innate and adaptive immunity. In the kidney exist 2 major subsets of cDCs: CD11b+ cDCs and CD103+ cDCs. We investigated their function in the most widely used model of experimental glomerulonephritis (GN) in mice: nephrotoxic nephritis (NTN). Consistent with a role for cDCs in nephrotoxic nephritis, depletion of ZBTB46+ cells (all cDCs) attenuated kidney injury, while deficiency of the CD103+ subset of cDCs accelerated injury via a mechanism that involved increased neutrophils. This RNAseq was performed to analyze transcriptional changes in FACS-sorted renal CD11b+ and CD103+ cDCs under healthy conditions and at day 7 of NTN to reveal why both subsets have different functions in GN. Overall design: The study was performed with total of 6 mice (wildtype, male, age 8-12 weeks). 3 mice were sacrificed in the healthy situation, 3 mice were sacrificed 7 days after injection of the nephrotoxic nephritis antiserum (NTN). From each mouse CD11b+ and CD103+ DCs were sorted, resulting in 4 experimental conditions with 3 biological replicates each: CD103_healthy, CD11b_healthy, CD103_NTN, CD11b_NTN.
Opposing Roles of Dendritic Cell Subsets in Experimental GN.
Sex, Specimen part, Treatment, Subject
View SamplesPrimary pneumocytes from KRas;Atg5fl/+ and KRas;Atg5fl/fl littermates were cultured for 48 hours and infected with AdCre-GFP to induce expression of the KrasG12D oncogene and concomitant Atg5 deletion. The transcriptional profile of those cells was determined by mRNA sequencing and uncovered differential expression in cellular movement, inflammatory response and oxidative stress response. Overall design: Comparison of transcriptomes from KRas;Atg5fl/+ and KRas;Atg5fl/fl pneumocytes
A dual role for autophagy in a murine model of lung cancer.
Specimen part, Subject
View Samples