Project description:Transcription profiling of fission yeast /yox1/ deletion and genome wide location analysis of Yox1p and Cdc10p transcription factors reveal a negative feedback interaction: /yox1 /is transcriptionally activated by MBF, and Yox1p in turn transcriptionally represses the MBF target genes (including /yox1 /itself).
Project description:In order to identify novel regulators of haematopoietic stem cell emergence in the mouse embryonic AGM region, 3 different types of gene expression comparisons were performed. In the first one, whole dorsal aortas from embryonic day (E) 11 embryos and E9 embryos were dissected, tissues from each time point were pooled and their expression profile compared (whole aorta (WA) E9 vs E11). 3 different pools/biological replicas were obtained for each time point, a dye swap was included in each experiment, a technical replicate was performed for each pool (including the dye swap) and 2 self-self hybridisation controls were included. In the second comparison, GFP+ cells were isolated by fluorescence-activated cell sorting from pooled E9 and E11 aortas from transgenic embryos that express GFP under the regulatory elements of the Ly-6A gene. We obtained one pool for each time point and compared their transcription profile (GFP E9 vs E11). This comparison also contained a dye swap and a technical replicate (also of the dye swap). In the third comparison, the E11 aorta was cut into 3 roughly equal parts. Pools were obtained of the middle region ("m") and the two outer regions, rostral and caudal, together ("r+c") and their expression profile compared (11AO m vs r+c). There was one pool of each population, and the experiment included a dye swap and a technical replicate (also of the dye swap) and one self-self hybridisation control. Isolated RNA was amplified by T7-mediated in vitro transcription, with one round for the first and the third comparison and two rounds for the second comparison (sorted GFP+ cells). Probes were labelled during the reverse transcription step. Within array normalisation was achieved with Lowess Smooth and between arrays with Z scores. ANOVA analysis identified differentially expressed genes and K means clustering was used to segregate them into upregulated and downregulated genes.
Project description:To understand how metabolic and nutritional factors governing adaptation to the host niche contribute to the virulence of Aspergillus fumigatus we compared transcriptomes of developmentally matched A.fumigatus isolates following laboratory culture or initiation of infection in the neutropenic murine lung.
Project description:One major effect of PI3-kinase activation downstream of the serine/threonine kinase Akt is the phosphorylation of the transcription factor FOXO1 and its neutralization. FOXO1 has several ubiquitous targets genes in many cell types that control cell quiescence, oxydative stress or apoptosis. However, it has been demonstrated that FOXO1 also has specific targets depending of the cellular context. The role of FOXO1 to regulate specific genes in T lymphocytes has not been investigated yet. To examine this point, we used the CD4+ leukemia Jurkat T-cell line, in which the PI3K pathway is constitutively turned-on and FOXO1 transcriptional activity strongly repressed. These cells were transduced with lentiviruses coding for a constitutively active form of FOXO1 fused to GFP and having the three AKT phosphorylation sites mutated to alanine (FOXO1-AAA-GFP) to restore its transcriptional activity. GFP-transduced cells were used as a control and the gene activation levels in the two cell populations analyzed 48 hours post-infection.
Project description:Acute kidney Injury (AKI) following open-heart surgery with cardiopulmonary bypass (CPB) is associated with an increased morbidity and mortality. The goal of this study is to explore patterns of gene expression in human AKI following CPB. This prospective observational study was approved by the Institutional Review Board Human Subjects Committee. Design and methods: Peripheral blood RNA samples were collected prospectively from a cohort of 30 patients at Caritas St. Elizabeths Medical Center in Boston, MA, undergoing CPB at time points immediately prior, and two and 24 hours following CPB. Gene expression patterns of four patients who developed AKI (cases) following CPB will be compared with six patients matched for clinical characteristics, who did not develop AKI (controls). AKI was defined by an incremental increase in serum creatinine of 30% within 72 hours following CPB. For peripheral blood RNA sampling, 2.5 ml of venous whole blood was collected into PAXgene Blood RNA Tube collection tubes (PreAnalytiX GmbH, Hombrechtikon, CH). Whole blood RNA was extracted with the PAXgene Blood RNA extraction Kit according to the manufacturers instructions (PAXgene RNA Kit Handbook, 06/2005). All biological samples are stored at 80° C.
Project description:Anaemia negatively affects the prognosis of patients with head and neck squamous cell carcinoma (HNSCC). This study investigates whether anaemia is associated to genetic changes and whether these changes predict the clinical outcome.
Project description:Placental villous tissue from women at term presenting various degrees of villitis of unknown etiology were analysed using Affymetrix microarrays. The severity of inflammation was graded using four levels: 0(no inflammation), 1(mild inflammation - multifocal low grade villitis), 2(moderate inflammation - patchy high grade villitis), and 3(severe inflammation - diffuse high grade villitis).
Project description:Abstract: Testicular cancers in young adult men derive from an inborn precursor lesion, called carcinoma in situ (CIS) of the testis. CIS cells are believed to arise from primordial germ cells or gonocytes that fail to differentiate into the male germ cell lineage, and show a remarkable resemblance to embryonic stem cells (ESC). With this study we aimed at further elucidating the origin of CIS using microarray analysis of genome-wide gene expression. CIS cells only constitute a small fraction of the tissue and therefore, to reduce contaminating RNA from other cell types, we developed a fast (160 sec) staining procedure specific for CIS and fetal germ cells. Highly enriched cell populations were obtained by laser microdissectioning. The expression profiles of CIS cells were compared to microdissected fetal gonocytes, oogonia and cultured ESC with and without genomic amplifications. The cell type most similar to CIS was the gonocytes indicating malignant transformation at this stage. The enriched cell populations allowed us to find unique expression patterns for the developmentally very related cell types. Analyses of the similarities and differences among the cell types may help us understand when and how the malignant transformation to CIS occurs.