Analysis of global gene expression of a human T-ALL cell lines, comparing GATA3low ETP-ALL (i.e. PER-117) and "typical" T-ALL
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ABSTRACT: Cell lines representing human T-ALL were analyzed to compare GATA3low ETP-ALL (i.e. PER-117) with "typical" T-ALL. Moreover, changes in global gene expression were assessed comparing GATA3low ETP-ALL (i.e. PER-117) and GATA3high ETP-ALL (i.e. Loucy) upon treatment with Decitabine, a hypomethylating agent. Analysis of 6 human cell lines representing "typical" T-ALL (BE13, Jurkat, Molt4, RPM18402) and ETP-ALL (Loucy, PER-117). Additionally, global gene expression was assessed before and after treatment of ETP-ALL cell lines with Decitabine
Project description:Cell lines representing human T-ALL were analyzed to compare GATA3low ETP-ALL (i.e. PER-117) with "typical" T-ALL. Moreover, changes in global gene expression were assessed comparing GATA3low ETP-ALL (i.e. PER-117) and GATA3high ETP-ALL (i.e. Loucy) upon treatment with Decitabine, a hypomethylating agent.
Project description:Two mice in which luciferase-positive LOUCY ETP-ALL cells were xenografted were treated with either vehicle or ABT-199 (50 mg/kg) for 11 days. Afterwards, single cells suspensions were made from bone marrow and spleen of both mice for 10x Genomics scRNA-seq. Following quality control, a total of 13,681 spleen cells and 11,442 bone marrow cells of the control mouse and 15,955 spleen cells and 9,443 bone marrow cells of the ABT 199 treated mouse were processed for further analysis. Our data show that ABT-199 triggers transcriptional changes in T-cell differentiation genes in leukemic cells obtained from the spleen microenvironment. These results are in line with our previous findings that mature typical T-ALL are resistant to ABT-199.
Project description:Tristetraprolin (TTP) binds to specific AU-rich elements in the 3'UTR of certain transcripts and regulates post-transcriptional gene expression by increasing the rate of mRNA turnover. In this study, we evaluated the effects of TTP deficiency on the overall gene expression of spleen tissue, in order to discover tissue specific targets of TTP under normal physiologic conditions. We utilized "Triple KO" (Zfp36-/-/TNFR1-/-/TNFR2-/-) mice that are deficient in TTP and two TNF receptors and compared the transcriptomic changes to "Double KO" (TNFR1-/-/TNFR2-/-) and WT mice. Spleen mRNA from four WT, four "Double KO", and four "Triple mice" was subjected to RNA-Seq in two phases. All the animals used in this study were males between the ages of 12-14 weeks and were on a mixed (75% C57BL/6NTac, 25% 129/SVEV) background. Examination of splenic gene expression difference between "Triple KO"-WT and "double KO"-WT data sets
Project description:Decitabine have a known DNA demethylating activity but also cause cytotoxicity through DNA damage. The two differing mechanisms of action confound studies investigating the effect of DNA demethylation in cancer treatment. The novel DNA methyltransferase 1 specific inhibitor GSK-3685032 causes loss of DNA methylation without DNA damage and offers the promising possibility to examine the molecular consequences of global loss of DNA methylation. To examine the transcriptional effect of global loss of methylation we performed total RNA sequencing of LOUCY and SUP-T1 cells treated with 10 nM Decitabine for 3 days or 300 nM of GSK3685032 for 3 and 7 days.
Project description:Decitabine (DEC) has a known DNA demethylating activity but also cause cytotoxicity through DNA damage. The two differing mechanisms of action confound studies investigating the effect of DNA demethylation in cancer treatment. The novel DNA methyltransferase 1 specific inhibitor GSK-3685032 causes loss of DNA methylation without DNA damage and offers the possibility to examine the molecular consequences of global loss of DNA methylation. EM-seq was used to evaluate the DNA demethylating effects of DEC and GSK-3685032 in LOUCY and SUP-T1 cells treated with 10 nM Decitabine for 3 days or 300 nM of GSK-3685032 for 3 and 7 days.
Project description:In molecular biology, the design of mechanistic experiments has to be optimized by considering statistical and biological principles. In contrast to statistical principles, biological principles of experimental design are not universally formulated. In an attempt to pinpoint generally acceptable rules, we investigated the importance of determining the optimal ranges of scale of i.e. dose and time in gene expression experiments. We propose a protocol for executing small scale, genome wide, range finding studies, covering a wide range of the potentially relevant part of the design space to find the optimal ranges of experimentation. This protocol is executed and a proof-of-concept is presented, where this approach is tested for both an in-vitro and an in-vivo study that aim to unravel DNA repair mechanisms provoked after UV radiation. We identified four challenges of range finding studies in omics experimentation; (1) the modularity of biological processes, (2) their dynamics, (3) the extent to which end-points indicate biological processes, and (4) the costs associated with the assays, which are all addressed by our approach. 48 MEF samples having various combinations of 9 timepoints and 6 UV-C Doses without replication were used
Project description:Oral Cavity Cancer Comparative analysis of gene expression using Affymetrix U133 plus 2.0 microarrays to compare differences between oral cancer samples (most HPV-) and adjacent "normal" mucosa. We have 24 matched pairs of tumor and normal from the same patient
Project description:In molecular biology, the design of mechanistic experiments has to be optimized by considering statistical and biological principles. In contrast to statistical principles, biological principles of experimental design are not universally formulated. In an attempt to pinpoint generally acceptable rules, we investigated the importance of determining the optimal ranges of scale of i.e. dose and time in gene expression experiments. We propose a protocol for executing small scale, genome wide, range finding studies, covering a wide range of the potentially relevant part of the design space to find the optimal ranges of experimentation. This protocol is executed and a proof-of-concept is presented, where this approach is tested for both an in-vitro and an in-vivo study that aim to unravel DNA repair mechanisms provoked after UV radiation. We identified four challenges of range finding studies in omics experimentation; (1) the modularity of biological processes, (2) their dynamics, (3) the extent to which end-points indicate biological processes, and (4) the costs associated with the assays, which are all addressed by our approach. 57 skin biopt samples taken from 12 individual mice on 8 timepionts and for 6 different UV-B doses. Per mouse 5 skin biopts were samples in time
Project description:We examined the transcriptomes of murine "expandable hemangioblasts" (eHBs) and their blood and endothelial progeny, comparing them to the transcriptomes of murine embryonic stem (ES) cells, primary murine endothelial cells isolated from E11.5 yolk sacs or embryos, and E14.5 fetal liver hematopoietic stem cells. Total RNAs were purified from lysates of cultured or primary cells, reverse transcribed, and sequenced on an Illumina HiSeq 2500.