RNA-seq of mouse alveolar macrophages sorted from CD11cCre/+ Tgfbr2fl/fl and control Tgfbr2fl/fl neonatal mice
Ontology highlight
ABSTRACT: Alveolar macrophages (AMs) are lung resident phagocytes. They derive from fetal liver monocytes, which colonize the lung during embryonic development and give rise to fully mature AMs perinatally. We have identified TGF- signaling as an indispensible regulator during this process. To analyze the impact of TGF- on the entire transcriptome of AMs, we performed RNA-seq on AMs deficient of Tgfbr2 in CD11cCre/+ Tgfbr2fl/fl mice at P3 with Tgfbr2fl/fl littermates as a control.
Project description:Organoids derived from small intestine epithelial cells of C57BL/6 wild-type mice were cultured in the presence or absence of IL-28 (1000 ng/ml) for 24 hours. The experimental setup included 8 samples in a pairwise design (4 unstimulated vs 4 stimulated). Profiling of whole-genome transcriptomic patterns was performed by RNA-Seq on an Illumina HiSeq 2500 platform using total RNA at the Next Generation Sequencing core unit of the University Hospital Erlangen, Germany. Demultiplexed reads were quality filtered against rRNAs, tRNAs, mt-rRNAs and mt-tRNAs. Alignment against the mus musculus reference genome (Ensemble V.84 for GRCm38) was performed with RNA-seq aligner STAR (V.2.5.1b) and statistical analysis of unique mappings (HTseq count) was computed in R using the DESeq2 1.10.1.
Project description:The epigenetic dysregulation of tumor suppressor genes is a major driver of human carcinogenesis. We have combined genome-wide methylation analyses with functional screening to identify novel candidate tumor suppressor genes in diffuse large B-cell lymphoma (DLBCL). We find that the dual-specificity phosphatase DUSP4 is aberrantly silenced in nodal and extranodal DLBCL due to promoter hypermethylation; ectopic expression of wild type DUSP4, but not of a phosphatase-deficient mutant, dephosphorylates c-JUN N-terminal kinase (JNK) and induces apoptosis in DLBCL cells. JNK inhibition prevents DLBCL survival in vitro and in vivo, and synergizes strongly with inhibitors of chronic active B-cell receptor signaling. Our results provide a mechanistic basis for the clinical development of JNK inhibitors in DLBCL, alone or in synthetic lethal combinations. A methylation profiling data set related to this experiment was also deposited at ArrayExpress under accession number E-MTAB-2926: http://www.ebi.ac.uk/arrayexpress/experiments/E-MTAB-2926/
Project description:Background: Inducing apoptosis of autoreactive lymphocytes is part of the therapeutic strategy for Crohn’s disease (CD) patients. Failure to respond to medical therapies upon inflammatory bowel disease could result from insufficient apoptosis. To date there are no useful molecular factors for the prediction of clinical relapse. Study Aims: Characterization of the BCL-2 family-related risk of therapy resistance and verifying its usefulness as parameter for the prediction of clinical relapse could be helpful in deciding which medical therapy to recommend. The project is directly targeted to develop rapidly therapeutic improvement for patients with IBD. A long-term goal is the development of new therapeutic options for the treatment of IBD via a physiologic homeostasis and turnover of lymphocytes. Study Design: We propose to characterize the BCL-2 family-related risk of therapy resistance and its usefulness as parameter for the prediction of clinical relapse upon medical therapy. Samples will be used for next generation sequencing, qPCR, WB, immunohistochemistry and immunofluorescence. The hypothesis is considered confirmed if expression of BCL-2 family members in human peripheral blood is significantly changed between patient groups and correlates with the number of lymphocyte subpopulations.
Project description:The aim of the study was to investigate which genes are up- and down-regulated in response to hyperthermia and combined thermoradiotherapy in HSP70 proficient and deficient canine osteosarcoma cell line. Canine osteosarcoma cell line Abrams was transfected with negative control siRNA or siRNA targeting HSP70. Cells were treated with hyperthermia (42C, 1 hour, HT), radiotherapy (6Gy, RT) and thermoradiotherapy (HTRT). RNA was extracted 24 hours after treatment and used for RNA sequencing. Three independent experiments were performed, each with negative control siRNA (Neg) and HSP70 knockdown (HSP70kd) cells. Cells were treated with radiotherapy (RT), hyperthermia (HT), thermoradiotherapy (HTRT) or untreated (ctrl). One experiment consists of 8 samples (ctrl, RT, HT, HTRT in neg and HSP70kd cells), total 24 samples were sequenced.
Project description:Mouse embryonic stem cells (mESCs), derived from pre-implantation blastocyst cells, can be maintained in vitro in defined N2B27 medium supplemented with two chemical inhibitors for GSK3 and MEK (2i) and the cytokine leukemia inhibitory factor (LIF), which act synergistically to promote self-renewal and pluripotency. Many efforts have been devoted to identify genes that promote exit from the pluripotent state and the transition to a primed state of differentiation. One of the first identified players in this process was the Wnt/b-catenin effector TCF7L1 (previously referred to as TCF3), belonging to the family of four TCF/LEF transcription factors, which acts as pro-differentiation factor by repressing pluripotency genes. Of note, there is little evidence that the genetic abrogation of the mechanisms required for the exit from the pluripotent state is sufficient to enable self-renewal in the absence of 2iL. Here, we found that complete loss-of-function of Tcf7, Lef1, Tcf7l1 and Tcf7l2, the genes encoding for the four TCF/LEF transcription factors, (refered to as qKO) allows mESCs to become fully 2iL-independent and to propagate in basal N2B27. To understand the genetic program that allows qKO cells to achieve 2iL-independent self-renewal, we performed RNA sequencing (RNA-seq) of qKO and wild type mESCs.
Project description:Tendon fascicles were extracted from tails of freshly euthanized mice and cultured for 6 days ex vivo in serum containing medium (10%) at either 3% oxygen and 29 degrees celsius or 21% oxygen and 37 degrees celsius.
Project description:Tendon fascicles were analysed directly after isolation from the tail of freshly euthanized mice and after of after ex vivo culture for 6 days. Fascicles were cultured in either serum containing medium (10%) or in serum-free medium at 21% oxygen and 37 degrees celsius.