Lung cancer is a leading cause of deaths in the world. There is a need to improve an understanding of mechanisms of malignant transformation and to develop genetic markers of disease for better and targeted therapies.
Cancer genomics identifies regulatory gene networks associated with the transition from dysplasia to advanced lung adenocarcinomas induced by c-Raf-1.
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View SamplesBackground: Information on the carcinogenic potential of chemicals is only availably for High Production Volume products. There is however, a pressing need for alternative methods allowing for the chronic toxicity of substances, including carcinogenicity, to be detected earlier and more reliably. Here we applied advanced genomics to a cellular transformation assay to identify gene signatures useful for the prediction of risk for carcinogenicity. Methods: Genome wide gene expression analysis and qRT-PCR were applied to untransformed and transformed Balb/c 3T3 cells that exposed to 2, 4-diaminotoluene (DAT), benzo(a)pyrene (BaP), 2-Acetylaminoflourene (AAF) and 3-methycholanthrene (MCA) for 24h and 120h, at different concentrations, respectively. Furthermore, various bioinformatics tools were used to identify gene signatures predicting for the carcinogenic risk. Results: Bioinformatics analysis revealed distinct datasets for the individual chemicals tested while the number of significantly regulated genes increased with ascending treatment concentration of the cell cultures. Filtering of the data revealed a common gene signature that comprised of 13 genes whose regulation in cancer tissue has already been established. Strikingly, this gene signature was already identified prior to cell transformation therefore confirming the predictive power of this gene signature in identifying carcinogenic risks of chemicals. Comparison of fold changes determined by microarray analysis and qRT-PCR were in good agreement. Conclusion: Our data describes selective and commonly regulated carcinogenic pathways observed in an easy to use in vitro carcinogenicity assay. Here we defined a set of genes which can serve as a simply assay to predict the risk for carcinogenicity by use of an alternative in vitro testing strategy.
Toxicogenomics applied to in vitro carcinogenicity testing with Balb/c 3T3 cells revealed a gene signature predictive of chemical carcinogens.
Cell line, Treatment, Time
View SamplesRANK-positive and RANK-negative luminal progenitor cells were isolated by FACS from histologically normal human breast tissue from wild-type human donors. RNA-seq gene expression profiling was used to find differentially expressed genes between the RANK-positive and RANK-negative cell populations. Overall design: Cells were isolated from 4 human patients. A paired analysis was used to compare RANK-positive and RANK-negative cells within patients.
RANK ligand as a potential target for breast cancer prevention in BRCA1-mutation carriers.
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Alternative generation of CNS neural stem cells and PNS derivatives from neural crest-derived peripheral stem cells.
Specimen part
View SamplesNeural crest-derived neural stem cells (NCSCs) from the embryonic PNS can be reprogrammed in neurosphere culture (NS) to rNCSCs that produce CNS progeny, including myelinating oligodendrocytes. Using global gene expression analysis we now demonstrate that rNCSCs completely lose their previous PNS characteristics and acquire the identity of neural stem cells derived from embryonic spinal cord (SCSCs). Reprogramming proceeds rapidly and results in a homogenous population of Olig2-, Sox3- and Lex-positive CNS stem cells. Low-level expression of pluripotency inducing genes Oct4, Nanog and Klf4 argues against a transient pluripotent state during reprogramming. The acquisition of CNS properties is prevented in the presence of BMP4 (BMP NCSCs) as shown by marker gene expression and the potential to produce PNS neurons and glia. In addition, genes characteristic for mesenchymal and perivascular progenitors are expressed, which suggests that BMP NCSCs are directed towards a pericyte progenitor/mesenchymal stem cell (MSC) fate. Adult NCSCs from mouse palate, an easily accessible source of adult NCSCs, display strikingly similar properties. They do not generate cells with CNS characteristics but lose the neural crest markers Sox10 and p75 and produce MSCs. These findings show that embryonic NCSCs acquire a full CNS identity in neurosphere culture. In contrast, MSCs are generated from adult pNCSCs and BMP NCSCs, which reveals that postmigratory NCSCs are a source for MSCs up to the adult stage.
Alternative generation of CNS neural stem cells and PNS derivatives from neural crest-derived peripheral stem cells.
Specimen part
View SamplesNeural crest-derived neural stem cells (NCSCs) from the embryonic PNS can be reprogrammed in neurosphere culture (NS) to rNCSCs that produce CNS progeny, including myelinating oligodendrocytes. Using global gene expression analysis we now demonstrate that rNCSCs completely lose their previous PNS characteristics and acquire the identity of neural stem cells derived from embryonic spinal cord (SCSCs). Reprogramming proceeds rapidly and results in a homogenous population of Olig2-, Sox3- and Lex-positive CNS stem cells. Low-level expression of pluripotency inducing genes Oct4, Nanog and Klf4 argues against a transient pluripotent state during reprogramming. The acquisition of CNS properties is prevented in the presence of BMP4 (BMP NCSCs) as shown by marker gene expression and the potential to produce PNS neurons and glia. In addition, genes characteristic for mesenchymal and perivascular progenitors are expressed, which suggests that BMP NCSCs are directed towards a pericyte progenitor/mesenchymal stem cell (MSC) fate. Adult NCSCs from mouse palate, an easily accessible source of adult NCSCs, display strikingly similar properties. They do not generate cells with CNS characteristics but lose the neural crest markers Sox10 and p75 and produce MSCs. These findings show that embryonic NCSCs acquire a full CNS identity in neurosphere culture. In contrast, MSCs are generated from adult pNCSCs and BMP NCSCs, which reveals that postmigratory NCSCs are a source for MSCs up to the adult stage.
Alternative generation of CNS neural stem cells and PNS derivatives from neural crest-derived peripheral stem cells.
Specimen part
View SamplesPurpose: Analyze the function of the transcription factors Hand2 and Gata3 in adult sympathetic neurons by induced knockout and RNAseq analysis Overall design: Method and Result: Hand2flx/del::DbhCreERT2 (referred to as mutant) and Hand2flx/+::DbhCreERT2 (referred to a control) were injected for 10 days with tamoxifen to activate Cre. Animals were killed, sympathetic ganglia (SCG+Stellate) collected and processed for RNA isolation and RNAseq (Stanzel et al., 2016). Gata3flx/flx::DbhCerERT2 (mutant) and Gata3flx/+::DbhCreERT2 (controls) were treated as Hand2 animals. 16 ganglia from 4 mice were pooled for Hand2 mutant and controls and Gata3 controls. As only rudimentary ganglia were present in Gata3 mutant mice (Tsarovina et al., 2010) ganglia from 8 mice were pooled. The specific effects of the Hand2 knockout are decribed in Stanzel et al., 2016. The analysis of ganglion rudiments in the Gata3 knockout revealed that Gata3 was not reduced, indicating that the remaining cells had escaped the knockout.
Distinct roles of hand2 in developing and adult autonomic neurons.
Specimen part, Subject
View SamplesExtracorporeal shockwave treatment was shown to improve orthopaedic diseases, wound healing and to stimulate lymphangiogenesis in vivo. The aim of this study was to investigate in vitro shockwave treatment (IVSWT) effects on lymphatic endothelial cell (LEC) behavior and lymphangiogenesis. We analyzed migration, proliferation, vascular tube forming capability and marker expression changes of LECs after IVSWT compared with HUVECs. Finally, transcriptome- and miRNA analyses were conducted to gain deeper insight into the IVSWT-induced molecular mechanisms in LECs. The results indicate that IVSWT-mediated proliferation changes of LECs are highly energy flux density-dependent and LEC 2D as well as 3D migration was enhanced through IVSWT. IVSWT suppressed HUVEC 3D migration but enhanced vasculogenesis. Furthermore, we identified podoplaninhigh and podoplaninlow cell subpopulations, whose ratios changed upon IVSWT treatment. Transcriptome- and miRNA analyses on these populations showed differences in genes specific for signaling and vascular tissue. Our findings help to understand the cellular and molecular mechanisms underlying shockwave-induced lymphangiogenesis in vivo.
Molecular and cellular effects of in vitro shockwave treatment on lymphatic endothelial cells.
Specimen part
View SamplesNeuroblastoma is an embryonal neoplasm that remains of dramatic prognosis in its aggressive forms. Activating mutations of the ALK tyrosine kinase receptor have been identified in sporadic and familial cases of this cancer. We generated knock-in mice carrying the two most frequent Alk mutations observed in neuroblastoma patients. We used microarrays to detail the global programme of gene expression underlying the impact of ALK mutations on neuroblastoma formation in a MYCN amplified background.
Activated Alk triggers prolonged neurogenesis and Ret upregulation providing a therapeutic target in ALK-mutated neuroblastoma.
Specimen part
View SamplesDifferent human mTEC subsets (MUC1, CEACAM5 and SGLT1) were purified by sequential enzymatic digestion (collagenase/dispase, trypsin) followed by enrichment using magnetic beads (CD45 beads, Miltenyi Biotech) and FACS sorting. Cells of the surface phenotype CD45-, CDR2-, EpCAM+ were further subdivided into MUC1+/MUC1-, CEACAM5+/CEACAM5- and SGLT1+/SGLT1- fractions. RNA was isolated using MACS SuperAmp protocol (Miltenyi Biotec) and hybridized to Illumina Whole-Genome Expression Beadchips. Gene expression of Antigen-positive and Antigen-negative mTEC subsets was compared.
Overlapping gene coexpression patterns in human medullary thymic epithelial cells generate self-antigen diversity.
Specimen part
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