Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Expression data from melanoma subpopulations

ABSTRACT: We aimed at identifying lymphangiogenic subpopulations by comparative analysis of single cell clones derived from a melanoma of a single patient. Selected clones were grafted into SCID mice, where they induced lymphangiogenesis and metastasized into sentinel nodes, whereas non-lymphangiogenic clones from the same patient did not metastasize. RNA isolated from primary SCID mouse tumors were used for transcriptome analysis. A total of 16 Samples were analysed, 4 per group. Parental pool MCM1(C), nonlymphangiogenic MCM1G(G), lymphangiogenic MCM1D(D1), lymphangiogenic and very metastatic MCM1DLN(D2).

ORGANISM(S): Homo sapiens

SUBMITTER: Mario Mikula

PROVIDER: E-GEOD-36035 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

ACCESS DATA

Publications

MET expression in melanoma correlates with a lymphangiogenic phenotype.

Swoboda Alexander A Schanab Oliver O Tauber Stefanie S Bilban Martin M Berger Walter W Petzelbauer Peter P Mikula Mario M

Human molecular genetics 20120507 15

Melanomas contain high frequencies of tumorigenic cells and their tumorigenic capacity resides in several distinct subpopulations within melanoma. Since their metastatic potential is linked to their ability to recruit lymphatic vessels, we aimed at identifying lymphangiogenic subpopulations by comparative in vitro analysis of single cell clones derived from a melanoma of a single patient. Selected lymphangiogenic clones were then grafted into severe combined immunodeficient mice, where they indu ...[more]

PMID: 22570180

Similar Datasets

Project description:Tumor cell invasion and metastasis are hallmarks of malignancy. Despite recent advances in the understanding of lymphatic spread, the mechanisms by which tumors metastasize to sentinel/distant lymph nodes and beyond are poorly understood. To gain new insights into this complex process, we established a highly metastatic melanoma cell line (B16F1-variant) by in vivo passaging the B16 parental cell line through the lymphatic system. Here, we characterized morphology, rate of cell proliferation, colony formation, migration, tumorogenicity, lymph flow, and capacities to induce tumor- and sentinel lymph node- lymphangiogenesis. Furthermore, microarray-based comparative analysis bewteen parental and passaged cell lines was performed to identify specific gene expression profiles. The most differentially expressed gene was SPP (osteopontin), a secreted glycophosphoprotein which is known to be involved in cancer metastasis. Overexpression of osteopontin in B16 F1-variant was confirmed by Western blot and quantitative RT-PCR. Treatment of cultured lymphatic endothelial cells (LEC) with osteopontin promoted cell migration mediated by the integrin α9 pathway. Our results identify osteopontin as a novel lymphangiogenic factor. B16 and B16 variants were cultured in DMEM supplemented with 20% fetal bovine serum (FBS; Invitrogen, Grand Island, NY) supplemented with antibiotic-antimycotic solution. Every B16 variant was cultured in duplicates. Total cellular RNA was isolated using the Trizol reagent (Invitrogen, Carlsbad, CA) extracted with chloroform, precipitated with isopropanol, washed with 70% ethanol, and dissolved in DNase-free/RNase-free distilled water. The concentration of RNA was measured using NanoDrop ND-1000 spectrophotometer (Witec AG, Littau, Switzerland) and RNA quality was assessed using 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA). Digoxigenin-UTP–labeled cRNA was generated and amplified from 500ng of total RNA using the NanoAmp RT-IVT Labeling Kit (Applied Biosystems, Foster City, CA) following the manufacturer's protocol, and was hybridized to Applied Biosystems Mouse Genome Survey Microarrays V2.0. Chemiluminescence detection, image acquisition, and analysis were performed using the Chemiluminescence Detection Kit (Applied Biosystems) and the Applied Biosystems 1700 Chemiluminescent Microarray Analyzer following the manufacturer's protocol. A total of two biological replicates were generated for each B16 variant and each cell line data consisted of pooled RNA of two diffferent passage numbers.

Project description:Gene transfer into HSCs by gammaretroviral vectors (RV) is an effective treatment for inherited blood disorders, although potentially limited by the risk of insertional mutagenesis. We evaluated the genomic impact of RV integration in T-lymphocytes from adenosine deaminase (ADA)-Severe combined immunodeficiency (SCID) patients 10 to 30 months after infusion of autologous, genetically-corrected CD34+ cells. Expression profiling on ex vivo T-cell bulk population revealed no difference with respect to healthy controls. To assess the effect of vector integration on gene expression at the single cell level, primary T-cell clones were isolated from two patients. T-cell clones harboured either one or two vector copies per cell and displayed partial to full correction of ADA expression, purine metabolism and TCR-driven functions. Analysis of retroviral integration sites (RIS) indicated a high diversity in T-cell origin, consistent with the polyclonal TCR-Vbeta repertoire. Quantitative transcript analysis of 120 genes within a 200kb-window around RIS showed modest (2.8- to 5.2-fold) disregulation of 5.8% genes in 18.6% of the T-cell clones compared to controls. Nonetheless, affected clones maintained a stable phenotype and normal functions in vitro. These results confirm that RV-mediated gene transfer for ADA-SCID is safe, and provide crucial information for the development of future gene therapy protocols. Global gene expression profiling was performed on CD4+ and CD8+ T-cell subsets purified ex vivo from three ADA-SCID patients at different times after gene therapy. The microarray analysis showed a substantial overlap with the expression patterns of T-cells from controls, indicating the absence of gross abnormalities in the development and function of T-cells derived from genetically corrected hematopoietic stem/progenitor cells. Experiment Overall Design: Transcript profiling was carried out in CD4+ and CD8+ T-cells purified with immunomagnetic beads (Miltenyi Biotec, Germany) from the peripheral blood lymphocytes of ADA-SCID patients (Pt1, 3 and 4) 10-30 months after autologous transplantation with genetically corrected CD34+ cells. At the indicated time points, the percentage of vector-positive T-cells by qPCR was >75%. Transcript profiles were determined using the Affymetrix HG-U133A microarray and compared to those of age-matched healthy controls (n=4; C1, C2, C3, C4).

Dataset Information

Expression data from melanoma subpopulations

Publications

MET expression in melanoma correlates with a lymphangiogenic phenotype.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets