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MicroRNA expression profiles in bone marrow fractions from gastric cancer patients

ABSTRACT: Both disseminated tumor cells and noncancerous host cells contributed to cancer progression cooperatively in bone marrow. Bone marrow samples were obtained from 4 gastric cancer patients, and were separated into 3 fractions (CD45 positive, CD45 negative/EpCAM positive, and CD14 positive fractions) by the automagnetic-activated cell separation (AutoMACS) system using CD45, EpCAM, and CD14 microbeads (Miltenyi Biotec, Germany). microRNA expression profiles in each fractions were evaluated in order to identify candidate prognostic markers for gastric cancer patients.

ORGANISM(S): Merkel cell polyomavirus Mus musculus Rattus norvegicus Human gammaherpesvirus 8 JC polyomavirus Betapolyomavirus macacae Human immunodeficiency virus 1 Murid gammaherpesvirus 4 Homo sapiens Human betaherpesvirus 5 Human alphaherpesvirus 2 Betapolyomavirus hominis Human alphaherpesvirus 1 human gammaherpesvirus 4 Mus musculus cytomegalovirus 2 Murid betaherpesvirus 1

PROVIDER: GSE40325 | GEO | 2013/08/23

SECONDARY ACCESSION(S): PRJNA173541

REPOSITORIES: GEO

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microRNA expression profiles in bone marrow fractions from gastric cancer patients

Project description:Both disseminated tumor cells and noncancerous host cells contributed to cancer progression cooperatively in bone marrow. Bone marrow samples were obtained from 4 gastric cancer patients, and were separated into 3 fractions (CD45 positive, CD45 negative/EpCAM positive, and CD14 positive fractions) by the automagnetic-activated cell separation (AutoMACS) system using CD45, EpCAM, and CD14 microbeads (Miltenyi Biotec, Germany). microRNA expression profiles in each fractions were evaluated in order to identify candidate prognostic markers for gastric cancer patients. In 4 patients with gastric cancer, bone marrow samples (40 mL) were obtained from iliac bones. Nucleated cells were collected by gradient centrifugation using Ficoll-Paque PREMIUM (GE Healthcare Life Science, USA) and Leucosep (Greiner Bio-One, Germany) according to the manufacturer’s instructions. Next, we separated bone marrow cells into 3 fractions using MACS: CD45 positive (CD45+), CD45 negative/EpCAM positive (CD45-/EpCAM+), and CD14 positive (CD14+). microRNA expression levels of whole bone marrow cells and each fractions were measured by the miRCURY™ LNA array microarray (6th gen-hsa, mmu & rno#208402, Exiqon, Vedbaek, Denmark). The miRCURY™ LNA array microarray slides were scanned using the Agilent G2505C Microarray Scanner System (Agilent Technologies, Inc., USA) and the data analysis was carried out using the Feature Extraction 10.7.3.1 (Agilent Technologies, Inc., USA).

2013-08-23 | E-GEOD-40325 | biostudies-arrayexpress

MicroRNA expression signatures of CD14-CD45-EpCAM+ cells in human bone marrow for colorectal cancer liver metastasis

Project description:To further development of our gene expression approach to biodosimetry, we have employed microRNA microarray expression profiling to identify genes with the potential to distinguish liver metastasis related microRNA. Colorectal cancer patients were administered anesthesia and 20 mL BM was taken from the right and left anterior iliac crests before surgery. Mononucleated cells were collected using a standard Ficoll-Hypaque gradient technique. To enrich for EpCAM+ cells, CD14+ cells were removed from the whole bone marrow using auto MACSTM pro (Milteny Biotec, Bergisch Gladbach, Germany) with anti-CD14 immunomagnetic beads (clone; TÜK4, Milteny Biotec). Next, CD45+ cells were removed by treatment with anti-CD45 immunomagnetic beads (clone; 5B1; Milteny Biotec). The residual CD14−CD45− cells were then incubated with FcR blocking reagent (Milteny Biotec), followed by incubation with anti-EpCAM immunomagnetic beads (clone; HEA-125, Milteny Biotec), and the CD14−CD45−EpCAM+ cells were taken up. Total RNA of these cells we analyzed the microRNA levels of CD14−CD45−EpCAM+ cells obtained from non-metastasis patients (n = 12) and liver metastasis patients (n = 7). Ten-microRNA consensus signature was identified that distinguished between CD14−CD45−EpCAM+ cells from liver metastasis patients and CD14−CD45−EpCAM+ cells from non-liver metastasis patients.

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Project description:Parallel DNA and RNA profiling of EpCAM-positive cells in bone marrow and primary tumor tissue with positive disseminated tumor cell (DTC) count via immunomagnetic Enrichment/Flow Cytometry (IE/FC) of metastatic breast cancer (MBC) patients confirm their malignant nature We developed a novel approach to isolate tumor cells with high purity from bone marrow which was subjected to immunomagnetic enrichment using EpCAM beads followed by fluorescence activated cell sorting (IE/FACS) to isolate EpCAM-positive cells away from leukocytes (CD45+). For DNA profiling, sorted cells were subjected to BAC array comparative genomic hybridization analysis following whole genome amplification. For RNA profiling, QPCR analysis was performed on sixty four (64) cancer-related genes using Taqman® low density arrays. For non-tumor controls, RNA profiling was performed on matched leukocytes (CD45+) isolated from the same enriched bone marrow samples.

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Frequent derepression of the mesenchymal transcription factor gene in acute myeloid leukemia (human)

Project description:Bone marrow samples from normal adult male donors were collected into EDTA. Red cells were removed by ammonium chloride lysis. Leukocytes were washed in SM buffer and CD34+ cells were separated from CD34- cells using an AutoMACS device and anti-CD34 immunomagnetic beads (Miltenyi Biotec), according to manufacturer’s instructions. For mature cell populations, CD34- cells were FACS purified according to the following immunophenotypes, with 7-AAD used to exclude dead cells: Neutrophils: side scatter high CD15+ CD16+. Monocytes: side scatter low-intermediate CD14+ CD16- CD15-. See also Huang et al., 2014.

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