MiRNA expression profiles of TIE2 expressing Monocytes (TEMs)
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE68322 | GEO | 2017/05/19
SECONDARY ACCESSION(S): PRJNA282423
REPOSITORIES: GEO
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Project description:To identify the gene expressing profiles of TIE2 expressing Monocytes(TEMs), we have employed the Agilent lncRNA Gene Expression 4×180K(Design ID:042818) microarray. Human peripheral blood mononuclear cells (PBMCs) in venous blood from healthy donors were isolated by Lymphoprep (Axis-Shield, Norway). Human monocytes in PBMCs, identified as cells that expressed CD14, were enriched by positive immunomagnetic selection using anti-CD14 MicroBeads (Miltenyi, Germany). TEMs (TIE2+CD14+) and TIE2-Monocytes (Tie2-CD14+) were then isolated by FACS-sorting (Aria II, BD Biosciences) using FITC-conjugated anti-CD14 (BD Biosciences, USA) and APC-conjugated anti-TIE2 (R&D System, USA) antibodies.Three TEMs samples together with their paried TIE2-Monocytes were detected.Expressions of sixteen genes (CDKN1A, FDXR, SESN1, BBC3 and PHPT1) from this signature was quantified in the same RNA samples by real-time PCR, confirming low variability between donors as well as the predicted radiation response pattern.
2017-05-17 | GSE57783 | GEO
Project description:To identify the gene expressing profiles of TIE2 expressing Monocytes(TEMs), we have employed the Agilent lncRNA Gene Expression 4Ã180K(Design ID:042818) microarray. Human peripheral blood mononuclear cells (PBMCs) in venous blood from healthy donors were isolated by Lymphoprep (Axis-Shield, Norway). Human monocytes in PBMCs, identified as cells that expressed CD14, were enriched by positive immunomagnetic selection using anti-CD14 MicroBeads (Miltenyi, Germany). TEMs (TIE2+CD14+) and TIE2-Monocytes (Tie2-CD14+) were then isolated by FACS-sorting (Aria II, BD Biosciences) using FITC-conjugated anti-CD14 (BD Biosciences, USA) and APC-conjugated anti-TIE2 (R&D System, USA) antibodies.Three TEMs samples together with their paried TIE2-Monocytes were detected.Expressions of sixteen genes (CDKN1A, FDXR, SESN1, BBC3 and PHPT1) from this signature was quantified in the same RNA samples by real-time PCR, confirming low variability between donors as well as the predicted radiation response pattern. The gene expressions of three independent paried TEMs and TIE2- Monocytes samples from different donors were measured.
2017-05-17 | E-GEOD-57783 | biostudies-arrayexpress
Project description:Th1/Th17-type T-cell responses are upregulated in Behcet’s disease (BD). However, signaling pathways associated with this aberrant immune response are not clarified. Whole-genome microarray profiling was performed with human U133 (Plus 2.0) chips using mRNA of isolated CD14+ monocytes and CD4+ T-cells from PBMC in patients with BD (n=9) and healthy controls (HC) (n=9). Flow cytometric analysis of unstimulated (US) and stimulated (PHA) STAT3 and pSTAT3 expressions of PBMCs were also analysed (BD and HC, both n=26). JAK1 was observed to be upregulated in both CD14+ monocytes (1.94 fold) and CD4+ T-lymphocytes (1.40 fold) of BD patients. Using canonical pathway enrichment analysis, JAK/STAT signaling was identified as activated in both CD14+ monocytes (p=2.95E-06) and in CD4+ lymphocytes (p=8.13E-04) in BD. Interferon (p=1.02E-07) and IL-6 (p=8.91E-03) signaling pathways were also prominent in CD14+ monocytes. Basal unstimulated total STAT3 expression was significantly higher in BD (1.2 vs 3.45, p<0.05). The JAK1/STAT3 signaling pathway is activated in BD, possibly through the activation of Th1/Th17-type cytokines such as IL-2, IFNγ, IL-6, IL-17 and IL-23.
2014-12-31 | GSE61399 | GEO
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Project description:Transcriptional profile of control and VEGF overexpressing FACS-isolated CD34+ Cancer stem cells from DMBA/TPA induced skin tumours Tumours were digested in collagenase I (Sigma) for 2 hours at 37°C on a rocking plate. Collagenase I activity was blocked by addition of EDTA (5mM) and then rinced in PBS supplemented with 2%FCS. After tumour digestion, cells were first incubated in PBS complemented with 30% FCS to block Fc receptors for 15 min at room temperature. Immunostaining was performed using biotin-conjugated anti-CD34 (clone RAM34; BD Pharmingen), FITC-conjugated anti-α6-integrin (clone GoH3; BD Pharmingen), PE-conjugated anti-CD45 (clone 30F11, eBiosciences), PE-conjugated anti CD31 (clone MEC13.3; BD Pharmingen), PE-conjugated anti-CD140a (clone APA5; eBiosciences), APC-Cy7-conjugated anti-Epcam (clone G8.8; Biolegend) by incubation for 30min on ice. Cells were washed and stained using APC-conjugated Streptavidin (BD Pharmingen) for 20min on ice. Living tumour cells were selected by forward scatter, side scatter and by Hoechst dye exclusion. Fluorescence-activated cell sorting analysis was performed using FACSAria and FACSDiva software (BD Biosciences). Sorted cells were collected in the lysis buffer provided by the manufacturer (Microprep kit, RNeasy, Stratagene) and RNA extraction was performed according to the manufacturer's protocol. Total RNA was analysed using Mouse whole genome 430 2.0 array from Affymetrix at the VIB microarray facility (KU – Leuven, Belgium). We analyzed CD34+ TECs isolated by FACS from a K14CreER:Rosa-VEGF-164 and control mice. Analysis of the microarray was perfored by the VIB microarray facility (KU – Leuven, Belgium).
2011-09-06 | E-GEOD-31465 | biostudies-arrayexpress
Project description:Purpose: identify differentially expressed genes between ex-Treg obtained 5, 10 or 15 days post-adoptive transfer into a lymphopenic environment compared to ex vivo Treg in order to identify Treg subset with heightened instability. Method: Treg from Foxp3YFP-Cre Rosa26RFP (PMID: 18387831, PMID: 17171761) were co-injected with congenically-disparated naive T cells in a 1:1 ratio into Rag1KO mice and ex-Treg were sorted at 15 days, 10 days or 5 days post-transfer. As a control, ex vivo Treg were sorted from Foxp3YFP-Cre Rosa26RFP (d0 sample). Next, samples were stained with multiplexing antibodies using the BD Single-Cell Multiplexing Kit (BD biosciences), then pooled and stained with the following oligonucleotide-conjugated antibodies: anti‑CD25 (PC61), anti-CD44 (IM7), anti-CD62L (MEL-14), anti-GITR (DTA-1), anti-CD4 (RM4-5), anti‑TCRb (H57-597), anti‑CD69 (H1.2F3), anti‑TIGIT (1G9), anti‑CD95 (Jo2), anti‑CD122 (TM‑1β), anti‑Tim-3 (5D12), anti‑CCR7 (4B12), anti‑CD103 (M290), anti‑CD279 (J43), anti‑CD274 (MIH5), anti‑CD233 (C9B7W), anti‑CD71 (C2), anti‑CD278 (7E7G9), anti‑ITGB7 (M293), anti‑CD137 (1AH2), anti‑CD40 (3/23), anti‑CD3e (145-2C) from BD Biosciences. Single-cell capture and cDNA library preparation was performed using the BD Rhapsody Single-cell analysis system (BD Biosciences), according to the manufacturer’s instructions, using a custom gene panel (591 genes). Sequencing was performed on an Illumina NextSeq500 instrument using a Mid‑Output kit v2.5 (150 cycles, paired-end). Sample demultiplexing, barcode processing, alignment, filtering, UMI counting were done using the standard BD Biosciences Rhapsody analysis pipeline on Seven Bridges (www.sevenbridges.com). Result: In this study, we showed that ex-Treg downregulate Treg core-specific genes immediatly upon adoptive transfer into a lymphopenic environment. Further, we identified a Treg population enriched for unstable Trg clones. Conclusion: Our study was able to identify a Treg subset enriched for unstable Treg with plastic potential. This Treg subset appears highly enriched for naïve peripheral-induced Treg.
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