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Transcriptional profiles of human CD14+ monocytes conditioned with melanoma exosomes [gene expression]

ABSTRACT: We analyzed gene and microRNA expression profiles of purified CD14 monocytes isolated from blood of 5 healthy donors after 24 h of in vitro co-culture with exosomes exosomes isolated from the INT12 melanoma cell line generated in our laboratory from a human melanoma specimen. Differentially expressed genes and miRNAs were identified between treated and untreated monocytes.

ORGANISM(S): Homo sapiens

PROVIDER: GSE102009 | GEO | 2018/10/26

REPOSITORIES: GEO

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Transcriptional profiles of human CD14+ monocytes conditioned with melanoma exosomes [miRNA expression]

Project description:We analyzed gene and microRNA expression profiles of purified CD14 monocytes isolated from blood of 5 healthy donors after 24 h of in vitro co-culture with exosomes exosomes isolated from the INT12 melanoma cell line generated in our laboratory from a human melanoma specimen. Differentially expressed genes and miRNAs were identified between treated and untreated monocytes.

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Project description:Serum-free Fibrocytes, Serum-containing Fibrocytes, CD14++CD16- Monocytes, CD14++CD16+ Monocytes, CD14+CD16++ Monocytes, Macrophages were all generated from up to 3 biological replicates from each of 3 separate donors. RNA was extracted (Ambion RNAqueous), labelled with cy3, mixed with cy5 labelled human reference (Stratagene), and hybridised to slides printed with Human AROS v4.0 oligonucleotides (Operon). Slides were scanned using a Perkin Elmer GX plus, and the data then normalised with GEPAS v4.0 and collated. Final data analysis was carried out using TMEV 4.0. SAM was performed using a 0.1% FDR. PCA were plotted from this list, and interrogation carried out using DAVID to determine pathway enrichment.

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SuperSAGE evidence for CD14++CD16+ monocytes as a third monocyte subset

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Project description:Myeloid-derived suppressor cells (MDSC) is a heterogeneous population of cells that can negatively regulate T-cell function. As opposed to murine MDSC, which are characterized as Gr-1+CD11b+ cells, human MDSC are not so clearly defined due to lack of specific markers. Our lab has previously identified a new subset of MDSC as CD14+HLA-DR-neg/low cells from PBMC. CD14+HLA-DR-neg/low MDSC not only suppress proliferation and IFN-gamma secretion of autologous T cells, but also induce CD25+Foxp3+ regulatory T cells that are suppressive in vitro, whereas the counterpart CD14+HLA-DR-high monocytes don’t have the effect. In this study, we compare the immune-related gene expression between CD14+HLA-DR-neg/low MDSC and CD14+HLA-DR-high monocytes to better characterize the difference between these two populations and to find new potential specific marker for human MDSC. PBMC were isolated from fresh blood healthy donor by density centrifugation. CD14+ cells were isolated by AutoMACS CD14 microbeads using a AutoMACS (Miltenyi), and then stained with CD14 and HLA-DR antibodies. MDSC and monocytes control cells were sorted as CD14+ HLA-DR-neg/low and CD14+HLA-DR-high cells respectively. The sorted two populations were immediately frozen in liquid nitrogen and shipped to the company on dry ice for RNA isolation and further microarray.

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