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.
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.
Project description:In order to study the influence of melanoma-derived exosomes in the lymphatic vasculature within the lymph nodes, we have profile the transcriptional changes occurring in human lymphatic endothelial cells treated with melanoma-secreted exosomes or with the conditioned medium from melanoma cells depleted of exosomes.
Project description:As part of our study in understanding the role of SP140 in inflammatory pathways in macrophages, we inhibited SP140 mRNA using siRNA. Peripheral blood mononuclear cells (PBMCs) were obtained from whole blood of healthy donors (from Sanquin Institute Amsterdam or from GSK Stevenage Blood Donation Unit) by Ficoll density gradient (Invitrogen). CD14+ monocytes were positively selected from PBMCs using CD14 Microbeads according to the manufacturer’s instructions (Miltenyi Biotec). CD14+ cells were differentiated with 20 ng/mL of macrophage colony-stimulating factor (M-CSF) (R&D systems) for 3 days followed by 3 days of polarization into classically activated (inflammatory) M1 macrophages (100 ng/mL IFN-γ; R&D systems). M1 macrophages were transfected with siGENOME human smartpool SP140 siRNA or non-targeting scrambled siRNA for 48h with DharmaFECT™ transfection reagents according to manufacturer’s protocol (Dharmacon). The cells were left unstimulated or stimulated with 100 ng/mL LPS (E. coli 0111:B4; Sigma) for 4h (for qPCR) or 24h (for Elisa). The cells were lysed (ISOLATE II RNA Lysis Buffer RLY-Bioline) for RNA extraction.150 ng total RNA was labelled using the cRNA labelling kit for Illumina BeadArrays (Ambion) and hybridized with Ref8v3 BeadArrays (Illumina). Arrays were scanned on a BeadArray 500GX scanner and data were normalized using quantile normalization with background subtraction (GenomeStudio software; Illumina). This submission only contains processed data
Project description:Transcriptional profiling of human mesenchymal stem cells comparing normoxic MSCs cells with hypoxic MSCs cells. Hypoxia may inhibit senescence of MSCs during expansion. Goal was to determine the effects of hypoxia on global MSCs gene expression.
