Lineage relationship in human dendritic cells
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE32400 | GEO | 2011/12/01
SECONDARY ACCESSION(S): PRJNA147933
REPOSITORIES: GEO
Similar Datasets
Project description:The lineage relationships and fate of human blood and tissue dendritic cells (DC) has significance for a number of diseases including HIV where both blood and tissue DC may be infected. We used gene expression profiling of monocyte and DC sub-populations sorted directly from blood and skin and compared this to monocyte derived DC (MDDC) and MUTZ3 Langerhans cells (LCs) to define the lineage relationships. Hierarchical clustering analysis showed that plasmacytoid DCs formed the most discrete cluster. The ex vivo derived myeloid cells formed two separate clusters of cells derived from blood, and skin. Separate and specific DC populations could be determined within the sub-clusters. During overnight culture CD14+ dermal DCs (DDC) converted to CD1a+ expressing cells in situ consistent with origin of the CD1a+ DDC from a local precursor rather than from circulating blood DC or monocyte precursors. The in vitro derived MDDC and MUTZ3 populations grouped within the skin DC cluster and MDDCs clustered most closely to CD14+ DDC consistent with the proposed similarity between these two cell types. We identified differential expression of novel genes in particular DC subsets including genes related to DC surface receptors (including C-type lectin receptors, toll-like receptors and galectins). Total RNA was extracted and hybridised to 24 bead arrays. Dendritic cells and monocytes from human blood and skin using magnetic bead and flow cytometry based cell sorting both before and after culture for 24 hours
2011-12-01 | E-GEOD-32400 | biostudies-arrayexpress
Project description:The lineage relationships and fate of human blood and tissue dendritic cells (DC) has significance for a number of diseases including HIV where both blood and tissue DC may be infected. We used gene expression profiling of monocyte and DC sub-populations sorted directly from blood and skin and compared this to monocyte derived DC (MDDC) and MUTZ3 Langerhans cells (LCs) to define the lineage relationships. Hierarchical clustering analysis showed that plasmacytoid DCs formed the most discrete cluster. The ex vivo derived myeloid cells formed two separate clusters of cells derived from blood, and skin. Separate and specific DC populations could be determined within the sub-clusters. During overnight culture CD14+ dermal DCs (DDC) converted to CD1a+ expressing cells in situ consistent with origin of the CD1a+ DDC from a local precursor rather than from circulating blood DC or monocyte precursors. The in vitro derived MDDC and MUTZ3 populations grouped within the skin DC cluster and MDDCs clustered most closely to CD14+ DDC consistent with the proposed similarity between these two cell types. We identified differential expression of novel genes in particular DC subsets including genes related to DC surface receptors (including C-type lectin receptors, toll-like receptors and galectins). Total RNA was extracted and hybridised to 62 cDNA arrays. Dendritic cells and monocytes from human blood and skin using magnetic bead and flow cytometry based cell sorting both before and after culture for 24 hours
2011-12-01 | E-GEOD-32648 | biostudies-arrayexpress
Project description:The lineage relationships and fate of human blood and tissue dendritic cells (DC) has significance for a number of diseases including HIV where both blood and tissue DC may be infected. We used gene expression profiling of monocyte and DC sub-populations sorted directly from blood and skin and compared this to monocyte derived DC (MDDC) and MUTZ3 Langerhans cells (LCs) to define the lineage relationships. Hierarchical clustering analysis showed that plasmacytoid DCs formed the most discrete cluster. The ex vivo derived myeloid cells formed two separate clusters of cells derived from blood, and skin. Separate and specific DC populations could be determined within the sub-clusters. During overnight culture CD14+ dermal DCs (DDC) converted to CD1a+ expressing cells in situ consistent with origin of the CD1a+ DDC from a local precursor rather than from circulating blood DC or monocyte precursors. The in vitro derived MDDC and MUTZ3 populations grouped within the skin DC cluster and MDDCs clustered most closely to CD14+ DDC consistent with the proposed similarity between these two cell types. We identified differential expression of novel genes in particular DC subsets including genes related to DC surface receptors (including C-type lectin receptors, toll-like receptors and galectins).
2011-12-01 | GSE32648 | GEO
Project description:In this study transcriptome profiling of dendritic cell subtypes was performed using various human dendritic cells. Monocyte-derived DC: CD14+ monocytes were obtained from buffy coats by Ficoll - gradient centrifugation and immunomagnetic cell separation using anti-CD14-conjugated microbeads. Monocytes were resuspended into 6-well culture dishes at a density of 1.5 x 10 million cells/ml and cultured in RPMI 1640 supplemented with 10% FBS containing 800 U/ml GM-CSF and 500 U/ml IL-4. Cells were cultured for 5 days and the IL-4 and GM-CSF addition was repeated at day 2. Dermal DC (DDC) and Langerhans Cells (LC) from skin: Human skin specimens were obtained from healthy donors undergoing corrective breast or abdominal plastic surgery after informed consent. Thick slices (3 mm) of skin containing the epidermis and the dermis were cut by use of a dermatome. Slices of skin were cut in pieces of 1 cm2 and incubated with Dispase II for 30â60 min at 37°C. The epidermis and dermis were separated with tweezers and washed with PBS. To isolate LC, the epidermal sheets were incubated with PBS with 0.05% trypsin for 10 min at 37°C, and a single-cell suspension was prepared by pushing the tissue through 100 µm pore nylon cell strainers with a plunger of a 2-ml syringe. Epidermal cell suspension was enriched for LC by density centrifugation over Lymphoprep and CD1a-guided MACS (Miltenyi Biotec). To isolate DDC, the dermis was incubated with PBS containing Dispase and Collagenase A at 37°C for 2 h, after which, a single-cell suspension was prepared by pushing through 100 µm pore nylon cell strainers with a plunger of a 2-ml syringe. Cell suspension was enriched for DDC by CD1a-guided MACS. Blood Cd1c+ DC: Blood CD1c+ DC were isolated from periferal blood. BDCA1+ (CD1c) DCs were incubated with a lineage-specific PE-labeled Ab mixture, HLA-DR-allophycocyanin, and BDCA1-FITC. Lineage-negative and HLA-DRhigh DCs, positive for BDCA1 were sorted on a FACS and collected in tubes containing 1 ml of 100% FBS. A total of 40,000â100,000 cells was isolated (>98% pure). Dendritic cell subtypes were differentiated in vitro from monocytes and primary dendritic cells were isolated from human skin and blood.
2010-10-27 | E-GEOD-23618 | biostudies-arrayexpress
Project description:In this study transcriptome profiling of dendritic cell subtypes was performed using various human dendritic cells. Monocyte-derived DC: CD14+ monocytes were obtained from buffy coats by Ficoll - gradient centrifugation and immunomagnetic cell separation using anti-CD14-conjugated microbeads. Monocytes were resuspended into 6-well culture dishes at a density of 1.5 x 10 million cells/ml and cultured in RPMI 1640 supplemented with 10% FBS containing 800 U/ml GM-CSF and 500 U/ml IL-4. Cells were cultured for 5 days and the IL-4 and GM-CSF addition was repeated at day 2. Dermal DC (DDC) and Langerhans Cells (LC) from skin: Human skin specimens were obtained from healthy donors undergoing corrective breast or abdominal plastic surgery after informed consent. Thick slices (3 mm) of skin containing the epidermis and the dermis were cut by use of a dermatome. Slices of skin were cut in pieces of 1 cm2 and incubated with Dispase II for 30–60 min at 37°C. The epidermis and dermis were separated with tweezers and washed with PBS. To isolate LC, the epidermal sheets were incubated with PBS with 0.05% trypsin for 10 min at 37°C, and a single-cell suspension was prepared by pushing the tissue through 100 µm pore nylon cell strainers with a plunger of a 2-ml syringe. Epidermal cell suspension was enriched for LC by density centrifugation over Lymphoprep and CD1a-guided MACS (Miltenyi Biotec). To isolate DDC, the dermis was incubated with PBS containing Dispase and Collagenase A at 37°C for 2 h, after which, a single-cell suspension was prepared by pushing through 100 µm pore nylon cell strainers with a plunger of a 2-ml syringe. Cell suspension was enriched for DDC by CD1a-guided MACS. Blood Cd1c+ DC: Blood CD1c+ DC were isolated from periferal blood. BDCA1+ (CD1c) DCs were incubated with a lineage-specific PE-labeled Ab mixture, HLA-DR-allophycocyanin, and BDCA1-FITC. Lineage-negative and HLA-DRhigh DCs, positive for BDCA1 were sorted on a FACS and collected in tubes containing 1 ml of 100% FBS. A total of 40,000–100,000 cells was isolated (>98% pure).
2010-10-27 | GSE23618 | GEO
Project description:Identification of MEK-ERK or p38MAPK dependent genes in human monocyte derived dendritic cells. Dendritic cells (DC) promote tolerance or immunity depending on their maturation state. Previous studies have revealed that DC maturation is enhanced or accelerated upon MEK-ERK signaling pathway inhibition. We have now determined the contribution of MEK-ERK activation to the profile of gene expression of human immature monocyte-derived dendritic cells (MDDC) and peripheral blood myeloid DC. ERK inhibition altered the expression of genes that mediate CCL19-directed migration (CCR7) and LDL binding (CD36, SCARB1, OLR1, CXCL16) by immature DC. Besides, ERK upregulated CCL2 expression while impaired the expression of DC maturation markers (RUNX3, ITGB7, IDO1). MEK-ERK-regulated genes exhibited an over-representation of cognate sequences for the Aryl Hydrocarbon Receptor (AhR) transcription factor, and we show that AhR mediates some of the ERK-dependent transcriptional effects in DC. Therefore, MEK-ERK signaling pathway regulates antigen capture, lymph node homing and the acquisition of maturation-associated genes, and its contribution to the maintenance of the immature state of MDDC and myeloid DC is partly dependent on the activity of AhR. Since pharmacological modulation of the MEK-ERK signaling pathway has been proposed as a potential therapeutic strategy for cancer, our findings indicate that ERK inhibitors might influence the generation of anti-tumor responses through regulation of critical DC effector functions. Human peripheral blood monocytes from three independent healthy donors (DC4, DC5 and DC7) were isolated by anti-CD14-labeled magnetic microbeads. CD14+ monocytes were cultured for 5 days in RPMI 10% FCS containing GM-CSF and IL-4 to generate immature monocyte-derived dendritic cells (MDDC). Immature MDDC were exposed to MEK inhibitor, U0126, or p38MAPK inhibitor, SB203580 for 1 hour and a final dose of GM-CSF and IL-4 were added to the culture. Cells were collected for analysis after 4, 10 or 24 hours.Total RNA from each condition was extracted using the All prep DNA/RNA/protein mini kit (Qiagen) and hybridized to an Agilent Human Whole Genome (4x44) Oligo Microarray. All experimental procedures were performed following manufacturer instructions.
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