Project description:To directly compare the SLE monocyte transcriptional program with that of blood mDC precursors, we purified lineage HLA-DRhighCD11chigh mDCs and CD14+ monocytes from the blood of five healthy donors. Their gene expression profiles were then compared to those of blood SLE monocytes. An unsupervised clustering analysis of transcripts present in >20% of the samples classified healthy monocytes, SLE monocytes and healthy mDCs into three well defined groups. A supervised analysis was then performed to find genes: 1) differentially expressed in healthy mDCs compared to monocytes; 2) shared by healthy blood mDCs and SLE blood monocytes. To directly compare the SLE monocyte transcriptional program with that of blood mDC precursors, we purified lineage HLA-DRhighCD11chigh mDCs and CD14+ monocytes from the blood of five healthy donors. Their gene expression profiles were then compared to those of blood SLE monocytes. An unsupervised clustering analysis of transcripts present in >20% of the samples classified healthy monocytes, SLE monocytes and healthy mDCs into three well defined groups. A supervised analysis was then performed to find genes: 1) differentially expressed in healthy mDCs compared to monocytes; 2) shared by healthy blood mDCs and SLE blood monocytes.
Project description:To directly compare the SLE monocyte transcriptional program with that of blood mDC precursors, we purified lineage HLA-DRhighCD11chigh mDCs and CD14+ monocytes from the blood of five healthy donors. Their gene expression profiles were then compared to those of blood SLE monocytes. An unsupervised clustering analysis of transcripts present in >20% of the samples classified healthy monocytes, SLE monocytes and healthy mDCs into three well defined groups. A supervised analysis was then performed to find genes: 1) differentially expressed in healthy mDCs compared to monocytes; 2) shared by healthy blood mDCs and SLE blood monocytes.
Project description:In our study, MegaClust - an unsupervised, data-driven algorithm - barely described mDCs and pDC subsets were identified. To confirm these findings we performed RNA sequencing Facs sorted of mDCs (CD123-CD11c+CD4+HLA-DR+), pDCs (CD123+CD11c-CD4+HLA-DR+) and monocytes (CD14+) from healthy donors and compared these with publicly available data.
Project description:Expression data from LCH lesion subpopulations and healthy donors' peripheral blood specimens Langerhans cell histiocytosis (LCH) is a myeloproliferative disorder that is characterized by the inflammatory lesions with pathogenic CD1a+CD207+ dendritic cells (DCs). BRAFV600E and other somatic activating MAPK gene mutations have been identified in differentiating bone marrow and blood myeloid cells, but the origin of the LCH lesion CD1a+CD207+DCs and mechanisms of lesion formation remain incompletely defined. In order to identify candidate LCH CD1a+CD207+DCs’ precursor populations, gene expression profiles of LCH lesion CD1a+CD207+DCs were first compared to established gene signatures from human myeloid cell subpopulations. Interestingly, the CD1c+ myeloid DC (mDC) gene signature was most enriched in the LCH CD1a+CD207+DC’ transcriptome. Additionally, the BRAFV600E allele was not only localized to CD1a+CD207-DCs and CD1a+CD207+DCs, but it was also identified in CD1c+mDCs in LCH lesions. Transcriptomes of CD1a+CD207-DCs were nearly indistinguishable from CD1a+CD207+DCs (both CD207low and CD207high subpopulations). Transcription profiles of LCH lesion CD1a+CD207+DCs and peripheral blood CD1c+mDCs from healthy donors were compared to identify potential LCH DC-specific biomarkers. HLADQB2 expression was significantly increased in LCH lesion CD1a+CD207+DCs compared to circulating CD1c+mDCs from healthy donors, and HLA-DQB2 antigen was identified on LCH lesion CD1a+CD207- and CD1a+CD207+DCs as well as on CD1c+(CD1a+CD207-) mDCs, but not in any other lesion myeloid subpopulations. Interestingly, HLADQB2 expression was specific to peripheral blood of patients with BRAFV600E+ peripheral blood mononuclear cells (PBMC), and HLA-DQB2+CD1c+blood cells were highly enriched for the BRAFV600E in these patients. These data support a model where blood CD1c+mDCs with activated ERK migrate to lesion sites where they differentiate into pathogenic CD1a+CD207+ DCs.
Project description:Expression data from LCH lesion subpopulations and healthy donors' peripheral blood specimens Langerhans cell histiocytosis (LCH) is a myeloproliferative disorder that is characterized by the inflammatory lesions with pathogenic CD1a+CD207+ dendritic cells (DCs). BRAFV600E and other somatic activating MAPK gene mutations have been identified in differentiating bone marrow and blood myeloid cells, but the origin of the LCH lesion CD1a+CD207+DCs and mechanisms of lesion formation remain incompletely defined. In order to identify candidate LCH CD1a+CD207+DCs’ precursor populations, gene expression profiles of LCH lesion CD1a+CD207+DCs were first compared to established gene signatures from human myeloid cell subpopulations. Interestingly, the CD1c+ myeloid DC (mDC) gene signature was most enriched in the LCH CD1a+CD207+DC’ transcriptome. Additionally, the BRAFV600E allele was not only localized to CD1a+CD207-DCs and CD1a+CD207+DCs, but it was also identified in CD1c+mDCs in LCH lesions. Transcriptomes of CD1a+CD207-DCs were nearly indistinguishable from CD1a+CD207+DCs (both CD207low and CD207high subpopulations). Transcription profiles of LCH lesion CD1a+CD207+DCs and peripheral blood CD1c+mDCs from healthy donors were compared to identify potential LCH DC-specific biomarkers. HLADQB2 expression was significantly increased in LCH lesion CD1a+CD207+DCs compared to circulating CD1c+mDCs from healthy donors, and HLA-DQB2 antigen was identified on LCH lesion CD1a+CD207- and CD1a+CD207+DCs as well as on CD1c+(CD1a+CD207-) mDCs, but not in any other lesion myeloid subpopulations. Interestingly, HLADQB2 expression was specific to peripheral blood of patients with BRAFV600E+ peripheral blood mononuclear cells (PBMC), and HLA-DQB2+CD1c+blood cells were highly enriched for the BRAFV600E in these patients. These data support a model where blood CD1c+mDCs with activated ERK migrate to lesion sites where they differentiate into pathogenic CD1a+CD207+ DCs.
Project description:human blood monocytes were isolated, activated and harvested at several timepoints In this study, we identified genes that were differentially expressed in human monocytes activated with eiter NOD2L and/or TLR2/1L. human blood monocytes were purified from healthy donors by Ficoll, Percoll and adherence. Monocytes were activated using NOD2L (MDP) and the TLR2/1L (19kD, triacylated peptide). Cells were harvested before activation (0h) and 6h and 24h after stimulation with ligands.