Project description:To explore the full extent of IFN-regulated transcriptional changes, we exposed monocytes from two healthy donors to recombinant type I IFN (IFN-M-NM-12b) in vitro. RNA was extracted at different incubation times (1, 6, 24, 48 and 72 hrs) and the expression data was normalized to that of monocytes cultured with medium. Blood monocytes isolated from healthy volunteers were incubated with 20% autologous serum alone or in the presence of 1000 U/ml of IFNM-NM-12b (Schering Plough, Kenilworth, NJ) in 6-well plates at a concentration of 106 monocytes per well in 3 ml of media. After incubation for one hour at 37 M-bM-^AM-0C, cells were harvested and RNA was extracted. Identical experiments were done after the following incubation time points: six hours, twenty four hours, two days, and three days.
Project description:Transcriptional profiling of human T cells analyzing the impact of race on the responsiveness to IFNa in healthy blood donors. Control and IFNa-treated samples derived from healthy Caucasian American vs. African American blood donors are compared.
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:Transcriptional profiling of human T cells analyzing the impact of race on the responsiveness to IFNa in healthy blood donors. Control and IFNa-treated samples derived from healthy Caucasian American vs. African American blood donors are compared. Two-condition experiment, Control vs. IFNa-treated. Paired design; untreated and treated samples for each donor.
Project description:We report cell specific responses to IFNa in 11 different peripheral immunocyte populations in the mouse. These cells represent the core ImmGen immunocyte lineage panel. Profiles from these were used to analyze cell specific responses to IFNa. In general a core set of ISG transcripts are induced in all cells. Smaller sets of ISGs were induced in a cell specific manner. In particular, splenic granulocytes and dendritic cells show restriced indcution of sets of ISGs. Male C57BL/6 mice were injected subcutaneously with 10,000U IFNa according to time course profile. Splenocytes/Peritoneal lavage suspensions were stained in multicolor antibody panels that were used to distinguish 11 different immunocyte populations (Immgen 11 cell set). These cells were sorted by FACS directly into TriZo and gene expression profiling was performed on Affymetrix Mouse Gene 1.0 ST Arrays.
Project description:Expression profiles at different time points during dendritic cell differentiation (induced by specific culture conditions) including monocytes as well as expression profiles between monocytes and completely differentiated cells (macrophages at day7 and dendritic cells at day7, respectively) were compared. Monocyte-derived dendritic cells (DC) were obtained by culturing elutriated monocytes with 20U/ml IL-4, 280U/ml GM-CSF and 10% FCS; monocyte-derived macrophages (MAC) were obtained by culturing elutriated monocytes with 2% AB serum. Three to seven biological replicates that are derived from independent healthy donors were included. One-color based gene expression. 2 datasets: dendritic cell kinetic study and comparison of monocyte, macrophage, and dendritic cells
Project description:Background: Circulating periostin levels are increased in systemic sclerosis (SSc) patients and correlates positively with disease severity. Additionally, monocytes/macrophages are one of the key cells in SSc pathogenesis. However, the effect of periostin on immune cells, particularly monocytes and macrophages, has not been well investigated. Objectives: To examine the effect of periostin on monocytes and monocyte-derived macrophages (MDMs) in SSc. Methods: Monocytes were purified from blood samples of nine diffuse cutaneous SSc patients and five healthy controls (HCs) and were differentiated into macrophages. Cell surface markers were assessed by flow cytometry. RNA sequencing analysis was performed on recombinant periostin-stimulated monocytes from three HCs. Cytokine/chemokine and extracellular matrix (ECM) protein expressions in MDMs differentiated supplemented with recombinant periostin were assessed by quantitative real-time polymerase chain reaction and bead-based immunoassays in six HCs. Moreover, we evaluated whether periostin was involved in monocyte migration by Transwell assay. Results: The modified Rodnan total skin thickness score correlated positively with the proportion of CD80-CD206+ M2 cells. The M2 macrophage proportion was reduced in rPn-stimulated MDMs in HCs, but not in SSc patients. mRNA expression levels of pro-fibrotic cytokine, chemokine, and ECM protein genes were significantly upregulated in recombinant periostin-stimulated monocytes and MDMs as compared to those from control monocytes and MDMs. Similar trends for their protein levels were also observed in MDMs. Moreover, the ratio of migrated cells in recombinant periostin-stimulated monocytes was significantly higher than that in control monocytes.