Figure 4

Lipopolysaccharide-induced intracellular cytokine expression in monocytes and myeloid dendritic cells in sepsis patients. Lipopolysaccharide (LPS)-induced intracellular cytokine expression in monocytes and in myeloid dendritic cells (MDCs) in patients with sepsis compared with healthy controls. (a) Unstimulated blood was stained for CD14, CD33 and CD16, and then monocyte subsets (region (R) 1, CD14^brightCD33^bright; R2, CD14^dimCD33^dim) and MDCs (R3, CD14^dimCD33^bright) were gated in a CD14/CD33 scatter plot for subsequent analysis of CD16 expression. This staining strategy allowed identification of CD16^positive (CD14^dimCD33^dim) monocytes (R2) despite the loss of CD16 expression after LPS stimulation. (b) Blood was drawn from healthy controls and from sepsis patients, and was stimulated with LPS (6 hours) in the presence of the secretion blocker Brefeldin A. After surface staining for CD14, HLA-DR and CD33, cells were intracellularly stained for TNFα, IL-1β, IL-6, and IL-10 using PE-labeled cytokine-specific monoclonal antibody or murine IgG₁-PE as control. The percentage of cytokine-positive CD14^brightCD33^positive monocytes, CD14^dimCD33^dim monocytes and CD14^negativeCD33^bright MDCs is given. *P < 0.05, **P < 0.01.