CD14, CD16 and HLA-DR reliably identifies human monocytes and their subsets in the context of pathologically reduced HLA-DR expression by CD14hi/CD16neg monocytes: Expansion of CD14hi/CD16pos and contraction of CD14lo/CD16pos monocytes in acute liver failure

Changes in monocytes and their subsets (CD14hi/CD16neg, CD14hi/CD16pos and CD14lo/CD16pos) have been described in several diseases. The combination of CD14, CD16 and HLA-DR has been suggested to discriminate monocytes from the CD16pos/HLA-DRneg NK-cells and neutrophils but no data exist whether this strategy can be used in situations when monocyte HLA-DR expression is pathologically reduced. Monocytes and their subsets were concurrently identified through negative (exclusion of CD66bpos neutrophils, CD56pos NKcells, CD19pos B-cells, and CD3pos T-cells) and positive gating (inclusion of monocytes by expression of CD14, CD16, and HLA-DR) strategies on 30 occasions [9 healthy controls (HC) and 21 patients with conditions associated with low monocyte HLA-DR expression]. Bland-Altman and Passing and Bablok regression statistics did not demonstrate any significant measurement bias between the two strategies of monocyte identification. Monocyte subset phenotype was then compared in 18 HC and 41 patients with acute liver failure (ALF). Compared with HC, in ALF, the percentage of CD14hi/CD16pos monocytes was higher (7% vs 4%) whilst the percentage of CD14lo/CD16pos was lower (1.9% vs. 7%) (P = 0.001); HLA-DR and CD86 MFIs on all monocyte subsets were lower, whilst CCR5, CD64, and CD11b MFIs were higher (P < 0.05). The relative expression by monocyte subsets of HLA-DR, CCR2, CCR5, CX3CR1, and CD11a was similar in ALF patients and HCs. Repeat analysis of an identical antibody-fluorochrome backbone targeting HLA-DR, CD14, and CD16 was assessed in 189 samples across 5 different experiments. There was excellent agreement in the results obtained using the positive gating strategy (interclass correlation coefficients > 0.8). Monocytes and their subsets can be reliably identified using an antibody-fluorochrome backbone of HLA-DR, CD14, and CD16. CD16pos monocytes continue to constitutively express HLA-DR even in conditions where HLA-DR is pathologically reduced on CD14hi/CD16neg monocytes. Understanding the changes in monocyte pheontype in ALF and similar clinico-pathological diseases may allow the development of novel biomarkers or therapeutic strategies. (C) 2012 International Society for Advancement of Cytometry