Differential cytokine expression in human blood monocyte subpopulations: a polymerase chain reaction analysis

Abstract

Abstract The subpopulation of strongly CD14-positive (CD14++) monocytes and monocytes coexpressing the CD16 antigen and low levels of CD14 (CD14+/CD16+ cells) were isolated by fluorescence-activated cell sorting (FACS) followed by stimulation with lipopolysaccharide (LPS) at 1 micrograms/mL. Polymerase chain reaction (PCR) after reverse transcription of isolated mRNA (RT-PCR) revealed similar levels of tumor necrosis factor (TNF) transcripts in both subpopulations. By contrast, transcripts for interleukin-10 (IL-10) were only detectable in CD14++ monocytes, whereas CD14+/CD16+ cells produced no detectable IL-10 transcripts after 4 hours. Only after 16 hours of LPS stimulation was a low level of IL-10 transcripts discernible in CD14+/CD16+ monocytes. The same pattern was seen at the protein level in that TNF in LPS-stimulated supernatants was comparable for both subpopulations, whereas IL-10 was detected in CD14++ monocytes but not in CD14+/CD16+ cells. To avoid interference of cell activation by CD14 and CD16 antibodies, cells were also isolated based on the high and low level of CD33 antigen expression. Again, weakly CD33-positive cells, which comprise the CD14+/CD16+ cells, showed no or only minimal IL-10 mRNA. When comparing blood monocyte subpopulations with alveolar macrophages (AM), AM showed high levels of LPS-stimulated TNF, whereas IL-10 transcripts were undetectable. Our data show that CD14+/CD16+ blood monocytes produce high levels of proinflammatory cytokines like TNF, whereas the anti-inflammatory IL-10 is low or absent, a pattern similar to what is seen in AM.