Based on CD14 and CD16 expression, human peripheral blood monocytes (MO) can be divided into a major CD14(high) CD16(-) population and two minor CD14(high) CD16(+) and CD14(dim) CD16(+) subpopulations. CD14(dim) CD16(+) MO are well characterized and regarded as pro-inflammatory because upon stimulation produce TNF-alpha but little, if any, IL-10. By contrast, little is known about CD14(high) CD16(+) MO. We investigated the surface expression of selected determinants by CD16(+) MO subpopulations, cytokine production, phagocytosis and antigen presentation. We found that both CD16(+) subpopulations had a higher expression of HLA-DR, CD86, CD54 and a lower expression of CD64 than CD14(high) CD16(-) population. In addition, CD14(high) CD16(+) MO showed a higher expression of CD11b and TLR4 than CD14(dim) CD16(+) and CD14(high) CD16(-) subpopulations. CD14(high) CD16(+) MO exhibited an increased phagocytic activity and a decreased antigen presentation in comparison with CD14(dim) CD16(+). As expected, lipopolysaccharide (LPS)-stimulated CD14(dim) CD16(+) MO produced TNF-alpha but little IL-10. By contrast, LPS-stimulated CD14(high) CD16(+) subpopulation produced significantly more IL-10 than CD14(dim) CD16(+) and CD14(high) CD16(-) MO. In conclusion, our data show that human peripheral blood CD16(+) MO are heterogeneous in function and consist of two subpopulations: CD14(dim) CD16(+) pro-inflammatory and CD14(high) CD16(+) with anti-inflammatory potential.