Divergent regulation of cell surface protease expression in HL-60 cells differentiated into macrophages with granulocyte macrophage colony stimulating factor or neutrophils with retinoic acid

Abstract

Taking advantage of the recently demonstrated presence of N-aminopeptidases and the serine protease dipeptidyi aminopeptidase IV (DPP IV) at the surface of human myeloblastic HL-60 cells, the regulation of these protease activities in HL-60 cell differentiation has been assessed using combined spectrophotometric and flow cytometric assays. Addition of human recombinant granulocyte macrophage colony stimulating factor (rHu-GM-CSF) to HL-60 cells to induce differentiation into macrophages led to a time and dose-dependent increase in both cell surface N-aminopeptidase and DPP IV activities. Protease up-regulation was due to an enhancement in cell surface protease number, associated with a slight rise in apparent affinities of the enzymes for their substrates. In contrast, in HL-60 cells induced to differentiate into neutrophils in the presenceof retinoic acid, expression of cell surface N-amlnopeptidases was almost completely abolished in a time-and dose-dependent fashion, and this down-regulation was accompanied by a weak but significant decrease in affinity. However, no noticeable difference was seen in serine DPP IV expression between retinoic acid-treated and untreated HL-60 cells. Retinoic acid treatment also reduced soluble protease activity in vitro indicating that down-regulation of membrane aminopeptldases was not due to their proteolytic clip. No modulation in the activity of any of the enzymes tested was seen with human recombinant tumor necrosis factor-α or retinol which do not induce HL-60 cell differentiation. The up-regulation of cell surface protease expression in HL-60 cells differentiated into macrophages was similar to that observed in monocytes isolated from peripheral blood: both DPP IV and N-aminopeptidase activities strictly increased on cells that undergo macrophage maturation (up to 5-fold) and independently of the nature of the differentiation inducer. Thus, the distinctive patterns of N-aminopeptidase and DPP IV expression that are seen in differentiating neutrophils and macrophages appear to be relatedto differences in stage of myeloid maturation. Because cell surface proteases are crucially involved in leukocyte functions, the data presented suggest that alterations in cell surface protease expression are associated with events controlling the differentiation of immature cells.