Altered sialidase expression in human myeloid cells undergoing apoptosis and differentiation (original) (raw)

To gain insight into sialic acid biology and sialidase/neuraminidase (NEU) expression in mature human neutrophil (PMN)s, we studied NEU activity and expression in PMNs and the HL60 promyelocytic leukemic cell line, and changes that might occur in PMNs undergoing apoptosis and HL60 cells during their differentiation into PMN-like cells. Mature human PMNs contained NEU activity and expressed NEU2, but not NEU1, the NEU1 chaperone, protective protein/cathepsin A(PPCA), NEU3, and NEU4 proteins. In proapoptotic PMNs, NEU2 protein expression increased > 30.0-fold. Granulocyte colonystimulating factor protected against NEU2 protein upregulation, PMN surface desialylation and apoptosis. In response to 3 distinct differentiating agents, dimethylformamide, dimethylsulfoxide, and retinoic acid, total NEU activity in differentiated HL60 (dHL60) cells was dramatically reduced compared to that of nondifferentiated cells. With differentiation, NEU1 protein levels decreased > 85%, PPCA and NEU2 proteins increased > 12.0-fold, and 3.0-fold, respectively, NEU3 remained unchanged, and NEU4 increased 1.7-fold by day 3, and then returned to baseline. In dHL60 cells, lectin blotting revealed decreased α2,3-linked and increased α2,6-linked sialylation. dHL60 cells displayed increased adhesion to and migration across human bone marrow-derived endothelium and increased bacterial phagocytosis. Therefore, myeloid apoptosis and differentiation provoke changes in NEU catalytic activity and protein expression, surface sialylation, and functional responsiveness. Polymorphonuclear leukocyte (PMN)s are part of the first line of host defenses against invasive prokaryotic pathogens 4 . Myeloid progenitors undergo maturation within the bone marrow into mature PMNs 1,2 . These motile and highly deformable cells are capable of percolating through and exiting the bone marrow sinusoids, circulating through the intravascular compartment, and recognizing and engaging adhesion molecules surface-expressed on the endothelium. PMNs undergo profound shape changes permitting these cells to squeeze through the small caliber microvasculature and interendothelial cell junctions to enter extravascular tissues 3 , where they adhere to and engulf bacteria for successful phagocytosis and intracellular killing 4 . The HL60 promyelocytic leukemia cell line has been used as a model for myelopoiesis, and its differentiation towards PMN-like cells has been used as a surrogate for short-lived, difficult to manipulate PMNs 5-8 . For example, these same HL60 cells are regularly used in opsonophagocytic assays to establish opsonic activity of vaccine-induced antibodies 9 . Given the critical role of sialic acid (SA), neuraminidase/sialidase (NEU), and sialyltransferase (ST) expression in PMN function 10-13 , we examined their impact on HL60 cellular responsiveness. SAs comprise a family of 9-carbon sugars, each carboxylated on the C1 position . These SA residues are almost always located at the terminus of glycan chains, where they are strategically positioned to influence intermolecular and cell-cell interactions. SA on the cell surface imparts a negative surface charge that generates