Functional properties of HL60 cells matured with all- trans-retinoic acid and DMSO: Differences in response to interleukin-8 and fMLP

Abstract

All- trans-retinoic acid (ATRA) causes granulocyte differentiation in patients with acute promyelocytic leukemia. HL60 cells are frequently used as an in vitro model for studying granulocytes during maturation. We have previously studied actin polymerization in response to fMLP in HL60 cells undergoing DMSO induced maturation, and reported that IL-8 causes actin polymerization in neutrophils in a manner similar to fMLP. We now compare chemotaxis and actin polymerization in response to IL-8 and fMLP, and nitroblue tetrazolium (NBT) reduction in HL60 cells matured with ATRA and DMSO. Cells cultured for 4 days with ATRA and DMSO showed morphologic evidence of maturation. NBD-phallacidin staining and flow cytometry were used to measure changes in F-actin content in response to IL-8 and fMLP. Uninduced cells were not capable of actin polymerization or chemotaxis. Cells matured with ATRA exhibited a 2.6-fold increase in F-actin content in response to IL-8, but only a 1.2-fold increase in response to fMLP. Cells matured with DMSO responded to both IL-8 and fMLP in an equal manner with 1.6-fold increases in F-actin. The 2 h migration for ATRA induced cells was 124 μm in response to IL-8, 107 μm with fMLP, and 105 μm in buffer. DMSO induced cells migrated 89 μm in response to IL-8, 106 μm with fMLP, and 66 μm in buffer. With maturation, 65% of the ATRA induced cells reduced NBT compared with only 15% of the DMSO induced cells. In summary, HL60 cells cultured in ATRA develop greater functional maturity than those cultured in DMSO, and a greater responsiveness to IL-8 than fMLP, a finding distinct from previously reported work in neutrophils.