Phenotypic and Functional Differentiation of Promyelocytic Cell Line HL-60 by N-N-dimethylformamide

Kim, Kyung Hyo; Seoh, Ju Young

Abstract

PURPOSE: HL-60 is a promyelocytic cell line. Fc receptors and complement receptor 3 (CR3) play important role in the protective response of granulocytes and monocytes against microbial infection. We quantified the expression of Fc I, Fc II, Fc III, and CD11b/CD18 during differentiation using HL-60 cells by N-N-dimethylformamide (DMF). Functional studies, such as phagocytic activity, respiratory burst and ADCC, were also performed.
METHODS
HL-60 cells were induced to differentiate by adding 0.8% DMF. On the 4th and 7th day after stimulation as well as before stimulation, the cells were analyzed for phenotypic and functional differentiaton. Phenotypic analysis was performed by flow cytometry after staining the cells with PE-conjugated anti-human CD64, CD32, CD16, CD11b, CD18, and isotype controls. And the measured fluorescent intensity was transformed into Molecules of Equivalent Soluble Fluorochromes (MESF). Phagocytic activity was also measured by flow cytometry after incubation with fluorochrome-conjugated beads. Respiratory burst was measured by chemiluminescence assay of cells incubated with luminol after stimulation with PMA. ADCC was measured by hemoglobin release assay.
RESULTS
The expression of CD11b, CD18 and CD64 on HL-60 cells markedly increased on the 4th day and slightly decreased on the 7th day. Expression of CD32 was already induced before differentiation induction and slightly increased by DMF. CD16 was not expressed during differentiation. In phagocytic assay, the phagocytic cell fraction increased by stimulation on 4th and 7th day. Chemiluminescence showed the DMF increased the respiratory burst of HL-60 cells on the 4th and 7th day. In ADCC, DMF increased the target cell lysis continuously.
CONCLUSION
HL-60 cells which were differentiated with DMF for are good models for studying opsonophagocytic assay of immunized sera.