Effect of HL-60 cells stimulated by IFN-γ and IL-10 to express B cell activating factor via transcriptional pathway
Objective To explore the effect of IFN-γ, IL-10 and IL-4 on B cell activating factor (BAFF) expression in human HL-60 cells, a kind of myeloid tumor cell lines, and its possible regulation mechanism. Methods Cultured human HL-60 cells were treated with IFN-γ, IL-10 and IL-4 for 1-3 days. The expression of membrane-bound BAFF on HL-60 cells was examined by flow cytometry, the amount of soluble BAFF was detected by ELISA assay, and the level of BAFF mRNA was tested by real-time PCR method. A functional 1021 bp fragment of the 5'-tlanking region of the human BAFF gene (-1349 to -329 bp) was cloned and investigated with serial 5'-deletion. The 5'-deleted promoters were recombinated with chloramphenicol acetyltransferase (CAT) as reporter gene. These five recombinant plasmids were transiently transfected to HL-60 cells with liposomal transfectian method. Promoters activities were determined by CAT reporter gene assay(CAT-ELISA) in those transfected cells treated with different cytokines. Results The results showed that the expression of membrane-bound BAFF, soluble BAFF and BAFF mRNA in human HL-60 cells were significantly elevated (P ＜ 0. 05) after incubated with IFN-γ and IL-10. In addition, IFN-γ and IL-10 showed significantly (P ＜ 0. 05) increased effects on promoter activity in human BAFF gane. And the cytokines-responsive sequences were located between -929 and -719 bp of the BAFF promoter region. Conclusion The enhancement of IFN-γ and IL-10 on BAFF expression and synthesis were regnla-ted by promoter activation. Our in vitro studies also raise the possibility to investigate the mechanisms regula-ting BAFF expression in other tumor cells of myeloid origin under pathological circumstances. 
 
Key words: 
Cyokines;  B cell activating factor;  HL-60 cells;  Promoter