The study of human myeloid differentiation using bromodeoxyuridine (BrdU)

Abstract

Little is known concerning the mechanism of myeloid differentiation. A human promyelocytic cell line (HL-60) differentiates to granulocytes or macrophage-like cells when cultured with a variety of agents. How these agents trigger myeloid differentiation is not understood. This study shows that 1.0-10.0 micrograms/ml bromodeoxyuridine (BrdU) induced myeloid differentiation of HL-60 in liquid culture. After 7 days, BrdU (3.0 micrograms/ml) produced only moderate inhibition of HL-60 growth, but induced myeloid maturation with 40% of the cells becoming morphologically more mature; 41% developed the ability to reduce nitroblue tetrazolium (NBT); 19% phagocytized Candida albicans; and 18% developed Fc receptors. The action of BrdU was mimicked by 5-iodo-deoxyuridine. Thymidine (Td) (1- to 10-fold excess) competitively inhibited incorporation of [3H]BrdU into DNA of HL-60 and inhibited the triggering of HL-60 differentiation by BrdU. The BrdU-induced maturation of HL-60 correlated with the incorporation of BrdU into DNA of HL-60. DNA buoyant density studies showed that about 46% of the Td was replaced by BrdU in each DNA strand of HL-60 as the cells differentiated in culture containing 3 micrograms/ml BrdU for 7 days. We established 20 thymidine kinase (TK)-deficient HL-60 clones. The HL-60 TK-deficient cells were unable to phosphorylate Td, to incorporate either [3H]Td or [3H]BrdU or differentiate in the presence of BrdU (1-1000 micrograms/ml). The HL-60 TK-deficient cells retained the ability to differentiate in the presence of other HL-60 inducers. Taken together, the studies suggest myeloid differentiation of HL-60 is triggered because of incorporation of BrdU into DNA of the cells.

Characterization of the metabolic forms of 6‐thioguanine responsible for cytotoxicity and induction of differentiation of HL‐60 acute promyelocytic leukemia cells

“…Thus, the differentiation inducing ability of xylosyladenine in the presence of an inhibitor of adenosine deaminase is abolished in an adenosine kinase-deficient clone of the Friend erythroleukemia (Lacour et al, 1980). Similarly, bromodeoxyuridine, a moderately effective initiator of the maturation of HL-60 cells, is unable to induce differentiation in a thymidine kinase deficient clone of this tumor (Koeffler et al, 1983).…”

Spontaneous and induced differentiation of human melanoma cells

“…BUdR has been shown to induce differentiation in other cell systems, including human myeloid leukemia cells (Koeffler et al, 1983) and neuroblastoma cells (Feyles et al, 1991), without significantly altering cellular function or viability. The ability of BUdR to modulate differentiation may be related to its incorporation into DNA.…”

Increased levels of the translation initiation factor eIF4E in differentiating epithelial lung tumor cell lines

“…In order to investigate if exposure of cell lines to BrdU results in a general increase in eIF4E expression, we have used the non‐epithelial, leukaemic cell line, HL‐60. In HL‐60 cells, BrdU induces myeloid differentiation (Koeffler et al, 1983; Yen and Forbes, 1990). eIF4E protein levels have been found to decrease during phorbol ester‐induced myeloid differentiation (Johnston et al, 1998).…”

“…We have used BrdU to study the differentiation of epithelial lung cancer cell lines in culture, where BrdU (a bromine‐substituted thymidine analogue) competes with naturally occurring thymidine for incorporation into DNA. The induction or inhibition of differentiation by BrdU depends on the particular cell type, inducing differentiation in human SCLC (Feyles et al, 1991), HL‐60 cells (Koeffler et al, 1983; Yen and Forbes, 1990) and neuroblastoma (Ross et al, 1995), while causing de‐differentiation in human melanomas (Thomas et al, 1993) and chondrocytes (Farrell and Leukens, 1995). It also selectively modulates the expression of a class of regulatory genes, each member of which is important for the development of a different cell lineage, either through altered promoter structure or increased/decreased affinity of transcriptional regulatory factors for substituted DNA.…”