Changes in glycosaminoglycan synthesis during the differentiation of HL-60 cells induced by 12-O-tetra-decanoylphorbol-13-acetate.

Abstract

The changes in glycosaminoglycan (GAG) synthesis during the differentiation of HL-60 cells to macrophage-like cells by treatment with 12-O-tetradecanoylphorbol-13-acetate (TPA) were studied by using 35S-sulfate or 3H-glucosamine as a tracer. The incorporation of 35S-sulfate into GAGs of HL-60 cells treated with TPA (TPA-treated HL-60 cells) decreased by 30% compared with untreated HL-60 cells (HL-60 cells). The profiles of the Sepharose CL-6B chromatography showed that the molecular weight size of proteoglycans (PGs) in the medium fraction of TPA-treated HL-60 cells was larger than that of HL-60 cells although the PGs in the cell fractions of both HL-60 and TPA-treated HL-60 cells were of the same molecular weight size. On the other hand, the molecular weight sizes of GAGs in both the medium and the cell fractions of TPA-treated HL-60 cells were larger than those of HL-60 cells. In order to examine the synthesis of GAG chains, HL-60 and TPA-treated HL-60 cells were incubated with 35S-sulfate in the presence of 4-methylumbelliferyl-.beta.-D-xyloside, an exogenous initiator. The incorporation of 35S-sulfate into total GAGs of the TPA-treated HL-60 cells exposed to the .beta.-D-xyloside increased 3-fold over that of HL-60 cells. HL-60 cells synthesized core protein-initiated GAGs and .beta.-D-xyloside-initiated GAGs but TPA-treated HL-60 cells synthesized .beta.-D-xyloside-initiated GAGs only. .beta.-D-Xyloside-initiated GAGs synthesized by both cells were obtained in the same fraction by Sepharose CL-6B chromatography. In addition, alterations in the chain length and synthetic activity of GAGs gradually occurred with the morphological changes and came to constant values 36 h after treatment with TPA. These results suggest a very close relationship with the monocytic differentiation of HL-60 cells. However, it is not possible at present to decide whether such changes in PG synthesis result from chain length only or induction of different PGs.