Alteration in glycosphingolipid pattern during phorbol-12-myristate-13-acetate-induced cell differentiation in human T-lymphoid leukemia cells.

We analyzed the patterns of glycosphingolipids (GSLs) from a line of cells derived from a clone of the human T-cell leukemia cells (CEM) that had been induced to differentiate by phorbol-12-myristate-13-acetate (PMA) into cells with a suppressor-like phenotype. We characterized the differentiation state of the cells by immunofluorescence by using anti-cell surface differentiation-specific monoclonal antibodies (OKT3, OKT4, OKT6, and OKT8). The GSLs were extracted and separated by thin-layer chromatography and the individual bands were quantitated by a dual-wavelength densitometer or by autoradiography of GSLs labeled with [14C]glucosamine and [14C]galactose. Treatment of the CEM cells with 0.16-16 nM PMA for 6 h to 6 days resulted in a dose- and time-dependent increase in the amount of two neutral GSLs [ceramide monohexoside and ceramide dihexoside] and three gangliosides [monosialoganglioside (GM3), sialosylparagloboside, and disialoganglioside (GD3)]. The increase in the neutral GSLs after PMA treatment reached its maximum at 30 h while GM3 peaked at 96 h. The increases in GM3 and sialosylparagloboside are presumably due to an increase in their synthesis levels because PMA promoted an elevated incorporation of glucosamine and galactose into these GSLs. The increase in the amount of GD3, on the other hand, is due to either a decrease in its degradation or use in other metabolic pathways because no detectable increase in glucosamine and galactose incorporation into this ganglioside could be found. Incubation of control or PMA-induced CEM cells with GM3 fractions purified from either CEM cells, human brain, or dog erythrocytes caused a reduction in cell growth and prevented the increase in reactivity of the induced cells with the OKT3 antibody. Incubation with semisynthetic ceramide dihexoside, however, prevented the decrease in reactivity with the OKT4 antibody. The observed changes in GSL patterns during PMA-induced differentiation of the CEM cells into suppressor-like cells and the inhibition of CEM cell growth by GM3 fractions suggest that the GSLs play a role in the control of cell growth and differentiation in the PMA-treated CEM cells.