Ascorbate Increases the 1,25 Dihydroxyvitamin D 3 -Induced Monocytic Differentiation of HL-60 Cells

Abstract

1,25 Dihydroxyvitamin D3 (calcitriol) induces differentiation of HL-60 leukemia cells. We studied the in vitro effect of a physiological concentration of ascorbate as potentiator of 1,25 dihydroxyvitamin D3 [(OH)2D3] activity by determining different markers of differentiation: nitroblue tetrazolium reduction, nonspecific esterase activity, and the expression of CD11b and CD14 surface antigens. Nitroblue tetrazolium reduction and nonspecific esterase activity increased up to 50% in the presence of both 1,25 (OH)2D3 plus 0.2 mM ascorbate (ASC), compared with (OH)2D3 as a unique agent. ASC also increased the expression of specific surface antigens (CD11b and CD14) during differentiation induced by 1,25 (OH)2D3, the effect being more pronounced after 48 hours of treatment with 10(-8) M 1,25 (OH)2D3. Furthermore, 1,25 (OH)2D3 alone increased intracellular cAMP level during differentiation, and the addition of ASC increased its concentration from 60 to 100% above the level reached with 1,25 (OH)2D3 as unique agent. ASC did not enhance the antiproliferative effect of calcitriol, suggesting that it only affects the ability of 1,25 (OH)2D3 to promote differentiation of HL-60 cells.

Cellular redox state and activating protein-1 are involved in ascorbate effect on calcitriol-induced differentiation

“…For unsupplemented control treatments, the absolute values were 4.37 _+ 1.9 and 5.37 _+ 1.2 for NBTand CD14-positive cells, respectively; for unsupplemented la,25(OH)zD3 treatments they were 11.5_+ 1.1 and 50.3 +7.2, respectively NADPH oxidase by the NBT-reducing test and the presence of CD14 in the plasma membrane of cells. Ascorbate addition to lc~,25(OH)2D3-induced cultures doubled the percentage of NBT-positive cells and increased CD14-positive cells in a 50% in agreement with our earlier results (Quesada et al 1996, LdpezLluch et al 1998. In both differentiation markers, neither D HA nor Asc-2-P affected the 10t,25 (OH)2O3-induced differentiation.…”

“…As we have shown previously, ascorbate increases the monocytic differentiation induced by la,25 (OH)2D3 in HL-60 cells (Quesada et al 1996). Ascorbate-related compounds such as the fully oxidized form, DHA, and a stable analogue of ascorbate, Asc-2-R which is slowly oxidized in solution, were unable to modify the rate of differentiation induced by 10~,25(OH)zD3 (Table 1).…”

“…We have previously demonstrated that ascorbate increases the differentiation induced by lcz,25-dihydroxyvitamin D3 (lc~,25(OH)2D3) in HL-60 cells as determined by the increase in expression of cell surface antigens such as CDllb and CD14, and the appearance of NADPH oxidase and nonspecific esterase activities as mature cells markers (Quesada et al 1996). Another antioxidant, (z-tocopherol, also increases the differentiation induced by lcz,25(OH)2D3 in HL-60 cells (Sokoloski et al 1997a,b).…”

The CD300a (IRp60) inhibitory receptor is rapidly up-regulated on human neutrophils in response to inflammatory stimuli and modulates CD32a (FcγRIIa) mediated signaling

“…In this model, treatment of undifferentiated HLA-60 cells with RA induces differentiation as detected by the expression of the CD11b receptor, but not the CD14 receptor. When undifferentiated HL-60 cells are induced to differentiate into monocytes both CD11b and CD14 expression is induced (Quesada et al, 1996). Untreated HL-60 cells do not express the CD300a receptor on the cell surface; however, when we induced neutrophil differentiation by adding RA a dramatic increase in CD300a cell surface expression was observed, indicating that the CD300a receptor expression is developmentally regulated (Fig.…”

Translational study of vitamin D differentiation therapy of myeloid leukemia: effects of the combination with a p38 MAPK inhibitor and an antioxidant

“…11 Yet another strategy to obviate this obstacle is to combine relatively low doses of deltanoids with the administration of another agent, such as an antioxidant, which augments the differentiation-inducing action of deltanoids, but does not enhance the levels of circulating calcium. For instance, it was shown that ascorbic acid and vitamin E enhance 1,25D 3 -induced monocytic differentiation of HL60 cells, 12,13 and that the antioxidants carnosic acid (CA), curcumin, ebselen and silibinin potentiate monocytic differentiation induced by low concentrations of 1,25D 3 . 14 Further, the inhibition of the signaling kinase p38 MAPK by its specific inhibitors SB203580 (SB203) or SB202190 (SB202) markedly accelerates monocytic differentiation of HL60 cells induced by low concentrations 1,25D 3 .…”