Monocytic Differentiation of Acute Promyelocytic Leukemia Cells in Response to 1,25-Dihydroxyvitamin D3 Is Independent of Nuclear Receptor Binding

Abstract

We have shown that 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) primes NB4 cells, the only available acute promyelocytic leukemia cell line, for 12-O-tetradecanoyl-phorbol-13-acetate-induced monocytic differentiation. Here, we have used isomers of 1,25(OH)2D3 to investigate the role of 1,25(OH)2D3 and its putative nuclear receptor (VDR) in NB4 cell monocytic differentiation. 1 beta,25-dihydroxyvitamin D3 (HL), a specific antagonist of only the nongenomic signals of 1,25(OH)2D3, attenuated the priming effect of 1,25(OH)2D3. The 6-cis conformer of 1,25(OH)2D3 (HF), which is unable to bind to VDR, was 20 times more potent than 1,25(OH)2D3 as a priming agent for monocytic differentiation. This response was also blocked by the HL antagonist. Unlike myelocytic HL-60 cells, which respond to 1,25(OH)2D3 with increases in VDR expression and monocytic differentiation, neither HF nor 1,25(OH)2D3 regulated VDR expression in NB4 cells. In the monocytic differentiation of acute promyelocytic leukemia cells, 1,25(OH)2D3 appears to signal through a pathway independent of VDR/VDRE action.

Expression of NADPH oxidase is induced by all-trans retinoic acid but not by phorbol myristate acetate and 1,25 dihydroxyvitamin D3 in the human promyelocytic cell line NB4

“…8,16,17,20 This cell line is not effectively induced into mature monocytic cells when these agents are used alone, but effective maturation has been observed when they are combined. 8,17,18,20,27 Here, we show that NB4 cells exposed to PMA or PMA + VD3 were unable to generate O − 2 resulting from the lack of expression of p22phox and p91phox, the membrane-associated subunits of the oxidase. Unlike HL60 cells which respond to VD3 through monocytic differentiation, O − 2 production 28 and increase in vitamin D receptor expression, 18 VD3 or VD3 + PMA-induced monocytic differentiation of NB4 cells occurs independently of VD3 binding to its nuclear receptor.…”

“…Unlike HL60 cells which respond to VD3 through monocytic differentiation, O − 2 production 28 and increase in vitamin D receptor expression, 18 VD3 or VD3 + PMA-induced monocytic differentiation of NB4 cells occurs independently of VD3 binding to its nuclear receptor. 18 Since VD3 does not induce the expression of NADPH oxidase activity in NB4 cells but does induce it in HL60 cells, 28 this suggests that the presence of a functional NADPH oxidase in leukemic cells exposed to VD3 is the result of genomic actions of VD3.…”

“…[16][17][18][19][20] PMA was solubilized in DMSO (1 mg/ml) and kept at −80°C. 1,25-dihydroxy vitamin D3 (kindly provided by U Fischer and P Weber, Hoffmann-La Roche, Basel, Switzerland) was solubilized in ethanol (240 M) and kept in glass tubes under nitrogen at −20°C.…”

“…Several recent reports mention that NB4 cells show bilineage potential with expression of monocyte-macrophage markers in response to inducers of monocytic differentiation such as 1,25 dihydroxyvitamin D3 (VD3) and phorbol myristate acetate (PMA). [16][17][18][19][20] We have therefore addressed the question of whether NADPH oxidase is expressed and functional in NB4 cells exposed to ATRA or to inducers of monocytic differentiation.…”

Characterization of the chimeric retinoic acid receptor RARα/VDR

“…66,67 Recent evidence suggests any D 3 effects in NB4 cells are independent of nuclear receptor binding. 68 The subline NB4.306, is resistant to both tRA and D 3 , 66 and does not express a functional PML/RAR␣. 69 However, NB4.306 cells do express RAR␣ and appear to retain some responsiveness to tRA, such as CD18 expression.…”