Synergistic Differentiation of U937 Cells by All-Trans Retinoic Acid and 1α,25-Dihydroxyvitamin D3 Is Associated with the Expression of Retinoid X Receptor α

1994 ◽  
Vol 203 (1) ◽  
pp. 272-280 ◽  
Author(s):  
H. Defacque ◽  
T. Commes ◽  
C. Sevilla ◽  
C. Rochetteegly ◽  
J. Marti
Keyword(s):  
Retinoic Acid ◽  
Retinoid X Receptor ◽  
U937 Cells ◽  
Dihydroxyvitamin D3 ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid

The Role of Transcriptional Coactivator TRAP220/MED1 in Nuclear Receptor-Mediated Myelomonocytic Differentiation.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2727-2727
Author(s):  
Mitsuhiro Ito ◽  
Norinaga Urahama ◽  
Akiko Sada ◽  
Kimikazu Yakushijin ◽  
Katsuya Yamamoto ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Nuclear Receptors ◽  
Nuclear Receptor ◽  
Precursor Cells ◽  
Mediator Complex ◽  
Dihydroxyvitamin D3 ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Hematopoietic Precursor

Abstract The TRAP/Mediator complex, the metazoan counterpart of the yeast Mediator complex, is master transcriptional regulatory complex composed of approximately 30 subunits. It was originally isolated as a thyroid hormone receptor (TR)-associated protein (TRAP) complex that mediates TR-activated transcription from DNA templates in vitro and probably acts in vivo after the action of other receptor-interacting coactivators involved in chromatin remodeling. The TRAP220/MED1 subunit of the TRAP/Mediator complex is proposed to act on a variety of major and specific biological events, including growth, differentiation and homeostasis, through physical interaction with nuclear receptors. The vitamin D receptor (VDR) and retinoic acid receptor (RAR), coupled with retinoid X receptor (RXR), are nuclear receptors which have important roles for monopoiesis and granulopoiesis, respectively. In this study, we present the functional role of TRAP220/MED1 in nuclear receptor-mediated monopoiesis and granulopoiesis. Since TRAP220 knockout (Trap220-/-) mice were mortalities during the early embryonic period before definitive hematopoiesis within the hepatic primordia becomes dominant, the function of TRAP220/MED1 in adult hematopoiesis was mostly unknown. However, these embryos appeared to have normal composition of nucleated erythroid cells. Therefore, the E9.0 yolk sac-derived hematopoietic precursor cells were used to differentiate into definitive hematopoietic colony forming units within the methylcellulose blood cell culture. The number of monocytic colonies (CFU-M) was significantly lower in knockouts than in wild type controls, while the numbers of other types of colonies (CFU-GEMM, CFU-GM, CFU-G and CFU-E) were comparable. Hence, TRAP220/MED1 appeared to be indispensable for optimal monocytic differentiation. Next, the HL-60 acute promyelocytic leukemia cells were used to elucidate directly and mechanistically the roles of TRAP220/MED1 in RAR- and VDR-dependent differentiation of the hematopoietic precursor cells into granulocytic and monocytic lineage cells. The expression of the TRAP220/MED1 subunit as well as other TRAP/Mediator subunits was induced when the cells were treated with their ligands, all-trans retinoic acid and 1,25-dihydroxyvitamin D3. Flow cytometric analyses showed that HL-60 cells, wherein TRAP220/MED1 was down-regulated, did not differentiate efficiently into monocytes and granulocytes by stimulation with 1,25-dihydroxyvitamin D3 and all-trans retinoic acid, correspondingly. The expression of direct target genes of VDR or RAR, as well as the differentiation marker genes, was low in the knockdown cells by stimulation with these ligands. In contrast, 12-O-tetradecanoylphorbol-13-acetate (TPA)- and dimethylsulphoxide (DMSO)-mediated monocytic and myeloid differentiation, which bypasses nuclear receptor-mediated signaling pathways, was not affected in knockdown cells. Collectively, these results indicated an indispensable role of TRAP220/MED1 in the optimal VDR- and RAR-mediated myelomonocytic differentiation processes in mammalian hematopoiesis.

Cooperative Action of 1alpha,25-Dihydroxyvitamin D3 and Retinoic Acid in NB4 Acute Promyelocytic Leukemia Cell Differentiation Is Transcriptionally Controlled.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4247-4247
Author(s):  
Jean-Noel Bastie ◽  
Nicole Balitrand ◽  
Isabelle Guillemot ◽  
Christine Chomienne ◽  
Laurent Delva
Keyword(s):  
Retinoic Acid ◽  
Transcriptional Activity ◽  
Target Genes ◽  
Leukemia Cell ◽  
Leukemic Cells ◽  
Dihydroxyvitamin D3 ◽  
Nb4 Cells ◽  
Cooperative Action ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid

Abstract All-trans-retinoic acid (RA) and 1alpha,25-dihydroxyvitamin D3 (1,25D3) are involved in the control of hematopoiesis and have been suggested to play a role in cellular differentiation and are as such potent inducers of differentiation of myeloid leukemia cells. In this study, we have shown that in promyelocytic NB4 cells, addition of 1,25D3 enhances terminal granulocytic RA-dependent differentiation concomitant with the enhanced activation of the RA-transcriptional activity through an RARbeta promoter. By EMSA and ChIP assays, we further demonstrate that while both VDR and RAR are bound to the RARbeta promoter in NB4 cells, addition of 1,25D3 increases VDR binding to this promoter while that of RA induces the release of VDR and increases the binding of RAR. Thus, contrary to normal myeloid cells, 1,25D3 does not act as a transrepressor of RA-transcriptional activity in leukemic cells suggesting that transcriptional regulation of RA-target genes may be modified in malignant cells. In promyelocytic leukemic cells the combination of 1,25D3 and RA results in both enhanced transactivation and differentiation.

Up-regulation of steroid sulphatase activity in HL60 promyelocytic cells by retinoids and 1α,25-dihydroxyvitamin D3

2001 ◽  
Vol 355 (2) ◽  
pp. 361-371 ◽  
Author(s):  
Philip J. HUGHES ◽  
Lucy E. TWIST ◽  
Jennifer DURHAM ◽  
M. Ansar CHOUDHRY ◽  
Mark DRAYSON ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Colon Adenocarcinoma ◽  
Cell Types ◽  
Retinoid Receptor ◽  
Dihydroxyvitamin D3 ◽  
Hl60 Cells ◽  
Steroid Sulphatase ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid ◽  
Multiple Cell

HL60 promyeloid cells express both classes of oestrogen receptor (ERα and ERβ). We show that hydrolysis of oestrone sulphate by steroid sulphatase is a major source of oestrone in HL60 cells, and that most of the released oestrone is not metabolized further to 17β-oestradiol. Treatment of HL60 cells with retinoids or 1α,25-dihydroxyvitamin D3 increased steroid sulphatase mRNA and activity in parallel with the induction of CD11b, an early marker of myeloid differentiation that is expressed before the differentiating cells stop proliferating. Use of agonists and antagonists against retinoid receptor-α and retinoid receptor-X revealed that both classes of retinoid receptor can drive steroid sulphatase up-regulation. Steroid sulphatase activity fluctuates during the cell cycle, being highest around the transition from G1 to S phase. During the differentiation of HL60 cells induced by all-trans-retinoic acid or 1α,25-dihydroxyvitamin D3, there is increased conversion of 17β-oestradiol into oestrone by an oxidative 17β-hydroxysteroid dehydrogenase. Treatment of Caco-2 colon adenocarcinoma cells with all-trans-retinoic acid or 1α,25-dihydroxyvitamin D3 also increases 17β-oestradiol oxidation to oestrone. An increase in local oestrone production therefore occurs in multiple cell types following treatment with retinoids and 1α,25-dihydroxyvitamin D3. The possible involvement of locally produced oestrogenic steroids in regulating the proliferation and differentiation of myeloid cells is discussed.

scholarly journals Cooperative Effects of a Dnmt Inhibitor and All-Trans Retinoic Acid in an AML Cell Line Model Lacking PML-Rara

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 1359-1359
Author(s):  
Ruth Meier ◽  
Gabriele Greve ◽  
Christoph Niemöller ◽  
Heiko Becker ◽  
Tobias Ma ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Colony Formation ◽  
Study Drug ◽  
Interferon Response ◽  
Cdna Libraries ◽  
U937 Cells ◽  
Go Analysis ◽  
Retinoic Acid Response Element ◽  
Trans Retinoic Acid ◽  
All Trans Retinoic Acid

Abstract Introduction: All-trans retinoic acid (ATRA, RA) has powerful activity in acute promyelocytic leukemia (APL); its efficacy in non-APL acute myeloid leukemia (AML) is still unclear, but may be enhanced by epigenetic drugs such as azanucleoside DNMT inhibitors (Blagitko-Dorfs et al. PLoS ONE 2013). In a randomized phase II study (DECIDER trial, NCT00867672) the addition of RA to decitabine (DAC) in newly diagnosed non-fit older AML patients resulted in a clinically meaningful extension of survival. We hypothesize that the add-on of RA to DAC results in cooperative transcriptome changes (possibly associated with demethylation) which may, at least in part, explain these clinical results. Materials and Methods: U937 cells were treated in triplicates with daily pulses of 200 nM DAC, 1 µM RA was administered after 48 hours (hr). Cells were harvested after 72 and 120 hr of treatment. Proliferation and apoptosis were determined by acridine orange / propidium iodide staining. For the colony formation assay (CFA), 100 cells were seeded after 120 hr treatment in methylcellulose, colonies were counted after 21 days. Stranded cDNA libraries were sequenced on an Illumina HiSeq2000 with 60 million 125 bp paired-end reads. Read quality was checked with FastQC (per base Q-score >38), alignment to the GencodeCompV24 reference genome with RNAStar, generation of count matrices with ht-seq and differential expression testing with DeSeq2. Transcript changes were considered significant with an adjusted FDR p-value < 0.01. GO analysis was performed with Metascape. Methylomes were generated using Infinium Human Methylation 450K BeadChip arrays and differential methylation data was obtained using the RnBeads package for R (Assenov et al. Nature Methods 2014). Results: In suspension culture, RA alone had no effect on the proliferation of U937 cells, DAC alone reduced proliferation by 74 %, the DAC+RA combination by 81 % (with viability always ≥ 90 %). RA alone also had no effect on colony growth, DAC reduced colony formation by 85 %, and DAC+RA decreased colony numbers by 96 %. To determine transcriptional add-on effects of DAC+RA, transcriptomes of cells treated for 72 hr were analyzed by RNA-Seq. RA alone had a modest effect on transcription, whereas DAC alone induced / downregulated 2538 and 620 transcripts, DAC+RA induced / downregulated 2848 and 1173 transcripts, respectively (Table 1, Fig. 1). 29.6 % (1192 transcripts) of all DAC+RA-altered transcripts were uniquely regulated by the combination and not by DAC or RA alone. GO analysis of the DAC+ATRA alone upregulated genes (664 transcripts) revealed a significant enrichment for immune system and cellular decay terms, downregulated genes were enriched for transcription and cell cycle terms. Of the transcripts already up- or downregulated by DAC or RA alone, 55.5 % of were further enhanced by DAC+RA. Among those, the retinoic acid receptors RARA, RARB and the retinoic acid response element (RARE)-containing tumor suppressor HIC-1 (Hassan et al. Cell Reports 2017) were induced 8.6-, 8.3- and 179.8-fold, respectively, compared to 2.4- and 5.3-fold (RARA and RARB, DAC alone) and 26.5-fold (HIC-1, RA alone) (validated by qRT-PCR). Among the most regulated uniquely induced transcripts by DAC+RA, the RA-regulating cytochrome P450 member CYP26A1 (regulated by multiple RAREs) was upregulated 347.3-fold. In order to further investigate this add-on effect of RA on DAC-induced transcriptome changes, DNA methylation was analyzed (72 and 120 hr). As expected, DAC significantly reduced CpG methylation in gene bodies and promoters, without further demethylation upon RA add-on after 72 hr (after 120 hr, a trend towards further demethylation was noted with DAC+ATRA). Conclusion: In vitro, the combination of DAC with RA resulted in enhanced growth inhibition and reduced colony formation compared to DAC alone. Transcriptome analysis revealed a high number of uniquely regulated transcripts, and an enhancement of regulation by the combination treatment. As expected, this also included genes with RAREs. The enrichment for genes involved in immune response (e.g. multiple interferon-response genes) encourages the combination of DNMTi-based therapies with immunotherapeutic agents. Therefore, further transcriptome analyses of transposable elements, when reactivated, may trigger an interferon-mediated immune reaction, are currently ongoing. Disclosures Lübbert: TEVA: Other: Study drug; Celgene: Other: Travel Support; Cheplapharm: Other: Study drug; Janssen: Honoraria, Research Funding.

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