tretinoin and Leukemia--Basophilic--Acute

Leukotriene C(4) synthase (LTC(4) S) is a pivotal enzyme for generation of cysteinyl-leukotrienes (cysLTs). LTC(4) S activity in rat basophilic leukemia-1 (RBL-1) cells increased after culture in the presence of retinoic acid (RA) analogues, which was inhibited by cycloheximide or actinomycin D (ACD). Unexpectedly, the co-addition of a low dose of ACD with RA further potentiated the upregulation of the LTC(4) S activity. Daunorubicin and mitomycin C also had a similar effect. When stimulated with calcium ionophore A23187, control cells did not produce cysLTs, but RA-treated cells generated cysLTs and the co-addition of ACD further increased. While LTC(4) S mRNA and protein increased in the cells treated with RA, the co-addition of ACD further potentiated both in proportion to the LTC(4) S activity. The effect of ACD was considered to enhance the transcription rate of LTC(4) S gene, but not the mRNA-stability. The addition of methylprednisolone (MP) inhibited generation of cysLTs from the cells with A23187-stimulation and also did LTC(4) S activity, but did not inhibit 5-lipoxygenase (5-LOX). The suppression of LTC(4) S with MP showed a dependent manner on the time-point and duration of MP-treatment after RA-addition which was correlated with reduction in LTC(4) S mRNA and protein. The cells cultured with RA plus ACD contained more histamine, chymase activity, and granules in the cytoplasm than the control cells, suggesting differentiation to mature mast cells. In consideration of RA-differentiation therapy, it may be of pathophysiological relevance that the antineoplastic agents potentiate RA-induced, steroid-sensitive, induction of LTC(4) S in RBL-1 cells.

Topics: Animals; Calcimycin; Cell Differentiation; Cell Line; Cell Size; Chymases; Cycloheximide; Dactinomycin; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Glutathione Transferase; Histamine; Interleukin-4; Leukemia, Basophilic, Acute; Lipoxygenase; Methylprednisolone; Rats; RNA Stability; RNA, Messenger; Serine Endopeptidases; Solubility; Time Factors; Transcription, Genetic; Tretinoin

CD43 (leukosialin, sialophorin), a cell-surface associated mucin that is constitutively expressed at high levels on most leukocytes, is thought to be involved in cell activation and adhesion. We here provide evidence that the vitamin A metabolites all-trans and 13-cis retinoic acid up-regulate CD43 on human leukemic (HMC-1) mast cells, as determined by flow cytometry, Western blot analysis, and by semiquantitative reverse transcriptase-polymerase chain reaction. Enhanced CD43 expression was accompanied by a strong increase in anti-CD43-mediated, LFA-1-dependent homotypic aggregation of HMC-1 cells, demonstrating that newly synthesized CD43 is functionally active in transmitting signals across the plasma membrane which result in enhanced cellular adhesion. CD43 expression was also enhanced in response to retinoic acids on isolated human skin mast cells and human monocytes, but not on cells of the basophilic cell line KU-812 and promyelocytic HL-60 cells, indicating that these agents might act in a cell-type specific manner. These combined result-point to a novel aspect in the regulation of CD43. Possibly, vitamin A metabolites act directly on the CD43 gene, since putative retinoic acid response elements have been detected within its regulatory regions.

Topics: Antigens, CD; Cell Aggregation; Cells, Cultured; HL-60 Cells; Humans; Isotretinoin; Leukemia, Basophilic, Acute; Leukemia, Mast-Cell; Leukosialin; Lymphocyte Function-Associated Antigen-1; Mast Cells; Monocytes; Sialoglycoproteins; Skin; Tretinoin; Tumor Cells, Cultured; Up-Regulation

We investigated inhibitory actions of dexamethasone (DEX) on retinoic acid (RA)-induced enhancement of leukotriene C4 (LTC4) synthesis in rat basophilic leukemia-1 (RBL-1) cells. Cultured cells were preincubated with RA for 16 h with or without DEX, and generation of LTC4 was measured by high performance liquid chromatography in cell-free and intact cell systems. RA (0.1 microgram/ml) significantly potentiated calcium ionophore-stimulated production of LTC4 synthesis. DEX inhibited the RA-induced enhancement of LTC4 synthesis by up to approximately 95% in intact cells when stimulated with calcium ionophore. RA-induced LTC4 synthase activity, which was determined by enzyme assay, was also inhibited by DEX by 65% in a cell-free system. This discrepancy of inhibition between the intact and cell-free systems was due to a partial inhibition of phospholipase A2 activity by DEX in the intact cells. These results indicate that the production of LTC4 is predominantly regulated at a level of LTC4 synthase. The induction of new LTC4 synthase activity by RA and inhibition of the RA-induced activity by DEX are important regulatory mechanisms of LTC4 synthesis.

Topics: Animals; Arachidonate 5-Lipoxygenase; Arachidonic Acid; Calcimycin; Dexamethasone; Glutathione Transferase; Kinetics; Leukemia, Basophilic, Acute; Leukotriene C4; Phospholipases A; Phospholipases A2; Rats; Tretinoin; Tumor Cells, Cultured

Overnight (10-16 h) incubation of retinoic acid (RA), a derivative of vitamin A, specifically induced LTC4 synthase activity (5 to 10-fold), but not LTA4 hydrolase activity in the lysate of rat basophilic leukemia-1 (RBL-1) cells. A time course study revealed that the increase of LTC4 synthase activity was time dependent and that the peak value was obtained after a 24-hour incubation with RA. The induction of enzyme activity was specifically localized to the microsomal fraction. Glutathione (GSH) S-transferase activity measured by using the same cell lysate as an enzyme source and 1-chloro-2,4-dinitrobenzene (DNCB) as a substrate was not influenced by RA treatment, indicating that the induction by RA is specific for membrane-bound LTC4 synthase. The induction of LTC4 synthase may be an important regulatory mechanism of peptide-LT synthesis in allergy and inflammatory diseases.

Topics: Animals; Enzyme Induction; Epoxide Hydrolases; Glutathione Transferase; Leukemia, Basophilic, Acute; Rats; Subcellular Fractions; Time Factors; Tretinoin; Tumor Cells, Cultured

Overnight incubation of rat basophilic leukemia-1 (RBL-1) cells with retinoic acid enhanced calcium ionophore-stimulated syntheses of LTC4 by more than 28-times (from 5.91 +/- 0.31 to 168.31 +/- 22.66 ng/2.10(6) cells) nd LTD4 by more than 7-times (from 5.27 +/- 0.12 to 39.38 +/- 14.89 ng/2.10(6) cells). The stimulatory action first appeared after a 10 h incubation with retinoic acid and was completely abolished by concomitant presence of low concentration cycloheximide (0.5 micrograms/ml) in the medium. However, LTB4 synthesis was dose-dependently inhibited by incubation with retinoic acid. Reduced form of glutathione significantly increased the synthesis of LTC4, but showed no action on the syntheses of LTD4 or LTB4. These results indicate a new enzyme synthesis of LTC4 synthetase.

Topics: Animals; Calcimycin; Cycloheximide; Glutathione; Leukemia, Basophilic, Acute; Leukotriene B4; Rats; SRS-A; Tretinoin; Tumor Cells, Cultured