thapsigargin and Leukemia--Myeloid

Topics: Animals; Biological Transport; Calcium; Calcium Channels; Calcium-Transporting ATPases; Cells, Cultured; Humans; Islets of Langerhans; Leukemia, Myeloid; Models, Biological; Pituitary Gland; Signal Transduction; Terpenes; Thapsigargin

Using DNA microarray and cluster analysis of expressed genes in a cloned line (M1-t-p53) of myeloid leukemic cells, we have analyzed the expression of genes that are preferentially expressed in different normal tissues. Clustering of 547 highly expressed genes in these leukemic cells showed 38 genes preferentially expressed in normal hematopoietic tissues and 122 other genes preferentially expressed in different normal nonhematopoietic tissues, including neuronal tissues, muscle, liver, and testis. We have also analyzed the genes whose expression in the leukemic cells changed after activation of WT p53 and treatment with the cytokine IL-6 or the calcium mobilizer thapsigargin. Of 620 such genes in the leukemic cells that were differentially expressed in normal tissues, clustering showed 80 genes that were preferentially expressed in hematopoietic tissues and 132 genes in different normal nonhematopoietic tissues that also included neuronal tissues, muscle, liver, and testis. Activation of p53 and treatment with IL-6 or thapsigargin induced different changes in the genes preferentially expressed in these normal tissues. These myeloid leukemic cells thus express genes that are expressed in normal nonhematopoietic tissues, and various treatments can reprogram these cells to induce other such nonhematopoietic genes. The results indicate that these leukemic cells share with normal hematopoietic stem cells the plasticity of differentiation to different cell types. It is suggested that this reprogramming to induce in malignant cells genes that are expressed in different normal tissues may be of clinical value in therapy.

Topics: Animals; Cell Line, Tumor; Cluster Analysis; Gene Expression; Gene Expression Profiling; Genes, p53; Interleukin-6; Leukemia, Myeloid; Mice; Neoplastic Stem Cells; Oligonucleotide Array Sequence Analysis; Thapsigargin; Tissue Distribution

Overexpression of wild-type p53 in M1 myeloid leukemia cells induces apoptotic cell death that was suppressed by the calcium ionophore A23187 and the calcium ATPase inhibitor thapsigargin (TG). This suppression of apoptosis by A23187 or TG was associated with suppression of caspase activation but not with suppression of wild-type-p53-induced expression of WAF-1, mdm-2, or FAS. In contrast to suppression of apoptosis by the cytokines interleukin 6 (IL-6) and interferon gamma, a protease inhibitor, or an antioxidant, suppression of apoptosis by A23187 or TG required extracellular Ca2+ and was specifically abolished by the calcineurin inhibitor cyclosporin A. IL-6 induced immediate early activation of junB and zif/268 (Egr-1) but A23187 and TG did not. A23187 and TG also suppressed induction of apoptosis by doxorubicin or vincristine in M1 cells that did not express p53 by a cyclosporin A-sensitive mechanism. Suppression of apoptosis by A23187 or TG was not associated with autocrine production of IL-6. Apoptosis induced in IL-6-primed M1 cells after IL-6 withdrawal was not suppressed by A23187 or TG but was suppressed by the cytokines IL-6, IL-3, or interferon gamma. The results indicate that these Ca2+-mobilizing compounds can suppress some pathways of apoptosis suppressed by cytokines but do so by a different mechanism.

Topics: Animals; Antineoplastic Agents; Apoptosis; Calcimycin; Calcium; Cell Survival; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; Cyclosporine; Cysteine Endopeptidases; Cytokines; Doxorubicin; Enzyme Inhibitors; fas Receptor; Gene Expression Regulation, Neoplastic; Genes, p53; Kinetics; Leukemia, Myeloid; Mice; Neoplasms, Experimental; Nuclear Proteins; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-mdm2; Recombinant Proteins; Thapsigargin; Transfection; Tumor Cells, Cultured; Tumor Suppressor Protein p53; Vincristine

Sphingosine-1-phosphate (SPP), a metabolite of sphingolipids, has been implicated as a second messenger in cell growth regulation and signal transduction via calcium mobilization from internal stores. This study shows that SPP mobilizes intracellular calcium in U937 cells and demonstrates for the first time the ability of SPP to activate the transcription factor NF-kappa B in these cells. Furthermore, calcium release from the internal stores by thapsigargin (TG), an inhibitor of the endoplasmic reticulum Ca2+ pump, was associated with activation of NF-kappa B. Moreover, we have shown that while an intracellular calcium chelator BAPTA/AM was able to inhibit both SPP- and TG-induced NF-kappa B activation, it had no effect on TNF-induced NF-kappa B activation. In addition, SPP-induced NF-kappa B activation was blocked both by cyclosporin A, known to inhibit calcineurin phosphatase activity, and by the antioxidant butylated hydroxyanisole. These observations suggest that intracellular calcium mobilization is required for SPP-induced NF-kappa B activation, which may involve calcineurin- and redox-dependent mechanisms.

Topics: Antioxidants; Butylated Hydroxyanisole; Calcium; Chelating Agents; Cyclosporine; DNA Probes; Egtazic Acid; Electrophoresis, Polyacrylamide Gel; Enzyme Inhibitors; Fluorescent Dyes; Genes, Reporter; Humans; Indoles; Leukemia, Myeloid; Luciferases; Lysophospholipids; NF-kappa B; Sphingolipids; Sphingosine; Thapsigargin; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha