d-609 and Leukemia--T-Cell

We found that SAPK/JNK was phosphorylated during X-ray-induced rapid cell death of MOLT-4 cells and that acid Sphingomyelinase inhibitor D609 suppressed the rapid cell death as well as phosphorylation of SAPK/JNK. Also C2-ceramide caused phosphorylation of SAPK/JNK, followed by rapid cell death. Further we isolated X-ray-resistant radiation-hybrid clones from MOLT-4 and 50 Gy irradiated mouse FM3A cells by repeated selections with 3 Gy irradiation. One of them named Rh-1a was found resistant to X-ray- as well as C2-ceramide-induced rapid cell death. Rh-1a cells had mouse DNA but no increase in either mouse or human Bcl-2 determined by Western blotting. Accumulation of p53 after X-irradiation was similarly observed in both parental MOLT-4 and Rh-1a cells. However, contrasting to prolonged and prominent phosphorylated status of SAPK/JNK in MOLT-4 cells, Rh-1a cells exhibited short transient increase and FM3A cells showed no increase of phosphorylated status SAPK/JNK after X-irradiation. Therefore, SAPK/JNK activation is considered important in X-ray-induced rapid cell death or apoptosis of MOLT-4 cells.

Topics: Animals; Blotting, Southern; Blotting, Western; Bridged-Ring Compounds; Cell Death; Cell Line; Dose-Response Relationship, Radiation; Enzyme Inhibitors; Humans; Leukemia, T-Cell; Mice; Mitogen-Activated Protein Kinase 8; Mitogen-Activated Protein Kinases; Norbornanes; Phosphodiesterase Inhibitors; Phosphorylation; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; Sphingomyelin Phosphodiesterase; Sphingosine; Thiocarbamates; Thiones; Time Factors; Transfection; Tumor Cells, Cultured; Tumor Suppressor Protein p53; X-Rays

In normal lymphocytes an inside-out signal up-regulating integrin adhesion is followed by a ligand-mediated outside-in cell spreading signal. Protein kinase C (PKC) inhibition blocks lymphocyte adherence to and spreading on fibronectin. In contrast, putative PLC inhibitors yield distinct differences with respect to adhesion and morphology. The phosphatidylinositol-specific phospholipase C (PLC) inhibitor neomycin blocked spreading of CD3/CD28-activated T cells on fibronectin by disrupting adhesion. Furthermore, when an additional inside-out signal for fibronectin adhesion is unnecessary such as with HPB-ALL T leukemic or phorbol-myristate-acetate-treated normal T cells, neomycin treatment does not alter adhesion or morphology. However, the phosphatidylcholine-specific PLC inhibitor D609 abrogates cell spreading without affecting adhesion to fibronectin in these cells as well as the CD3/CD28-activated T cells. These results strongly suggest that inside-out signaling for the integrin alpha4beta1 in lymphocytes proceeds through phosphatidylinositol-specific PLC and PKC, whereas the outside-in signal utilizes phosphatidylcholine-specific PLC and PKC.

Topics: Alkaloids; Benzophenanthridines; Bridged-Ring Compounds; Cell Adhesion; Enzyme Inhibitors; Fibronectins; Humans; Integrins; Leukemia, T-Cell; Lymphocyte Activation; Neomycin; Norbornanes; Phenanthridines; Phosphodiesterase Inhibitors; Protein Kinase C; Protein Synthesis Inhibitors; Sensitivity and Specificity; Signal Transduction; Substrate Specificity; T-Lymphocytes; Thiocarbamates; Thiones; Tumor Cells, Cultured; Type C Phospholipases