ucn-1028-c and Prostatic-Neoplasms

Compelling evidence indicates that activation of the JNK/SAPK signaling pathway is obligatory for apoptosis induction by multiple cell stresses that activate the sphingomyelin cycle. Moreover, ectopic expression of bcl-2 can impair apoptosis signaling by most of the cell stresses that activate the ceramide/JNK pathway. Here we show that enforced expression of bcl-2 protects prostate carcinoma cells against the induction of apoptosis by exogenous C2-ceramide. Moreover, enforced bcl-2 expression blocked the capacity of C2-ceramide to activate JNK1, indicating bcl-2 functions at the level of JNK1 or upstream of JNK1 in the ceramide/JNK pathway. The contribution of bcl2 to the regulation of the arachidonate pathway for prostate carcinoma cell survival was also investigated using highly selective inhibitors of arachidonate metabolism. Our results indicate bcl-2 can protect cells against diminished availability of arachidonic acid, 12-HETE, and 15-HETE. Finally, arachidonic acid substantially suppresses the induction of apoptosis by C2-ceramide, providing evidence for the opposing influences of these lipid signaling pathways in the mediation of prostate carcinoma cell survival. These results provide evidence for opposing influences of the ceramide and arachidonate signaling pathways in the mediation of cell death and cell survival, respectively, in prostate carcinoma cells and suggest a dual role for bcl-2 in this context.

Topics: Animals; Apoptosis; Arachidonic Acid; Arachidonic Acids; Calcium-Calmodulin-Dependent Protein Kinases; Cell Survival; Enzyme Activation; Enzyme Inhibitors; Humans; Hydroxyeicosatetraenoic Acids; JNK Mitogen-Activated Protein Kinases; Lipid Metabolism; Male; Mitogen-Activated Protein Kinases; Naphthalenes; Prostatic Neoplasms; Protein Kinase C; Proto-Oncogene Proteins c-bcl-2; Rats; Signal Transduction; Sphingosine; Transfection; Tumor Cells, Cultured

Prostate carcinoma has become the second most fatal cancer in American men. In rat Dunning prostate adenocarcinoma cells, increased cellular motility has been associated positively with their increased metastatic potential. However, the mechanism(s) responsible for regulation of tumor cell motility is poorly understood. We have reported that a lipoxygenase metabolite of arachidonic acid 12(S)-hydroxyeicosatetraenoic acid [12(S)-HETE] augments tumor cell metastatic potential through activation of protein kinase C (PKC). We report here that 12(S)-HETE increased the motility of AT2.1 cells and this 12(S)-HETE increased motility was inhibited by PKC inhibitor calphostin C. Western blot analysis revealed that AT2.1 cells expressed the Ca(2+)-dependent PKC isoform alpha and Ca(2+)-independent PKC isoform delta. Pretreatment of cells with a Ca2+ chelator BAPTA blocked the 12(S)-HETE increased motility. Further, the motility of AT2.1 cells was increased in a dose dependent manner by thymelea toxin, a selective PKC alpha activator. Our data demonstrate that 12(S)-HETE augments the motility of AT2.1 cells via its selective activation of PKC alpha which may serve as a key target for the development of antimetastatic drugs useful for combating prostate cancers.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Adenocarcinoma; Animals; Calcium; Cell Movement; Chelating Agents; Egtazic Acid; Enzyme Activation; Enzyme Inhibitors; Humans; Isoenzymes; Male; Naphthalenes; Prostatic Neoplasms; Protein Kinase C; Protein Kinase C-alpha; Protein Kinase C-delta; Rats; Tumor Cells, Cultured

Prostate carcinoma has become the second most fatal cancer in American men. In Dunning R3327 rat prostate adenocarcinoma cells, elevated invasiveness positively correlates with metastatic potential. However, the mechanism(s) responsible for regulation of tumor cell motility and invasion is poorly understood. We have reported that a lipoxygenase metabolite of arachidonic acid, 12(S)-hydroxyeicosatetraenoic acid [12(S)-HETE], augments tumor cell metastatic potential through activation of protein kinase C (PKC).. We proposed to determine the effect of 12(S)-HETE on the motility and invasion of low-metastatic rat prostate AT2.1 tumor cells and the effect of 12(S)-HETE activation of specific PKC isoform(s) in these processes.. The motility of AT2.1 cells was determined by the colloidal gold phagokinetic track assay and the invasiveness measured as their ability to invade through basement membrane Matrigel-coated filters. Expression of PKC isoforms was determined by Western blotting of the whole cell lysate with isoform-specific anti-PKC antibodies. Cytosol and membrane fractions were prepared and the subcellular distribution of PKC was analyzed by Western blotting and activity assay. The effect of 12(S)-HETE on cell proliferation was examined. Data were analyzed for significance of difference with the two-sampled, two-sided Student's t test.. 12(S)-HETE increased the motility and invasion of AT2.1 cells, and this 12(S)-HETE-increased motility and invasion were inhibited by a selective PKC inhibitor, calphostin C, as well as a Ca2 chelator, bis-(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid/tetra(acetoxy-methyl)ester. AT2.1 cells expressed the PKC isoforms alpha and delta, and 12(S)-HETE increased the membrane association of PKC alpha but not delta. Further, the motility and invasion of AT2.1 cells were increased by thymelea toxin, a selective activator of PKC alpha over PKC delta.. 12(S)-HETE augments the invasiveness of AT2.1 cells via selective activation of PKC alpha.. 12(S)-HETE modulation of PKC alpha invasiveness may be an important mechanism of action for the regulation of the invasive potential of rat prostate carcinoma cells, and the 12-lipoxygenase enzyme and/or PKC alpha may serve as key targets for the development of anti-invasive agents useful for combating the spread of prostate cancer.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; 3T3 Cells; Adenocarcinoma; Animals; Antibiotics, Antineoplastic; Blotting, Western; Cell Division; Cell Movement; Egtazic Acid; Electrophoresis, Polyacrylamide Gel; Hydroxyeicosatetraenoic Acids; Isoenzymes; Male; Mice; Naphthalenes; Neoplasm Invasiveness; Polycyclic Compounds; Prostatic Neoplasms; Protein Kinase C; Rats; Tumor Cells, Cultured