oligomycins and Prostatic-Neoplasms

In normal prostate epithelial cells low m-aconitase activity decreases citrate oxidation leading to citrate accumulation. In prostate cancer cells m-aconitase activity is increased and citrate content is lower. The effect of inhibition of m-aconitase on ATP production by prostate cancer cells (PC3) is not known nor is the contribution of glycolysis versus respiration.. ATP content of PC3 cells as affected by inhibition of m-aconitase (fluoroacetate (FA), zinc), inhibition of glycolysis (2DxG), or respiration (DNP, oligomycin) was determined. The ability to maintain ATP using glucose or glutamine as sole substrate was also determined. Intermediates including ATP, lactate, glucose, and glutamine were assayed in neutralized perchloric acid (PCA) cell extracts, virgin, and conditioned medium by enzymatic fluorometry.. Data show that inhibition of m-aconitase, glycolysis, or respiration alone did not decrease ATP content. Inhibition of both glycolysis and respiration were required to decrease ATP content. PC3 cells were able to produce ATP with either glucose or glutamine as sole substrate. Though FA clearly inhibited m-aconitase there was no evidence that zinc had a similar effect.. PC3 cells can support ATP production when m-aconitase is inhibited by using glycolysis or oxidation of substrate (e.g., glutamine) entering the TCA cycle distal to citrate.

Topics: Aconitate Hydratase; Adenosine Triphosphate; Antimetabolites; Cell Line, Tumor; Citric Acid; Deoxyglucose; Dinitrophenols; Enzyme Inhibitors; Fluoroacetates; Glycolysis; Humans; Male; Oligomycins; Oxygen Consumption; Prostatic Neoplasms; Zinc

Ca(2+) plays a fundamental role in the control of a variety of cellular functions, in particular, in energy metabolism and apoptosis. In this study, we show that TGF-beta at concentrations of 0.1-1.0 ng/ml transiently increases the level of intracellular Ca(2+) ([Ca(2+)](in)) in human prostate carcinoma, PC-3U, cells. Experiments with mitochondrial inhibitors (oligomycin and antimycin A) and an inhibitor of endoplasmic reticulum Ca(2+) uptake (BHQ) implied that the effect of TGF-beta1 was due to an effect on the mitochondria. TGF-beta1 treatment resulted in a decrease in ATP synthesis and to a depolarisation, leading to a release of Ca(2+) from mitochondria and decreased activity of the Ca(2+) pumps. Analysis of the mitochondria within the PC-3U cells by polarography and membrane potential-sensitive dye (Rhodamine 123) confirmed that under these experimental conditions, TGF-beta1 inhibited ATP synthesis and depolarised the mitochondria. The results implicate that TGF-beta1 affects the function of the mitochondria and may be of significance for the understanding of the proapoptotic effect of TGF-beta1 in these cells.

Topics: Antimycin A; Calcium; Calcium-Transporting ATPases; Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; Carcinoma; Cell Line; Endoplasmic Reticulum; Energy Metabolism; Enzyme Inhibitors; Homeostasis; Humans; Male; Membrane Potentials; Mitochondria; Oligomycins; Oxygen; Prostatic Neoplasms; Transforming Growth Factor beta; Transforming Growth Factor beta1; Uncoupling Agents

The effects of mitochondrial uncoupling on the calcium homeostasis of prostatic cells were investigated using the prostatic cancer cell line LNCaP and indo-1 spectrofluorimetry. Carbonyl cyanide m-chloro-phenylhydrazone (CCCP) was used as uncoupler. Resting LNCaP cells responded to CCCP by a biphasic increase in [Ca2+]i. The first phase of increase which corresponded to the release of a mitochondrial CCCP-sensitive Ca2+ store was followed by a second increase phase consisting of Ca2+ influx through the plasma membrane. The relationship between the CCCP- and the InsP3-sensitive stores was investigated using thapsigargin (TG). The release part of the Ca2+ response to TG was reduced in a time-dependent manner by previous exposure of the cells to CCCP, suggesting that CCCP also acts on non-mitochondrial stores. Our results show that CCCP releases Ca2+ from both mitochondrial and non-mitochondrial stores in prostatic cells. The possible mechanisms of these effects are discussed.

Topics: Adenosine Triphosphate; Antimycin A; Calcium; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Cell Membrane; Enzyme Inhibitors; Fluorescent Dyes; Humans; Indoles; Male; Mitochondria; Oligomycins; Prostatic Neoplasms; Spectrometry, Fluorescence; Thapsigargin; Time Factors; Tumor Cells, Cultured; Uncoupling Agents