Prooxidant-antioxidant shift induced by androgen treatment of human prostate carcinoma cells.

BACKGROUND Prostate cancer is a disease associated with aging. Also commonly associated with increasing age is a shift in the prooxidant-antioxidant balance of many tissues toward a more oxidative state, i.e., increased oxidative stress. We hypothesize that androgen exposure, which has long been associated with the development of prostate cancer, may be a means by which the prooxidant-antioxidant balance of prostate cells is altered. PURPOSE Using established prostate carcinoma cell lines, we studied the effect of androgens on various parameters of oxidative state (e.g., generation of hydrogen peroxide and hydroxyl radicals, lipid peroxidation, and oxygen consumption) and antioxidant defense mechanisms (e.g., the glutathione system and catalase). METHODS The androgen-responsive LNCaP and the androgen-independent DU145 prostate carcinoma cell lines were exposed to 5 alpha-dihydrotestosterone (DHT) and to the synthetic androgen R1881. The cellular proliferation responses were measured by use of a fluorometric assay to quantitate the amount of DNA. The generation of reactive oxygen species was measured by use of 2',7'-dichlorofluorescin diacetate, a dye that fluoresces in the presence of hydrogen peroxide or hydroxyl radicals. Lipid peroxidation was quantitated by use of a chromogen specific for malonaldehyde and 4-hydroxy-2(E)-nonenal. General mitochondrial activity was determined by assaying 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction. A Clark-type electrode was used to assess oxygen consumption per cell. Intracellular glutathione concentrations and the activities of catalase and gamma-glutamyl transpeptidase were measured spectrophotometrically. All P values resulted from two-sided tests. RESULTS DHT at less than 1 to 100 nM (a concentration range encompassing the physiologic levels of DHT considering all ages) and R1881 at 0.1-1 nM concentrations were effective in inducing in LNCaP cells comparable proliferative responses and changes in oxidative stress. In contrast, neither DHT nor R1881 had any effect on the oxidative stress in DU145 cells. The mitochondrial activity in LNCaP cells, as measured by MTT reduction, was significantly elevated above the levels of the untreated controls by DHT (0.1-1000 nM) and R1881 (0.05-1 nM) (P < .001 in both). Oxygen consumption and catalase activity were increased in LNCaP cells in the presence of 1 nM R1881 by 60% and 40%, respectively, over the values in the untreated control cells (P < .03 and P < .01, respectively). The same concentration of R1881 resulted in a decrease in intracellular glutathione concentrations and an increase in gamma-glutamyl transpeptidase activity in LNCaP cells. Treatment with the oxidizing agents H2O2 and menadione produced an increase in gamma-glutamyl transpeptidase activity in LNCaP cells, whereas treatment with the antioxidant compound ascorbic acid (100 mM) reduced the oxidative stress produced in LNCaP cells by 1 nM R1881 and completely blocked the gamma-glutamyl transpeptidase activity. CONCLUSIONS Physiologic levels of androgens are capable of increasing oxidative stress in androgen-responsive LNCaP prostate carcinoma cells. The evidence suggests that this result is due in part to increased mitochondrial activity. Androgens also alter intracellular glutathione levels and the activity of certain detoxification enzymes, such as gamma-glutamyl transpeptidase, that are important for maintenance of the cellular prooxidant-antioxidant balance.