Green tea extract attenuates cyclosporine A-induced oxidative stress in rats.

Cyclosporine A (CsA) nephrotoxicity underweighs the therapeutic benefits of such a powerful immunosuppressant. Whether oxidative stress plays a role in such toxicity is not well delineated. We investigated the potential of green tea extract (GTE) to attenuate CsA-induced renal dysfunction in rats. Three main groups of Sprague-Dawley rats were used: CsA, GTE, and GTE plus CsA-receiving animals. Corresponding control groups were also used. CsA was administered in a dose of 20mg kg(-1) day(-1), i.p., for 21 days. In the GTE/CsA groups, the rats received different concentrations of GTE (0.5, 1.0 and 1.5%), as their sole source of drinking water, 4 days before and 21 days concurrently with CsA. The GTE group was treated with 1.5% concentration of GTE only for 25 days. A concomitant administration of GTE, to CsA receiving rats, markedly prevented the generation of thiobarbituric acid-reacting substances (TBARS) and significantly attenuated CsA-induced renal dysfunction as assessed by estimating serum creatinine, blood urea nitrogen, uric acid and urinary excretion of glucose. A considerable improvement in terms of reduced glutathione content and activity of antioxidant enzymes in the kidney homogenate of the GTE/CsA-receiving rats was observed. The activity of lysosomal enzymes, N-acetyl-beta-glucosaminidase, beta-glucuronidase and acid phosphatase was significantly inhibited following GTE co-administration. Our data prove the role of oxidative stress in the pathogenesis of CsA-induced kidney dysfunction. Supplementation of GTE could be useful in reducing CsA nephrotoxicity in rats. However, clinical studies are warranted to investigate such an effect in human subjects.

Possible role of hydroxyl radicals in the oxidation of dichloroacetonitrile by Fenton-like reaction.

Dichloroacetonitrile (DCAN), is a member of haloacetonitrile group and detected in drinking water supplies as a by-product of chlorination process. The mechanism of DCAN-induced carcinogenesis is believed to be mediated by oxidative bioactivation of DCAN molecules. The present study was designed to investigate if reactive oxygen species (ROS), similar to that generated in biological systems, are capable of oxidative activation of DCAN. A model ROS generation system (Fenton-like reaction; Fe2+ and H2O2) that predominantly produces hydroxyl radical (OH*) was used. DCAN oxidation was monitored by the extent of cyanide (CN-) release. The results indicate that DCAN was markedly oxidized by this system, and the rate of oxidation was dependent on DCAN concentration. Four-fold increase in H2O2 concentration (50-200 mM) resulted in a 35-fold increase in CN- release. The rates of DACN oxidation in presence of various transition metals were in the following order; iron>copper>titanium. DCAN oxidation was enhanced significantly by the addition of vitamin C and sulfhydryl compounds such as glutathione, N-acetyl-L- cysteine, and dithiothreitol (10 mM) to 140, 130, 145 and 136% of control, respectively. Addition of H2O2 scavenger; catalase or iron chelator; desferrioxamine (DFO) resulted in a significant decrease in CN- release 47 and 41% of control, respectively. Addition of various concentrations of the free radical scavengers, DMSO, or mannitol, to the incubation mixtures caused a significant decrease in DCAN oxidation, 32 and 50% of control, respectively. Michaelis-Menten kinetic analysis of the rates of this reaction, with or without inhibitors, indicated that ROS mediated oxidation of DCAN was inhibited by catalase (Ki = 0.01 mM)>DFO (0.02 mM) > mannitol (0.09 mM) > DMSO (0.12 mM). In conclusion, our results indicate that DCAN is oxidized by a ROS-mediated mechanism. This mechanism may have an important role in DCAN bioactivation and DCAN-induced genotoxicity at target organs where multiple forms of ROS generating systems are abundant.

Chloroacetonitrile-induced toxicity and oxidative stress in rat gastric epithelial cells.

Chloroacetonitrile (CAN), is a disinfectant by-product of chlorination of drinking water. Epidemiological studies indicate that exposure to CAN via drinking water might present a potential hazard to human health. The objective of the present work was to investigate the cytotoxic effects as well as the oxidative stress induced by CAN in cultured rat gastric epithelial cells (GECs). GECs were exposed in vitro to different concentrations of CAN (5-40 microm) for 60 min. Also, GECs were incubated with CAN (10 microm) for different time intervals extending to 180 min. Cytotoxicity was determined by assessing cell viability and lactate dehydrogenase (LDH) release, glutathione (GSH) level and lipid peroxidation as indicated by malondialdehyde (MDA) production. Exposure of GECs CAN (10 microm) for 60 min caused a 50% decrease in cell viability and induced an eightfold increase of LDH leakage. In the same experiment, CAN caused a decrease in cellular GSH content to approximately 50% and significantly enhanced MDA accumulation (approx. sevenfold). These toxic responses to CAN were dependent on both concentration and duration of exposure to CAN. There was a good correlation between LDH release and GSH depletion (r =0.96, P<0.05). Treatment of GECs with 5 m mN -acetyl- l -cysteine (NAC) prior to exposure to CAN afforded some degree of protection as indicated by a significant decrease in the LDH leakage (32% of total leakage) and lipid peroxidation (54%) as compared to CAN alone-treated cells. Also, pretreatment of GECs with vitamin C (1 m m) or vitamin E (10 microm) significantly inhibited LDH leakage (20 and 36% of total leakage, respectively). Preincubation with 1 m m desferroxiamine (DFO), a ferric iron chelator, or 10 microm phenanthroline (PHE), a ferrous iron chelator, diminished CAN-induced LDH leakage by 16 and 21% of total leakage, respectively and MDA production by 40 and 44%, respectively. In conclusion, our results suggest that CAN has a potential cytotoxic effect in rat GECs; and thiol group-donors, antioxidants and iron chelators can play a critical role against CAN-induced cellular damage.