Copper Toxicity and Lipid Peroxidation in Isolated Rat Hepatocytes: Effect of Vitamin E

Abstract

ABSTRACT. We investigated the role of lipid peroxidation as the mechanism mediating copper toxicity in isolated rat hepatocytes and the modulating effect of vitamin E. Hepatocytes, isolated from rats fed diets containing deficient (E-), sufficient (E+), and excess (E++) amounts of vitamin E, were incubated with CuClz (0-2400 pM) for 150 min. Dose and time-dependent decreases in hepatocyte viability (determined by trypan blue exclusion and lactate dehydrogenase release) due to copper toxicity correlated with production of malonyldialdehyde in E-and E+ hepatocytes. However, malonyldialdehyde generation did not accompany copper toxicity in E++ cells. Copper toxicity was enhanced in E-compared to E+ and E++ hepatocytes as assessed by cell viability studies and ultrastructural plasma membrane bleb formation.

Evidence for involvement of oxygen free radicals in bile acid toxicity to isolated rat hepatocytes

“…Although this oxygen concentration in the gas mixture may appear high and nonphysiological, it must be pointed out that the oxygen delivered to the hepatocyte in uiuo is mainly ( > 90%) delivered from hemoglobin-bound oxygen (which is absent in our system) and not from that dissolved in plasma. For these reasons, the standard gas mixture used to maintain isolated hepatocyte integrity in in uitro suspensions has been a 95% 0,/5% CO, mixture (58), even in studies of free radical-induced hepatocyte injury (21,(59)(60)(61)(62). However, because of concern that hepatocytes were exposed to hyperoxia under these conditions, experiments were conducted under lower, more physiological 0, concentrations.…”

“…Under subdued light, 4 x lo6 hepatocytes were washed in tocopherol-free incubation medium containing BSA, then washed twice in BSA-free, tocopherol-free PBS and stored in a 2 : 1 mixture of PBS and 1% ascorbic acid (in ethanol) at -70" C and protected from light. Within 2 mo of storage, after saponification with KOH and extraction into hexane, a-and y-tocopherol contents of hepatocfles were measured on HPLC with fluorescence detection (205 nm for excitation and 340 nm for emission wavelengths), as previously described (21,25). Hepatocyte a-tocopherol content was stable for at least In a second set of experiments, a series of free radical scavengers and inhibitors were individually added to 50-ml aliquots of hepatocytes for 30-min preincubation before addition of bile acids.…”

“…Aliquots of hepatocytes were removed from each incubation every 30 to 60 min and analyzed for viability (trypan blue exclusion) and for generation of lipid peroxidation products by a modification of the thiobarbituric acid (TBA)-reacting substances (TBARS) method (24), as previously described (21). For trypan blue exclusion, 200 cells were examined microscopically on a hemocytometer for uptake of trypan blue into the nucleus after 2 min incubation of 50 p1 cell suspension in 50 pl of 0.2% trypan blue.…”

Antioxidant, pro‐oxidant effect of the thiosemicarbazone derivative Schiff base (4‐(1‐phenylmethylcyclobutane‐3‐yl)‐2‐(2‐hydroxybenzylidenehydrazino) thiazole) and its metal complexes on rats

“…48 Copper coordination compounds with TSC and SB ligands have been extensively studied in recent years. 18,[49][50][51][52][53][54][55][56][57][58][59][60][61] Some of them are known to produce severe cytotoxicity with generation of reactive oxygen species, 56,57 others have been used as efficient catalysts for the oxidation of various substrates.…”

The Effect of the Serum Amino Acid Levels Thiosemicarbazone Derivatives and its Metal Complexes on Rats

“…Copper is an essential element that contributes to important intracellular metabolic events [34], but is also a latent toxic element that leads to chemical element poison by excess accumulation [35] and is a potent hepatotoxin [36].…”