Differential expression of the lenticular antioxidant system in laboratory animals: a determinant of species predilection to oxidative stress-induced ocular toxicity?

PURPOSE: Various animal species have been used to study oxidative stress-induced cataractogenesis; however, given that differences in the expression of the lens antioxidant system may influence species susceptibility to oxidative stress, we compared and contrasted a broad spectrum of components of the lens antioxidant system in dog, rat, marmoset, and rabbit. METHODS: Lenses collected from beagle dogs, Sprague-Dawley rats, marmosets, and New Zealand white rabbits were assayed for reduced glutathione (GSH), and activities of copper-zinc and manganese superoxide dismutase (CuZn-SOD; Mn-SOD), catalase (CAT), glutathione peroxidase (GPX), gamma-Glutamylcysteine synthetase (GCS), glutathione reductase (GR), glutathione-S-transferases (GST), and glucose-6-phosphate dehydrogenase (GPDH), and Trolox equivalent antioxidant capacity (TEAC). RESULTS: Expression of the lens antioxidant system varied considerably between species. Marmoset lens contained the highest levels of GSH, its respective biosynthetic and recycling enzymes GCS and GR, and the associated H(2 )O(2)-dismutation enzyme GPX. Activities of both SOD isoforms were also highest in marmoset lens. However, activities of the xenobiotic-conjugating enzyme GST and NADPH-generating enzyme GPDH were relatively low. In contrast, dog lens had the lowest levels of GSH, GCS, GR, and Cu-Zn SOD (1/2, 1/2 and 1/33, and 1/63 that in marmoset) but highest levels of GST and GPDH. Rabbit lens contained the highest CAT activity, at up to 3.5-fold that for marmoset and rat. CONCLUSION: These results demonstrate substantial variation in lens antioxidant systems between different laboratory animal species. Given that such variation may affect relative susceptibility to oxidative stress-mediated ocular toxicity, our findings may provide useful information when choosing different animal species for lens research.

Validation of MRI measurement of cardiac output in the dog: the effects of dobutamine and minoxidil.

The use of magnetic resonance imaging (MRI) for the measurement of cardiac output parameters in anesthetized adult male beagle dogs has been validated against a widely accepted thermodilution method. Using a multislice cine gradient echo MRI method to acquire images of the entire heart, left ventricular lumen volumes were measured at systole and diastole in seven animals. Cardiac output correlated well (R 2 = 0.88) with thermodilution measurements made in a parallel manner, both before and during acute stimulation with the inotrope dobutamine. In a chronic study of changes in cardiac morphology and function brought about by the antihypertensive minoxidil, MRI reliably detected the expected increases in stroke volume (28%) and cardiac output (58%) resulting from neural reaction to decreased blood pressure. Left ventricular lumen enlarged as well in response to fluid retention and plasma volume increase. Two in four minoxidil-treated animals also developed clear MRI-visible pericardial effusion.

Advantages of glutamate dehydrogenase as a blood biomarker of acute hepatic injury in rats.

In a recent study in rats, alanine aminotransferase (ALT), the preferred plasma biomarker of hepatocellular injury in rats, was ineffective at detecting marked hepatic necrosis produced by acetaminophen (Human and Experimental Toxicology 19, 277-83, 2000). In contrast, glutamate dehydrogenase (GLDH) was markedly elevated. Accordingly, these enzymes were comprehensively evaluated as plasma biomarkers of hepatocellular injury in rats using several other models of hepatic injury, including partial hepatectomy and exposure to methapyrilene, dexamethasone, cyproterone, isoniazid, lead nitrate, and Wyeth-14643. Other enzymes also evaluated were aspartate aminotransferase (AST), sorbitol dehydrogenase (SDH), and the hepatobiliary marker alkaline phosphatase (ALP). Compared to plasma ALT increases, plasma GLDH increases were up to 10-fold greater, up to 3-fold more persistent, and occurred at times following hepatocellular injury when plasma ALT was not increased. Plasma GLDH activity was not inhibited by the test compounds, whereas ALT was substantially inhibited by both isoniazid and lead nitrate. While plasma GLDH activity was unaffected by induction, ALT was induced by cyproterone and dexamethasone, and ALP was induced by Wyeth-14643 and partial hepatectomy. GLDH was concluded to be a more effective biomarker of acute hepatic injury than ALT, AST, SDH or ALP in the rat, based primarily on the large increase following hepatocellular injury, prolonged persistence in the blood following injury, high sensitivity for detection of injury (including pre-necrotic injury), high tissue specificity, and lower susceptibility to inhibition or induction.

Effect of diquat on the antioxidant system and cell growth in human neuroblastoma cells.

Oxidative stress elicits an adaptive antioxidant response, which varies with tissue type. Diquat, a potent redox cycler that generates reactive oxygen species, has been used to study oxidative stress; however, its effect on the antioxidant system has not been characterized in neuronal cells. Accordingly, we measured antioxidant parameters and cell growth in human neuroblastoma SH-SY5Y cells cultured for 48 h in medium containing 5, 10, or 25 microM diquat dibromide or phosphate-buffered saline. Viable cells were assayed for glutathione (GSH) and activities of catalase (CAT), superoxide dismutase (SOD), glutathione reductase (GR), glutathione peroxidase (GPX), and glucose-6-phosphate dehydrogenase (GPDH). Mitochondrial function was evaluated by glutamate dehydrogenase (GDH) activity and MTT reduction. Diquat caused a marked concentration-related decrease in viable cell count ( by 26, 51, and 87% at 5, 10, and 25 microM diquat). Cell viability was only affected at 10 and 25 microM diquat and did not fully account for the decreased viable cell count. Concentration-related increases also occurred with GSH levels and a majority of antioxidant enzymes activities; however, the mode and magnitude varied with parameter. Increases in GSH, CAT, SOD, and GR were maximal at 25 microM diquat (to 3-, 6-, 2-, and 1.5-fold control values, respectively). GPDH activity was maximal at 10 microM diquat and then decreased to 86% of control activity at 25 microM diquat. GPX activity showed a concentration-related decrease (to 35% of control). Activity of the mitochondrial enzyme GDH increased 3-fold at 25 microM diquat, along with a lesser increase in MTT reduction. We conclude that diquat reduces cell growth in neuroblastoma cells and induces an adaptive antioxidant response, which are concentration dependent and occur at sublethal concentrations. At higher concentrations, diquat alters mitochondrial function and becomes increasingly toxic.