M. Kihlström

Age- and sex-related differences in lipid peroxidation of mouse cardiac and skeletal muscles

1. The purpose of the present study was to characterize age- and sex-related changes in lipid peroxidation capacities and enzymatic antioxidants of cardiac and skeletal muscles in NMRI-mice (Mus musculus). 2. Lipid peroxidation rates (unstimulated and enzymatic/iron-stimulated) strongly decreased in skeletal muscle during ageing. 3. Unstimulated lipid peroxidation rate but not that of stimulated, also decreased in cardiac muscle. 4. The total level of Fe2+/ascorbate-stimulated non-enzymatic lipid peroxidation was not, however, affected by ageing. 5. The activity of catalase slightly increased in cardiac muscle and that of glutathione peroxidase in skeletal muscle during ageing. 6. Unstimulated lipid peroxidation rate was significantly higher in the skeletal muscle of male than female mice. 7. Correspondingly, the Fe2+/ascorbate-stimulated lipid peroxidation capacities of microsomal and mitochondrial fractions of skeletal muscle were significantly higher in male mice. 8. The activity of glutathione peroxidase as well as the concentration of lipofuscin were higher in the cardiac muscles of female than male mice.

Chronic hydrogen peroxide intake and peroxide metabolizing enzyme activities in some tissues of mice and rats.

Chronic daily intake of 0.5% H2O2 in drinking water decreased Se-dependent glutathione peroxidase (Se-GSHPx) activity in rat skeletal muscle, kidney and liver. Non-Se GSHPx activity decreased in kidney. Deprivation of drinking water decreased Se-GSHPx activity in kidney and non-Se GSHPx activity in kidney and liver. H2O2 intake decreased activity of catalase in rat skeletal muscle. H2O2 intake or water deprivation caused no changes in these enzyme activities in mice.

Prednisolone decreases exercise-induced acid hydrolase response in mouse skeletal muscle.

SummaryMale NMRI-mice were subjected to exhaustive treadmill exercise. 3 and 6 days after the exertion, quadriceps femoris muscles were examined histologically and analyzed for acid hydrolases in order to follow the degree and progress of injuries. Prednisolone (PRED), an anti-inflammatory corticosteroid, was given to some of the animals in order to modify the exercise response. The PRED administration began 14 h before exercise and continued until the end of the experiment (6 days). The doses were 25 and 50 mg·kg−1 i.p. twice a day. The activities of both arylsulphatase andΒ-glucuronidase increased significantly in the exercise control group after 3 and 6 days. The increase in activity correlated with fibre necrosis and an abundant infiltration of inflammatory cells, and was greatest after 3 days. After 6 days the inflammatory response decreased and regenerating muscle fibres were seen. PRED decreased the exercise-induced acid hydrolase response. The decrease was most prominent after 3 days with PRED 50 mg·kg−1·day−1. PRED also diminished degeneration and inflammation. The results suggest that the decrease in acid hydrolase activities was due to a lesser infiltration of inflammatory cells to the injured area.

Enzymatic and nonenzymatic lipid peroxidation capacities and antioxidants in hypoxic and reoxygenated rat myocardium

The effects of 60 min hypoxia and subsequent reoxygenation for 30 min on enzymatic (NADPH-dependent) and nonenzymatic (Fe2+/ascorbate-induced) lipid peroxidation capacities and on antioxidant levels were studied using Langendorff-perfused rat hearts. The assays were done on the myolayer of the right ventricle (RV) and on the subepi- and subendomyolayers of the left ventricle (epi/endo LV) after normoxic, hypoxic, and reoxygenation phases. The region injured by hypoxia/reoxygenation was located mainly in endo LV, seen as a lesser penetration of the fluorescent dye fluorescein in the myocardium. The electron microscopic findings after reoxygenation revealed swelling of the mitochondria, amorphous mitochondrial structures, and formation of paracrystallines. The myofibrillar structure of the cells was disrupted and the cells showed marked fluid accumulation. Membrane structures were marginated and formed blebs and multilamellar bodies. Ultrastructural changes were most prominent in endo LV, especially after reoxygenation. The increase in leakage of lactate in the perfusate revealed the onset of anaerobic metabolism. Abrupt release of the cytoplasmic enzymes lactate dehydrogenase and creatine kinase at the beginning of the reoxygenation phase suggested cell membrane injury. The capacity for Fe2+/ascorbate-induced lipid peroxidation slightly increased in RV and that for NADPH-dependent, enzymatic lipid peroxidation in endo LV after reoxygenation. Catalase, glutathione peroxidase, and superoxide dismutase activities remained unchanged, whereas glucose-6-phosphate dehydrogenase activity decreased after reoxygenation in RV.(ABSTRACT TRUNCATED AT 250 WORDS)

Increased susceptibility to lipid peroxidation in skeletal muscles of dystrophic hamsters

The results showed that the total content of lipids, which could be peroxidized with Fe(2+)/ascorbate stimulation in vitro, was 45.4% and 53.7% higher than normal in the dystrophic hamster muscle at the age of 1 and 3 months, respectively. Correspondingly, the susceptibility to lipid peroxidation (stimulated by ADP-chelated iron at 37°C) was 38.6–74.3% higher in dystrophic muscles. The increases were not related to necrotic lesions and inflammation observed. The activities of glucose-6-phosphate dehydrogenase, glutathione reductase, thioredoxin reductase and catalase were increased in dystrophic muscles but those of superoxide dismutases and glutathione peroxidase were unaffected.

Activities of some antioxidative and hexose monophosphate shunt enzymes of skeletal muscle in neuromuscular diseases

ABSTRACT The activities of some antioxidative and hexose monophosphate shunt enzymes, as well as of 2 hydrolases were studied in skeletal muscle biopsy specimens taken from 39 patients with neuromuscular diseases and from 15 controls. The activity of Se‐dependent glutathione peroxidase was higher in patients with congenital myotonia, whereas in the other diagnostic groups this enzyme activity was the same as in the controls. The Se‐independent and total glutathione peroxidase activity of patients in the various diagnostic groups did not differ from the controls. Moreover, no difference were observed in catalase activity between the patient groups and the controls. The activities of the rate limiting enzymes of hexose monophosphate shunt, glucose‐6‐phosphate dehydrogenase and 6‐phosphogluconate dehydrogenase of muscle biopsy samples of various patient groups did not show any significant difference from controls. The activity of a lysosomal hydrolase, β‐N‐acetylglucosaminidase, was increased in patients with polyneuropathy and the activity of a nonlysosomal protease, alkaline protease, was high in patients with Charcot‐Marie‐Tooth disease. The activities of Se‐dependent glutathione peroxidase, 6‐phosphogluconate dehydrogenase and of both hydrolases showed a significant correlation to the magnitude of muscle atrophy.

Acid proteolytic activities in mouse liver and muscle tissues after treatment with protease inhibitor leupeptin

The activities of acid proteolytic enzymes were assayed in the liver and muscular tissues of mice (Mus musculus) 1, 6 and 24 hr after the administration of a protease inhibitor leupeptin (i.p., 15.5 mg/kg body wt). Leupeptin administration induced a strong inhibition of cathepsin B and a moderate inhibition of cathepsin C and acid autolytic rate in mouse liver 1 hr after injection. Thereafter the inhibition reduced and disappeared during 24 hr. The activity of cathepsin D was increased in liver 6 and 24 hr after injection. The activity of beta-glucuronidase was not affected by the leupeptin treatment. The administration of leupeptin did not affect the rate of acid autolysis and the activities of cathepsin C and D in cardiac and skeletal muscles. A slight increase in cathepsin B activity was observed 1 hr after leupeptin treatment in calf muscles. The cause of both tissue and enzyme specific changes after leupeptin treatment is discussed.

Effects of free fatty acids, lysophosphatides and phospholipase treatment on lipid peroxidation of myocardial homogenates and membrane fractions

The effects of various free fatty acids, lysophosphatides and phospholipase treatments on the enzymatic and the non-enzymatic lipid peroxidation capacities in the heart homogenates and subcellular fractions were studied. The results showed a dose related inhibition of both the enzymatic and non-enzymatic lipid peroxidation with free fatty acids. A significant inhibition occurred as early as at the concentration of 25-50 microM of several fatty acids both in homogenates and in organelle fractions. In general, the inhibition was greatest with cis-unsaturated, long-chain fatty acids. The inhibition was also induced by the pretreatment of the homogenates with phospholipase A2 but not with phospholipase C. The lysophosphatidyl cholines (16:0 and 18:1) had a stimulatory effect on the enzymatic lipid peroxidation capacity at the physiological concentrations. The results show that the stimulatory/inhibitory effect of various lipid amphiphiles on lipid peroxidation is strongly structure linked and the mitochondrial fraction is the most susceptible to the injury induced by lipid amphiphiles.

Endurance training decreases the alkaline proteolytic activity in mouse skeletal muscles

SummaryAlkaline and myofibrillar protease activities of rectus femoris, soleus, and tibialis anterior muscles and the pooled sample of gastrocnemius and plantaris muscles were analyzed in male NMRI-mice during a running-training program of 3, 10, or 20 daily 1-h sessions. The activity of citrate synthase increased during the endurance training, reflecting the increased oxidative capacity of skeletal muscles. The activities of alkaline and myofibrillar proteases continually decreased in the course of the training program in all muscles studied. Instead, the activity ofΒ-glucuronidase (a marker of lysosomal hydrolases) increased in all muscles. The highest activities were observed at the beginning of the training program. Present results, together with our earlier observations, show that the type of training, running as opposed to swimming, modulates the training responses in alkaline protease activities. Further, diverse adaptations in the activities of alkaline proteases and a lysosomal hydrolase suggest differences in the function of different proteolytic systems.

Selective effects of some anesthetics and detergents on lipid peroxidation of mouse heart homogenates

1. The effects of some anesthetics and detergents on the Fe2+/ascorbate-stimulated non-enzymatic lipid peroxidation potential and on the NADPH-dependent enzymatic lipid peroxidation capacity were characterized in mouse heart homogenates. 2. Chlorpromazine turned out to be the most efficient inhibitor, causing a 50% inhibition at a concentration of 0.03 mM in the non-enzymatic assay, and at a concentration of 0.02 mM in the enzymatic assay. 3. Tetracaine was about a 10-times weaker inhibitor with IC50-values of 0.25 mM. High concentration of dibucaine (1 mM) exerted a 60% inhibition in the non-enzymatic assay, but lidocaine and procaine had no prominent effect with the concentrations used. 4. In the non-enzymatic, Fe(2+)-stimulated system, a 50% inhibition was obtained by using SDS, Triton X-100, and deoxycholic acid at concentrations of 0.004, 0.03, and 0.15%, respectively. 5. In the NADPH-dependent enzymatic lipid peroxidation system, corresponding concentrations were 0.02, 0.04 and 0.1%. Deoxycholate and Triton X-100 even stimulated (10-20%) the enzymatic lipid peroxidation at the lowest concentrations (0.005-0.01%). Saponin was the least effective of these detergents. 6. It is suggested that anesthetics and detergents induce structural rearrangements in the myocardiac membranes which result in the unavailability of phospholipid substrates to lipid peroxidation.