Jiankang Liu

Age-associated changes in superoxide dismutase activity, thiobarbituric acid reactivity and reduced glutathione level in the brain and liver in senescence accelerated mice (SAM): a comparison with ddY mice

The antioxidant defense alteration in young and old senescence accelerated mice (SAM) was studied by examining superoxide dismutase (SOD) activity, thiobarbituric acid (TBA) reactivity, and reduced glutathione (GSH) level in brain and liver tissues. The changes were compared with those in age-paired ddY mice, a strain exhibiting normal aging. SAM mice showed an age-dependent increase in SOD activity in liver, and an age-dependent increase in TBA reactivity in both the brain and the liver; they also showed an age-dependent decrease in the GSH level in the brain and the liver. When compared with ddY mice, SAM mice showed a higher SOD activity in the brain (at both 3 and 11 months old), a lower GSH level in the liver (at 3 months old), and a higher TBA reactivity in the liver (at 3 months old). These findings suggest that the mechanism of senescence acceleration in SAM mice is to some extent related to free radical damage.

Antioxidant action of guilingji in the brain of rats with FeCl3-induced epilepsy.

The effects of Guilingji, an antiaging prescription comprised of traditional Chinese medicinal herbs and animal components, on the brain level of thiobarbituric acid reactive substances (TBARS) and superoxide dismutase (SOD) activity in the brain of rats with FeCl3-induced epilepsy were examined with fluorophotometry and electron spin resonance (ESR) spectrometry, respectively. The results showed that pretreatment of rats with Guilingji decreased the levels of TBARS in the left cortex, midbrain, and olfactory lobe, and increased the SOD activity in the midbrain and hypothalamus. These results are the first to demonstrate that Guilingji may possess an antiepileptic effect further to its proposed antiaging action, because decreasing the TBARS elevation and increasing the SOD attenuation in the brain are suggested to be important characteristics of antiepileptic agents. These results suggest that both the proposed antiaging and antiepileptic effects of Guilingji may operate through the mechanism of antioxidation.

Oxidative Damage in the Senescence‐accelerated Mouse

ABSTRACT: The senescence‐accelerated mouse (SAM) exhibited a shortened life span (about 18 months) and early manifestation of various signs of senescence, including changes in physical activity, skin, and spinal curvature. The mechanism of senescence acceleration in SAM is thought to be related to free radical damage. Oxidative phosphorylation was estimated in liver mitochondria from SAMPS and the senescence‐resistant subtrain, SAMR1. The respiratory control ratio decreased during aging, and the ATP/O, an index of ATP synthesis, was depressed at 18 months of age in SAMPS. DNP‐dependent uncoupled respiration in liver mitochondria was markedly decreased, and active uptake of calcium was markedly dysfunctional with aging. These findings suggest that the functional disorders in mitochondria may be closely related to the shorter life span of SAMPS. White‐footed (WF) mice can live at least to 5.5 years, when some animals are still capable of reproducing and their external body condition remains healthy. The mitochondrial functions were examined in the same way as in the SAM experiments. However, no particular finding responsible for their longevity was observed in WF mice at 3 and 12 months old. More comprehensive examinations on more aged WF mice are needed for explanation of their greater longevity.

Adenosines scavenged hydroxyl radicals and prevented posttraumatic epilepsy

Intracortical injection of iron ions has been used as a model of posttraumatic epilepsy. Oxidation of lipids in neural membranes by reactive oxygen species, especially hydroxyl radicals (OH), is involved in the mechanisms responsible for iron-induced seizures. We examined the scavenging effects of adenosine (Ado) and 2-chloroadenosine (Cl-Ado) on OH radicals and superoxide (O2.-) using an electron spin resonance (ESR) spectrometer, and the occurrence of epileptic discharges in electrocorticogram (ECoG) induced by FeCl3 injection into the sensorimotor cortex of rats. Though DMPO-O2.- spin adducts generated by the hypoxanthine-xanthine oxidase system were not quenched by Ado or Cl-Ado, 5 mM of each showed a quenching effect on DMPO-OH spin adducts (5.3 x 10(16) spins/ml) generated by the Fenton reagent. In ECoG of rats, spike discharges appeared 15-45 min after FeCl3 injection (500 nmol) into the sensorimotor cortex, and polyspikes and/or ictal patterns were observed 70-90 min after the injection. Cl-Ado (1 mg/kg) or Ado (5 mg/kg) injected intraperitoneally 30 min prior to the FeCl3 injection suppressed or delayed the occurrence of epileptic discharges induced by FeCl3. Cl-Ado and Ado may suppress the occurrence of epileptic discharges by scavenging OH and by their anticonvulsant effect.

Involvement of reactive oxygen species in emotional stress: A hypothesis based on the immobilization stress-induced oxidative damage and antioxidant defense changes in rat brain, and the effect of antioxidant treatment with reduced glutathione

We examined the oxidative damage and antioxidant defense changes with immobilization-induced emotional stress in the rat brain. Though superoxide dismutase activity remained unchanged, brain peroxidation was significantly accelerated by the immobilization stress. Membrane fluidity study with spin labeling in brain cortical membrane showed that immobilization stress induced an increase in microviscosity of membrane layer near the surface and in the ordering of membrane proteins but a decrease in microviscosity at the core of the membrane bilayer. The Na, K-ATPase activity decreased whereas the levels of some monoamines and their metabolites increased along with their metabolic rate. The administration of reduced glutathione showed a protective effect on the immobilization stress-induced stomach bleeding, oxidative damage and abnormal changes in the brain antioxidant defenses. Based on these results and on previous reports, we hypothesize that immobilization stress may induce the formation of reactive oxygen species which weakens the brain antioxidant defenses and induces oxidative damage. The antioxidant administration of reduced glutathione provides further evidence to support the above hypothesis, and also may provide clues in the search for a rational therapy to emotional stress. A possible correlation of emotional stress to aging is also discussed.

Probucol scavenged 1,1-diphenyl-2-picrylhydrazyl radicals and inhibited formation of thiobarbituric acid reactive substances

Probucol is suggested to have antioxidant properties. The direct scavenging action of probucol on hydroxyl radicals, superoxide and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radicals were examined using electron spin resonance (ESR) spectrometry. Probucol scavenged DPPH radicals dose dependently but showed no effect on hydroxyl radicals and superoxide generated by Fenton reaction and by hypoxanthine-xanthine oxidase system, respectively. It inhibited the formation of thiobarbituric acid reactive substances (TBARS) in rat cortex homogenate induced by ascorbic acid and FeCl2 at low dose, but it increased TBARS formation at high doses. Probucol showed no effect on the carbon centered radicals. Iron injection into the rat cortex, which is an experimental model for traumatic epilepsy, increased TBARS level in the cortex, hippocampus, striatum and cerebellum, but pretreatment with probucol inhibited the increase in these brain parts except for the hippocampus. These results suggest that the antioxidant property of probucol is partly due to its free radical scavenging effect.

Direct detection of circulating free radicals in the rat using electron spin resonance spectrometry

We developed a new technique for directly observing in vivo free radical formation in the circulating blood of living rats using electron spin resonance (ESR) spectrometry without any labeling or trapping agents. It was found that a doublet peak spectrum was obtained following ferric citrate and ascorbic acid injection. The signals were confirmed in different ways to be due to ascorbic acid radicals. These results provide evidence to support the involvement of free radical intermediates in iron-ascorbic acid reactions, and further confirm the suggested mechanisms of both the adverse and protective effects of ascorbic acid in biological systems. Furthermore, this method of direct observation is a new application of ESR spectrometry to living animals.

Age-related increases in superoxide dismutase activity and thiobarbituric acid-reactive substances: Effect of Bio-catalyzer in aged rat brain

This study describes, using electron spin resonance spectrometry/spin trapping technique, the increase superoxide dismutase (SOD) activity in the mitochondrial and cytosolic fraction of the cortex, midbrain, pons-medulla oblongata and cerebellum, and in thiobarbituric acid-reactive substances (TBARS) in the cortex, cerebellum and hippocampus of the aged rats. The results show that corresponding to the increased life span and improved physical conditions observed after peroral long-term treatment with Bio-catalyzer, a commercial natural fermented health food supplement marketed in Japan and in the Philippines and earlier reported to be a hydroxyl radical scavenger with weaker scavenging activity on superoxide radical (O2−), SOD which is involved in the metabolic degradation of O2− was further increased, whereas TBARS decreased. These findings suggest that the increased SOD activity in the brain as a defense mechanism against age-related accumulation of reactive oxygen species, in particular superoxide radicals, was enhanced with Biocatalyzer treatment while age-related peroxidation of neuronal membrane, as measured by TBARS, was decreased.

Partitioning of vasoactive intestinal polypeptide into lipid bilayers

Incubation of radiolabeled vasoactive intestinal polypeptide (VIP) with preformed lipid vesicles composed of phosphatidylcholine, phosphatidylglycerol and cholesterol resulted in reversible and saturable association of the peptide with the lipid bilayer. The pH-optimum for the reaction was in the physiological range. The vesicle-associated peptide displayed enhanced stability to proteolytic digestion, it was efficiently released into the supernatant by detergent-solubilization of the vesicles but remained vesicle-associated during treatment with agents that disrupt ionic interactions. Peptide binding by electrically neutral vesicles was lower than that by negative vesicles. Electron spin resonance studies with 5-doxylstearic acid or 16-doxylstearic acid labeled vesicles suggested that VIP decreased the fluidity close to the surface of the bilayer and increased the fluidity in its hydrophobic core. These observations suggest that VIP can bind and penetrate lipid bilayers.

Free radicals, lipid peroxides and antioxidants in blood of patients with myotonic dystrophy.

We studied the levels of free radicals, lipid peroxides and antioxidants, as well as superoxide dismutase (SOD) activity in the blood of six patients with myotonic dystrophy (MyD) (mean age 52.8, SD 5.0 years) and seven controls (mean age 48.8, SD 6.3 years). Electron spin resonance was used to assess the free radicals by the spin-trapping method using 5,5-dimethyl-l-pyrroline-l-oxide. The levels of C centre radical (P < 0.05) and H radical (P < 0.05) in blood from the six MyD patients were significantly higher than those in the seven controls. The SOD activities in red blood cells and serum from the six MyD patients showed no significant difference from those in the seven controls. The serum lipid peroxide concentration was increased in five of the MyD patients and tended to increase further as the disease progressed. The serum vitamin E level was low in two patients and in the low normal range in three. Serum coenzyme Q10 was decreased in four patients. The serum selenium level was decreased in two patients and that of serum albumin was decreased in three. Therefore we conclude that increased levels of free radicals and lipid peroxides and decreased antioxidant levels play an important role in the pathogenesis of MyD.