Rapid Communication Muscle Mitochondrial ATP Production in Progressive Supranuclear Palsy

Abstract: Six patients with progressive supranuclear palsy (PSP) and 12 age-matched disease-free subjects participated in this study designed to compare rates of ATP production by intact mitochondria from biopsied skeletal muscle. When pyruvate and malate were used as metabolic substrates, rates of ATP production were 0.184 ± 0.025 μmol/min/U of citrate synthase (CS) activity (a mitochondrial marker) in control subjects and 0.131 ± 0.051 μmol/min/U of CS in PSP patients. In the presence of succinate, rates of ATP formation were 0.137 + 0.02 μmol/min/U of CS in controls and 0.109 ± 0.04 /4mUmol/min/U of CS in patients. With N,N,N',N' -tetramethyl- p -phenylenediamine (TMPD) and ascorbate, rates were 0.034 ± 0.008 μm Umol/min/U of CS in controls and 0.022 ± 0.01 μmol/min/U of CS in PSP subjects. Differences between the control and PSP populations reached statistical significance with pyruvate/malate and TMPD/ascorbate. No differences in either muscle histopathology or histochemistry were found between patient and control subjects. Results of this study suggest that oxidative phosphorylation defects occur in muscle mitochondria from patients with PSP.     

New Species of Human Tyrosine Hydroxylase mRNA Are Produced in Variable Amounts in Adrenal Medulla and Are Overexpressed in Progressive Supranuclear Palsy

Abstract: Alternative splicing of human tyrosine hydroxylase (TH) pre-mRNA produces four mRNAs leading to four different TH isoforms and is thought to have important regulatory functions. We show that the diversity of TH mRNAs is greater than previously described in the autonomous nervous system: New splice junctions corresponding to the skipping of exon 3 were identified by amplification of cDNA synthesized from pheochromocytoma RNA. In all cases the reading frame was maintained. These species were assayed by RNase protection experiments; their abundance (4–6%) was comparable to that of the previously identified human TH-3 and -4 species in normal adrenal medulla. However, higher levels (11–34%) of these species were found in adrenal medullas of patients suffering from progressive supranuclear palsy. Whether such changes are specific to the disease or the consequences of the stress associated with this severe neurodegeneration remains to be established.     

β-Amyloid Neurotoxicity In Vitro: Evidence of Oxidative Stress but Not Protection by Antioxidants

Abstract: Recent data from several groups suggest that the primary mechanism of β-amyloid neurotoxicity may be mediated by reactive oxygen species. To evaluate this hypothesis, we first compared the efficacy of antioxidant agents in preventing toxicity caused by oxidative insults (iron, hydrogen peroxide, and tert -butyl hydroperoxide) and β-amyloid peptides in cultured rat hippocampal neurons. Tested antioxidants (propyl gallate, Trolox, probucol, and promethazine) generally provided significant protection against oxidative insults but not β-amyloid peptides. Next, we examined whether β-amyloid causes oxidative stress, by comparing levels of lipid peroxidation after exposure to either iron or β-amyloid. In a cell-free system, iron but not β-amyloid generated lipid peroxidation. In culture, both insults caused rapid increases in lipid peroxidation, with iron inducing higher levels at later time points. Pretreatment with the antioxidant probucol significantly reduced lipid peroxidation caused by both insults but only attenuated iron toxicity, suggesting that lipid peroxidation does not contribute directly to cell death induced by β-amyloid. Finally, we observed that increasing basal levels of oxidative stress by pretreating cultures with subtoxic doses of iron significantly increased neuronal vulnerability to β-amyloid. The ability of β-amyloid to induce oxidative stress and the demonstration that oxidative stress potentiates β-amyloid toxicity support the clinical use of antioxidants for AD. However, these data do not support the theory that the primary mechanism of β-amyloid toxicity involves oxidative pathways, indicating a continued need to identify additional cellular responses to β-amyloid that underlie its neurodegenerative actions.     

Serum and urine malondialdehyde levels in NIDDM patients with and without hyperlipidemia

Malondialdehyde (MDA), a marker of lipid peroxidation, was measured as thiobarbituric acid reactive substance (TBARS) in 78 noninsulin-dependent diabetic patients, 38 hyperlipidemic patients, and 28 healthy subjects. Diabetic patients were divided into groups and subgroups according to the existence of hyperlipidemia and other complications. Serum and urine MDA concentrations were significantly higher in diabetic and nondiabetic patient groups than in the control group. By contrast to urine MDA level, serum MDA level was significantly higher in hyperlipidemic diabetics than that of normolipidemic diabetics. Serum MDA levels in the hyperlipidemic diabetic group and urine MDA levels in both diabetic groups were significantly higher than those in hyperlipidemic nondiabetic group. In both diabetic groups, the existence of complications didn't affect serum and urine MDA levels. No correlation existed between serum and urine MDA levels in both patient groups and control subjects. This study confirmed the existence of lipid peroxidation disorders in diabetic patients.     

Epidermal Oxidative Stress in Vitiligo

Epidermal levels of enzymatic and non-enzymatic antioxidants such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), vitamin E (Vit E), ubiquinol (CoQ₁₀H₂), and reduced glutathione (GSH), as well as polyunsaturated fatty acids of phospholipids (PL-PUFA), were evaluated in the affected epidermis of 15 patients with active vitiligo (AVP) and in the corresponding epidermis of 15 healthy phototype matched controls. The epidermal levels of CoQ₁₀H₂, Vit E, GSH, and CAT activity were significantly reduced in AVP and were associated with a marked increase of oxidized glutathione, whereas SODs and GSH-Px activities and ubiquinone concentration remained similar to control values. Antioxidant deficiency, in particular the decline of lipophilic antioxidants, i. e., CoQ₁₀H₂ and Vit E, accounts well for PL-PUFA reduction observed in vitiligo epidermis, mainly affecting C18: 3 n-3, C20: 3 n-6, C20: 4 n-6, and C22: 6 n-3 fatty acids and suggesting the occurrence of a lipoperoxidative process. In conclusion, both an imbalance of the intracellular redox status and a significant depletion of enzymatic and non-enzymatic antioxidants feature the epidermis of AVP, and represent a fingerprint of an abnormal oxidative stress leading to epidermal cell injury.     

Basal Lipid Peroxidation in Substantia Nigra Is Increased in Parkinson''s Disease

Polyunsaturated fatty acid (PUFA) levels (an index of the amount of substrate available for lipid peroxidation) were measured in several brain regions from patients who died with Parkinson's disease and age-matched control human postmortem brains. PUFA levels were reduced in parkinsonian substantia nigra compared to other brain regions and to control tissue. However, basal malondialdehyde (MDA; an intermediate in the lipid peroxidation process) levels were increased in parkinsonian nigra compared with other parkinsonian brain regions and control tissue. Expressing basal MDA levels in terms of PUFA content, the difference between parkinsonian and control substantia nigra was even more pronounced. Stimulating MDA production by incubating tissue with FeSO4 plus ascorbic acid, FeSO4 plus H2O2, or air alone produced lower MDA levels in the parkinsonian substantia nigra, probably reflecting the lower PUFA content. These results may indicate that an increased level of lipid peroxidation continues to occur in the parkinsonian nigra up to the time of death, perhaps because of continued exposure to excess free radicals derived from some endogenous or exogenous neurotoxic species.     

Oxidative stress-related mechanisms affecting response to aspirin in diabetes mellitus

Type 2 diabetes mellitus (T2DM) is a major cardiovascular risk factor. Persistent platelet activation plays a key role in atherothrombosis in T2DM. However, current antiplatelet treatments appear less effective in T2DM patients vs nondiabetics at similar risk. A large body of evidence supports the contention that oxidative stress, which characterizes DM, may be responsible, at least in part, for less-than-expected response to aspirin, with multiple mechanisms acting at several levels. This review discusses the pathophysiological mechanisms related to oxidative stress and contributing to suboptimal aspirin action or responsiveness. These include: (1) mechanisms counteracting the antiplatelet effect of aspirin, such as reduced platelet sensitivity to the antiaggregating effects of NO, due to high-glucose-mediated oxidative stress; (2) mechanisms interfering with COX acetylation especially at the platelet level, e.g., lipid hydroperoxide-dependent impaired acetylating effects of aspirin; (3) mechanisms favoring platelet priming (lipid hydroperoxides) or activation (F₂-isoprostanes, acting as partial agonists of thromboxane receptor), or aldose-reductase pathway-mediated oxidative stress, leading to enhanced platelet thromboxane A₂ generation or thromboxane receptor activation; (4) mechanisms favoring platelet recruitment, such as aspirin-induced platelet isoprostane formation; (5) modulation of megakaryocyte generation and thrombopoiesis by oxidative HO-1 inhibition; and (6) aspirin–iron interactions, eventually resulting in impaired pharmacological activity of aspirin, lipoperoxide burden, and enhanced generation of hydroxyl radicals capable of promoting protein kinase C activation and platelet aggregation. Acknowledgment of oxidative stress as a major contributor, not only of vascular complications, but also of suboptimal response to antiplatelet agents in T2DM, may open the way to designing and testing novel antithrombotic strategies, specifically targeting oxidative stress-mediated mechanisms of less-than-expected response to aspirin.     

丘脑底核高频和低频电刺激治疗帕金森病的临床效果评价

目的:观察和比较丘脑底核高频电刺激与低频电刺激治疗帕金森病(PD)的临床效果。方法:对入选的31名PD患者行双侧丘脑底核电刺激手术,术后1个月,在高频刺激条件下,进行UPDRS运动评分,同时对震颤、强直、运动迟缓、中轴症状进行评分;术后6个月,在关闭刺激、高频刺激和低频刺激三种刺激条件,同样进行相关评分。结果:术后1个月和术后6个月,除中轴症状外,UPDRS运动评分和震颤、强直、运动迟缓评分均较术前明显降低(P0.05)。术后6个月,HFS、LFS刺激条件下,UPDRS运动评分和震颤、强直、运动迟缓评分均较OFF降低(P0.05),但中轴症状评分无明显降低(P0.05)。术后6个月,LFS较HFS,各项评分均无明显差异。结论:丘脑底核高频和低频电刺激均能改善PD的运动功能,对震颤、强直和运动迟缓疗效明显,但对中轴症状均无明显治疗效果。     

Increased carbonylation of the lipid phosphatase PTEN contributes to Akt2 activation in a murine model of early alcohol-induced steatosis

The production of reactive aldehydes such as 4-hydroxynonenal (4-HNE) is a key event in the pathogenesis of alcoholic liver disease (ALD), which ranges from simple steatosis to fibrosis. The lipid phosphatase PTEN plays a central role in the regulation of lipid metabolism in the liver. In this study, the effects of chronic ethanol feeding and carbonylation on the PTEN signaling pathway were examined in a 9-week mouse feeding model for ALD. Chronic ethanol consumption resulted in altered redox homeostasis as evidenced by decreased GSH, decreased Trx1, and increased GST activity. Both PTEN expression and PTEN phosphorylation were significantly increased in the livers of ethanol-fed mice. Carbonylation of PTEN increased significantly in the ethanol-fed mice compared to pair-fed control animals, corresponding to decreased PTEN 3-phosphatase activity. Concomitantly, increased expression of Akt2 along with increased Akt phosphorylation at residues Thr³⁰⁸, Thr⁴⁵⁰, and Ser⁴⁷³ was observed resulting in increased Akt2 activity in the ethanol-fed animals. Akt2 activation corresponded to a decrease in cytosolic SREBP and ChREBP. Subsequent LC/MS/MS analysis of 4-HNE-modified recombinant human PTEN identified Michael addition adducts of 4-HNE on Cys⁷¹, Cys¹³⁶, Lys¹⁴⁷, Lys²²³, Cys²⁵⁰, Lys²⁵⁴, Lys³¹³, Lys³²⁷, and Lys³⁴⁴. Computational-based molecular modeling analysis of 4-HNE adducted to Cys⁷¹ near the active site and Lys³²⁷ in the C2 domain of PTEN suggested inhibition of enzyme catalysis via either stearic hindrance of the active-site pocket or prevention of C2 domain-dependent PTEN function. We hypothesize that 4-HNE-mediated PTEN inhibition contributes to the observed activation of Akt2, suggesting a possible novel mechanism of lipid accumulation in response to increased reactive aldehyde production during chronic ethanol administration in mice.     

Biomarkers of diabetes-associated oxidative stress and antioxidant status in young diabetic patients with or without subclinical complications

The aims of the study were to ascertain the potential role of oxidative stress in the onset of disease-related pathophysiological complications in young type 1 diabetes patients. Indicative parameters of lipoperoxidation, protein oxidation, and changes in antioxidant defense system status were measured in blood samples from 26 young diabetic patients with recently diagnosed (< 6 months) microangiopathy (+DC), 28 diabetic patients without complications (−DC), and 40 healthy age-matched controls (CR). Both diabetic groups presented similar fructosamine and glycated hemoglobin (HbA1c) values. Results showed erythrocyte glutathione peroxidase activity, glutathione content, and plasma β-carotene to be significantly lower in diabetic patients compared with control subjects, but with no significant differences between −DC and +DC groups. Antioxidant enzyme superoxide dismutase activity was significantly higher in the erythrocytes of diabetic patients independently of the presence of microvascular complications. However, the plasma -tocopherol/total lipids ratio was significantly diminished in +DC group compared with −DC (p = .008). Lipid peroxidation indices measured in plasma included malondialdehyde, lipid hydroperoxides, and lipoperoxides, which were significantly elevated in our diabetic patients regardless of the presence of complications. Evidence of oxidative damage to proteins was shown both through the quantification of plasma protein carbonyl levels, which were significantly higher in −DC (0.61 ± 0.09 mmol/mg prot), and higher still in the +DC patients (0.75 ± 0.09 mmol/mg prot) compared with those of controls (0.32 ± 0.03 mmol/mg prot; p < .01) and immunoblot analysis of protein-bound carbonyls. Additionally, a marked increase in protein oxidation was observed in +DC patients through assessment of advanced oxidation protein products (AOPP) considered to be an oxidized albumin index; AOPP values were significantly higher in +DC than in −DC patients (p < .01) and CR (p < .0001). These results point to oxidatively modified proteins as a differential factor possibly related to the pathogenesis of diabetic complications.     

Markers for oxidative stress associated with soft rots in French beans (Phaseolus vulgaris) infected by Botrytis cinerea

The role of active oxygen species has been studied in spreading soft-rot lesions caused by the necrotrophic fungal pathogen Botrytis cinerea Pers.:Fr. in leaves of four genotypes of French bean (Phaseolus vulgaris L.). Large increases were observed for the aldehydic end-products of oxidative damage, malondialdehyde and 4-hydroxy-2-nonenal, as a result of infection in each of the genotypes studied. Similar increases were found in a stable free radical and g=4.27 Fe(III) signals, but not Mn(II) signals, in electron paramagnetic resonance spectra. These changes were accompanied by large decreases in ascorbic acid levels, with changes in the antioxidant glutathione being genotype dependent. Received: 3 May 2000 / Accepted: 13 July 2000     

Progressive supranuclear palsy as differential diagnosis of Parkinson's disease in the elderly

IntroductionProgressive supranuclear palsy (PSP) is a syndrome characterized by progressive parkinsonism with early falls due to postural instability, typically vertical gaze supranuclear ophthalmoplegia, pseudobulbar dysfunction, neck dystonia and upper trunk rigidity as well as mild cognitive dysfunction. Progressive supranuclear palsy must be differentiated from Parkinson's disease taking into account several so-called red flags.Materials and methodsWe report a case series hallmarked by gait abnormalities, falls and bradykinesia in which Parkinson's disease was the initial diagnosis.ResultsDue to a torpid clinical course, magnetic resonance imaging (MRI) was performed demonstrating midbrain atrophy, highly suggestive of progressive supranuclear palsy.ConclusionThe neuroradiological exams (magnetic resonance imaging, single photon emission computer tomography, and positron emission tomography) can be useful for diagnosis of PSP. Treatment with levodopa should be considered, especially in patients with a more parkinsonian phenotype.     

Direct Evidence for the Involvement of Free Radicals in Ischemic Insult to the Intestine

Ischemia of rat intestine was induced in vivo by occlusion of the superior mesenteric artery (SMA) for 15 min. Sodium salicylate, 100 mg/kg, given IP, 30 min prior to the ischemic event served as a specific trap for hydroxyl radicals and provided direct evidence for the involvement of free radicals during the ischemic insult. Portions of the bowel were sequentially isolated and removed. The hydroxylation products. dihyd-roxybenzoic acid (DHBA) derivatives were isolated, identified and qunatified by HPLC coupled with electrochemical detection (ECD). The level of 2,5-DHBA (Mean ± SE, ng/g tissue) in the preischemic bowel (N = 21) was 241.8 ± 10.0. It rose significantly to 313.3 ± 15.5 in the ischemic specimen (p = 0.0129) and remained unchanged in the reperfusion period (322.8 ± 15.5). The histological examination correlated well with these levels: mild villi damage in the ischemic period with no further damage in the reperfusion period.     

Antioxidants in the serum of children with insulin-dependent diabetes mellitus

To determine whether alteration in serum antioxidant status is related to the increased oxidative stress as a cause of diabetic angiopathy, we measured both the antioxidant activity (AOA) and total peroxyl radical-trapping antioxidant parameter (TRAP), and their component individual antioxidants in serum of children with insulin-dependent diabetes mellitus (IDDM). The AOA was measured as the ability to inhibit lipid autoxidation in brain homogenates. TRAP was assayed as the ability to delay lipid peroxidation induced by an azo initiator. Antioxidants measured were ceruloplasmin, transferrin, and albumin components of AOA; and ascorbic acid, uric acid, protein sulfhydryl, and alpha-tocopherol as components of TRAP. Serum AOA appeared to be decreased in the diabetics in relation to poor glycemic control, corresponding to the decrease in transferrin and albumin. Serum haptoglobin level was also decreased in the diabetics. Similarly, the directly measured TRAP value was decreased in the diabetic serum mainly due to the decreased contribution of unidentified chain-breaking antioxidants, despite the increase in ascorbic acid and alpha-tocopherol. The decrease in both types of antioxidant activity in the diabetic serum, as new findings, suggests that a defective serum antioxidant status contributes to the increased oxidative stress in IDDM.     

Kupffer Cell Function in Thyroid Hormone-Induced Liver Oxidative Stress in the Rat

The influence of thyroid hormone (L-3, 3', 5-triiodothyronine, T₃) on Kupffer cell function was studied in the isolated perfused rat liver by colloidal carbon infusion. Rates of carbon uptake were determined from the influent minus effluent concentration difference and the flow rate, and the respective carbon-induced respiratory activity was calculated by integration of the area under the O₂ curves during carbon infusion. In the concentration range of 0.2 to 2.0 mg of carbon/ml, livers from euthyroid rats exhibited a sigmoidal-type kinetics of carbon uptake, with a Vmax of 4.8 mg/g liver/min and a concentration of 0.82 mg/ml for half-maximal rate; carbon-induced O₂ uptake presented a hyperbolic-type kinetics, with a Vmax of 4.57 μmol of O₂/g liver and a Km of 0.74 mg of carbon/ml, which significantly correlates with the carbon uptake rates. Light-microscopy showed that carbon was taken up exclusively by non-parenchymal cells, predominantly by Kupffer cells. Thyroid calorigenesis was found in parallel with increased rates of hepatic O₂ consumption and thiobarbituric acid reactive substances (TBARS) formation, glutathione (GSH) depletion, and higher sinusoidal lactate dehydrogenase (LDH) efflux compared to control values. In the concentration range of 0.25 to 0.75 mg/ml, carbon infusion did not modify liver LDH efflux in control rats, while it was significantly enhanced in T₃-treated animals. In this latter group, higher carbon concentrations (1 and 1.3 mg/ml) led to loss of viability of the liver. At 0.25 to 0.75 mg of carbon/ml, both the rates of carbon uptake and the associated carbon-induced respiratory activities were significantly increased by T₃ treatment, effects that were abolished by pretreatment of the rats with gadolinium chloride (GdCl₃). In addition, GdCl₃ decreased by 50% the changes induced by T₃ in hepatic GSH content and TBARS formation. It is concluded that hyperthyroidism enhances Kupffer cell function, correlated with the increased number of liver macrophages observed histologically, which may represent an alternate source of reactive O₂ species to that induced in parenchymal cells, thus contributing to the enhanced oxidative stress status developed.     

Spinal Morphine Administration Reduces the Fatty Acid Contents in Spinal Cord and Brain by Increasing Oxidative Stress

It is well known that oxidative stress damages bimolecules such as DNA and lipids. No study is available on the morphine-induced oxidative damage and fatty acids changes in brain and spinal tissues. The aim of this work was to determine the effects of morphine on the concentrations and compositions of fatty acid in spinal cord segments and brain tissues in rabbits as well as lipid peroxidation (LP) and glutathione (GSH) levels in cortex brain. Twelve New Zealand albino rabbits were used and they were randomly assigned to two groups of 6 rabbits each. First group used as control although morphine administrated to rats in second group. Cortex brain and (cervical, thoracic, lumbar) samples were taken. The fatty acids between n:18.0 and 21.0 were present in spinal cord sections and n:10 fatty acids in control animals were present in the brain tissues. Compared to n:20.0–24.0 fatty acids in spinal cord sections and 8.0 fatty acids in the brain tissues of drug administered animals. The concentration and composition of the fatty acid methyl esters in spinal cord and brain tissues was decreased by morphine treatments. LP levels in the cortex brain were increased although GSH levels were decreased by the morphine administration. In conclusion, unsaturated fatty acids contents in brain and spinal cord sections and GSH were reduced by administrating spinal morphine although oxidative stress as LP increased. The inhibition oxidative damage may be a useful strategy for the development of a new protection for morphine administration as well as opiate abuse.     

Weak antioxidant defenses make the heart a target for damage in copper-deficient rats

Copper deficiency causes more salient pathologic changes in the heart than in the liver of rats. Although oxidative stress has been implicated in copper deficiency-induced pathogenesis, little is known about the selective toxicity to the heart. Therefore, we examined the relationship between the severity of copper deficiency-induced oxidative damage and the capacity of antioxidant defense in heart and liver to investigate a possible mechanism for the selective cardiotoxicity. Weanling rats were fed a purified diet deficient in copper (0.4 μg/g diet) or one containing adequate copper (6.0 μg/g diet) for 4 weeks. Copper deficiency induced a 2-fold increase in lipid peroxidation in the heart (thiobarbituric assay) but did not alter peroxidation in the liver. The antioxidant enzymatic activities of superoxide dismutase, catalase, and glutathione peroxidase were, respectively, 3-, 50- and 1.5-fold lower in the heart than in the liver, although these enzymatic activities were depressed in both organs by copper deficiency. In addition, the activity of glutathione reductase was 4 times lower in the heart than in the liver. The data suggest that a weak antioxidant defense system in the heart is responsible for the relatively high degree of oxidative damage in copper-deficient hearts.     

Estrogens Attenuate and Corticosterone Exacerbates Excitotoxicity, Oxidative Injury, and Amyloid β-Peptide Toxicity in Hippocampal Neurons

Abstract: Steroid hormones, particularly estrogens and glucocorticoids, may play roles in the pathogenesis of neurodegenerative disorders, but their mechanisms of action are not known. We report that estrogens protect cultured hippocampal neurons against glutamate toxicity, glucose deprivation, FeSO₄ toxicity, and amyloid β-peptide (Aβ) toxicity. The toxicity of each insult was significantly attenuated in cultures pretreated for 2 h with 100 n M -10 µ M 17β-estradiol, estriol, or progesterone. In contrast, corticosterone exacerbated neuronal injury induced by glutamate, FeSO₄, and Aβ. Several other steroids, including testosterone, aldosterone, and vitamin D, had no effect on neuronal vulnerability to the different insults. The protective actions of estrogens and progesterone were not blocked by actinomycin D or cycloheximide. Lipid peroxidation induced by FeSO₄ and Aβ was significantly attenuated in neurons and isolated membranes pretreated with estrogens and progesterone, suggesting that these steroids possess antioxidant activities. Estrogens and progesterone also attenuated Aβ- and glutamate-induced elevation of intracellular free Ca²⁺ concentrations. We conclude that estrogens, progesterone, and corticosterone can directly affect neuronal vulnerability to excitotoxic, metabolic, and oxidative insults, suggesting roles for these steroids in several different neurodegenerative disorders.