Synthesis and structure–activity relationships study of dithiolethiones as inducers of glutathione in the SH-SY5Y neuroblastoma cell line

Parkinson’s disease is a neurodegenerative disorder that involves the degeneration of nigrostriatal dopaminergic neurons. Elevated levels of reactive oxygen species have been shown to deplete cellular levels of the ubiquitous antioxidant glutathione, leading to oxidative stress and eventual neuronal cell death. Dithiolethiones, a class of sulfur-containing heterocyclic molecules, have been shown to induce cellular production of glutathione in a variety of tissues, but have not been extensively evaluated in neurons. Herein, we report the synthesis and preliminary structure–activity relationships study of several substituted dithiolethiones. Three molecules were identified (D3T, CPDT, and 2d) that potently induced cellular glutathione in the SH-SY5Y neuroblastoma cell line. Furthermore, these compounds were found to provide neuroprotection in the 6-hydroxydopamine model of neurotoxicity. This study suggests that dithiolethione-mediated neuroprotection may have potential as a disease-modifying antiparkinsonian therapy.     

Phenylbutyrate up-regulates the DJ-1 protein and protects neurons in cell culture and in animal models of Parkinson disease

Parkinson disease is caused by the death of midbrain dopamine neurons from oxidative stress, abnormal protein aggregation, and genetic predisposition. In 2003, Bonifati et al. (23) found that a single amino acid mutation in the DJ-1 protein was associated with early-onset, autosomal recessive Parkinson disease (PARK7). The mutation L166P prevents dimerization that is essential for the antioxidant and gene regulatory activity of the DJ-1 protein. Because low levels of DJ-1 cause Parkinson, we reasoned that overexpression might stop the disease. We found that overexpression of DJ-1 improved tolerance to oxidative stress by selectively up-regulating the rate-limiting step in glutathione synthesis. When we imposed a different metabolic insult, A53T mutant α-synuclein, we found that DJ-1 turned on production of the chaperone protein Hsp-70 without affecting glutathione synthesis. After screening a number of small molecules, we have found that the histone deacetylase inhibitor phenylbutyrate increases DJ-1 expression by 300% in the N27 dopamine cell line and rescues cells from oxidative stress and mutant α-synuclein toxicity. In mice, phenylbutyrate treatment leads to a 260% increase in brain DJ-1 levels and protects dopamine neurons against 1-methyl 4-phenyl 1,2,3,6-tetrahydropyridine (MPTP) toxicity. In a transgenic mouse model of diffuse Lewy body disease, long-term administration of phenylbutyrate reduces α-synuclein aggregation in brain and prevents age-related deterioration in motor and cognitive function. We conclude that drugs that up-regulate DJ-1 gene expression may slow the progression of Parkinson disease by moderating oxidative stress and protein aggregation.     

Rapid Communication: Oxidative Stress Induces Apoptosis in Embryonic Cortical Neurons

Abstract: Glutamate-induced glutathione depletion in immature embryonic cortical neurons has been shown to lead to oxidative stress and cell death. We have used this in vitro model to investigate the mechanism(s) by which free radicals induce neuronal degeneration. We find that glutathione depletion leads to hypercondensation and fragmentation of chromatin into spherical or irregular shapes, a morphologic signature of apoptosis. These morphologic changes are accompanied by laddering of DNA into multiple oligonucleosomal fragments and can be prevented by the antioxidants idebenone and butylated hydroxyanisole. Cell death induced by glutathione depletion can also be prevented by inhibitors of macromolecular synthesis. Taken together, these observations suggest that oxidative stress can induce apoptosis in neurons.     

Diabetes,oxidative stress,and antioxidants: a review

Increasing evidence in both experimental and clinical studies suggests that oxidative stress plays a major role in the pathogenesis of both types of diabetes mellitus. Free radicals are formed disproportionately in diabetes by glucose oxidation, nonenzymatic glycation of proteins, and the subsequent oxidative degradation of glycated proteins. Abnormally high levels of free radicals and the simultaneous decline of antioxidant defense mechanisms can lead to damage of cellular organelles and enzymes, increased lipid peroxidation, and development of insulin resistance. These consequences of oxidative stress can promote the development of complications of diabetes mellitus. Changes in oxidative stress biomarkers, including superoxide dismutase, catalase, glutathione reductase, glutathione peroxidase, glutathione levels, vitamins, lipid peroxidation, nitrite concentration, nonenzymatic glycosylated proteins, and hyperglycemia in diabetes, and their consequences, are discussed in this review. In vivo studies of the effects of various conventional and alternative drugs on these biomarkers are surveyed. There is a need to continue to explore the relationship between free radicals, diabetes, and its complications, and to elucidate the mechanisms by which increased oxidative stress accelerates the development of diabetic complications, in an effort to expand treatment options.     

Oxidative stress and neuroinflammation in the story of Parkinson’s disease: Could targeting these pathways write a good ending?

Although, current medications for Parkinson’s disease can control and relief symptoms of the disease efficiently, they are unable to either prevent progression of the disease or maintain their controlling ability as a long-term medication. To find suitable adjuvant and/or alternative treatments, researchers have investigated antioxidative and anti-inflammatory approaches, since emerging evidence consider oxidative stress and neuroinflammation as leading causes of the development of Parkinson’s disease. Here, how oxidative stress and neuroinflammation take part in Parkinson’s disease pathogenesis was discussed based on featured studies in this context. Then, preclinical and clinical trial studies, which evaluated antioxidative and anti-inflammatory compounds’ ability to treat Parkinson’s disease, were reviewed.     

Mitochondrial dysfunction and oxidative damage in the molecular pathology of Parkinson’s disease

Parkinson’s disease is a complex disorder that is characterized by progressive degeneration of nigrostriatal dopaminergic neurons. Its development is determined by the interaction between the genetic constitution of a body and environmental factors. Analysis of the genes associated with monogenic forms of Parkinson’s disease implicated proteasomal degradation, differentiation of dopaminergic neurons, mitochondrial dysfunction, and oxidative damage in its pathogenesis. The review considers ample data that suggest a key role for mitochondrial dysfunction and oxidative stress.     

Nitrones as therapeutics

Nitrones have the general chemical formula X-CH=NO-Y. They were first used to trap free radicals in chemical systems and then subsequently in biochemical systems. More recently several nitrones, including alpha-phenyl-tert-butylnitrone (PBN), have been shown to have potent biological activity in many experimental animal models. Many diseases of aging, including stroke, cancer development, Parkinson disease, and Alzheimer disease, are known to have enhanced levels of free radicals and oxidative stress. Some derivatives of PBN are significantly more potent than PBN and have undergone extensive commercial development for stroke. Recent research has shown that PBN-related nitrones also have anti-cancer activity in several experimental cancer models and have potential as therapeutics in some cancers. Also, in recent observations nitrones have been shown to act synergistically in combination with antioxidants in the prevention of acute acoustic-noise-induced hearing loss. The mechanistic basis of the potent biological activity of PBN-related nitrones is not known. Even though PBN-related nitrones do decrease oxidative stress and oxidative damage, their potent biological anti-inflammatory activity and their ability to alter cellular signaling processes cannot readily be explained by conventional notions of free radical trapping biochemistry. This review is focused on our studies and others in which the use of selected nitrones as novel therapeutics has been evaluated in experimental models in the context of free radical biochemical and cellular processes considered important in pathologic conditions and age-related diseases.     

Oxidative stress and neurodegeneration: the involvement of iron

Many evidences indicate that oxidative stress plays a significant role in a variety of human disease states, including neurodegenerative diseases. Iron is an essential metal for almost all living organisms due to its involvement in a large number of iron-containing proteins and enzymes, though it could be also toxic. Actually, free iron excess generates oxidative stress, particularly in brain, where anti-oxidative defences are relatively low. Its accumulation in specific regions is associated with pathogenesis in a variety of neurodegenerative diseases (i.e., Parkinson’s disease, Alzheimer’s disease, Huntington’s chorea, Amyotrophic Lateral Sclerosis and Neurodegeneration with Brain Iron Accumulation). Anyway, the extent of toxicity is dictated, in part, by the localization of the iron complex within the cell (cytosolic, lysosomal and mitochondrial), its biochemical form, i.e., ferritin or hemosiderin, as well as the ability of the cell to prevent the generation and propagation of free radical by the wide range of antioxidants and cytoprotective enzymes in the cell. Particularly, ferrous iron can act as a catalyst in the Fenton reaction that potentiates oxygen toxicity by generating a wide range of free radical species, including hydroxyl radicals (·OH). The observation that patients with neurodegenerative diseases show a dramatic increase in their brain iron content, correlated with the production of reactive oxigen species in these areas of the brain, conceivably suggests that disturbances in brain iron homeostasis may contribute to the pathogenesis of these disorders. The aim of this review is to describe the chemical features of iron in human beings and iron induced toxicity in neurodegenerative diseases. Furthermore, the attention is focused on metal chelating drugs therapeutic strategies.     

Antioxidant and pro-oxidant effect of the thiolic compounds N-acetyl-L-cysteine and glutathione against free radical-induced lipid peroxidation

The antioxidant ability of thiol compounds has been the subject of much of the current research about oxidative stress. The direct scavenging of hydroxyl radicals by thiols has been suggested as their protection mechanisms. Nevertheless, the interaction of thiols with reactive radicals can generate thiyl radicals, which, in turn, may impart a pro-oxidant function. The purpose of this study has been to establish the effect of the thiol compounds N -acetyl- l -cysteine (NAC) and glutathione (GSH) against the peroxidative processes involving membrane lipids. The results obtained support the ability of NAC and GSH to suppress the 2,2'-azobis-(2-amidinopropane) dihydrochloride (AAPH)-dependent or to enhance the Fe 2+ /H 2 O 2 -dependent oxidative actions. The evaluation of thiobarbituric acid reactive substances (TBARS) production, the study of the influence of oxidants on membrane fluidity and the measurements of the changes in the fluorescence of bilayer probes, such as 3-( p -(6-phenyl)-1,3,5-hexatrienyl)phenylpropionic acid (DPH-PA), have shown the antioxidant and pro-oxidant effects of both NAC and GSH. Also their dependence on the nature of the radicals generated by the oxidative systems used has been shown. The use of ESR spectroscopy has allowed us to establish the ability of these compounds to scavenge the AAPH-derived radicals, to determine the formation of thiyl radicals in the iron-mediated oxidation and to evaluate the enhanced production of hydroxyl radicals by NAC and GSH.     

Oxidation of 3,4-dihydroxyphenylacetaldehyde, a toxic dopaminergic metabolite, to a semiquinone radical and an ortho-quinone

The oxidation and toxicity of dopamine is believed to contribute to the selective neurodegeneration associated with Parkinson disease. The formation of reactive radicals and quinones greatly contributes to dopaminergic toxicity through a variety of mechanisms. The physiological metabolism of dopamine to 3,4-dihydroxyphenylacetaldehyde (DOPAL) via monoamine oxidase significantly increases its toxicity. To more adequately explain this enhanced toxicity, we hypothesized that DOPAL is capable of forming radical and quinone species upon oxidation. Here, two unique oxidation products of DOPAL are identified. Several different oxidation methods gave rise to a transient DOPAL semiquinone radical, which was characterized by electron paramagnetic resonance spectroscopy. NMR identified the second oxidation product of DOPAL as the ortho-quinone. Also, carbonyl hydration of DOPAL in aqueous media was evident via NMR. Interestingly, the DOPAL quinone exists exclusively in the hydrated form. Furthermore, the enzymatic and chemical oxidation of DOPAL greatly enhance protein cross-linking, whereas auto-oxidation results in the production of superoxide. Also, DOPAL was shown to be susceptible to oxidation by cyclooxygenase-2 (COX-2). The involvement of this physiologically relevant enzyme in both oxidative stress and Parkinson disease underscores the potential importance of DOPAL in the pathogenesis of this condition.     

Amyloid precursor protein-mediated free radicals and oxidative damage: implications for the development and progression of Alzheimer's disease

Alzheimer's disease (AD) is a late-onset dementia that is characterized by the loss of memory and an impairment of multiple cognitive functions. Advancements in molecular, cellular, and animal model studies have revealed that the formation of amyloid beta (Abeta) and other derivatives of the amyloid precursor protein (APP) are key factors in cellular changes in the AD brain, including the generation of free radicals, oxidative damage, and inflammation. Recent molecular, cellular, and gene expression studies have revealed that Abeta enters mitochondria, induces the generation of free radicals, and leads to oxidative damage in post-mortem brain neurons from AD patients and in brain neurons from cell models and transgenic mouse models of AD. In the last three decades, tremendous progress has been made in mitochondrial research and has provided significant findings to link mitochondrial oxidative damage and neurodegenerative diseases such as AD. Researchers in the AD field are beginning to recognize the possible involvement of a mutant APP and its derivatives in causing mitochondrial oxidative damage in AD. This article summarizes the latest research findings on the generation of free radicals in mitochondria and provides a possible model that links Abeta proteins, the generation of free radicals, and oxidative damage in AD development and progression.     

自由基稳衡性动态

虽然自由基具有很活泼的化学性质 ,但在需氧生物的进化中却一直保持着自由基稳衡性动态的特征 ,体现于某些生物程序 ,包括 :在生理情况下履行其生理作用 ;维持其产生与清除于接近平衡 ;余剩的自由基引发生物大分子的损伤及其损伤可被修复。营养素及其代谢物和“必需”抗氧化剂对自由基稳衡态的正常维持起着关键性作用。谷胱甘肽及其它生物物质的稳衡性动态与自由基稳衡性动态有着协调的相互关系。在各种生活条件下 ,不同年龄的健康人体内自由基稳衡性动态应维持良好 ,以预防衰老前氧化应激与氧化损伤的发生     

Apport des biomarqueurs au diagnostic de la maladie de Parkinson

Neurodegenerative diseases have a high prevalence in the elderly. Progress in medical care depends on the accuracy and precocity of the diagnosis. Biomarkers have proved to be a valuable tool for the diagnosis of Alzheimer’s disease and a similar development is expected for Parkinson’s disease. Recent data suggest that markers of α-synuclein aggregation and oxidative stress in blood and cerebrospinal fluid could contribute to the diagnosis of Parkinson’s disease.     

Dopamine-derived quinones affect the structure of the redox sensor DJ-1 through modifications at Cys-106 and Cys-53

The physiological role of DJ-1, a protein involved in familial Parkinson disease is still controversial. One of the hypotheses proposed indicates a sensor role for oxidative stress, through oxidation of a conserved cysteine residue (Cys-106). The association of DJ-1 mutations with Parkinson disease suggests a loss of function, specific to dopaminergic neurons. Under oxidative conditions, highly reactive dopamine quinones (DAQs) can be produced, which can modify cysteine residues. In cellular models, DJ-1 was found covalently modified by dopamine. We analyzed the structural modifications induced on human DJ-1 by DAQs in vitro. We described the structural perturbations induced by DAQ adduct formation on each of the three cysteine residues of DJ-1 using specific mutants. Cys-53 is the most reactive residue and forms a covalent dimer also in SH-SY5Y DJ-1-transfected cells, but modification of Cys-106 induces the most severe structural perturbations; Cys-46 is not reactive. The relevance of these covalent modifications to the several functions ascribed to DJ-1 is discussed in the context of the cell response to a dopamine-derived oxidative insult.     

Oxidative stress in malaria parasite-infected erythrocytes: host-parasite interactions

Experimenta naturae, like the glucose-6-phosphate dehydrogenase deficiency, indicate that malaria parasites are highly susceptible to alterations in the redox equilibrium. This offers a great potential for the development of urgently required novel chemotherapeutic strategies. However, the relationship between the redox status of malarial parasites and that of their host is complex. In this review article we summarise the presently available knowledge on sources and detoxification pathways of reactive oxygen species in malaria parasite-infected red cells, on clinical aspects of redox metabolism and redox-related mechanisms of drug action as well as future prospects for drug development. As delineated below, alterations in redox status contribute to disease manifestation including sequestration, cerebral pathology, anaemia, respiratory distress, and placental malaria. Studying haemoglobinopathies, like thalassemias and sickle cell disease, and other red cell defects that provide protection against malaria allows insights into this fine balance of redox interactions. The host immune response to malaria involves phagocytosis as well as the production of nitric oxide and oxygen radicals that form part of the host defence system and also contribute to the pathology of the disease. Haemoglobin degradation by the malarial parasite produces the redox active by-products, free haem and H(2)O(2), conferring oxidative insult on the host cell. However, the parasite also supplies antioxidant moieties to the host and possesses an efficient enzymatic antioxidant defence system including glutathione- and thioredoxin-dependent proteins. Mechanistic and structural work on these enzymes might provide a basis for targeting the parasite. Indeed, a number of currently used drugs, especially the endoperoxide antimalarials, appear to act by increasing oxidant stress, and novel drugs such as peroxidic compounds and anthroquinones are being developed.     

Proteins protect lipid membranes from oxidation by thiyl radicals

Oxidation of polyunsaturated fatty acids by thiyl radicals derived from GSH or Cys is believed to be responsible for some of the biological damage resulting from lipid oxidation under oxidative stress. However, this has not been demonstrated in complex biological systems. In this study, we measured the formation of lipid hydroperoxides in liposomes exposed to radicals generated by gamma radiation from GSH, GSSG, GSMe, Cys and Met. In the absence of proteins, the radicals oxidized the liposome lipids. In the presence of proteins, the thiyl radicals failed to react with the liposomes, even though the protein radicals efficiently oxidized the S-compounds. It appears that the thiyl and other S-radicals were effectively scavenged by the protein before initiating lipid oxidation. The results suggest that membrane lipid oxidation in vivo by thiyl radicals is unlikely to be a significant event.     

铁与帕金森病关系研究进展

大量研究证实帕金森患者黑质铁异常沉积,脑内铁代谢紊乱,铁促进氧化应激是神经细胞损伤的重要病理机制。本文综述了铁代谢异常与帕金森病关系的研究概况,指出从中药中寻找金属螯合活性成分治疗PD是极其诱人的研究方向。     

The oxidation of glutathione by cobalt/tungsten carbide contributes to hard metal-induced oxidative stress

The occupational exposure to cobalt/tungsten carbide (Co/WC) dusts causes asthma and interstitial fibrosis. The International Agency for Research on Cancer (IARC) recently classified the mixture Co/WC as probably carcinogenic to humans (group 2A). The mechanism of action of Co/WC involves particle driven generation of Reactive Oxygen Species (ROS) with consequent oxidative damage. The present study evaluates the reactivity of Co/WC dust toward glutathione (GSH) and cysteine (Cys). Co/WC oxidized thiols through a mechanism involving the generation of sulphur-centred radicals. The results are consistent with the oxidation taking place at surface active sites, a part of which is accessible only to Cys S-H groups, but not to GSH ones. Such a reaction, with consequent irreversible depletion of antioxidant defenses of cells, will potentiate the oxidative stress caused by particle and cell generated ROS.     

Glutathione precursors replenish decreased glutathione pool in cystinotic cell lines

Cystinosis is an inherited disorder due to mutations in the CTNS gene which encodes cystinosin, a lysosomal transmembrane protein involved in cystine export to the cytosol. Both accumulation of cystine in the lysosome and decreased cystine in the cytosol may participate in the pathogenic mechanism underlying the disease. We observed that cystinotic cell lines have moderate decrease of glutathione content during exponential growth phase. This resulted in increased solicitation of oxidative defences of the cell denoted by concurrent superoxide dismutase induction, although without major oxidative insult under our experimental conditions. Finally, decreased glutathione content in cystinotic cell lines could be counterbalanced by a series of exogenous precursors of cysteine, denoting that lysosomal cystine export is a natural source of cellular cysteine in the studied cell lines.     

Oxidative Stress Induces a Form of Programmed Cell Death with Characteristics of Both Apoptosis and Necrosis in Neuronal Cells

Abstract: Oxidative stress is implicated in a number of neurological disorders including stroke, Parkinson's disease, and Alzheimer's disease. To study the effects of oxidative stress on neuronal cells, we have used an immortalized mouse hippocampal cell line (HT-22) that is particularly sensitive to glutamate. In these cells, glutamate competes for cystine uptake, leading to a reduction in glutathione and, ultimately, cell death. As it has been reported that protein kinase C activation inhibits glutamate toxicity in these cells and is also associated with the inhibition of apoptosis in other cell types, we asked if glutamate toxicity was via apoptosis. Morphologically, glutamate-treated cells underwent plasma membrane blebbing and cell shrinkage, but no DNA fragmentation was observed. At the ultrastructural level, there was damage to mitochondria and other organelles although the nuclei remained intact. Protein and RNA synthesis inhibitors as well as certain protease inhibitors protected the cells from glutamate toxicity. Both the macromolecular synthesis inhibitors and the protease inhibitors had to be added relatively soon after the addition of glutamate, suggesting that protein synthesis and protease activation are early and distinct steps in the cell death pathway. Thus, the oxidative stress brought about by treatment with glutamate initiates a series of events that lead to a form of cell death distinct from either necrosis or apoptosis.