The role of oxidative stress in physiological and pathological processes in the thyroid gland; possible involvement in pineal-thyroid interactions

Reactive oxygen species (ROS) play an important role in physiological processes, but - when being in excess - ROS cause oxidative damage to molecules. Under physiological conditions, the production and detoxification of ROS are more-or-less balanced. Also in the thyroid, ROS and free radicals participate in physiological and pathological processes in the gland. For example, hydrogen peroxide (H2O2) is crucial for thyroid hormone biosynthesis, acting at different steps of the process. Additionally, H2O2 is believed to participate in the Wolff-Chaikoff's effect, undergoing in conditions of iodide excess in the thyroid. Much evidence has been accumulated indicating that oxidative stress is involved in pathomechanism of thyroid disease, e.g., Graves' disease, goiter formation or thyroid cancer. Melatonin (N-acetyl-5-methoxytryptamine) - the main secretory product of the pineal gland - is a well-known antioxidant and free radical scavenger, widely distributed in the organism. Mutual relationships between the pineal gland and the thyroid have - for a long time - been a subject of intensive research. The abundant to-date's evidence relates mostly to the inhibitory action of melatonin on the thyroid growth and function and - to a lesser extent - to the stimulatory effects of thyroid hormones on the pineal gland. It is highly probable that under physiological conditions melatonin and, possibly, other antioxidants regulate ROS generation for thyroid hormone synthesis. We believe that melatonin may protect against extensive oxidative damage in the course of certain thyroid disorders or in case of a harmful action of some external factors on the thyroid. Thus, oxidative damage and the protective action of antioxidants, melatonin included, may occur during both physiological and pathological processes in the thyroid, however, this assumption, requires further studies.     

A new technique for the quantitative assessment of 8-oxoguanine in nuclear DNA as a marker of oxidative stress. Application to dystrophin-deficient DMD skeletal muscles

This is the first report on the development of an immunohistochemical technique, combined with quantitative image analysis, for the assessment of oxidative stress quantitatively in nuclear DNA in situ, and its application to measure DNA damage in Duchenne muscular dystrophic (DMD) muscles. Three sequential staining procedures for cell nuclei, a cell marker, and a product of oxidative DNA damage, 8-oxoguanine (8-oxoG), were performed. First, the nuclei in muscle sections were stained with Neutral Red followed by the capture of their images with an image analysis system used for absorbance measurements. Second, the same sections were then immunostained for laminin in basement membranes as the cell marker. Next, the sections were treated with 2 N HCl to remove the bound Neutral Red and to denature tissue DNA. Third, the sections were immunostained for 8-oxoG in DNA, using diaminobenzidine (DAB) to reveal the antibody complex. This was followed by capture of the images of the immunostained sections as previously. The absorbances at 451.2 nm of bound Neutral Red and DAB polymer oxides, the final product of 8-oxoG immunostaining, were measured in the same myonuclei in the sections. Analysis of these absorbances permitted indices of the 8-oxoG content, independent of the nuclear densities, to be determined in nuclear DNA in single myofibres and myosatellite cells surrounded by basement membranes. We found that the mean index for the myonuclei in biceps brachii muscles of 2- to 7-year-old patients was 14% higher than that in age-matched normal controls. This finding of the increased oxidative stress in the myonuclei in young DMD muscles agrees with the previous reports of increased oxidative stress in the cytoplasm in the DMD myofibres and myosatellite cells. The present technique for the quantitative assessment of oxidative stress in nuclear DNA in situ is applicable not only in biomedical research but also in the development of effective drugs for degenerative diseases related to oxidative stress.     

The involvement of nitric oxide in maneb- and paraquat-induced oxidative stress in rat polymorphonuclear leukocytes

Oxidative stress plays a crucial role in the manifestations of maneb (MB) and paraquat (PQ)-induced toxicity including MB+PQ-induced Parkinson's disease (PD). Polymorphonuclear leukocytes (PMNs) actively participate in the oxidative stress-mediated inflammation and organ toxicity. The present study was undertaken to investigate the MB- and/or PQ-induced alterations in the indices of oxidative stress in rat PMNs. Animals were treated with or without MB and/or PQ in an exposure time dependent manner. In some sets of experiments, the animals were pre-treated with NOS inhibitors N(G)-nitro-L-arginine methyl ester (L-NAME) and aminoguanidine (AG) along with respective controls. A significant increase in myeloperoxidase (MPO), superoxide dismutase (SOD), nitric oxide, iNOS expression and lipid peroxidation (LPO) was observed in PMNs of MB- and/or PQ-treated animals, while catalase and glutathione S-transferase (GST) activities were attenuated. L-NAME and AG significantly reduced the augmented nitrite content, iNOS expression and MPO activity to control level in MB and PQ exposed animals. Although the augmented LPO was also reduced significantly in L-NAME and AG treated rat PMNs, the level was still higher as compared with controls. Alterations induced in SOD and GST activities were not affected by NOS inhibitors. The results thus suggest that MB and/or PQ induce iNOS-mediated nitric oxide production, which in turn increases MPO activity and lipid peroxidation, thereby oxidative stress.     

The involvement of nitric oxide in maneb- and paraquat-induced oxidative stress in rat polymorphonuclear leukocytes

Oxidative stress plays a crucial role in the manifestations of maneb (MB) and paraquat (PQ)-induced toxicity including MB+PQ-induced Parkinson's disease (PD). Polymorphonuclear leukocytes (PMNs) actively participate in the oxidative stress-mediated inflammation and organ toxicity. The present study was undertaken to investigate the MB- and/or PQ-induced alterations in the indices of oxidative stress in rat PMNs. Animals were treated with or without MB and/or PQ in an exposure time dependent manner. In some sets of experiments, the animals were pre-treated with NOS inhibitors N^G-nitro-L-arginine methyl ester (L-NAME) and aminoguanidine (AG) along with respective controls. A significant increase in myeloperoxidase (MPO), superoxide dismutase (SOD), nitric oxide, iNOS expression and lipid peroxidation (LPO) was observed in PMNs of MB- and/or PQ-treated animals, while catalase and glutathione S-transferase (GST) activities were attenuated. L-NAME and AG significantly reduced the augmented nitrite content, iNOS expression and MPO activity to control level in MB and PQ exposed animals. Although the augmented LPO was also reduced significantly in L-NAME and AG treated rat PMNs, the level was still higher as compared with controls. Alterations induced in SOD and GST activities were not affected by NOS inhibitors. The results thus suggest that MB and/or PQ induce iNOS-mediated nitric oxide production, which in turn increases MPO activity and lipid peroxidation, thereby oxidative stress.     

The contribution of DNA repair and antioxidants in determining cell type-specific resistance to oxidative stress

The aims of this study were; (i) to elucidate the mechanisms involved in determining cell type-specific responses to oxidative stress and (ii) to test the hypothesis that cell types which are subjected to high oxidative burdens in vivo, have greater oxidative stress resistance. Cultures of the retinal pigment epithelium (RPE), corneal fibroblasts, alveolar type II epithelium and skin epidermal cells were studied. Cellular sensitivity to H2O2 was determined by the MTT assay. Cellular antioxidant status (CuZnSOD, MnSOD, GPX, CAT) was analyzed with enzymatic assays and the susceptibility and repair capacities of nuclear and mitochondrial genomes were assessed by QPCR. Cell type-specific responses to H2O2 were observed. The RPE had the greatest resistance to oxidative stress (P>0.05; compared to all other cell types) followed by the corneal fibroblasts (P < 0.05; compared to skin and lung cells). The oxidative tolerance of the RPE coincided with greater CuZnSOD, GPX and CAT enzymatic activity (P < 0.05; compared to other cells). The RPE and corneal fibroblasts both had up-regulated nDNA repair post-treatment (P < 0.05; compared to all other cells). In summary, variations in the synergistic interplay between enzymatic antioxidants and nDNA repair have important roles in influencing cell type-specific vulnerability to oxidative stress. Furthermore, cells located in highly oxidizing microenvironments appear to have more efficient oxidative defence and repair mechanisms.     

The relationship between oxidative/nitrative stress and pathological inclusions in Alzheimer's and Parkinson's diseases

Alzheimer's (AD) and Parkinson's diseases (PD) are late-onset neurodegenerative diseases that have tremendous impact on the lives of affected individuals, their families, and society as a whole. Remarkable efforts are being made to elucidate the dominant factors that result in the pathogenesis of these disorders. Extensive postmortem studies suggest that oxidative/nitrative stresses are prominent features of these diseases, and several animal models support this notion. Furthermore, it is likely that protein modifications resulting from oxidative/nitrative damage contribute to the formation of intracytoplasmic inclusions characteristic of each disease. The frequent presentation of both AD and PD in individuals and the co-occurrence of inclusions characteristic of AD and PD in several other neurodegenerative diseases suggests the involvement of a common underlying aberrant process. It can be surmised that oxidative/nitrative stress, which is cooperatively influenced by environmental factors, genetic predisposition, and senescence, may be a link between these disorders.     

Impaired mitochondrial oxidative phosphorylation in skeletal muscle of the dystrophin-deficient mdx mouse

The mdx mouse, an animal model of the Duchenne muscular dystrophy, was used for the investigation of changes in mitochondrial function associated with dystrophin deficiency. Enzymatic analysis of skeletal muscle showed an approximately 50% decrease in the activity of all respiratory chain-linked enzymes in musculus quadriceps of adult mdx mice as compared with controls, while in cardiac muscle no difference was observed. The activities of cytosolic and mitochondrial matrix enzymes were not significantly different from the control values in both cardiac and skeletal muscles. In saponin-permeabilized skeletal muscle fibers of mdx mice the maximal rates of mitochondrial respiration were about two times lower than those of controls. These changes were also demonstrated on the level of isolated mitochondria. Mdx muscle mitochondria had only 60% of maximal respiration activities of control mice skeletal muscle mitochondria and contained only about 60% of hemoproteins of mitochondrial inner membrane. Similar findings were observed in a skeletal muscle biopsy of a Duchenne muscular dystrophy patient. These data strongly suggest that a specific decrease in the amount of all mitochondrial inner membrane enzymes, most probably as result of Ca²⁺ overload of muscle fibers, is the reason for the bioenergetic deficits in dystrophin-deficient skeletal muscle.     

Proteomic profiling of naturally protected extraocular muscles from the dystrophin-deficient mdx mouse

Duchenne muscular dystrophy is the most frequent neuromuscular disorder of childhood. Although this x-linked muscle disease is extremely progressive, not all subtypes of skeletal muscles are affected in the same way. While extremities and trunk muscles are drastically weakened, extraocular muscles are usually spared in Duchenne patients. In order to determine the global protein expression pattern in these naturally protected muscles we have performed a comparative proteomic study of the established mdx mouse model of x-linked muscular dystrophy. Fluorescence difference in-gel electrophoretic analysis of 9-week-old dystrophin-deficient versus age-matched normal extraocular muscle, using a pH 4-7 gel range, identified out of 1088 recognized protein spots a moderate expression change in only seven protein species. Desmin, apolipoprotein A-I binding protein and perilipin-3 were found to be increased and gelsolin, gephyrin, transaldolase, and acyl-CoA dehydrogenase were shown to be decreased in mdx extraocular muscles. Immunoblotting revealed a drastic up-regulation of utrophin, comparable levels of β-dystroglycan and key Ca²⁺-regulatory elements, and an elevated concentration of small stress proteins in mdx extraocular muscles. This suggests that despite the lack of dystrophin only a limited number of cellular systems are perturbed in mdx extraocular muscles, probably due to the substitution of dystrophin by its autosomal homolog. Utrophin appears to prevent the loss of dystrophin-associated proteins and Ca²⁺-handling elements in extraocular muscle tissue. Interestingly, the adaptive mechanisms that cause the sparing of extraocular fibers seem to be closely linked to an enhanced cellular stress response.     

Combining redox-proteomics and epigenomics to explain the involvement of oxidative stress in psychiatric disorders

Psychiatric disorders affect approximately 10% of adults in North-America. The complex nature of these illnesses makes the search for their pathophysiology a challenge. However, studies have consistently shown that mitochondrial dysfunction and oxidative stress are common features across major psychiatric disorders, including bipolar disorder and schizophrenia. Nevertheless, little is known about specific targets of oxidation in the brain. The search for redox sensors (protein targets for oxidation) will offer information about which pathways are regulated by oxidation in psychiatric disorders. Additionally, DNA is also a target for oxidative damage and recently, studies have suggested that oxidation of cytosine and guanosine can serve as an epigenetic modulator by decreasing or preventing further DNA methylation. Therefore, this review aims to discuss how we can use redox-proteomics and epigenomics to help explain the role of oxidative damage in major psychiatric disorders, which may ultimately lead to the identification of targets for development of new medications.     

The involvement of serotonin metabolism in cigarette smoke-induced oxidative stress in rat lung in vivo

Abstract

Recently, we have reported the dysregulation of circulating serotonin (5-hydroxytryptamine, 5-HT) homeostasis in patients with chronic obstructive pulmonary disease (COPD). An increase in metabolism of 5-HT has been reported to induce oxidative stress via monoamine oxidase (MAO)-dependent pathway. The present study aimed at investigating the effect of cigarette smoke exposure on the systemic circulation and local airway 5-HT levels as well as MAO-mediated oxidative pathway using a cigarette smoke-exposed rat model. Male Sprague-Dawley rats (150–200 g) were exposed to either sham air or 4% (v/v, smoke/air) cigarette smoke for 1 hour daily for 56 consecutive days. Sera, bronchoalveolar larvage (BAL) and lung tissues were collected 24 hours after the last exposure. We found a significant reduction in the reduced glutathione (rGSH) and an elevation in advanced oxidation protein products (AOPP), a protein oxidation marker, in the lung of cigarette smoke-exposed group (p?<?0.05). A significant increase in 5-HT was found in serum (p?<?0.05), but not in the BAL or lung, after cigarette smoke exposure. MAO-A activity was significantly elevated in the lung of cigarette smoke-exposed group (p?<?0.05). Furthermore, increased superoxide anion levels were found in lung homogenates of cigarette smoke-exposed rats after incubation with 5-HT (p?<?0.05), which was positively associated with the increase in MAO-A activity (r?=?0.639, p?<?0.05). Our findings supported the presence of GSH disruption and protein oxidation in the lung after cigarette smoke exposure. The metabolism of 5-HT by MAO-A in the lung enhanced cigarette smoke-induced superoxides, which might contribute to the pathogenesis of COPD.     

OxyR介导的细菌氧化胁迫应答调控机制研究进展

OxyR属于LysR型转录因子家族的氧化胁迫调控蛋白,是细菌抵抗氧化胁迫压力的重要调控因子。OxyR能够通过调控过氧化氢酶和过氧化物酶等抗氧化基因的表达清除H₂O₂、参与铁代谢控制胞内过氧化物的产生以及修复生物大分子氧化损伤,从而抵抗氧化胁迫。OxyR的基因表达调控功能依赖于其还原态和氧化态之间的转变,改变调控蛋白对下游基因调控区的亲和能力。氧化态OxyR识别启动子区的结合序列,激活或抑制过氧化氢酶等基因的表达。还原态和氧化态的转换依赖于在氧化状态下分子间二硫键的形成。本文综述了近年来细菌OxyR调控基因表达的最新研究进展,有助于深入理解OxyR在细菌抵抗氧化胁迫的作用方式,为相关致病菌的防治奠定分子基础。     

The effects of rosiglitazone on oxidative stress and lipid profile in left ventricular muscles of diabetic rats

We investigated the effect of rosiglitazone (RSG), a high-affinity ligand for the peroxisome proliferator-activated receptor gamma which mediates insulin-sensitizing actions, on the lipid profile and oxidative status in streptozotocin (STZ)-induced Type 2 diabetes mellitus (DM) rats. Wistar albino male rats were randomly divided into an untreated control group (C), a C + RSG group which was treated with RSG (4 mg kg(-1)) two times a day by gavage, a diabetic group (D) that was treated with a single intraperitoneal injection of STZ (45 mgkg(-1)), D + RSG group which were treated with RSG two times a day by gavage, respectively. Lipid profiles, HbA(1c) and blood glucose levels in the circulation and malondialdehyde (MDA) and 3-nitrotyrosine (3-NT) levels in left ventricular muscle were measured. Treatment of D rats with RSG resulted in a time-dependent decrease in blood glucose. We found that the lipid profile and HbA(1c) levels in D + RSG group reached the C rat values at the end of the treatment period. There was a statistically significant difference between the C + RSG and C groups in 3-NT levels. In group D, 3-NT and MDA levels were found to be increased when compared with C, C + RSG and D + RSG groups. In the D + RSG group, MDA levels were found to be decreased when compared with C and C + RSG. Our study suggests that the treatment of D rats with RSG for 8 weeks may decrease the oxidative/nitrosative stress in left ventricular tissue of rats. Thus in diabetes-related vascular diseases, RSG treatment may be cardioprotective.     

The latest progress on miR‐374 and its functional implications in physiological and pathological processes

Non‐coding RNAs (ncRNAs) have been emerging players in cell development, differentiation, proliferation and apoptosis. Based on their differences in length and structure, they are subdivided into several categories including long non‐coding RNAs (lncRNAs >200nt), stable non‐coding RNAs (60‐300nt), microRNAs (miRs or miRNAs, 18‐24nt), circular RNAs, piwi‐interacting RNAs (26‐31nt) and small interfering RNAs (about 21nt). Therein, miRNAs not only directly regulate gene expression through pairing of nucleotide bases between the miRNA sequence and a specific mRNA that leads to the translational repression or degradation of the target mRNA, but also indirectly affect the function of downstream genes through interactions with lncRNAs and circRNAs. The latest studies have highlighted their importance in physiological and pathological processes. MiR‐374 family member are located at the X‐chromosome inactivation center. In recent years, numerous researches have uncovered that miR‐374 family members play an indispensable regulatory role, such as in reproductive disorders, cell growth and differentiation, calcium handling in the kidney, various cancers and epilepsy. In this review, we mainly focus on the role of miR‐374 family members in multiple physiological and pathological processes. More specifically, we also summarize their promising potential as novel prognostic biomarkers and therapeutic targets from bench to bedside.     

An in vitro oxidative stress test for determining pollutant tolerance in algae

An in vitro oxidative stress test has been developed to assess pollutant tolerance in freshwater algae using Euglena gracilis as the test organism and FeSO₄ and NaCl as the pollutants. The test evaluates free radical-mediated oxidative stress through the concomitant application of three biochemical assays: (1) the non-invasive, gas chromatographic-volatile headspace analysis of hydroxyl radicals (OH) using dimethyl-sulphoxide as a radical trap; (2) the spectroscopic determination of total antioxidant activity; (3) a fluorescent microscopy viability test. In vitro pollutant testing was devised to simulate contaminant loadings that impact urban retention ponds. E. gracilis was found to be tolerant to FeSO₄ (2–10% (w/v)) and NaCl (10–5000 ppm) as indicated by high positive viabilities (ca. 100%) and low, or no OH production, as compared to controls. Total antioxidant activity increased with increasing pollutant loading suggesting that the organism has the capacity to enhance antioxidant defence in response to pollutant stress. This in vitro test provides a new approach to monitor the effects of water quality on the biological components of urban and/or polluted aquatic ecosystems. It also has a potential application in the identification of putative algal phytoremediators.     

The expression of myogenic regulatory factors and muscle growth factors in the masticatory muscles of dystrophin-deficient (MDX) mice

The activities of myogenic regulatory factors (MRF) and muscle growth factors increase in muscle that is undergoing regeneration, and may correspond to some specific changes. Little is known about the role of MRFs in masticatory muscles in mdx mice (the model of Duchenne muscular dystrophy) and particularly about their mRNA expression during the process of muscle regeneration. Using Taqman RT-PCR, we examined the mRNA expression of the MRFs myogenin and MyoD1 (myogenic differentiation 1), and of the muscle growth factors myostatin, IGF1 (insulin-like growth factor) and MGF (mechanogrowth factor) in the masseter, temporal and tongue masticatory muscles of mdx mice (n = 6 to 10 per group). The myogenin mRNA expression in the mdx masseter and temporal muscle was found to have increased (P < 0.05), whereas the myostatin mRNA expressions in the mdx masseter (P < 0.005) and tongue (P < 0.05) were found to have diminished compared to those for the controls. The IGF and MGF mRNA amounts in the mdx mice remained unchanged. Inside the mdx animal group, gender-related differences in the mRNA expressions were also found. A higher mRNA expression of myogenin and MyoD1 in the mdx massterer and temporal muscles was found in females in comparison to males, and the level of myostatin was higher in the masseter and tongue muscle (P < 0.001 for all comparisons). Similar gender-related differences were also found within the control groups. This study reveals the intermuscular differences in the mRNA expression pattern of myogenin and myostatin in mdx mice. The existence of these differences implies that dystrophinopathy affects the skeletal muscles differentially. The finding of gender-related differences in the mRNA expression of the examined factors may indicate the importance of hormonal influences on muscle regeneration.     

Aging, sex differences, and oxidative stress in human respiratory and limb muscles

Oxidative stress is involved in the sarcopenia of aging muscles. On the grounds that ventilatory muscles are permanently active, and their activity may even increase with aging, we hypothesized that the levels of oxidative stress would probably be increased in the external intercostals of elderly healthy individuals. We conducted a case-control study in which reactive carbonyl groups, malondialdehyde-protein adducts, 3-nitrotyrosine immunoreactivity, Mn-superoxide dismutase (Mn-SOD), and catalase were detected using immunoblotting in external intercostals and quadriceps (open muscle biopsies) obtained from 12 healthy elderly and 12 young individuals of both sexes. In elderly subjects, reactive carbonyls, malondialdehyde-protein adducts, 3-nitrotyrosine, Mn-SOD, and catalase were significantly greater in the external intercostals than in the young controls. A post hoc analysis, in which men and women from both groups were analyzed separately, revealed that the external intercostals of elderly women, but not those of elderly men, showed significantly increased levels of reactive carbonyls, malondialdehyde-protein adducts, 3-nitrotyrosine, and Mn-SOD compared to those of control females. This study suggests that differences in muscle activity might explain the differential pattern of oxidative stress observed in human respiratory and limb muscles with aging as well as the likely existence of a sex-related regulation of this phenomenon in these muscles.