Glutathione S-transferase Protein Levels Research Articles

Therapeutic Role of a Cysteine Precursor, OTC, in Ischemic Stroke Is Mediated by Improved Proteostasis in Mice.

Oxidative stress aggravates brain injury following ischemia/reperfusion (I/R). We previously showed that ubiquilin-1 (Ubqln1), a ubiquitin-like protein, improves proteostasis and protects brains against oxidative stress and I/R-induced brain injury. Here, we demonstrate that a small molecule compound, L-2-oxothiazolidine-4-carboxylic acid (OTC) that functions as a precursor of cysteine, upregulated Ubqln1 and protected cells against oxygen-glucose deprivation-induced cell death in neuronal cultures. Further, the administration of OTC either at 1h prior to ischemia or 3h after the reperfusion significantly reduced brain infarct injury and improved behavioral outcomes in a stroke model. Administration of OTC also increased glutathione (GSH) level and decreased superoxide production, oxidized protein, and neuroinflammation levels in the penumbral cortex after I/R in the stroke mice. Furthermore, I/R reduced both Ubqln1 and the glutathione S-transferase protein levels, whereas OTC treatment restored both protein levels, which was associated with reduced ubiquitin-conjugated protein level. Interestingly, in the Ubqln1 knockout (KO) mice, OTC treatment showed reduced neuroprotection and increased ubiquitin-conjugated protein level when compared to the similarly treated non-KO mice following I/R, suggesting that OTC-medicated neuroprotection is, at least partially, Ubqln1-dependent. Thus, OTC is a potential therapeutic agent for stroke and possibly for other neurological disorders and its neuroprotection involves enhanced proteostasis.

Biobreeding rat islets exhibit reduced antioxidative defense and N-acetyl cysteine treatment delays type 1 diabetes

Islet-level oxidative stress has been proposed as a trigger for type 1 diabetes (T1D), and release of cytokines by infiltrating immune cells further elevates reactive oxygen species (ROS), exacerbating β cell duress. To identify genes/mechanisms involved with diabetogenesis at the β cell level, gene expression profiling and targeted follow-up studies were used to investigate islet activity in the biobreeding (BB) rat. Forty-day-old spontaneously diabetic lymphopenic BB DRlyp/lyp rats (before T cell insulitis) as well as nondiabetic BB DR+/+ rats, nondiabetic but lymphopenic F344lyp/lyp rats, and healthy Fischer (F344) rats were examined. Gene expression profiles of BB rat islets were highly distinct from F344 islets and under-expressed numerous genes involved in ROS metabolism, including glutathione S-transferase (GST) family members (Gstm2, Gstm4, Gstm7, Gstt1, Gstp1, and Gstk1), superoxide dismutases (Sod2 and Sod3), peroxidases, and peroxiredoxins. This pattern of under-expression was not observed in brain, liver, or muscle. Compared with F344 rats, BB rat pancreata exhibited lower GST protein levels, while plasma GST activity was found significantly lower in BB rats. Systemic administration of the antioxidant N-acetyl cysteine to DRlyp/lyp rats altered abundances of peripheral eosinophils, reduced severity of insulitis, and significantly delayed but did not prevent diabetes onset. We find evidence of β cell dysfunction in BB rats independent of T1D progression, which includes lower expression of genes related to antioxidative defense mechanisms during the pre-onset period that may contribute to overall T1D susceptibility.

Proteome Analysis of Bronchoalveolar Lavage Fluid in Chronic Hypersensitivity Pneumonitis

Hypersensitivity pneumonitis (HP) is an immune-mediated lung disease induced by inhalation of numerous antigens. Pathologically, chronic HP tends to show usual interstitial pneumonia (UIP) and fibrotic nonspecific interstitial pneumonia (fNSIP) patterns. Patients with UIP pattern present insidious onset and a risk for acute exacerbations. To evaluate the proteomic differences of bronchoalveolar lavage fluid (BALF) between UIP and fNSIP patterns, BALF from seven patients with UIP pattern and four patients with fNSIP pattern was examined using two-dimensional gel electrophoresis and mass spectrometry. By individually comparing each BALF sample, we found that the protein levels of surfactant protein A (SP-A), immunoglobulin heavy chain α, α-2 heat shock glycoprotein, haptoglobin β, and immunoglobulin J chain were significantly higher in the patients with UIP pattern than those in the patients with fNSIP pattern. In contrast, the protein levels of glutathione s-transferase, vitamin D-binding protein, and β-actin were significantly higher in the patients with fNSIP pattern than those in the patients with UIP pattern. To confirm the results of SP-A in the BALF proteome, we performed enzyme-linked immunosorbent assay in a larger group. The concentrations of SP-A in BALF from the patients with UIP pattern were significantly higher than those from the patients with fNSIP pattern (2.331 ± 1.656 μg/ml vs. 1.319 ± 1.916 μg/ml, p = 0.034). We identified several proteins that may play roles in the development of pathological differences between UIP and fNSIP patterns of chronic HP.

Increased resistance to hydrogen peroxide‐induced cardiac contracture is associated with decreased myocardial oxidative stress in hypothyroid rats

The purpose of this study was to determine whether decreased oxidative stress would increase the resistance to cardiac contracture induced by H(2)O(2) in hypothyroid rats. Male Wistar rats were divided into two groups: control and hypothyroid. Hypothyroidism was induced via thyroidectomy. Four weeks post surgery, blood samples were collected to perform thyroid hormone assessments, and excised hearts were perfused at a constant flow with or without H(2)O(2) (1 mmol/L), being divided into two sub-groups: control, hypothyroid, control + H(2)O(2), hypothyroid + H(2)O(2). Lipid peroxidation (LPO) was evaluated by chemiluminescence (CL) and thiobarbituric acid reactive substances (TBARS) methods, and protein oxidation by carbonyls assay in heart homogenates. Cardiac tissue was also screened for superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) activities, and for total radical-trapping antioxidant potential (TRAP). Analyses of SOD and glutathione-S-transferase (GST) protein expression were also performed in heart homogenates. Hypothyroid hearts were found to be more resistant to H(2)O(2)-induced contracture (60% elevation in LVEDP) as compared to control. CL, TBARS, carbonyl, as well as SOD, CAT, GPx activities and TRAP levels were reduced (35, 30, 40, 30, 16, 25, and 33%, respectively) in the cardiac homogenates of the hypothyroid group as compared to controls. A decrease in SOD and GST protein levels by 20 and 16%, respectively, was also observed in the hypothyroid group. These results suggest that a hypometabolic state caused by thyroid hormone deficiency can lead to an improved response to H(2)O(2) challenge and is associated with decreased oxidative myocardial damage.

Thyme (Thymus vulgarisL.) Leaves and Its Constituents Increase the Activities of Xenobiotic-Metabolizing Enzymes in Mouse Liver

The effects of thyme (Thymus vulgaris L.) leaves and its phenolic compounds, thymol and carvacrol, on the activities of xenobiotic-metabolizing enzymes, i.e., phase I enzymes such as 7-ethoxycoumarin O-deethylase (ECOD) and phase II enzymes such as glutathione S-transferase (GST) and quinone reductase (QR), were investigated. Mice were fed with a diet containing thyme (0.5% or 2.0%) or treated orally with thymol (50-200 mg/kg) or carvacrol (50-200 mg/kg) once a day for 7 successive days, and then the enzyme activities in the livers were analyzed. Dietary administration of 2% thyme caused slightly but significantly higher ECOD, GST, and QR activities by 1.1-1.4-fold. Thymol (200 mg/kg) treatment resulted in significantly higher ECOD, GST, and QR activities by 1.3-1.9-fold, and carvacrol (200 mg/kg) treatment caused significantly higher ECOD, GST, and QR activities by 1.3-1.7-fold. Thymol-treated animals had significantly higher protein levels of GST alpha and GST micro, and carvacrol-treated animals had significantly higher levels of GST micro. These results imply that thyme contains bifunctional inducers (i.e., substances capable of inducing both phase I and phase II enzymes) and that thymol and carvacrol may account for the effects of thyme.

The influence of diet on the regional distribution of glutathioneS-transferase activity in channel catfish intestine

There is evidence that glutathione conjugates are the major metabolites formed following systemic uptake of carcinogenic contaminants from the intestine. The effect of commercial diet versus a semi-purified diet on the distribution of glutathione S-transferase (GST) activity was examined in proximal, medial, and distal sections of catfish intestine. The bulk of GST activity with 1-chloro-2,4-dinitrobenzene, ethacrynic acid, and 3H-benzo[a]pyrene-4,5-oxide, and the percent cytosolic protein cross-reacting with anti-catfish GST-pi were in the more proximal segments and dropped off distally in the two diet groups. However, the total GST-pi cross-reacting protein in the proximal section was significantly higher in fish fed a chow diet. Western blot analysis revealed pi-class GST to be expressed principally in the proximal intestine. Cytosol samples cross-reacted with antibodies to human GST-alpha, -mu, and -pi, but not -theta, classes. Alpha-like GST isoforms of MW 26,200 and 24,600, absent in sections from fish fed a purified diet, were differentially expressed only in the distal section of chow-fed fish. These results indicate that diet significantly elicits regional differences in GST protein levels, that components of the commercial chow affect GST protein expression in the distal intestine, and that maintenance diet should be taken into consideration during dietary exposure studies.

Enhancement of radiation-inducible hepatic glutathione-S-transferases Ya, Yb1, Yb2, Yc1, and Yc2 gene expression by oltipraz: possible role in radioprotection.

Previous studies have shown that radiation in combination with oltipraz enhances hepatic microsomal epoxide hydrolase expression. The effects of gamma-ray radiation exposure in combination with oltipraz on the expression of hepatic glutathione-S-transferase (GST) subunits Ya, Yb1, Yb2, Yc1, and Yc2 were examined in the rat. Northern RNA blot analyses revealed that GST mRNA levels were altered in response to daily 3- or 0.5-Gy doses of radiation. The hepatic GST mRNA levels were transiently decreased at 3 and 8 hr after a single 3-Gy dose of radiation. The GST Ya, Yb1, Yb2, Yc1, and Yc2 mRNA levels were increased by 2-4-fold at 15 and 24 hr after irradiation with 3 Gy, followed by return to the levels of untreated rats at 48 hr after treatment. The treatment of animals with oltipraz alone resulted in dose-related increases in the GST Ya, Yb1, Yc1, and Yc2 mRNA levels, whereas Yb2 mRNA levels were, minimally increased. Although a single dose of oltipraz (30 mg/kg orally) caused a minimal 2-fold elevation in the hepatic GST Ya mRNA level, exposure of animals to both oltipraz and 3-Gy radiation resulted in a 4-fold relative increase in GST Ya mRNA level, indicating that the Ya mRNA expression was additively enhanced by the combination treatment. The Yb1/2 and Yc1/2 mRNA expressions were also enhanced by oltipraz in combination with radiation. Multiple exposure of rats to daily 0.5-Gy radiation caused time-related increases in GST gene expression. The greatest enhancement in GST expression was observed at 24 hr after a single 0.5-Gy dose of radiation in conjunction with oltipraz (e.g., a 9-fold relative increase in GST Ya), whereas the relative additive increases in GST mRNA were less pronounced at day 3 or 5 after treatment. These increases in the GST mRNA levels were consistent with those in the immunochemically detectable GST protein levels. Histopathological examinations revealed that exposure of rats to radiation (0.5 Gy/day for 3-5 days) caused mild-to-moderate hepatocyte degeneration with sinusoidal congestion, whereas oltipraz (30 mg/kg/day for 3 days) was effective in blocking the radiation-induced liver injury. The enhanced expression of these GST isoforms by oltipraz may be associated in part with its hepatoprotective effect against the injury caused by ionizing radiation.