Browsing by Author "Mimic-Oka, Jasmina (56022732500)"

Purpose: We aimed to discern the role of glutathione (GSH) associated enzymes in maintaining high GSH levels in renal cell carcinoma (RCC) of the clear cell type and analyze RCC enzyme antioxidant capacity. Since changes in cellular redox balance in RCC might also be related to alterations of glutathione S-transferase (GST) phenotype, GST class α and π expression was also explored. Methods and materials: Human kidney specimens of tumor and distant nontumor regions were obtained from 15 patients with RCC at the time of surgery. The activities of GSH-replenishing enzymes, γ-glutamylcysteine synthetase (γ-GCS), γ-glutamyl transferase (γ-GT), and glutathione reductase (GR), as well as the activities of antioxidant enzymes glutathione peroxidase (GPX) and catalase (CAT) were determined spectrophotometrically. GST α and π class expression was determined by immunoblot. Results: In the course of renal cancerization, significant changes appear in the activities of GSH-replenishing and antioxidant enzymes. The activity of the key enzyme of GSH synthesis, γ-GCS, is up-regulated (P < 0.001), while the activities of γ-GT and GR are down-regulated in renal tumors compared to nontumor tissue (P < 0.001 and P < 0.05, respectively). Activities of GPX and CAT were also down-regulated (P < 0.001 and P < 0.05, respectively) in RCC. Changes in enzyme antioxidant capacity in RCC were associated with decreased GST class α (P < 0.001) and unchanged GST π expression at the protein level. Conclusions: Changes in redox status in RCC as a consequence of decreased enzyme antioxidant capacity, together with altered GST α expression, may be important factors in development and tumor growth. The up-regulation of γ-GCS and high levels of GSH in RCC may be an attempt to limit injury caused by oxidative stress. © 2008 Elsevier Inc. All rights reserved.

Background: The presence of glutathione transferase (GST) M1 null genotype (GSTM1-null) in end-stage renal disease (ESRD) patients is associated with lower overall survival rate in comparison to those with GSTM1-active variants. We examined association between GSTM1 and GSTT1 deletion polymorphisms as well as SNPs in GSTA1/rs3957357 and GSTP1/rs1695 genes with overall and cause-specific cardiovascular mortality in ESRD patients. Methods. Total of 199 patients undergoing hemodialysis were included in the study. Median value of time elapsed from dialysis initiation until the death, or the end of follow-up was 8 ± 5 years. The effect of GSTM1, GSTT1, GSTP1 and GSTA1 gene polymorphisms on predicting overall and specific cardiovascular outcomes (myocardial infarction, MI or stroke) was analyzed using Cox regression model, and differences in survival were determined by Kaplan-Meier. Results: GSTM1-null genotype in ESRD patients was found to be independent predictor of overall and cardiovascular mortality. However, after false discovery rate and Bonferroni corrections this effect was lost. The borderline effect modification by wild-type GSTA1*A/*A genotype on associations between GSTM1-null and analyzed outcomes was found only for death from stroke. Homozygous carriers of combined GSTM1*0/GSTA1*A genotype exhibited significantly shorter time to death of stroke or MI in comparison with carriers of either GSTM1-active or at least one GSTA1*B gene variant. The best survival rate regarding cardiovascular outcome was found for ESRD patients with combined GSTM1-active and mutant GSTA1*B/*B genotype. Conclusions: Combined GSTM1*0/GSTA1*A genotypes might be considered as genetic markers for cardiovascular death risk in ESRD patients, which may permit targeting of preventive and early intervention. © 2014 Suvakov et al.; licensee BioMed Central Ltd.

Purpose: To get more insight into molecular mechanisms underlying oxidative stress and its role in different types of seizure, in this study, oxidative byproducts of proteins, lipids and DNA, as well as, antioxidant enzyme activities were studied in adult patients with epilepsy. Methods: Study was performed in 60 patients with epilepsy and in 25 healthy controls. Plasma protein reactive carbonyl derivatives (RCD) and protein thiol groups (P-SH), byproducts of oxidative protein damage, as well as antioxidant enzyme activities, superoxide dismutase (SOD) and glutathione peroxidase (GPX) were studied spectrophotometrically. Urinary 8-epi-prostaglandin F2α (8-epi-PGF2α) and 8-hydroxy-2′-deoxyguanosine (8-OHdG), representative byproducts of lipid and DNA oxidative damage, respectively, were determined by enzyme immunoassay. Results: RCD levels were significantly increased (p = 0.001), while P-SH content was decreased in patients with first seizure (p = 0.052) compared to controls, independently of the seizure type. Urinary 8-epi-PGF2α and 8-OHdG were significantly increased in patients with epilepsy (p = 0.001 and p = 0.001). Rise in 8-epi-PGF2α was more pronounced in patients with generalized tonic-clonic seizure (GTCS) compared to those with partial seizure (PS). Both SOD and GPX activity were significantly increased in epileptic patients compared to controls (p = 0.001 and p = 0.001), but only SOD activity was significantly higher in patients with GTCS than in those with PS. Conclusions: Data on enhanced protein, lipid and DNA oxidation, together with upregulated antioxidant enzyme activities, confirm the existence of systemic oxidative stress in patients with epilepsy. It might be speculated that post-translational modification to existing functional proteins, particularly alterations to ion channels, might be at least partially responsible for acute early changes in neuronal networks. © 2010 British Epilepsy Association.

Purpose: To get more insight into molecular mechanisms underlying oxidative stress and its role in different types of seizure, in this study, oxidative byproducts of proteins, lipids and DNA, as well as, antioxidant enzyme activities were studied in adult patients with epilepsy. Methods: Study was performed in 60 patients with epilepsy and in 25 healthy controls. Plasma protein reactive carbonyl derivatives (RCD) and protein thiol groups (P-SH), byproducts of oxidative protein damage, as well as antioxidant enzyme activities, superoxide dismutase (SOD) and glutathione peroxidase (GPX) were studied spectrophotometrically. Urinary 8-epi-prostaglandin F2α (8-epi-PGF2α) and 8-hydroxy-2′-deoxyguanosine (8-OHdG), representative byproducts of lipid and DNA oxidative damage, respectively, were determined by enzyme immunoassay. Results: RCD levels were significantly increased (p = 0.001), while P-SH content was decreased in patients with first seizure (p = 0.052) compared to controls, independently of the seizure type. Urinary 8-epi-PGF2α and 8-OHdG were significantly increased in patients with epilepsy (p = 0.001 and p = 0.001). Rise in 8-epi-PGF2α was more pronounced in patients with generalized tonic-clonic seizure (GTCS) compared to those with partial seizure (PS). Both SOD and GPX activity were significantly increased in epileptic patients compared to controls (p = 0.001 and p = 0.001), but only SOD activity was significantly higher in patients with GTCS than in those with PS. Conclusions: Data on enhanced protein, lipid and DNA oxidation, together with upregulated antioxidant enzyme activities, confirm the existence of systemic oxidative stress in patients with epilepsy. It might be speculated that post-translational modification to existing functional proteins, particularly alterations to ion channels, might be at least partially responsible for acute early changes in neuronal networks. © 2010 British Epilepsy Association.

Objectives: Glutathione S-transferase P1 (GSTP1) provides an important link between activity of regulatory stress kinases and apoptotic pathways. It can be hypothesized that up-regulated GSTP1, in TCC, might enhance apoptosis inhibition. We aimed to establish whether relationship between GSTP1 expression and executive (pro-caspase 3, cleaved caspase 3) and regulatory (Bcl-2) apoptotic pathways in TCC exists. Materials and Methods: Samples were obtained from 84 TCC patients (41 consecutive patient with muscle noninvasive and 43 consecutive patients with muscle invasive TCC tumors), who underwent surgery at the Institute of Urology and Nephrology, Clinical Centre of Serbia, during 2006 and 2007. Expression of GSTP1, pro-caspase 3 (CPP32), and Bcl-2, as well as cleaved caspase-3 labeling index (LI) were determined by immunocytochemistry. Levels of expression were correlated with tumor stage, grade, and invasiveness. Results: GSTP1 protein expression was demonstrated in all tumor samples examined. According to GSTP1 status, all tumors were divided into groups with low, moderate, or high GSTP1 status. Expression of CPP32 and cleaved caspase 3 was positive in 80% of TCC patients. Their levels differed significantly between groups with various GSTP1 expression (P < 0.05), with the lowest CPP32 expression and cleaved caspase 3 LI in tumors with high GSTP1 status. Moreover, significant negative correlation was found between GSTP1 level and cleaved caspase 3 LI (r = -0.459, P = 0.041). The positive rate of Bcl-2 protein expression was 48%. Most of the Bcl-2 positive patients exhibited at the same time high GSTP1 positivity (P = 0.078). Significant association with tumor grade and stage was found for all examined parameters except for CPP32 regarding tumor grade. Conclusions: Based on results obtained, we conclude that enhanced GSTP1 expression in TCC of urinary bladder is associated with altered apoptotic pathways. Molecular interplay between GSTP1 and members of apoptotic cascade might, at least partially, play a role in development of invasive characteristics of TCC. © 2011 Elsevier Inc.

The efficiency of human recombinant epoetin in alleviating anemia in hemodialyzed patients has been well documented. However, the effects of rhEPO therapy in correction of antioxidant capacity are not completely explained. In this study we examined both extracellular (plasma) and intracellular (red blood cells) antioxidant potential in hemodialyzed patients before and after three and six months of epoetin treatment by evaluating markers of oxidative stress (malondialdehyde) and antioxidant capacity (thiol groups, superoxide dismutase, and glutathione peroxidase). Six months of treatment with epoetin was followed by significant increases in thiol groups, superoxide dismutase and glutathione peroxidase activities in both plasma and red blood cells of hemodialyzed patients. Hence, during accelerated erythropoiesis, an increase in the number of young hematopoietic cells may replenish erythrocyte superoxide dismutase and glutathione peroxidase activity. However, the consequences of an imbalance between enzymatic antioxidant system (higher superoxide dismutase and lower glutathione peroxidase activity) that exists in these patients are the very high red blood cell and plasma malondialdehyde levels. These results suggest that, in spite of epoetin treatment and improvement in red blood cells and plasma antioxidant capacity, the production of reactive oxygen species overwhelms the intracellular and extracellular antioxidant capacity.

BackgroundIncreased oxidative stress is a hallmark of end-stage renal disease (ESRD). Glutathione S-transferases (GST) are involved in the detoxification of xenobiotics and protection of oxidative damage. We hypothesized that genetic polymorphism in antioxidant enzymes GSTA1, GSTM1, GSTP1 and GSTT1 is more frequent in ESRD and modulates the degree of oxidative stress in these patients.MethodsGSTA1, GSTM1, GSTP1 and GSTT1 genotypes were determined in 199 ESRD patients and 199 age-and gender-matched controls. Markers of protein and lipid oxidative damage [thiol groups, carbonyl groups, advanced oxidative protein products, nitrotyrosine, malondialdehyde (MDA) and MDA adducts], together with total oxidant status and pro-oxidant-antioxidant balance were determined. Results Individual GST polymorphisms influence vulnerability to both protein and lipid oxidation, with GSTM1-null gene variant having the most pronounced effect. Furthermore, a strong combined effect of null/low-activity GSTM1, GSTT1, GSTA1 and GSTP1 genotypes in terms of susceptibility towards oxidative and carbonyl stress was found in ESRD patients. When patients were stratified according to GSTM1 and GSTT1, the highest oxidant damage was noted in those with the GSTM1-null/GSTT1-null genotype. The observed effect was even stronger in patients with the third low-activity GSTP1 or GSTA1 genotype. Finally, the level of oxidative and carbonyl stress was most pronounced in the subgroup of patients with all four null or low-activity GSTM1, GSTT1, GSTP1 and GSTA1 genotypes.ConclusionsAccording to the GST genotype, ESRD patients may be stratified in terms of the level of oxidative and carbonyl stress that might influence cardiovascular prognosis, but could also improve efforts towards individualization of antioxidant treatment. © 2013 The Author.

Objectives: Our aim was to perform a comprehensive analysis of the antioxidant capacity of transitional cell carcinoma (TCC) of urinary bladder and discern the role of enzymes associated with glutathione (GSH) in maintaining high GSH levels in these tumours. Because the redox-sensitive protein glutathione S-transferase P1 (GSTP1) might provide an important link between high antioxidant capacity and inhibition of apoptotic pathways, we also explored how the redox state in tumour cells interacts with the expression of GSTP1. Methods: We examined spectrophotometrically the specific activities of GSH-replenishing enzymes involved in GSH synthesis (γ-glutamylcysteine synthetase, γ-GCS), GSH regeneration (glutathione reductase, GR), and antioxidant protection (glutathione peroxidase, GPX; superoxide dismutase, SOD) in the cytosolic fraction of tumours and the surrounding normal tissue of 30 TCC patients. GSTP1-1 expression was also analyzed. Results: We found a significant increase in the activity of both GSH-replenishing and antioxidant enzymes as well as enhanced GSTP1-1 expression in tumours in comparison with adjacent normal uroepithelium. Mean γ-GCS and GR activities in tumours were about 4- and 2-fold higher, respectively, than in corresponding normal tissue. Expression of GSTP1 correlated significantly with GSH level and γ-GCS and GR activities. GPX and SOD activities in TCC were also markedly increased. Conclusions: Enhanced GSH-replenishing pathways account for increased GSH levels in TCC. Upregulated GPX and SOD also contribute to high antioxidant potential in TCC. Under such conditions, expression of redox-sensitive GSTP1 protein is upregulated. © 2007 European Association of Urology.

Exposure to potential carcinogens is an etiologic factor for renal cell carcinoma (RCC) and transitional cell carcinoma (TCC) of the urinary bladder. Cytosolic glutathione S-transferases (GSTs) are a superfamily of enzymes that protect normal cells by catalyzing conjugation reactions of electrophilic compounds, including carcinogens, to glutathione. Some GST enzymes possess antioxidant activity against hydroperoxides. The most well characterized classes have been named alpha (GSTA), mu (GSTM), pi (GSTP) and theta (GSTT); each of these classes contains several different isoenzymes. Several types of allelic variation have been identified within classes, with GSTM1-null, GSTT1-null and GSTP1-Ile105/Ile105 conferring impaired catalytic activity. The effects of GSTM1 and GSTT1 polymorphism on susceptibility to RCC depend on exposure to specific chemicals. Individuals with the GSTM1-null genotype carry a higher risk for TCC. The roles of GSTT1 polymorphism in TCC and GSTP1 polymorphisms in both cancers are still controversial. During kidney cancerization, expression of GSTA isoenzymes tends to decrease, which promotes the pro-oxidant environment necessary for RCC growth. In the malignant phenotype of TCC of the bladder, upregulation of various GST classes occurs. Upregulation of GSTT1 and GSTP1 might have important consequences for TCC growth by providing a reduced cellular environment and inhibition of apoptotic pathways. © 2009 Macmillan Publishers Limited. All rights reserved.

Objectives: Glutathione S-transferases (GSTs) are a family of enzymes involved in detoxification. Genes encoding for GSTA1, GSTM1, GSTP1, and GSTT1 proteins are polymorphic, which can result in complete or partial loss of enzyme activity. Previous studies have associated polymorphisms of GSTA1, GSTM1, and GSTP1 genes with a higher risk of bladder cancer, but this is still controversial. Potential role of GSTA1 polymorphism in susceptibility to bladder cancer in Whites is lacking. We examined association between GSTA1, GSTM1, GSTP1, and GSTT1 gene variants and bladder cancer risk and evaluated whether they were modified by smoking. Materials and methods: A hospital-based case-control study recruited 201 incidence cases and 122 age-matched controls. Deletion polymorphism of GSTM1 and GSTT1 was identified by polymerase chain reaction method. Single nucleotide polymorphism of GSTA1 and GSTP1 was identified by restriction fragment length polymorphism method. Uniconditional multivariate logistic regression was applied to model association between genetic polymorphisms and bladder cancer risk, as well as effect modification by smoking. Results: No significant difference was observed in the distributions of GSTM1, GSTT1, GSTA1, and GSTP1 gene variants between patients and controls. None of the examined polymorphisms was significantly associated with bladder cancer risk independently. The results of gene-smoking interaction analyses indicated a significant combined effect of smoking and all common GST polymorphisms tested (P for trend = 0.001). However, the most significant effect on bladder cancer risk was observed in smokers carrying lower activity GSTA1-AB/BB and GSTM-null genotype (OR = 3.5, P < 0.05) compared with GSTA1-AA and GSTM1-active non-smokers. Overall, the risk observed did not significantly differ with respect to quantity of cigarettes smoked. However, heavy smokers with GSTM1-null genotype had 2 times higher risk of bladder cancer than GSTM1-null light smokers (OR = 4.8 vs. OR = 2.0) when GSTM1-active non-smokers served as reference group. Smokers carrying both GSTM1-null and GSTA1-AB + BB genotypes exhibited the highest risk of bladder cancer (OR = 2.00, P = 0.123). Conclusions: Null or low-activity genotypes of the GSTA1, GSTM1, GSTT1, and GSTP1 did not contribute independently towards the risk of bladder cancer in our patients. However, in association with smoking, both low activity GSTA1 and GSTM1-null genotype increase individual susceptibility to bladder cancer. © 2013 .

Upregulated glutathione S-transferase P1 (GSTP1) plays an important role in the resistance to apoptosis in transitional cell carcinoma (TCC) of the urinary bladder (UB) and represents a potential target for chemotherapeutic agents. Our aim was to perform a systematic investigation of a glutathione S-transferase (GST) isoenzyme profile (GSTM, GSTP1, and GSTT1) in the upper urinary tract (UUT) TCC and compare it with the GST isoenzyme pattern of the UB TCC and normal urothelium. We examined GST activity spectrophotometrically by using substrates for the overall GST activity, GSTP1, and GSTT1 in the cytosolic fraction. GSTP1 and GSTM expression was analyzed by Western blotting. The results obtained have shown that the overall GST activity was significantly higher in UUT TCC in comparison with urothelium (P < 0.001), which gradually increases with tumor grade (P < 0.05). The mean GSTP1 and GSTT1 activities in UUT TCC were 2- and 3.6-fold higher, respectively, than in the normal urothelium (P < 0.001), and these values did not differ significantly from activities found in the UB TCC. GSTM was expressed in 42% of the UUT TCC and 50% of the UB TCC specimens. The level of GSTM expression was slightly increased in the UUT TCC specimens in comparison with normal urothelium (P > 0.05). We conclude that 3 major cytosolic GST classes, GSTM, GSTP1, and GSTT1, are expressed in the UUT TCC. The isoenzyme profile of GST in the UUT TCC is similar to that observed in the UB TCC; it shows essentially the same alteration of the GST phenotype in the course of cancerization. The association of GSTT1 and GSTP1 upregulation with the malignant phenotype of the UUT TCC might result in resistances to both chemotherapy and apoptosis. © 2010 Mosby, Inc. All rights reserved.

Background: We aimed to study the relationship between markers of oxidative lipid or protein damage and ventricular remodeling and the validity of 8-epi-prostaglandin F2α (8-epi-PGF2α) as an indicator of disease severity in patients with ischemic chronic heart failure (CHF). Patients and methods: We enrolled four groups of 12 patients with varying CHF according to the New York Heart Association (NYHA) classification and 25 controls. Urinary 8-epi-PGF2α and plasma malondialdehyde and protein thiol (P-SH) groups were correlated with echocardiographic indices of remodeling. The reliability of isoprostanes was analyzed by a receiver operating characteristics (ROC) curve. Results: NYHA class III and IV patients exhibited elevated 8-epi-PGF2α levels, increased malondialdehyde concentrations and decreased P-SH groups when compared to controls and NYHA I and II patients. 8-Epi-PGF2α and P-SH groups correlated significantly with indices of remodeling. The ROC curve drawn for 8-epi-PGF2α allowed us to differentiate NYHA class III and IV patients from NYHA class I and II patients with a sensitivity of 95.8% and specificity of 95.8% (cut off 0.84 ng/mg creatinine; area under curve 0.99; P < 0.001). Conclusions: Markers of oxidative damage are unlikely to play a significant role in early stages of CHF. However, they might become important in the course of CHF when their concentrations reach critical levels. Urinary 8-epi-PGF2α is a reliable indicator of symptomatic CHF. © 2008 W. S. Maney and Son Ltd.

Background: We aimed to study the relationship between markers of oxidative lipid or protein damage and ventricular remodeling and the validity of 8-epi-prostaglandin F2α (8-epi-PGF2α) as an indicator of disease severity in patients with ischemic chronic heart failure (CHF). Patients and methods: We enrolled four groups of 12 patients with varying CHF according to the New York Heart Association (NYHA) classification and 25 controls. Urinary 8-epi-PGF2α and plasma malondialdehyde and protein thiol (P-SH) groups were correlated with echocardiographic indices of remodeling. The reliability of isoprostanes was analyzed by a receiver operating characteristics (ROC) curve. Results: NYHA class III and IV patients exhibited elevated 8-epi-PGF2α levels, increased malondialdehyde concentrations and decreased P-SH groups when compared to controls and NYHA I and II patients. 8-Epi-PGF2α and P-SH groups correlated significantly with indices of remodeling. The ROC curve drawn for 8-epi-PGF2α allowed us to differentiate NYHA class III and IV patients from NYHA class I and II patients with a sensitivity of 95.8% and specificity of 95.8% (cut off 0.84 ng/mg creatinine; area under curve 0.99; P < 0.001). Conclusions: Markers of oxidative damage are unlikely to play a significant role in early stages of CHF. However, they might become important in the course of CHF when their concentrations reach critical levels. Urinary 8-epi-PGF2α is a reliable indicator of symptomatic CHF. © 2008 W. S. Maney and Son Ltd.

Objective: To address the role of serum γ-glutamyl transferase (GGT) as a marker of metastases in patients with renal cell carcinoma. Methods: Serum alkaline phosphatase and GGT were determined in 156 patients with localized renal cell carcinoma and 60 patients with metastases as proven by echosonography, computerized tomography and bone scan. The control group consisted of 50 healthy subjects matched for sex and age. Sensitivity and specificity of both enzymes as markers of metastatic disease were compared. In metastatic patients, enzyme activities were analyzed according to the site of metastases. Results: Both alkaline phosphatase and GGT activities were normal in majority of patients with localized renal cell carcinoma and increased in most of the patients with metastatic disease (80% and 70%, respectively). GGT did not significantly differ from alkaline phosphatase in terms of sensitivity (70% vs 80%) and specificity (89% vs 92%). Concerning the site of metastases, high frequencies of increased GGT and alkaline phosphatase were found in patients with liver-only metastases (80% and 90%, respectively). All of the patients with both liver and bone metastases exhibited increased activity of both enzymes. Despite the fact that bone cells do not express GGT, increased activity was found in patients with bone metastases-only (45%), suggesting that enzymes might be released from tumor cells. Conclusions: Our data provided evidence that GGT is a sensitive marker of metastatic renal cell carcinoma. However, findings of abnormal GGT activity cannot specify the site of involvement. © 2007 The Japanese Urological Association.