Oxidative Stress in Hemodialyzed Patients and the Long-Term Effects of Dialyzer Reuse Practice

Clinical Biochemistry, Vol. 30, No. 8, 601– 606, 1997 Copyright © 1997 The Canadian Society of Clinical Chemists Printed in the USA. All rights reserved 0009-9120/97 $17.00 1 .00

PII S0009-9120(97)00100-8

Oxidative Stress in Hemodialyzed Patients and the Long-Term Effects of Dialyzer Reuse Practice KADER KO¨SE,1 PAKIZE DOG˘AN,1 ZU¨BEYDE GU¨NDU¨Z,2 RUHAN DU¨S¸U¨NSEL,2 and CENGIZ UTAS¸2 Department of 1Biochemistry and 2Nephrology, Faculty of Medicine, Erciyes University, 38039 Kayseri, Turkey Objectives: To investigate the existence of an altered oxidant/ antioxidant balance in patients on regular hemodialysis treatment (RHT) and whether there is any effect of dialyzer reuse on oxidative damage and antioxidative mechanism. Design and methods: Malondialdehyde (MDA) levels and glutathione peroxidase (GPx) activities in both plasma and erythrocytes, plasma selenium (Se) levels, and erythrocyte superoxide dismutase (SOD) activities of RHT patients were determined at the beginning and end of 4-month reuse period. Results: When compared to healthy controls, both plasma and erythrocyte MDA levels were found to be significantly higher in RHT patients before the dialyzer reuse practice; whereas both plasma and erythrocyte GPx activities, erythrocyte SOD activity, and also plasma Se levels were lower in the same patient group than those of controls. When statistical comparison was made on RHT patients between before and after the reuse period, the decreases in MDA levels but increases in the enzyme activities and also an increase in plasma Se levels were observed after the reuse period. However, erythrocyte SOD activities and plasma Se levels measured after the reuse period were not found to be statistically different from the control values; MDA levels still remained elevated above the control values, and GPx activities were not attained to those of controls, after the reuse practice. In addition, positive correlations were found between activities of erythrocyte SOD and GPx enzymes, between GPx and Se levels and negative correlations between the activities of both enzymes and MDA levels in erythrocytes of patients on RHT. Conclusion: These findings may indicate that dialyzer reuse may provide, at least partly, an improvement on oxidative stress in patients on RHT.

KEY WORDS: hemodialysis; lipid peroxidation; malondialdehyde; superoxide dismutase; glutathione peroxidase; selenium; dialyzer reuse.

Introduction he occurrence of reactive oxygen species (ROS), known as oxidants, is an attribute of normal aerobic life. The steady-state formation of oxidants is balanced by a similar rate of their consumption by antioxidants (1). Mainly, antioxidant enzymes are

T

Correspondence: Kader Ko¨se, Department of Biochemistry, Faculty of Medicine, Erciyes University, 38039 Kayseri, Turkey. E-mail: [email protected] Manuscript received March 20, 1997; revised and accepted June 20, 1997. CLINICAL BIOCHEMISTRY, VOLUME 30, DECEMBER 1997

superoxide dismutase (SOD; E.C.1.15.1.1) and hydroperoxidases such as selenium (Se)-dependent glutathione peroxidase (GPx; E.C.1.11.1.9) (1,2). SOD catalyzes the conversion of superoxide anion (O•2 2 ) into hydrogen peroxide (H2O2); and GPx reduces all types of hydroperoxides to nontoxic alcohols (2). Oxidative damage inflicted by ROS is also referred to as “oxidative stress” and reflects of a shift in the oxidant/antioxidant balance in favor of the former (1). ROS-mediated oxidation of membrane lipids results in formation of lipid peroxidation products such as malondialdehyde (MDA); in addition, lipid peroxidation is generally believed to be a significant factor in various pathological events (3). Several studies have also suggested that the oxidant/antioxidant imbalance exists in patients on regular hemodialysis treatment (RHT) (4 –12), whereas the results have not been concordant for the time being. Recently, the growing practice of dialyzer reuse in many dialysis units is mainly based on medical and economic considerations: dialyzer reuse is associated with less intradialytic symptoms compared to first use (13,14), there is no increase in morbidity or mortality, and a substantial reduction in cost (14 –16). Although the beneficial effects of dialyzer reuse were reported in several studies (13–16), there are a few studies related to the effects of dialyzer reuse on the antioxidant activity and lipid peroxidation (17,18). The aim of the present study was to investigate the relationships between enzymatic mechanisms of radical detoxification, lipid peroxidation in patients on RHT and also long-term effects of dialyzer reuse practice on these phenomena. For this reason, MDA levels and GPx activities in both plasma and erythrocytes, plasma Se levels, and erythrocyte SOD activities of hemodialyzed patients were determined at the beginning and after 4 months of the reuse period, and compared to those levels of healthy controls. 601

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Materials and methods SUBJECTS The study included 12 patients (7 females and 5 males) with end-stage renal failure on RHT. Their ages ranged from 16 to 50 years (mean 6 SD, 32.8 6 12.7 years). Primary renal diseases included: chronic glomerulonephritis (six cases), chronic pyelonephritis (two cases), chronic interstitial nephritis (one case), polycystic kidney disease (one case), and two unknown etiologies. Exclusion criteria were patients with diabetes, chronic respiratory insufficiency, intercurrent infection, and hepatic disorders. None of them had received antioxidant drugs such as vitamin E, erythropoietin, etc., during the study period. The patients were initially on first use and the duration of hemodialysis treatment of the patients was between 10 and 46 months (mean 6 SD, 20.9 6 10.1) then shifted to reuse. The adequacy of dialysis was monitored by measuring serum urea nitrogen and creatinine levels of patients, monthly. Of these patients, five were dialyzed twice weekly and seven were dialyzed three times weekly. Hemodialysis was performed for 4 hours using cuprophan hollow-fiber dialyzers (surface area 1.2 m) with blood flow rates of 200 mL/min and the acetate dialysate flow rates of 500 mL/min. The patients included had never been treated by reuse before. The practice of dialyzer reuse was performed in the patients on RHT for 4 months with cuprophan dialyzers, which were washed by using the ECHO (Mesa Medical Inc., Wheat Ridge, Colorado 80033, USA) dialyzer reuse machine with 2% formaldehyde. Procedures of dialyzer reuse were adapted from Kant et al.’s study (15). During this practice, total 432 reuse processes were performed for 79 dialyzers (mean 6 SD, 6.8 6 4.1; range 1 to 15). A total of 12 healthy volunteers (5 females and 7 males, aged 15 to 44 years, mean age 29.4 6 8.3 years), without clinical or laboratory evidence of any disease were also included in the present study, as control group. This study received ethical committee approval and informed consent was obtained from all participants. METHODS Laboratory tests were carried out on 10 mL of blood drawn into preservative-free heparinized disposable syringes from the antecubital vein of control subjects, and the arteriovenous fistula of dialyzed patients before and after the reuse period. Whole blood, transported to polypropylene tubes free of trace elements, was kept at 4°C for 30 min, then centrifuged at 1500 3 g for 15 min to separate plasma and erythrocytes. The plasma was subdivided and stored at 270°C for a maximum of 15 days before analyses of GPx activity, MDA, and Se levels. 602

The erythrocyte pellet was washed three times with cold isotonic saline, and then diluted with saline to the original blood volume. The susceptibility of erythrocytes to lipid peroxidation was determined immediately; remaining packed cells were stored at 270°C for a maximum of 15 days before analyses of GPx and SOD activities. The degree of lipid peroxidation in erythrocytes and plasma samples was assessed by methods based on measuring the concentration of the pink chromogen compound, which forms when MDA couples to thiobarbituric acid (TBA) (19,20). A spectrophotometric assay, modified by Jain (19), was used to determine the susceptibility of erythrocytes to lipid peroxidation. Hemoglobin concentrations of erythrocyte suspensions were also measured with Drabkin’s reagent. The MDA levels of groups were expressed in nmol MDA per gram of hemoglobin (nmol MDA/g Hb). Plasma MDA levels were assayed by a spectrophotometrical method (20), which was partly modified in our laboratory, excluding the high pressure chromatography separation step. Briefly, the mixture of plasma, phosphoric acid, and TBA was boiled at 100°C for 60 min. After cooling, equal volumes of n-butanol were added to the medium and mixed vigorously. The butanol-phase was separated by centrifugation and absorbance was measured at 532 nm. The CV value of the modified method presented was found to be 3.54% in our laboratory. MDA levels were expressed in micromoles MDA per liter (mmol MDA/L). GPx activity in both plasma and erythrocytes was determined by the coupled assay described by Paglia and Valentine (21) using H2O2 as substrate. The assay kinetics were calculated by using a molar absorbtivity of NADPH of 6.200 M21 cm21 at 340 nm. The results were expressed as micromoles of NADPH oxidized per min and per gram of hemoglobin (U/g Hb) for erythrocyte GPx and as international units per liter (U/L) for plasma GPx. Plasma Se levels were assayed by a direct graphite furnace atomic absorbtion spectro-photometry (Hitachi Z-8000, Tokyo, Japan), using Zeeman-effect background correction (22). The results were expressed in micromoles of Se per liter (mmol Se/L). Erythrocyte SOD activity was detected by its ability to inhibit the reduction of nitroblue tetrazolium (NBT), with xanthine-xanthine oxidase used as a O•2 generator (23). One unit of SOD activity is 2 defined by the amount of the enzyme required to inhibit the rate of NBT reduction by 50% as determined from the standard curve generated with bovine SOD (Sigma). The results were expressed as specific activity, units per gram hemoglobin (U/g Hb). STATISTICAL

ANALYSIS

The values were expressed as the mean 6 SD (1 SD). Unpaired t-test was used in statistical comparison of the data from patients and controls. The significance of the differences between before and CLINICAL BIOCHEMISTRY, VOLUME 30, DECEMBER 1997

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TABLE 1 Plasma and Erythrocyte Values of Healthy Controls and Hemodialyzed Patients Before and After Reuse Patients (n 5 12) Before Reuse

After Reuse

Controls (n 5 12)

39.50 6 10.20a 19.79 6 2.90a 637.15 6 134.72a

17.28 6 3.30a,c 24.55 6 3.93a,d 781.32 6 96.21e

6.69 6 0.81 44.24 6 7.05 859.08 6 81.48

3.53 6 0.62a 292.59 6 43.94a 2.06 6 0.62b

2.63 6 0.40b,e 343.40 6 47.57b,d 2.49 6 0.50

2.22 6 0.32 399.08 6 67.23 2.72 6 0.47

Parameters Erythrocyte MDA (nmol/g Hb) GPx (U/g Hb) SOD (U/g Hb) Plasma MDA (mmol/L) GPx (U/L) Se (mmol/L)

Values are mean 6 SD. n, number of subjects. Between control and patients (control vs patients before reuse and controls vs. patients after reuse: ap , 0.001; bp , 0.05. Between patients before and after reuse: cp , 0.001; dp , 0.01; ep , 0.05.

after the reuse period was calculated by paired t-test. In addition, correlation coefficient was determined by linear regression analysis performed between biochemical parameters measured in all groups. P values # 0.05 were considered significant. Results The data of the present study are represented in Table 1. Although there were significant increases in both plasma and erythrocyte MDA levels in RHT patients before the dialyzer reuse practice when compared with controls, it was observed that the long-term trial of the dialyzer reuse led to the significant decreases in plasma and erythrocyte MDA levels in the same patients when compared with their initial levels. However, these decreases in MDA levels could not attained to control values. However, plasma and erythrocyte GPx activities were found to be lower in patients both before and after the reuse period than those of control group. But, GPx activities in both plasma and erythrocytes significantly increased with respect to its beginning level in patients after the dialyzer reuse process. Plasma Se levels were also found to be lower in hemodialyzed patients before reuse, whereas no

statistical difference was found for the plasma Se levels of patients after the reuse compared with controls, and also there was no significant difference between Se levels of patients before and after the reuse. Although there was significant decrease in erythrocyte SOD activity in patients before reuse compared with controls, SOD activity was found to be significantly higher after the practice of dialyzer reuse and there was no significant difference between SOD activities of patients after the reuse and of controls. When correlation analyses were performed, positive and significant correlations were observed between plasma GPx-Se; plasma GPx-erythrocyte GPx; erythrocyte GPx-Se; erythrocyte GPx-erythrocyte SOD parameters in both controls and patients before and after the reuse. However, negative and significant correlations were present between MDAGPx and MDA-SOD in erythrocytes in only patients group (Table 2). No correlation was observed among other parameters. Discussion Chronic renal failure (CRF) is accompanied by a complex pathology. Although the role of ROS in CRF

TABLE 2 Correlation Analyses, Between Parameters Measured in Patients and Controls Patients (n 5 12) Controls (n 5 12)

Before Reuse

After Reuse

Parameters

r

p

r

p

r

p

Plasma GPx-plasma Se Erc GPx-plasma Se Erc GPx-plasma GPx Erc GPx-Erc SOD Erc MDA-Erc GPx Erc MDA-Erc SOD

0.815 0.640 0.714 0.688 20.356 20.220

,0.001 ,0.05 ,0.01 ,0.05 NS NS

0.828 0.621 0.696 0.678 20.628 20.584

,0.001 ,0.05 ,0.05 ,0.05 ,0.05 ,0.05

0.814 0.725 0.672 0.780 20.694 20.765

,0.001 ,0.01 ,0.01 ,0.01 ,0.05 ,0.01

Erc, erythrocyte; n, number of subjects; NS, no significant. CLINICAL BIOCHEMISTRY, VOLUME 30, DECEMBER 1997

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is not definitely demonstrated, an increase in oxidative metabolism in blood granulocytes has been described in end-stage renal failure and in RHT patients (24). Possibly small amounts of plasma constituents such as IgG and complement components are bound to the dialysis membrane thereby creating a surface biologically active for granulocytes (25). Since the activation of peripheral blood granulocytes during hemodialysis may lead to increased generation of ROS (26,27). Increased plasma myeloperoxidase (MPO) activity in hemodialyzed patients (25,28), also shown in our previous study (29), describes the existence of neutrophil activation, which may lead to the lipid peroxidation. Indeed, lipid peroxidation has invariably been found to be accelerated in both plasma (4 – 6) and erythrocytes (6 –9) of patients on RHT, characterized by elevated levels of MDA. Our findings showing high MDA levels in both plasma and erythrocytes in patients group before reuse are in good agreement with these studies. Altered antioxidant enzymedefense mechanisms in the erythrocytes seem to be one of the important factors leading to peroxidation. However, the data on hemodialyzed patients about SOD and GPx activities, considered to be major determinants of the antioxidant capacity of erythrocytes, are controversial: SOD activity was found to be either decreased (5– 8), increased (9) or unchanged (4); GPx was also variably reported to be decreased (7–12) or unchanged (4) in erythrocytes of hemodialyzed patients. In agreement with some of these reports, erythrocyte SOD and GPx activities were found to be lower in our patients on RHT before reuse. Decreases in erythrocyte SOD activity could be explained by increased H2O2 concentrations (30). Because of the decreased GPx activity in erythrocytes in these patients, the accumulation of H2O2 can inhibit SOD (2). The increase in H2O2 may originate from either excess releasing of H2O2 from activated neutrophils or excess production of H2O2 in erythrocytes in patients on RHT. In agreement with the present study, blood Se levels are frequently reported to be lower in hemodialyzed patients (5,9). Decreased Se levels in hemodialyzed patients may be attributed to inadequate intake or to redistribution from the plasma pool into tissues as defense mechanism against oxidative processes. However, the integrity of GPx requires adequate intake of Se (1,2), and Se deficiency results not only in a decrease of GPx activity, but also in a decrease of GPx-protein (31). In addition, GPx may be inactivated during oxidative stress; also, superoxide anion itself can inhibit peroxidase function (32). Excess O•2 2 generation by activated neutrophils may directly lead to inactivation of GPx activity in hemodialyzed patients. Since decreased erythrocyte SOD activity seems to be together with low GPx levels in many studies related to hemodialyzed patients (7–11), GPx must be considered to be complementary to SOD. For this reason, the activity of GPx may be expected to 604

correlate with that of SOD. Indeed, in both patients and controls strong positive correlations were found between activities of these two erythrocyte enzymes in the present study. This might represent an appropriate response for continuing erythrocyte integrity. Significant negative correlations were also found between the activities of both enzymes and MDA levels in erythrocytes of patients on RHT. These findings may contribute to the consideration about the existence of decreased efficiency of the cellular defense mechanism against ROS, which lead to lipid peroxidation (5,7,8). The present study confirms that there was significant decrease in plasma GPx activity of patients on generation by activated RHT (5–12). Excess O•2 2 neutrophils (32), high plasma MDA levels (33) may be responsible for the decreased plasma GPx levels in these patients. Apart from these, Se deficiency may be a factor for low GPx activities. Since strong correlations between Se and GPx activities in both plasma and erythrocytes were found in the present study. According to the present findings, it might be suggested that chronic hemodialysis treatment may induce an increased generation of lipid peroxidation products, accompanied by a significant depression of GPx activities in both plasma and erythrocytes, and also of erythrocyte SOD activity in these patients. However, significant alterations were observed in these parameters after dialyzer reuse practice: increases in enzyme activities, but decreases in MDA levels in both plasma and erythrocytes were detected; although MDA levels remained elevated above healthy control values. Reuse practice seems to be beneficial with regard to oxidative stress. The present study shows the effects of long-term use of dialyzer reuse on oxidative metabolism in patients on RHT for the first time. To our current knowledge, there have been only two studies showing short-term effects of dialyzer reuse on oxygen radical production and lipid peroxidation in hemodialyzed patients; which were controversial (17,34). Markert et al. (34) have found depression of oxygen radical production by isolated peripheral blood neutrophils at 15 min after the start of hemodialysis during the first use of the cuprophan dialyzer; this situation was normalized upon the reuse of the membranes. In contrast to this study, Trznadel et al. (17) have reported that whole blood O•2 2 generation and erythrocyte SOD activity were decreased, plasma MDA levels were increased, but erythrocyte MDA levels were not changed with consecutive uses of the same dialyzer, compared to those before hemodialysis with first-used dialyzer. The findings of Trznadel et al. (17) seem to indicate that dialyzer reuse may exert beneficial effects on whole blood O•2 2 generation by reducing the activation of neutrophils, and protect erythrocyte membrane lipids from peroxidation. However, the practice of dialyzer reuse was performed for 4 months in the present study. At the end of this period, plasma and erythrocyte MDA levels were decreased significantly in CLINICAL BIOCHEMISTRY, VOLUME 30, DECEMBER 1997

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contrast to Trznadel’s study (17), while enzyme activities, GPx and especially SOD, were increased. During hemodialysis, in addition to complement activation (28,34) direct contact of the blood with an insufficiently biocompatible dialysis membrane may induce degranulation and increased oxidative metabolism of peripheral blood neutrophils, leading to augmented ROS generation (26,27). However, dialyzer reuse has been postulated to improve biocompatibility by attenuating the changes in a variety of blood constituents on exposure to the dialysis membrane (35). The significant reduction in the ability of the dialyzer to activate the release of complement constituents during the following uses of membranes has also been reported (13). Indeed, plasma proteins of various molecular weight, especially fibrinogen, form a layer by adsorbing on first-used dialyzer membrane (36), and prevent direct bloodcell-membrane interactions, which leading to improve membrane biocompatibility on following uses (13,36). Therefore, neutrophil activation was found to be proportional to the amount of protein adsorbed on dialyzer membrane (36). When all of these are taken into consideration, reuse practice appears to be more efficient in the reduction of activated neutrophils as well as in removing from plasma oxidants capable of altering erythrocyte integrity. In conclusion, the rises of SOD and GPx activities in hemodialyzed patients at the end of reuse period may imply improved antioxidant defense depending on the reduced oxidative damage by ROS, as reflected by decreases in MDA levels. Acknowledgement This study was supported by the Research Foundation of Erciyes University.

References 1. Sies H. Oxidative stress: from basic research to clinical application. Am J Med 1991; 91(suppl 3C): 31S– 37S. 2. Bast A, Haenen GRMM, Doelman CJA. Oxidants and antioxidants: state of the art. Am J Med 1991; 91(suppl 3C): 2S–13S. 3. Yagi K. Lipid peroxides and human diseases. Chem Phys Lipids 1987; 45: 337–51. 4. Asayama K, Shiki Y, Ito H, et al. Antioxidant enzymes and lipoperoxide in blood in uremic children and adolescents. Free Radic Biol Med 1990; 9: 105–9. 5. Richard MS, Arnaud J, Jurkovitz C, et al. Trace elements and lipid peroxidation abnormalities in patients with chronic renal failure. Nephron 1991; 57: 10 –15. 6. Trznadel K, Pawlicki L, Kedziora J, Luciak M, Blacszczyk J, Buczynski A. Superoxide anion generation, erythrocytes superoxide dismutase activity, and lipid peroxidation during hemoperfusion and hemodialysis in chronic uremic patients. Free Radic Biol Med 1989; 6: 393–7. 7. Tu´ri S, Nemeth I, Vargha I, Matkovies B, Dobos E. Erythrocyte defense mechanisms against free oxygen radicals in hemodialyzed uremic children. Pediatr Nephrol 1991; 5: 179 – 83. CLINICAL BIOCHEMISTRY, VOLUME 30, DECEMBER 1997

8. Paul JL, Sall ND, Soni T, et al. Lipid peroxidation abnormalities in hemodialyzed patients. Nephron 1993; 64: 106 –9. 9. Eiselt J, Racek J, Holocek V, Jerabek Z, Opatrny K. Lipid peroxidation and the antioxidant system in bicarbonate hemodialysis and acetate-free biofiltration. Vnitrni Lekarstvi 1995; 41: 235–9. ¨ , Bas¸es¸me E, Canbolat O, Kavutcu 10. Durak I, Akyol O M. Reduced erythrocyte defense mechanisms against free radical toxicity in patients with chronic renal failure. Nephron 1994; 66: 76 – 80. 11. Mimic-oka J, Simic T, Ekmescic V, Dragicevic P. Erythrocyte glutathione peroxidase and superoxide dismutase activities in different stages of chronic renal failure. Clin Nephrol 1995; 44: 44 – 8. 12. Saint-Georges MD, Bonnefont DJ, Bourely BA, et al. Correction of selenium deficiency in hemodialyzed patients. Kidney Int 1989; 27(suppl): 274 –7. 13. Shusterman NH, Feldman HI, Wasserstein A, Strom BL. Reprocessing of hemodialyzers: a critical appraisal. Am J Kidney Dis 1989; 15: 81–91. 14. Baris E, McGregor M. The reuse of hemodialyzers: an assessment of safety and potential savings. Can Med Assoc J 1993; 48: 175– 83. 15. Kant KS, Pollack VE, Cathey M, Goetz D, Berlin R. Multiple use of dialyzers: safety and efficiency. Kidney Int 1981; 19: 728 –38. 16. Pollack VE, Kant KS, Parnell SL, Levin NW. Repeated use of dialyzers is safe: long term observations on morbidity and mortality in patients with end-stage renal disease. Nephron 1986; 42: 217–23. 17. Trznadel K, Luciak M, Pawlicki L, Kedziora J, Blaszczyk J, Buczynski A. Superoxide anion generation and lipid peroxidation processes during hemodialysis with reused cuprophan dialyzers. Free Radic Biol Med 1990; 8: 429 –32. 18. Gu¨ndu¨z Z, Utas¸ C, Ko¨se K, Du¨s¸u¨nsel R, Dog˘an P. The effects of dialyzer reuse on lipid peroxidation in hemodialysis patients (Abstract). EDTA-ERA Congress April 1994, Vienna, Pp. 184. 19. Jain SK. Evidence for membrane lipid peroxidation during the in vivo aging of human erythrocytes. Biochim Biophys Acta 1988; 937: 205–10. 20. Wong SHY, Knight JA, Hopfer SM, Zaharia O, Leach CN, Sunderman FW. Lipoperoxides in plasma as measured by liquid-chromatographic seperation of malondialdehyde-thiobarbituric acid adduct. Clin Chem 1987; 33: 214 –20. 21. Paglia DE, Valentine WN. Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase. J Lab Clin Med 1967; 70: 158 – 69. 22. Jacobson BE, Lockitch G. Direct determination of selenium in serum by graphite-furnace atomic absorbtion spectrometry with deuterium background correction and a reduced palladium modifier: age specific reference ranges. Clin Chem 1988; 34: 709 –14. 23. Sun Yi, Oberley LW, Li Y. A simple method for clinical assay of superoxide dismutase. Clin Chem 1988; 34: 497–500. 24. Paul JL, Roch-Arveiller M, Man NK, Luong N, Moatti N, Raichuarg D. Influence of uremia on polymorphonuclear leukocytes oxidative metabolism in end-stage renal disease and dialyzed patients. Nephron 1991; 57: 428 –32. 25. Ha¨llgren R, Venge P, Danielson BG. Neutrophil and eosinophil degranulation during hemodialysis are mediated by the dialysis membrane. Nephron 1982; 32: 329 –34. 605

¨ SE ET AL. KO

26. Kuwahara T, Markert M, Wauters JP. Neutrophil oxygen radical production by dialysis membranes. Nephrol Dial Transplant 1988; 3: 661–5. 27. Luciak M, Trznadel K. Free oxygen species metabolism during hemodialysis with different membranes. Nephrol Dial Transplant 1991; 3(suppl): 66 –70. 28. Ho¨rl WH, Riegel W, Schollmeyer P, Rautenberg W, Neumann S. Different complement and granulocyte activation in patients dialyzed with PMMA dialyzers. Clin Nephrol 1986; 25: 304 –7. 29. Gu¨ndu¨z Z, Du¨s¸u¨nsel R, Ko¨se K, Utas¸ C, Dog˘an P. The effects of dialyzer reuse on plasma antioxidative mechanisms in patients on regular hemodialysis treatment. Free Radic Biol Med 1996; 21: 225–31. 30. Fridovich I. Superoxide dismutases. Annu Rev Biochem 1975; 44: 147–51. 31. Takahashi K, Newburger PE, Cohen HJ. Glutathione peroxidase protein. Absence in selenium deficiency states and correlation with enzymatic activity. J Clin Invest 1986; 77: 1402– 4.

606

32. Blum J, Fridovich I. Inactivation of glutathione peroxidase by superoxide radical. Arch Biochem Biophys 1985; 240: 500 – 8. 33. Arshad MAQ, Bhadia S, Cohen RM, Subbiah MTR. Plasma lipoprotein peroxidation potantial: a test to evaluate individual susceptibility. Clin Chem 1991; 37: 1756 – 8. 34. Markert M, Heierli C, Kawahara T, Frei J, Wauters JP. Dialyzed polymorphonuclear neutrophil oxidative metabolism during dialysis: a comparative study with 5 new and reused membranes. Clin Nephrol 1988; 29: 129 –36. 35. Pereira BJ, King AJ, Poutsika DD, Strom JA, Dinarello CA. Comparison of first use and reuse of cuprophan membranes on interleukin-1 receptor antagonist and interleukin-1 beta production by blood mononuclear cells. Am J Kidney Dis 1993; 22: 288 –95. 36. Kuwahara T, Markert M, Wauters JP. Proteins adsorbed on hemodialysis membranes modulate neutrophil activation. Artif Organs 1989; 13: 427–31.

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