Effect of Hemodialysis Michal Toborek,
on Lipid Peroxidation and Antioxidant With Chronic Renal Failure Tomasz Wasik, Marian Dr&dz,
Mariusz Klin, Krystyna
System in Patients
Magner-Wrbbel,
and Ewa Kopieczna-Grzebieniak Plasma lipid peroxidation, activity of erythrocyte superoxide dismutase (SOD) and catalase, and serum antioxidant activity (AOA) in uremic patients were examined before and after hemodialysis. An increased level of lipid peroxidation, a decreased serum AOA level, and elevated SOD and normal catalase activity before hemodialysis were observed in uremic patients compared with controls. Hemodialysis was found to produce increased lipid peroxidation, a simultaveous decrease of SOD activity, and lack of any changes in serum AOA and erythrocyte catalase. It is suggested that intensification of lipid peroxidation during hemodialysis could account for accelerated progress of atherosclerosis in patients with renal’insufficiency. Copyright 0 1992 by W.B. Saunders Company
H
IGH CARDIOVASCULAR mortality is commonly observed in patients with chronic renal failure requiring prolonged hemodialysis. l.? However, some doubts have been raised about whether atherosclerosis is accelerated in these patients, and/or whether long-term hemodialysis therapy itself is responsible for this process; there are some well-documented studies forlm3and against4” these contradictory concepts. Metabolic disturbances in uremia, which could contribute to development of atherosclerosis, are not yer well understood. Chronic renal failure is accompanied by hypertension, hyperlipidemia, hyperuricemia, and glucose intolerance. Patients treated by maintenance hemodialysis have a constant pattern of lipid abnormalities: hypertriglyceridemia. enrichment of intermediate-density lipoprotein (IDL) and low-density lipoprotein (LDL) with triglycerides, increased concentration of very-low-density lipoprotein (VLDL),’ and a marked decrease of highdensity lipoprotein (HDL) and a slight decrease of LDL levels.x Several abnormalities in apolipoproteins are also observed: decreased level of apolipoproteins A9 and C-II,‘O elevated concentration of apolipoproteins E and A-IV, presence of apolipoproteins A-IV in VLDL, IDL, and LDL. and presence of apolipoproteins B-48, C, and E in LDL.’ A diminished activity of lipoprotein lipase and hepatic lipase could account for some of these disturbances.” It is possible that other mechanisms are also involved in the development of atherosclerotic lesions in the course of chronic renal failure. An elevated concentration of plasma homocysteine-the sulfurous, atherogenic amino acid’?--found in uremic patients could be one of these factors.13 Trznadel et alI4 and Chauchan et alIs observed an elevated level of erythrocyte and plasma malondialdehyde (MDA), an indicator of lipid peroxidation, in patients with chronic uremia. It has been suggested that lipid peroxidation could participate in the development of anemia in the course of renal insufficiency.13 Free radical reactions, including lipid peroxidation, are considered to be important factors in the pathogenesis of a variety of diseases. Free radicals, especially oxygen-reactive species formed by univalent reduction of oxygen, are characterized by very high chemical activity. They can damage proteins, lipids, carbohydrates, and nucleic acids. Plasma membranes are critical targets of free radical reactions. Reactive oxygen species can easily produce Metabolism,
Vol 41, No 11 (November),
1992:
pp 1229.1232
injuries to cell membranes by initiation of polyunsaturated fatty acid peroxidation, inactivation of membrane enzymes and receptors, depolymerization of polysaccharides, and protein cross-linking and fragmentation. These disturbances result in changes of the membrane structure, fluidity, transport, antigenic character, etc.lh In a variety of cells, the endothelium is very susceptible to oxidative stress and free radical-mediated reactions; it is generally assumed that injury to the endothelium may trigger atherogenesis.” The aim of our report was to determine the effects of hemodialysis itself on plasma lipid peroxidation, activities of antioxidant enzymes, and serum antioxidant activity (AOA), and also to try to estimate whether processes occurring during hemodialysis could contribute to atherosclerosis in patients with chronic renal failure. MATERIALS AND METHODS
Fourteen male patients were included in the study; mean age was 38 + 11 years and mean duration of hemodialysis was 56 ‘-c 11 months. Chronic glomerulonephritis was found to be the primary renal disease in all patients. They were hemodialyzed three times a week, 5 hours daily. using cuprophan, hollow-fiber dialyzers that had an area of 1.2 rn? (Model 120: Gambro. Lund, Sweden). Blood flow was approximately 200 mL/min. and dialysate flow was 500 mlimin. Serum creatinine levels in patients were 1,079 + 210 mmol/L before and 573 2 110 mmol/L after hemodialysis. The control group consisted of 15 healthy, male blood donors having a mean age of 35 2 8 years and creatinine levels of less than 100 mmol/L. Blood was drawn into heparinized tubes before and after hemodialysis. Red blood cells were separated from plasma by centrifugation. washed, diluted four times with buffered normal saline, and hemolyzed. Plasma lipid peroxide levels were determined by the thiobarbituric acid test according to the method of Buege and Aust,‘s superoxide dismutase (SOD) activity was
From the Department of Biochemistr) and Chemistry and the Department of Pharmacology, Silesian Medical Academy. Medvkbw. Poland. Present address: M. T.. Department of Nutrition and Food Science, University of Kentucky 212 Funkhouser Building, Lexington, KY 40506-0054. Address reprint requests to Michal Toborek, MD, Department of Nutrition and Food Science, University of Kentucky, 212 Funkhouser Building, Lexington, KY40506-0054. Copyright 0 1992 by W. B. Saunders Cornpan! 0026.0495192l4111-0014$03.00/O
1230
TOBOREK ET AL
by the method of Misra and Fridovich,19 catalase activity was determined by the method described by Aebi,zo and serum AOA was determined according to the method of Stocks et ak21Serum AOA provides a measurement of the capacity of serum to inhibit spontaneous lipid peroxidation generated in standard brain homogenates. Each test serum is added to the brain homogenate, and MDA (an indicator of lipid peroxidation) levels are determined immediately and after a 60-minute incubation in a 37°C water bath. Each test value is compared with each control value in which buffer is added in place of serum. AOA is calculated using the following equation: AOA = 1 - (nmol MDA/mL [test] - nmol MDA/mL [0 time])/(nmol MDA/mL [control] nmol MDAimL [0 time]) x 100. Total protein content was determined by the modified Lowry method.22 Student’s f test was used to estimate differences between groups. determined
6
0
RESULTS
Control
The obtained results are shown in Figs 1 to 4. Prehemodialysis lipid peroxide levels in patients with renal failure were increased in comparison to those of controls, and their further intensification was noted during the course of hemodialysis (Fig 1). Erythrocyte SOD activity after hemodialysis was lower than at the start of the procedure, but was still higher than normal values (Fig 2). Serum AOA levels remained unchanged throughout hemodialysis, but mean values both before and after hemodialysis were significantly lower than those found in healthy subjects (Fig 3). Similar to serum AOA, erythrocyte catalase activity (Fig 4) was almost the same at the beginning and the end of hemodialysis. DISCUSSION
Although hemodialysis leads to improvement of several biochemical parameters, eg, creatinine and urea levels, blood pressure, and plasma lipid patterns, it can also evoke several harmful atherogenic effects due to bioincompatibil-
Before dialysis
Treatment
After dialysis
Groups
Fig 2. Erythrocyte SOD activity in patients with chronic renal failure before and after hemodialysis and in normal subjects. A unit of SOD is defined as quantity of enzyme required to produce 50% inhibition of the rate of adrenaline oxidation to adrenochrome. Statistical analysis: control versus before dialysis, P < .Ol; control versus after dialysis, P < .Ol; before versus after dialysis, P < .05.
ity of dialyzer components,23 accumulation of polyvinylchloride,24 aluminum,25 and acetate,26 and an altered zinc to copper ratio. 27The effect of hemodialysis on intensification of plasma lipid peroxidation found in the present study can be an additional factor inducing development of atherosclerosis in uremic patients. The increase of lipid peroxidation resulting from hemodialysis could be provoked by bioincompatibility of the dialyzer membrane. Contact with the membrane can lead to
80
60
Control Control
Before dialysis
Before dialysis
Treatment Treatment
After dialysis
After dialysis
Groups
Groups
Fig 1. Plasma lipid peroxide levels in patients with chronic renal failure before and after hemodialysis and in normal subjects [control). Lipid peroxide levels are expressed as nmol MDA/mL plasma. Statistical analysis: control versus before dialysis, P c .05; control versus after dialysis, P < .Ol; before versus after dialysis, P < .Ol.
Fig 3. Serum AOA in patients with chronic renal failure before and after hemodialysis relative to that of healthy subjects. Serum AOA is expressed in terms of percentage f%) inhibition of spontaneous autoxidation of brain homogenates. Statistical analysis: control versus before dialysis, P < .Ol; control versus after dialysis, P < .Ol; before versus after dialysis, NS.
HEMODIALYSIS
AND OXIDATIVE
1231
STRESS oxidation,
hemoprotein inactivation. and protein and nucleic acid degradation.28 Moreover, it seems that the increase of lipid peroxidation during hemodialysis could be intensified by a parallel decrease in the activity of SOD [Fig 21, an antioxidant enzyme that protects cells from the attack of 0;’ by
Control
Before dialysis
Treatment
After dialysis
Groups
Fig 4. Erythrocyte catalase activity in patients with chronic renal failure before and after hemodialysis compared with that of healthy subjects. A unit of cetalase (k) is the rate constant of a first-order reaction catalyzed by the enzyme. Statistical analysis: control versus before dialysis, NS; control versus after dialysis, NS; before versus after dialysis, NS.
allergic reactions arising from sensitization to the membrane component and to membrane-induced complement activation; the first mechanism dominates in the applied hollow-fiber dialyzer. IgA antibodies react against conjugates of serum albumin and residual ethylene oxide, and cause activation and degranulation of mast cells secreting a variety of active biological compounds including histamine, arachidonic acid metabolites, and platelet-activating factorsZ3 Activated neutrophils generate large amounts of superoxide anion radicals (0;‘) via the reaction catalyzed by reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, 202 + NADPH + 20, + NADP+ + H +. 0;’ could participate in the formation of other active oxygen species, eg, hydroxyl radical (OH’) and hydrogen peroxide (H202), of which the former can initiate peroxidation of polyunsaturated fatty acidsz8 However, lipid peroxidation could be not only the cause but also the result of ceil damage.29 When activated leukocytes come into contact with the dialyzer membrane, they release hydrolytic enzymes from lysosomes, thus causing injury to cell plasma membranesZ3 Stimulated neutrophils also release myeloperoxidase into extracellular medium; myeloperoxidase is an enzyme that catalyzes oxidation of a number of halides and pseudohalides. Chloride ion, due to its high extracellular concentration, is preferentially oxidized to hypochlorous acid, a potent oxidizing agent that is approximately 100-fold more reactive than H202. Hypochlorous acid can exert harmful effects on sulfhydryl
catalyzing the dismutation of 0;’ to yield Hz02 and 01.~~ Inhibition of SOD can result not only in an increase of OS’, but also in the elevation of other active oxygen species (eg, OH) and the intensification of lipid peroxidation processes. The reason that SOD activity decreases during hemodialysis could be connected with copper and zinc status in hemodialyzed patients suffering from chronic renal failure. We have found that hemodialysis decreases plasma copper and zinc levels to less than 50% of control values.30 Since copper and zinc are cofactors of cytoplasmic SOD, changes in their concentration can affect this enzymatic activity. We also found that the activity of SOD was much higher in uremic patients than in control healthy subjects of a similar age group (Fig 2). This observation is partially in agreement with the study of Chauchan et ali who found increased activity of SOD before hemodialysis and its normalization after hemodialysis; in contrast, Vanella et a131 and Trznadel et all4 found that erythrocyte SOD activity in patients with chronic renal failure was decreased. It is possible that these reported differences in SOD activity were connected with the severity of disease and frequency of hemodialyses. It may be that SOD activity could reach higher values after its decrease during hemodialysis in less severely ill patients, and that this protective process against 0;’ and lipid peroxidation does not exist in severe cases of renal failure. Decreased serum AOA in patients with chronic renal insufficiency and the lack of significant changes in its value during hemodialysis observed in our study agree with the results of Kuroda et aL3’ In conclusion, the present study confirmed increased levels of plasma lipid peroxides, elevated erythrocyte SOD, activity and decreased serum AOA in patients with chronic renal failure treated by maintenance hemodialysis. We found intensification of lipid peroxidation during hemodialysis and indicated that it was caused by decreased SOD activity. We suggest that hemodialysis can accelerate atherosclerosis by increasing lipid peroxidation in patients with renal insufficiency.
ACKNOWLEDGMENT The authors thank Dr Bernhard Hennig tions and for reviewing the manuscript.
for his useful
sugges-
REFERENCES 1. Wing AJ, Brunner FP, Brynger H, et al: Cardiovascularrelated causes of death and the fate of patients with renovascular disease. Contrib Nephrol41:306-311, 1984 2. Linder
A, Charra
B, Sherrard
DJ, et al: Accelerated
athero-
sclerosis in prolonged maintenance hemodialysis. N Engl J Med 290:697-701.1914 3. Kudoh Y, Imura 0: Study on the atherosclerosis mechanism in chronic hemodialysis. Jpn Circ J 51:631-641, 1987
1232 4. Rostand SG, Kirk KA, Rutsky EA: Relationship of coronary risk factor to hemodialysis-associated ischemic heart disease. Kidney Int 22:304-3081982 5. Burke JF Jr, Frances GC, Moore LL, et al: Accelerated atherosclerosis in chronic-dialysis patients-Another look. Nephron 21:181-1851978 6. Ritz E, Wiecek A, Gnasso A, et al: Is atherosclerosis accelerated in uremia? Contrib Nephrol52:1-9, 1986 7. Nestel PJ, Fidge NH, Than MH: Increased lipoproteinremnant formation in chronic renal failure. N Engl J Med 307:329-333, 1982 8. Felts JM, Zacherle B, Childress G: Lipoprotein spectrum analysis of uremic patients maintained on chronic hemodialysis. Clin Chim Acta 93:127-134. 1979 9. Savide E, Gibson JC, Steward JH, et al: High density lipoprotein in chronic renal failure and after renal transplantation. Br Med J 1:928-930,1979 10. Starpans I, Felts JM, Zacherle B: Apoprotein composition of plasma lipoproteins in uremic patients on hemodialysis. Clin Chim Acta 93:135-143, 1979 11. Mordasini R, Frey F, Flury W, et al: Selective deficiency of hepatic triglyceride lipase in uremic patients. N Engl J Med 29711362-1366, 1977 12. McCully KS, Olszewski AJ, Vezeridis MP: Homocysteine and lipid metabolism in atherogenesis: Effect of the homocysteine thiolactonyl derivatives, thioretinaco and thioretinamide. Atherosclerosis 83:197-206, 1990 13. Kang S-S, Wong PWK, Bidani A, et al: Plasma proteinbound homocyst(e)ine in patients requiring chronic hemodialysis. Clin Sci 65:335-336, 1983 14. Trznadel K, Pawlicki L, Kedziora J, et al: Activity of superoxide dismutase in erythrocytes and malondialdehyde concentration in erythrocytes and plasma in various periods of renal insufficiency. Pol Arch Med Wewn 80:261-266, 1988 15. Chauchan DP, Gupta PH, Nampoothiri MRN, et al: Determination of erythrocyte superoxide dismutase, catalase, glucose-6phosphate dehydrogenase, reduced glutathione and malondialdehyde in uremia. Clin Chim Acta 123:153-159, 1982 16. Slater TF: Free radical mechanisms in tissue injury. Biothem J 222:1-15, 1984 17. Hennig B, Chow CK: Lipid peroxidation and endothelial cell injury: Implication in atherosclerosis. Free Radical Biol Med 4:99-106, 1988 18. Buege JA, Aust SD: Microsomal lipid peroxidation, in
TOBOREK
ET AL
Colowick SP, Kaplan NO (eds): Methods in Enzymology, vol 52. New York, NY, Academic, 1978, pp 302-310 19. Misra HP, Fridovich I: The role of superoxide anion in the autoxidation of epinephrine: A simple assay for superoxide dismutase. J Biol Chem 247:3170-3175. 1972 20. Aebi HE: Catalase, in Bergmeyer HV (ed): Methods of Enzymatic Analysis, vol 3. Weinheim, Germany, Verlag Chemie, 1983, pp 272-286 21. Stocks J, Gutteridge JMC, Sharp RJ, et al: Assay using brain homogenate for measuring the antioxidant activity of biological fluids. Clin Sci Mol Med 47:215-222,1974 22. Hartree ET: Determination of protein. A modification of the Lowry method that gives a linear response. Anal Biochem 481422-427, 1972 23. Colton CK: Quantitation of membrane biocompatibility. Contrib Nephrol59:110-125,1987 24. Lewis LM, Flechtner TW, Kerkay J, et al: Determination of plasticizen levels in serum hemodialysis patients. Trans Am Sot Artif Intern Organs 23:566-572,1977 25. Milliner DS, Malekzadeh M, Liberman E, et al: Plasma aluminium levels in pediatric dialysis patients: Comparison of hemodialysis and continuous peritoneal dialysis. Mayo Clin Proc 621269-274,1987 26. Scribner BH: The role of acetate on atherogenesis in the patients on chronic hemodialysis. Proc Dial Transplant Forum 7:181-182,1977 27. Okuyama S, Mishina H, Hasegawa K, et al: Probable atherogenic role of zinc and copper as studied in chronic hemodialysis patients. Tohoku J Exp Med 138:227-279, 1982 28. Grisham MB, McCord JM: Chemistry and cytotoxicity of reactive oxygen metabolities, in Physiology of Oxygen Radicals. Baltimore, MD, American Physiological Society, 1986, pp 1-18 29. Kappus H: Lipid peroxidation: Mechanisms, analysis, enzymology and biological relevance, in Sies H (ed): Oxidative Stress. London, UK, Academic, 1985, pp 273-310 30. Danch A, Drozdz M, Kopieczna-Grzebieniak E, et al: Serum copper and zinc concentration in patients with chronic renal failure before and after hemodialysis. XXVI Meeting of the Polish Biochemical Society, Gdansk, Poland, September 1990 31. Vanella A, Geremia E, Pinturo R, et al: Superoxide dismutase activity and reduced glutathione content in erythrocytes of uremic patients on chronic dialysis. Acta Haematol 70:312-315, 1983 32. Kuroda M, Asaka S, Tofuku Y, et al: Serum antioxidant activity in uremic patients. Nephron 41:293-298.1985
on Lipid Peroxidation and Antioxidant With Chronic Renal Failure Tomasz Wasik, Marian Dr&dz,
Mariusz Klin, Krystyna
System in Patients
Magner-Wrbbel,
and Ewa Kopieczna-Grzebieniak Plasma lipid peroxidation, activity of erythrocyte superoxide dismutase (SOD) and catalase, and serum antioxidant activity (AOA) in uremic patients were examined before and after hemodialysis. An increased level of lipid peroxidation, a decreased serum AOA level, and elevated SOD and normal catalase activity before hemodialysis were observed in uremic patients compared with controls. Hemodialysis was found to produce increased lipid peroxidation, a simultaveous decrease of SOD activity, and lack of any changes in serum AOA and erythrocyte catalase. It is suggested that intensification of lipid peroxidation during hemodialysis could account for accelerated progress of atherosclerosis in patients with renal’insufficiency. Copyright 0 1992 by W.B. Saunders Company
H
IGH CARDIOVASCULAR mortality is commonly observed in patients with chronic renal failure requiring prolonged hemodialysis. l.? However, some doubts have been raised about whether atherosclerosis is accelerated in these patients, and/or whether long-term hemodialysis therapy itself is responsible for this process; there are some well-documented studies forlm3and against4” these contradictory concepts. Metabolic disturbances in uremia, which could contribute to development of atherosclerosis, are not yer well understood. Chronic renal failure is accompanied by hypertension, hyperlipidemia, hyperuricemia, and glucose intolerance. Patients treated by maintenance hemodialysis have a constant pattern of lipid abnormalities: hypertriglyceridemia. enrichment of intermediate-density lipoprotein (IDL) and low-density lipoprotein (LDL) with triglycerides, increased concentration of very-low-density lipoprotein (VLDL),’ and a marked decrease of highdensity lipoprotein (HDL) and a slight decrease of LDL levels.x Several abnormalities in apolipoproteins are also observed: decreased level of apolipoproteins A9 and C-II,‘O elevated concentration of apolipoproteins E and A-IV, presence of apolipoproteins A-IV in VLDL, IDL, and LDL. and presence of apolipoproteins B-48, C, and E in LDL.’ A diminished activity of lipoprotein lipase and hepatic lipase could account for some of these disturbances.” It is possible that other mechanisms are also involved in the development of atherosclerotic lesions in the course of chronic renal failure. An elevated concentration of plasma homocysteine-the sulfurous, atherogenic amino acid’?--found in uremic patients could be one of these factors.13 Trznadel et alI4 and Chauchan et alIs observed an elevated level of erythrocyte and plasma malondialdehyde (MDA), an indicator of lipid peroxidation, in patients with chronic uremia. It has been suggested that lipid peroxidation could participate in the development of anemia in the course of renal insufficiency.13 Free radical reactions, including lipid peroxidation, are considered to be important factors in the pathogenesis of a variety of diseases. Free radicals, especially oxygen-reactive species formed by univalent reduction of oxygen, are characterized by very high chemical activity. They can damage proteins, lipids, carbohydrates, and nucleic acids. Plasma membranes are critical targets of free radical reactions. Reactive oxygen species can easily produce Metabolism,
Vol 41, No 11 (November),
1992:
pp 1229.1232
injuries to cell membranes by initiation of polyunsaturated fatty acid peroxidation, inactivation of membrane enzymes and receptors, depolymerization of polysaccharides, and protein cross-linking and fragmentation. These disturbances result in changes of the membrane structure, fluidity, transport, antigenic character, etc.lh In a variety of cells, the endothelium is very susceptible to oxidative stress and free radical-mediated reactions; it is generally assumed that injury to the endothelium may trigger atherogenesis.” The aim of our report was to determine the effects of hemodialysis itself on plasma lipid peroxidation, activities of antioxidant enzymes, and serum antioxidant activity (AOA), and also to try to estimate whether processes occurring during hemodialysis could contribute to atherosclerosis in patients with chronic renal failure. MATERIALS AND METHODS
Fourteen male patients were included in the study; mean age was 38 + 11 years and mean duration of hemodialysis was 56 ‘-c 11 months. Chronic glomerulonephritis was found to be the primary renal disease in all patients. They were hemodialyzed three times a week, 5 hours daily. using cuprophan, hollow-fiber dialyzers that had an area of 1.2 rn? (Model 120: Gambro. Lund, Sweden). Blood flow was approximately 200 mL/min. and dialysate flow was 500 mlimin. Serum creatinine levels in patients were 1,079 + 210 mmol/L before and 573 2 110 mmol/L after hemodialysis. The control group consisted of 15 healthy, male blood donors having a mean age of 35 2 8 years and creatinine levels of less than 100 mmol/L. Blood was drawn into heparinized tubes before and after hemodialysis. Red blood cells were separated from plasma by centrifugation. washed, diluted four times with buffered normal saline, and hemolyzed. Plasma lipid peroxide levels were determined by the thiobarbituric acid test according to the method of Buege and Aust,‘s superoxide dismutase (SOD) activity was
From the Department of Biochemistr) and Chemistry and the Department of Pharmacology, Silesian Medical Academy. Medvkbw. Poland. Present address: M. T.. Department of Nutrition and Food Science, University of Kentucky 212 Funkhouser Building, Lexington, KY 40506-0054. Address reprint requests to Michal Toborek, MD, Department of Nutrition and Food Science, University of Kentucky, 212 Funkhouser Building, Lexington, KY40506-0054. Copyright 0 1992 by W. B. Saunders Cornpan! 0026.0495192l4111-0014$03.00/O
1230
TOBOREK ET AL
by the method of Misra and Fridovich,19 catalase activity was determined by the method described by Aebi,zo and serum AOA was determined according to the method of Stocks et ak21Serum AOA provides a measurement of the capacity of serum to inhibit spontaneous lipid peroxidation generated in standard brain homogenates. Each test serum is added to the brain homogenate, and MDA (an indicator of lipid peroxidation) levels are determined immediately and after a 60-minute incubation in a 37°C water bath. Each test value is compared with each control value in which buffer is added in place of serum. AOA is calculated using the following equation: AOA = 1 - (nmol MDA/mL [test] - nmol MDA/mL [0 time])/(nmol MDA/mL [control] nmol MDAimL [0 time]) x 100. Total protein content was determined by the modified Lowry method.22 Student’s f test was used to estimate differences between groups. determined
6
0
RESULTS
Control
The obtained results are shown in Figs 1 to 4. Prehemodialysis lipid peroxide levels in patients with renal failure were increased in comparison to those of controls, and their further intensification was noted during the course of hemodialysis (Fig 1). Erythrocyte SOD activity after hemodialysis was lower than at the start of the procedure, but was still higher than normal values (Fig 2). Serum AOA levels remained unchanged throughout hemodialysis, but mean values both before and after hemodialysis were significantly lower than those found in healthy subjects (Fig 3). Similar to serum AOA, erythrocyte catalase activity (Fig 4) was almost the same at the beginning and the end of hemodialysis. DISCUSSION
Although hemodialysis leads to improvement of several biochemical parameters, eg, creatinine and urea levels, blood pressure, and plasma lipid patterns, it can also evoke several harmful atherogenic effects due to bioincompatibil-
Before dialysis
Treatment
After dialysis
Groups
Fig 2. Erythrocyte SOD activity in patients with chronic renal failure before and after hemodialysis and in normal subjects. A unit of SOD is defined as quantity of enzyme required to produce 50% inhibition of the rate of adrenaline oxidation to adrenochrome. Statistical analysis: control versus before dialysis, P < .Ol; control versus after dialysis, P < .Ol; before versus after dialysis, P < .05.
ity of dialyzer components,23 accumulation of polyvinylchloride,24 aluminum,25 and acetate,26 and an altered zinc to copper ratio. 27The effect of hemodialysis on intensification of plasma lipid peroxidation found in the present study can be an additional factor inducing development of atherosclerosis in uremic patients. The increase of lipid peroxidation resulting from hemodialysis could be provoked by bioincompatibility of the dialyzer membrane. Contact with the membrane can lead to
80
60
Control Control
Before dialysis
Before dialysis
Treatment Treatment
After dialysis
After dialysis
Groups
Groups
Fig 1. Plasma lipid peroxide levels in patients with chronic renal failure before and after hemodialysis and in normal subjects [control). Lipid peroxide levels are expressed as nmol MDA/mL plasma. Statistical analysis: control versus before dialysis, P c .05; control versus after dialysis, P < .Ol; before versus after dialysis, P < .Ol.
Fig 3. Serum AOA in patients with chronic renal failure before and after hemodialysis relative to that of healthy subjects. Serum AOA is expressed in terms of percentage f%) inhibition of spontaneous autoxidation of brain homogenates. Statistical analysis: control versus before dialysis, P < .Ol; control versus after dialysis, P < .Ol; before versus after dialysis, NS.
HEMODIALYSIS
AND OXIDATIVE
1231
STRESS oxidation,
hemoprotein inactivation. and protein and nucleic acid degradation.28 Moreover, it seems that the increase of lipid peroxidation during hemodialysis could be intensified by a parallel decrease in the activity of SOD [Fig 21, an antioxidant enzyme that protects cells from the attack of 0;’ by
Control
Before dialysis
Treatment
After dialysis
Groups
Fig 4. Erythrocyte catalase activity in patients with chronic renal failure before and after hemodialysis compared with that of healthy subjects. A unit of cetalase (k) is the rate constant of a first-order reaction catalyzed by the enzyme. Statistical analysis: control versus before dialysis, NS; control versus after dialysis, NS; before versus after dialysis, NS.
allergic reactions arising from sensitization to the membrane component and to membrane-induced complement activation; the first mechanism dominates in the applied hollow-fiber dialyzer. IgA antibodies react against conjugates of serum albumin and residual ethylene oxide, and cause activation and degranulation of mast cells secreting a variety of active biological compounds including histamine, arachidonic acid metabolites, and platelet-activating factorsZ3 Activated neutrophils generate large amounts of superoxide anion radicals (0;‘) via the reaction catalyzed by reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, 202 + NADPH + 20, + NADP+ + H +. 0;’ could participate in the formation of other active oxygen species, eg, hydroxyl radical (OH’) and hydrogen peroxide (H202), of which the former can initiate peroxidation of polyunsaturated fatty acidsz8 However, lipid peroxidation could be not only the cause but also the result of ceil damage.29 When activated leukocytes come into contact with the dialyzer membrane, they release hydrolytic enzymes from lysosomes, thus causing injury to cell plasma membranesZ3 Stimulated neutrophils also release myeloperoxidase into extracellular medium; myeloperoxidase is an enzyme that catalyzes oxidation of a number of halides and pseudohalides. Chloride ion, due to its high extracellular concentration, is preferentially oxidized to hypochlorous acid, a potent oxidizing agent that is approximately 100-fold more reactive than H202. Hypochlorous acid can exert harmful effects on sulfhydryl
catalyzing the dismutation of 0;’ to yield Hz02 and 01.~~ Inhibition of SOD can result not only in an increase of OS’, but also in the elevation of other active oxygen species (eg, OH) and the intensification of lipid peroxidation processes. The reason that SOD activity decreases during hemodialysis could be connected with copper and zinc status in hemodialyzed patients suffering from chronic renal failure. We have found that hemodialysis decreases plasma copper and zinc levels to less than 50% of control values.30 Since copper and zinc are cofactors of cytoplasmic SOD, changes in their concentration can affect this enzymatic activity. We also found that the activity of SOD was much higher in uremic patients than in control healthy subjects of a similar age group (Fig 2). This observation is partially in agreement with the study of Chauchan et ali who found increased activity of SOD before hemodialysis and its normalization after hemodialysis; in contrast, Vanella et a131 and Trznadel et all4 found that erythrocyte SOD activity in patients with chronic renal failure was decreased. It is possible that these reported differences in SOD activity were connected with the severity of disease and frequency of hemodialyses. It may be that SOD activity could reach higher values after its decrease during hemodialysis in less severely ill patients, and that this protective process against 0;’ and lipid peroxidation does not exist in severe cases of renal failure. Decreased serum AOA in patients with chronic renal insufficiency and the lack of significant changes in its value during hemodialysis observed in our study agree with the results of Kuroda et aL3’ In conclusion, the present study confirmed increased levels of plasma lipid peroxides, elevated erythrocyte SOD, activity and decreased serum AOA in patients with chronic renal failure treated by maintenance hemodialysis. We found intensification of lipid peroxidation during hemodialysis and indicated that it was caused by decreased SOD activity. We suggest that hemodialysis can accelerate atherosclerosis by increasing lipid peroxidation in patients with renal insufficiency.
ACKNOWLEDGMENT The authors thank Dr Bernhard Hennig tions and for reviewing the manuscript.
for his useful
sugges-
REFERENCES 1. Wing AJ, Brunner FP, Brynger H, et al: Cardiovascularrelated causes of death and the fate of patients with renovascular disease. Contrib Nephrol41:306-311, 1984 2. Linder
A, Charra
B, Sherrard
DJ, et al: Accelerated
athero-
sclerosis in prolonged maintenance hemodialysis. N Engl J Med 290:697-701.1914 3. Kudoh Y, Imura 0: Study on the atherosclerosis mechanism in chronic hemodialysis. Jpn Circ J 51:631-641, 1987
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