The carboxyproxyl-derived spin trap (CP-H) is an appropriate detector-compound for oxidative stress.

Reperfusion of ischemic tissue disturbs the balance between reactive oxygen species (ROS) and the cellular antioxidative defense. This imbalance is known as oxidative stress. In this study the spin trap 3-carboxy-2,2,5,5-tetramethylpyrrolin-1-hydroxide (CP-H) with its ESR-detectable paramagnetic analogue 3-carboxy-2,2,5,5-tetramethylpyrrolin-1-oxyl (*CP) was analyzed in vitro and in vivo. In preliminary in vitro experiments we studied the interaction of CP-H with reactive compounds like hydroxyl radicals (*OH) and alkylperoxyl radicals (ROO*) which are formed during organ reperfusion or tissue reoxygenation. The increase in the peak intensity of the ESR signal of the *CP-radical was used as a measure for CP-H oxidation by the above-mentioned oxidizing radicals. It could be clearly shown that *OH as well as ROO* induce CP-H oxidation. The intensity of the ESR signal (*CP) depends on the concentration of the applied oxidant. In a further set of in vitro experiments we analyzed some factors influencing the stability of the generated *CP. Cellular reductants are able to interact with many radicals whereby their paramagnetic signal intensity decreases. We could show that glutathione (GSH) up to 5 mM does not influence *CP concentration. On the other hand, ascorbate at a concentration of 0.6 mM significantly reduces 55% of *CP within 60 min to the ESR-silent CP-H. At 1 mM ascorbate the *CP derived ESR signal is reduced within 60 min by 90%. Lower concentrations of ascorbate (0.1-0.3 mM) do not significantly decrease signal intensity within 1 h. Homogenization of ischemic rat kidney in the presence of an air-equilibrated buffer obviously induces the formation of oxidizing radicals which in turn are able to convert diamagnetic CP-H into paramagnetic *CP. The intensity of the formed *CP was analyzed in a 600 g supernatant with ESR spectroscopy at 25 degrees C. It could be demonstrated that at least 3.0 +/- 0.5 microM *CP is formed 15 min after starting tissue homogenization and reoxygenation. Subsequent measurements of the *CP concentration indicated that its signal intensity continuously decreases. After 75 min a residual *CP concentration of 0.7 +/- 0.3 microM was monitored. Removal of mitochondria from the homogenate by centrifugation at 6,000g decelerates the disappearance of *CP but does not block it completely. In summary it could be shown that the marker (CP-H) is able to indicate the formation of oxidizing radicals during reoxygenation of ischemic tissue. This method underestimates the amount of produced oxidizing radicals. One reason for this is the reduction of *CP by some cellular reductants. Other reasons will be discussed. We assume that the used method allows a nearly real-time determination of radical production during organ reoxygenation.

Did the gradual loss of GLUT2 cause a shift to diabetic disorders in the New Zealand obese mouse (NZO/Hl)?

The New Zealand obese mouse (NZO/Hl) is characterised by hereditary obesity and type-2 diabetes, including insulin resistance, hyperinsulinaemia, and glucose intolerance. In other diabetic models, it has been revealed that the proper functioning of the glucose transporter isoform 2 (GLUT2) is essential for adequate secretion of insulin. The aim of this study was to compare the distribution of islet cells and GLUT2, as well as the expression of GLUT2-mRNA, in the pancreas of NZO mice and metabolically unimpaired NMRI (Naval Medical Research Institute) mice. Pancreas tissue was obtained from different stages of development. For molecular determination of the expression level of GLUT2-mRNA, total-RNA was extracted from the pancreas and analysed by quantitative real-time RT-PCR. All investigated NZO mice displayed increased weight, elevated hyperinsulinaemia, and slightly enhanced blood glucose levels compared with the NMRI control mice. By means of immunofluorescence microscopy drastically reduced insulin levels were detected, which might be compensated by the observed islet cell hyperplasia and hypertrophy. Furthermore, the normally peripheral localisation of the alpha-cells within islets was disturbed. By contrast, there were no changes in somatostatin cell distribution. However, considerable differences appeared with regard to GLUT2: whereas the beta-cells of NMRI mice showed dense immunostaining of the GLUT2 transporter on the cell surface, in all age groups of NZO mice, GLUT2 on the plasma membranes was reduced and dispersed in the cytoplasm. These findings agree with the molecular biological results, which displayed decreased mRNA-expression of GLUT2. In summary, the observed alteration of islet morphology and of GLUT2 expression in diabetic mice complements our previous results from a superfusion protocol and further clarifies the mechanisms of diabetogenesis in NZO mice.

Formation of ascorbate radicals as a measure of oxidative stress: an in vitro electron spin resonance-study using 2,2-Azobis (2-amidinopropane) dihydrochloride as a radical generator.

Reactive oxygen species (ROS) are continuously formed in biological systems. Any increase in radical production or decrease in the defense against ROS induces oxidative stress. This imbalance between ROS formation and ROS detoxification is believed to be involved in a variety of pathogenic processes, including ischemia-reperfusion injury. Various markers indicating oxidative stress has been used in experimental and clinical studies. One of them is ascorbate free radical (AFR), electron spin resonance intensity of which correlates with the severity of radical formation. We investigated the impact of alkyl peroxyl radicals produced by 2,2-Azobis (2-amidinopropane) dihydrochloride decomposition on the magnitude of the AFR signal. Our data confirmed the principal applicability of AFR as a nontoxic marker of radical generation.

Functional response of a limited beta-cell mass to long-term hyperglycemia.

250 syngeneic islets were implanted either beneath the kidney capsule or into the liver of diabetic LEW 1.A rats to investigate the functional response of a limited mass of beta-cells to long-term hyperglycemia. The number of islets, per se, was expected to be insufficient to reverse the hyperglycemia. All animals were characterized by a substantial body weight gain. Unexpectedly, 29% of the rats with a subrenal and 40% of the animals with a portal islet graft normalized their plasma glucose in 64 +/- 13 and 75 +/- 12 days, respectively. Depending on the glycemic state of the recipients, there was an elevation of the graft insulin content after 120 days over the level at transplantation. The responsiveness of the implanted islets to different secretagogues was tested either in vitro by static incubation of the prepared grafts from the kidney or in situ by perfusion of the islet-containing liver. Grafts of normoglycemic rats showed a pronounced response, although the biphasic profile of the hormone release was lost. In principle, grafts exposed permanently to a hyperglycemic environment have kept their responsiveness, although the insulin outflow was considerably lower. The functional viability of the islets was not influenced by the site of transplantation. Long-term hyperglycemia does not necessarily result in destruction and loss of beta-cells even when their total mass is already limited, but it obviously impairs their functional responsiveness.

Lucigenin chemiluminescence in human seminal plasma.

Seminal plasma protects spermatozoa from the detrimental effects of reactive oxygen species such as hydrogen peroxide. We investigated the lucigenin-dependent chemiluminescence in cell-free seminal plasma from andrological patients. The seminal plasma was separated from cells by centrifugation. In all seminal plasmas studied lucigenin-dependent chemiluminescence (LCL) was detected. The LCL showed a strong pH-dependence. The signal was stable if samples were stored at +4 degrees C for up to 4 days or up to 8 days at -80 degrees C. Filtration of the samples (0.45 and 0.22 microm pore size) did not lower their luminescence. The addition of superoxide dismutase (SOD) and ascorbic acid oxidase (AAO) lowered LCL nearly to baseline values while trolox and desferal showed moderate effect, whereas allopurinol had no effect. Electron paramagnetic resonance spectroscopy demonstrated ascorbyl radicals in seminal plasma. Physiological concentrations of ascorbic acid yielded SOD-inhibitable lucigenin-chemiluminescence. The nitroblue-tetrazolium assay showed that ascorbic acid in buffer solution produced formazan. Superoxide-anion radicals were not detected in seminal plasma by the spin-trap DEPMPO due to their low steady state concentration. It is concluded that in seminal plasma ascorbate reacts with molecular oxygen yielding ascorbyl radicals and superoxide anion. If lucigenin is added to seminal plasma, reducing substances present, such as ascorbate, reduce lucigenin to the corresponding radical; this radical reacts with molecular oxygen and also forms O2-. So LCL in human seminal plasma results from the autoxidation of ascorbate and the oxidation of the reduced lucigenin. While the physiological relevance of the former mechanism is unknown, the latter is an artifact.

Comparison between xanthine oxidases from buttermilk and microorganisms regarding their ability to generate reactive oxygen species.

Xanthine oxidase (XO) forms uric acid from xanthine. It is assumed that at the same time oxygen is reduced by the XO to reactive oxygen species (ROS), mainly to .O2- and to H2O2. Under certain conditions such ROS can be highly damaging to cellular structures. Therefore, XO was frequently used as a model system, in which the impact of ROS on cellular compounds and structures has been investigated. In this in vitro study xanthine oxidases from buttermilk and from microorganisms were compared regarding their ability to generate ROS. It could be shown that both enzymes are able to transform xanthine to uric acid but differ significantly in their reductive properties to oxygen. XO from buttermilk reduces oxygen to both .O2- and H2O2 whereas XO from microorganisms generates H2O2, but fails to form .O2-. Since .O2- are involved in maintaining transition metal-mediated formation of hydroxyl radicals (.OH) from H2O2, we conclude that XO from microorganisms is therefore largely unsuitable in studies investigating just the interaction of .O2- with other ROS on cellular compounds.

Effects of twelve months ethanol feeding on rat liver metabolic activity.

Thirty male Wistar rats were fed with 15% ethanol (V/V) for one year. Thirty other male animals were the control group. To determine the possible metabolic disturbances caused by chronic ethanol feeding in blood we measured in blood metabolic parameters, and in a liver perfusion assay the hepatic insulin clearance and hepatic urea production in these animals. Between the ethanol-fed and the control animals there were significant differences in the following parameters: blood insulin concentration (47 vs. 2 microU/ml) and activities of amino acid transferases in liver homogenates at the end of the perfusion experiments (ASAT, 5950 vs. 70; ALAT, 3632 vs. 93 U/l). The other parameters were still normal in the ethanol-fed animals. Thus in these experiments after 12 months of 15% (V/V) ethanol feeding the rats still showed only a state of beginning metabolic disturbances in the liver. The results are discussed under consideration of the formation of acetaldehyde protein adducts in all rat organs investigated, namely, liver, kidney, heart and skeleton muscle, gut and spleen.

[Postischemic reperfusion injury. Biochemical and methodological principles]. / Der postischämische Reperfusionsschaden. Biochemische und methodische Grundlagen.

The term "ischemic reperfusion injury" encompasses all toxic events in a cell that occur during ischemia and subsequent reoxygenation. These reactions have a significant effect, for example, on the rate of organ survival in kidney transplantation. Reactive oxygen intermediates (ROI) play an important role in the process of postischemic reperfusion. The basic mechanisms of generation and detoxification of ROI as well as the possibilities for their registration and quantification under conditions of ischemic reperfusion injury in the rat kidney are demonstrated in this report. A prerequisite to developing cytoprotective strategies is understanding the precise course of these mechanisms to minimize damage caused by ischemia and the subsequent reperfusion, thus retaining the organ's function to the greatest extent.

Influence of oxygen concentration on redox cycling of alloxan and dialuric acid.

Alloxan, a chemical diabetogen, decays in the absence of reductants into alloxanic acid. In the presence of glutathione, it is reduced via the alloxan radical into dialuric acid, which autoxidizes back to alloxan. During this redox cycling process, reactive oxygen species are formed that destroy beta-cells in islets of Langerhans. Previous experiments were conducted with oxygen concentrations about ten times as high as within cells. The aim of our in vitro study was to evaluate the impact of different oxygen concentrations (0, 25, 250 micromol/l) at a given initial ratio of glutathione and alloxan on this redox cycling. Reduction of alloxan, oxidation of glutathione, and the formation of glutathiol (GSSG) were continuously recorded by HPLC for 90 minutes at 25 degrees C in air, calibration gas, or argon. In the absence of reductants, alloxan irreversibly decomposed into alloxanic acid regardless of oxygen presence. When the reaction system contained glutathione, decomposition was significantly retarded and therefore influenced by oxygen. In argon, decay could not be observed due to its reduction and the absence of oxygen. Increasing oxygen concentration enabled a redox cycling and therefore an ongoing decay. The highest decomposition along with the highest consumption of glutathione occurred at 250 micromol/l oxygen. The lower the oxygen, the more dialuric acid could be detected. After calculation, about 33 redox cycles per hour generates an amount of reactive oxygen species sufficient to damage pancreatic beta cells and induce insulin deficiency.

[Insulin extraction by the isolated perfused rat liver. The effect of portal insulin mass inflow and the duration of infusion on hepatic extraction and elimination rate of insulin]. / Insulinextraktion durch die isoliert perfundierte Rattenleber. Einfluss des portalen Insulinmasseflusses und der Infusionsdauer auf die hepatische Extraktion und die Eliminierungsrate von Insulin.

Rat liver from well fed Wistar-rats was perfused in vitro at 37 degrees C in a flow-through system with Krebs-Ringer-bicarbonate buffer, pH 7.4, in the presence of 20 vol.-% freshly prepared bovine erythrocytes. After 20 min of liver perfusion, insulin was infused into the portal vein either in form of a three-step profile at different insulin concentrations (every step lasting 20 min) or as a permanent infusion of insulin for 40 min. For each step the percentage of hepatic insulin extraction (%hIEx) and the hepatic insulin removal rate (hIRR) was calculated by means of radioimmunological insulin determination in both input and output of the liver. After a single passage about 53% of the portal insulin input of 68 ng insulin/min (step 1) were removed. The hIRR amounted to 4.1 ng insulin/min X g liver. If the insulin input was then increased to about 320 ng insulin/min (step 2), %hIEx decreased to 28%; under these conditions the hIRR amounted to 9,4 ng insulin/min X g liver. If the portal input of insulin again was reduced to 71 ng insulin/min (step 3), the %hIEx remained at about 28% despite a nearly identical input of insulin (compared with that of step 1). The hIRR was then only 2.2 ng insulin/min X g liver. During a 40 min permanent infusion of insulin (184 ng insulin/min) the %hIEx declines from 61% to about 20% within the first 20 min. Then %hIEx was stabilized at this level. hIRR shows a similar behaviour. These experiments show clearly that the %hIEX as well as the hIRR were changed significantly in dependence on portal insulin-input, on the duration of insulin infusion and in dependence on the extent of insulin load of the liver during perfusion.

[Insulin extraction by the isolated perfused rat liver using infusion conditions of a biphasic insulin concentration profile]. / Insulinextraktion durch die isoliert perfundierte Rattenleber unter Bedingungen der Infusion eines biphasischen Insulin-Konzentrationsprofils.

Rat liver was perfused in vitro at 37 degrees C in an open system with Krebs-Ringer-Bicarbonate buffer, pH 7.4, in the presence of 20 vol% freshly prepared bovine erythrocytes. A biphasic insulin concentration profile, imitating the glucose induced insulin secretion of the isolated perfused rat pancreas was infused by means of a gradient mixer into the portal vein of the liver. The quality of the infused gradient was tested in a series of preliminary experiments by dye-infusion into the portal vein. It could be shown by synchronous fractionation of medium samples immediatly prior and after liver passage and by subsequent radioimmunological insulin determination that the biphasic profile of the infused insulin gradient was preserved after a single liver passage. The insulin concentration in the portal vein under experimental conditions ranged from 0 to 3.2 ng insulin/ml perfusate. About 70-80% of the insulin was removed by the liver. It could be further demonstrated that the hepatic insulin removal rate is rising with increasing insulin concentration in the liver input. The highest hepatic insulin removal rate obtained under the used portal insulin concentrations amounted to 2 ng insulin/min X g liver ww.

Apoptosis in the heart: when and why?

Since mammalian cardiac myocytes essentially rely on aerobic energy metabolism, it has been assumed that cardiocytes die in a catastrophic breakdown of cellular homeostasis (i.e. necrosis), if oxygen supply remains below a critical limit. Recent observations, however, indicate that a process of gene-directed cellular suicide (i.e. apoptosis) is activated in terminally differentiated cardiocytes of the adult mammalian heart by ischemia and reperfusion, and by cardiac overload as well. Apoptosis or programmed cell death is an actively regulated process of cellular self destruction, which requires energy and de novo gene expression, and which is directed by an inborn genetic program. The final result of this program is the fragmentation of nuclear DNA into typical 'nucleosomal ladders', while the functional integrity of the cell membrane and of other cellular organelles is still maintained. The critical step in this regulated apoptotic DNA fragmentation is the proteolytic inactivation of poly-[ADP-ribose]-polymerase (PARP) by a group of cysteine proteases with some structural homologies to interleukin-1 beta-converting enzyme (ICE-related proteases [IRPs] such as apopain, yama and others). PARP catalyzes the ADP-ribosylation of nuclear proteins at the sites of spontaneous DNA strand breaks and thereby facilitates the repair of this DNA damage. IRP-mediated destruction of PARP, the 'supervisor of the genome', can be induced by activation of membrane receptors (e.g. FAS or APOI) and other signals, and is inhibited by activation of 'anti-death genes' (e.g. bcl-2). Overload-triggered myocyte apoptosis appears to contribute to the transition to cardiac failure, which can be prevented by therapeutic hemodynamic unloading. In myocardial ischemia, the activation of the apoptotic program in cardiocytes does not exclude their final destiny to catastrophic necrosis with release of cytosolic enzymes, but might be considered as an adaptive process in hypoperfused ventricular zones, sacrificing some jeopardized myocytes to regulated apoptosis, which may be less arrhythmogenic than necrosis with the primary disturbance of membrane function.

Effect of in vivo and in vitro application of glucagon, insulin and epinephrine on Ca++-transport properties of liver mitochondria.

In vivo administration of glucagon, insulin or epinephrine, respectively, gives rise to an increase of Ca++-retention time as well as of the Ca++-uptake rate in subsequently isolated rat liver mitochondria. Whereas the changes of Ca++-transport properties after pretreatment with glucagon or epinephrine occur already 6--15 min after their administration, the effect of insulin is observed not earlier than 30 min after its application. Under diabetic and starving conditions the Ca++-retention time of isolated liver mitochondria is prolonged, whereas no alteration of the uptake rate occurs. Since alloxan as well as streptozotocin induced qualitatively similar changes, a specific action of alloxan on liver mitochondria can be ruled out. Application of insulin 60--90 min prior to decapitation normalizes the changes of mitochondrial Ca++-transport observed under chronic alloxan diabetic conditions. Cycloheximide abolishes the prolongation of Ca++-retention in mitochondria from alloxan diabetic rats, but has no influence on the changes induced by glucagon pretreatment.

[Characterization of the functional state of the in vitro Hb-free perfused rat liver]. / Charakterisierung des funktionellen Zustandes der in vitro Hb-frei perfundierten Rattenleber.

Rat liver was perfused in vitro with Krebs-Henseleit-medium and albumin at 25 degrees C in a recirculating system without hemoglobin over a period of 120 min. The following basic parameters for characterization of isolated liver perfusion were recorded: medium-pO2 prior and after liver passage, flow-rate, pH, and hepatic O2-consumption. Beyond this, concentration of lactate and pyruvate, hepatic glucose production, activity of aspartate-aminotransferase, leucine-aminopeptidase and acylase as well as concentration of K+-ions in the perfusion fluid were measured. In dependence on nutrition state of liver donors (fed or starved rats) the rate of glycogenolysis or rate of lactate-stimulated gluconeogenesis was calculated. The endogenous glycogenolysis can be blocked by an in vivo injection of propranolol. The propranolol-inhibited glycogenolysis can be stimulated by an in vitro glucagon application.

[The isolated perfused rat liver--methodology and viability characteristics]. / Die isoliert perfundierte Rattenleber--Methodik und Vitalitätscharakteristik.

A simple method for in vitro perfusion of the rat liver is presented. The operative technique for mobilizing the liver, the equipment, the medium, and the conditions used for the perfusion were described in detail. Directions for cleaning and sterilising the system are mediate. The represented perfusion system is suitable not only for the system are recirculatory but also for non-recirculatory organ perfusions. The problems of the viability of the perfused liver are discussed and the different qualification of several viability parameters is debated.

[Proteolytic degradation of glucagon by human erythrocytes]. / Proteolytischer Glukagonabbau durch menschliche Erythrozyten.

Human erythrocytes possess a proteolytic activity degrading glucagon (and insulin) even at very low concentrations with a high degree of efficiency. The enzyme which likely belongs to the class of insulin-glucagon-proteinases, can be inhibited by chelating agents, such as ethylenediamine tetraacetic acid and o-phenanthroline, thiol blocking reagents, such as p-chloromercuribenzoate and N-ethylmaleimide as well as by proteinase inhibitors directed against serine proteinases, such as Contrykal and Trasylol. No inhibition could be shown by leupeptin. Insulin in an equimolar range is capable of inhibiting glucagon degradation competitively. Dithioerythritol stimulates the degrading activity. Co++, Zn++, Mn++ and Ca++ prevent the o-phenanthroline mediated inhibition of glucagon degrading activity, whereas Mg++, Cu++, Cd++ and Fe+++ have an inhibitory effect. The glucagon degradation exhibits a pH optimum at 7,1 with an apparent Km of 4.4 X 10(-6) mol/l. The insulin-glucagon-proteinase in human erythrocytes is supposed to have a regulatory influence within the carbohydrate pathway.

Transformation of barbituric acid into alloxan by hydroxyl radicals: interaction with melatonin and with other hydroxyl radical scavengers.

Barbituric acid (2,4,6-pyrimidinetrione) can be transformed by a non-enzymatic hydroxylation into alloxan (2,4,5,6-pyrimidinetetrone). This transformation can be used as a reaction indicating the formation of hydroxyl radicals (.OH). This conversion was detected using HPLC. Formation of .OH was demonstrated by electron spin resonance (ESR) spectroscopy combined with spin-trapping techniques. It was shown that .OH generated via the Fenton reaction abstracts first a hydrogen atom from barbituric acid (BA) and forms intermediately a paramagnetic derivative of BA. After a second attack by another .OH, the BA radical is transformed into dialuric acid (DA), which autoxidizes via the alloxan radical (.ALX) to ALX. Superoxide radicals (.O2-) are formed during autoxidation of DA and.ALX. They are able to regenerate ferrous ions. As a result, traces of iron salts are capable of catalyzing the conversion of large amounts of BA into ALX. Several scavengers of .OH were tested with regard to their efficiency in preventing the transformation of BA into ALX. Of all the scavengers analyzed, melatonin was shown to be one of the most potent compounds.

Does the anaerobic formation of hydroxyl radicals by paraquat monocation radicals and hydrogen peroxide require the presence of transition metals?

This in vitro study investigated the formation of hydroxyl radicals (*OH) under anaerobic conditions through the direct reaction between paraquat radicals (PQ(+)*) and hydrogen peroxide (H(2)O(2)) by quantitative UV-VIS and electron spin resonance (ESR) spectroscopy. PQ(+)* was formed by paraquat reduction using either sodium dithionite or the xanthine/xanthine oxidase reaction as electron donors. The anaerobic formation of PQ(+)* was quantified both by measuring light absorption at 605 nm or by ESR techniques respectively, using either the absorption coefficient or ultramarine as a stable spin standard. Detection of *OH took place with aid of the spin trap 5-diethoxyphosphoryl-5-methyl-1-pyrroline- N-oxide (DEPMPO). Generation or addition of H(2)O(2) to PQ(+)* eliminates the 35-line ESR signal of PQ(+)* and subsequently generates the 8-line ESR signal of the DEPMPO-OH adduct. The elimination of PQ(+)* as well as the formation of OH-DEPMPO adduct was not influenced by 1.0 mM deferoxamine, indicating that iron or other transition metals are, at least under anoxic conditions, not necessarily involved in the generation of the most aggressive reactive oxygen species *OH.

Secretion and elimination of insulin by the in vitro perfused pancreas and liver of rats with thioacetamide-induced liver cirrhosis.

Pancreatic secretion and hepatic removal of insulin have been measured in thioacetamide (TAA)-induced compensated rat liver cirrhosis in perfusion experiments. Peripheral plasma concentrations of glucose and insulin were slightly decreased in TAA-treated rats. Pancreatic secretion and hepatic removal of insulin remained unchanged by the TAA-treatment. Thus, even in morphologically and biochemically proven experimental liver cirrhosis, insulin secretion and removal may not be disturbed.

Experimental study showing a diminished cytosolic antioxidative capacity in kidneys of aged rats.

In response to the rising demand for renal transplantations, more and more marginal (e.g. older) organs are being transplanted with the result of decreasing graft survival rates. Ischemia-reperfusion injury via oxidative stress is thought to be the main pathogenetic factor for this phenomenon. The cytosolic antioxidative capacity (CAC; expressed as superoxide anion radical scavenging capacity and quantified as the amount of cytosol (=ID(50)), which scavenges 50% of superoxide anions generated by a defined xanthine oxidase activity in vitro) and the catalase activity were therefore quantified in renal tissues of young (10 weeks) and older (40 and 60 weeks) Wistar rats and compared to each other. CAC with an ID(50) of 0.064 microl in 10-week-old rats was significantly higher than in older rats (0.152 microl in 40- and 0.100 microl in 60-week-old rats; p < 0.01). The catalase activity in 10-week-old rats was 18, 200 +/- 3,500 U/g w/w and 18,900 +/- 850 U/g w/w in 40-week-old rats. In 60-week-old rats, however, catalase activity was found to be significantly less (7,500 +/- 175 U/g w/w; p < 0.01). In conclusion, the aforementioned significant decrease of the cytosolic antioxidative capacity of kidneys in older rats should be the rationale for extensive cytoprotective, antioxidative treatment trials especially after renal transplantation from aged donors.