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CuZn superoxide dismutase, Mn superoxide dismutase, catalase and glutathione peroxidase activities in lymphocytes and erythrocytes were studied in 9 children with insulin-dependent diabetes mellitus (IDDM) as well as in 21 healthy children. The mean erythrocyte CuZn superoxide dismutase and glutathione peroxidase were statistically significantly lower in the IDDM group compared with the controls although almost all IDDM results fell within the mean ± 2 sd limits of the controls. The small differences found can hardly be assigned biological significance. Erythrocyte catalase as well as lymphocyte CuZn superoxide dismutase and Mn superoxide dismutase did not differ from the controls.

1. Disulfiram has long been used in the treatment of chronic alcoholism. It is in vivo partially reduced to diethyldithiocarbamate, which is an efficient inhibitor of Cu, Zn-containing superoxide dismutase both in vitro and in vivo. The recently described extracellular superoxide dismutase is even more sensitive to diethyldithiocarbamate than Cu, Zn-superoxide dismutase. 2. To test for the possibility that long term treatment with disulfiram leads to inhibition of the superoxide dismutases, plasma extracellular superoxide dismutase and erythrocyte Cu, Zn-superoxide dismutase were determined in 12 disulfiram-treated alcoholics, and compared with 11 non-treated alcoholics and 19 healthy controls. 3. Plasma extracellular superoxide dismutase was moderately reduced (about 20%) in the disulfiram-treated alcoholics as compared with the non-treated alcoholics and the healthy controls. No effect of disulfiram treatment on erythrocyte Cu, Zn-superoxide dismutase activity was demonstrated.

The current work was initiated upon the finding of a positive correlation (0.05

Twelve patients with subjective symptoms, ascribed by the patients themselves to mercury released from dental restorations, were investigated. In addition to a general dental examination the following parameters were registered: the total number of amalgam surfaces in the mouth; potential and polarization of existing and accessible dental metallic restorations for calculation of intraoral currents. As regards the highest calculated intraoral current for each individual there was a statistically significant difference between the patient group and a control group consisting of 12 persons. An analysis of the amount of selenium, glutathione-peroxidase and mercury in the blood showed no differences between the patient and the control group. However, a statistically significant positive correlation could be seen between the total number of amalgam surfaces and the plasma-mercury level for patients and controls pooled together. The numerous other blood parameters analyzed did not reveal any differences between the groups.

The reference intervals of 18 blood components were established for an elderly population in the 8th decade of life. The most significant findings were a broader range of values for most of the components, lower values of plasma folate and plasma potassium and higher values of erythrocyte sedimentation rate, plasma creatinine and serum cholesterol, the lattermost in women only, compared to younger people.

1. The neuronal ceroid-lipofuscinoses is a group of diseases characterized by a widespread accumulation in the body of pigments believed to be end-products of lipid-peroxidation damaged organelles. It was recently shown that cerebrospinal fluid from patients with infantile and juvenile neuronal ceroid-lipofuscinosis were less protective against superoxide radical-induced hydroxyl radical formation compared with controls [3]. 2. The content of superoxide dismutase isoenzymes in cerebrospinal fluid and in plasma from patients with different forms of neuronal ceroid-lipofuscinosis was analysed. No significant difference from controls could be demonstrated in samples from patients with juvenile neuronal ceroid-lipofuscinosis. The few samples from patients with infantile and late infantile neuronal ceroid-lipofuscinosis analysed all fell within the range defined by the controls.

1. 1. A manganese containing superoxide dismutase from bovine heart mitochondria was isolated and characterized. 2. 2. It has a molecular weight of about 86,000 and is composed of 4 noncovalently bound subunits of equal size. 3. It appears to contain 2 mole manganese per mole enzyme. 4. The carbohydrate content is very low. 5. The specific activity and amino acid composition are similar to those of other mitoehondrial superoxide dismutases. 6. The enzyme forms complexes with ampholytes and can therefore not be analysed by isoelectric focusing.

The effect of benzylpenicillin (BPC) pretreatment on the kinetics and brain sensitivity for thiopental was studied in male rats using a previously developed electroencephalgrafic (EEG) threshold method. Thiopental was infused intravenously with constant infusion rate. The rats were killed by decapitation immediately after the first burst suppression of 1 sec. or more (the silent second) which was observed in the EEG-record during the infusion. Thiopental concentration in serum and in different brain regions was determined by a high pressure liquid chromatografic method. After pretreatment with 0.9 g/kg of BPC the dose of thiopental needed to induce the silent second was significantly reduced (- 20 per cent) when compared with saline treated controls. The serum concentration was also reduced (- 30 per cent) after this BPC pretreatment but the concentrations in the different brain regions were the same in both groups. After pretreatment with 1.2 g/kg of BPC almost all animals had convulsions, the dose needed to obtain the silent second was very much reduced and there were reduced concentrations of thiopental in the different brain regions. After both doses of BPC high negative correlations were found between BPC concentrations in brain tissue and thiopental concentrations in hippocampus and brainstem indicating an interaction between the drugs. The most probable site of this interaction is the organic acid transport system out of the CNS which could be used by both substances. Lipid solubility is not the only factor involved in the distribution of thiopental in the rat brain.

A single injection of alloxan1, or alloxan-like derivatives of uric acid2,3, kills the insulin-producing islet β-cells and causes diabetes mellitus in animals. This effect is probably mediated by a sequence of redox reactions involving the production of superoxide anion radicals in or near the β-cells4–8. Superoxide anions may also arise in inflammation9–15, and islet inflammation often accompanies the outbreak of human diabetes16, perhaps following the combined onslaught of viruses and chemical agents on the β-cells17. We now report that injections of superoxide dismutase, an enzyme removing superoxide anion radicals, act prophylactically against alloxan-induced diabetes.

CuZn Superoxide dismutase (SOD) and Mn SOD activities were analyzed in hypothalamus, nucleus caudatus, hippocampus and cortex gyrus cinguli from 12 chronic alcoholics and from 16 controls. The CuZn SOD activities were slightly lower and the Mn SOD activities were slightly higher in the brain pieces from chronic alcoholics compared to the controls. The slight differences found can hardly be assigned etiological importance in the degenerative processes in brain tissue connected with chronic alcoholism.

The secretory enzyme extracellular superoxide dismutase (EC-SOD) occurs in at least three forms, which differ with regard to heparin affinity: A lacks affinity, B has intermediate affinity, and C has relatively strong affinity. The affinity of EC-SOD C for various sulphated glycosaminoglycans (GAGs) was assessed (a) by determining the concentration of NaCl required to release the enzyme from GAG-substituted Sepharose 4B and (b) by determining the relative potencies of the GAGs to release EC-SOD C from heparan sulphate-Sepharose 4B. Both methods indicated the same order of affinity. Heparin bound EC-SOD C about 10 times as avidly as the studied heparan sulphate preparation, which in turn was 10 and 150 times as efficient as dermatan sulphate and chondroitin sulphate respectively. Chondroitin sulphate showed weak interaction with EC-SOD C at physiological ionic strength. Heparin subfractions with high or low affinity for antithrombin III were equally efficient. The binding of EC-SOD C to heparin-Sepharose was essentially independent of pH in the range 6.5-9; below pH 6.5 the affinity increased, and beyond pH 9.5 there was a precipitous fall in affinity. The inhibitory effect of NaCl on the binding of EC-SOD C to GAGs indicates that the interaction is of electrostatic nature. EC-SOD C carries a negative net charge at neutral pH, and it is suggested that the binding occurs between the negative charges of the GAG sulphate groups and a structure in the C-terminal end of the enzyme that has a cluster of positive charges. These results are compatible with the notion that heparan sulphate proteoglycans on cell surfaces or in the intercellular matrix may serve to bind EC-SOD C in tissues.

Human cupro-zinc superoxide dismutase is polymorphic in northern Sweden. The genetic variant type has a lower mobility at electrophoresis in alkaline buffer. The enzyme was isolated from erythrocytes from one of the rare homozygotes and its properties compared to those of the common type. The isoelectric point of the variant was higher (4.85) than that of the common type (4.7). Small differences in amino acid composition were found but no definite amino acid substitutions could be pointed out. The molecular weights were equal as judged from electrophoreses in polyacrylamide gels in the presence of dodecylsulphate. The ultraviolet spectra were similar. Parameters related to the active site of the enzyme were very similar; i.e. specific activity and sensitivity to inhibition by cyanide and by H2O2. These parameters, especially the latter two, differ widely between species. Both enzymes were stable for weeks at neutral pH at 37 degrees C, whereas the common type was significantly more stable at pH 4 and pH 11 and also at incubation in neutral buffer at 70 degrees C. It appears that the active site of the variant is conserved whereas the stability of the enzyme is affected.

Toxic oxygen-centred radicals have been linked to β-cell damage brought about by some chemicals and might conceivably also be of importance in the pathogenesis of spontaneous insulin-dependent diabetes mellitus (1DDM). In several model systems, superoxide dismutase has been protective. The major protector against superoxide radicals in the extracellular space appears to be a recently discovered superoxide dismutase, EC-superoxide dismutase. This enzyme was analysed in plasma from children with insulin-dependent diabetes mellitus. The specimens were obtained from 8 patients with disease of recent onset and from 15 patients with disease of longer duration. There was no significant difference in EC-superoxide dismutase between the patients and controls.

A complementary DNA clone from human placenta, encoding human extracellular superoxide dismutase (EC-SOD; superoxide:superoxide oxidoreductase, EC 1.15.1.1), has recently been isolated and characterized. An expression plasmid, based on the EC-SOD complementary DNA, was transfected into Chinese hamster ovary cells (CHO-K1). The transfected cells secreted human EC-SOD to the culture medium. The secreted recombinant (r) EC-SOD was isolated in high yield with a three-step procedure beginning with immobilized monoclonal anti-EC-SOD antibodies. The properties of the rEC-SOD were compared with native (n) EC-SOD isolated from human umbilical cords. The specific activities and amino-terminal amino acid sequences were identical. The amino acid compositions were virtually identical and very similar to the composition deduced from the complementary DNA sequence. Both rEC-SOD and nEC-SOD contained 4 Cu and 4 Zn atoms per molecule, and the presence of Zn in EC-SOD is thus now established. The rEC-SOD produced is type C, since its affinity for heparin-Sepharose was identical to that of nEC-SOD type C. Both enzymes bound to concanavalin A, lentil lectin, and wheat germ lectin and are thus glycoproteins. rEC-SOD and nEC-SOD seem to have the same subunit structure and composition as analyzed by polyacrylamide gel electrophoresis and gel chromatography.

— The ceroid-lipofuscin pigments found in neuronal ceroid-lipofuscinosis are believed to arise from lipid peroxidation-damaged organelle membranes. In several in vitro systems superoxide dismutase inhibits lipid peroxidation. Erythrocytes and lymphocytes were prepared from ten patients with juvenile neuronal ceroid-lipofuscinosis (Spielmeyer-Vogt-Batten's disease). Copper-zinc and manganese superoxide dismutases were selectively assayed by a direct method on homogenates of the cells. No significant differences were found between the patients, neurological controls and healthy adults.

We studied the influence of plasmapheresis on the plasma concentrations of proteins (IgG, IgA, beta 2-microglobulin) used for estimation of tumour cell mass in patients with multiple myeloma. Simultaneously, the effects of plasmapheresis on plasma concentrations of proteins (CRP, alpha 1-antitrypsin, haptoglobin) used for the assessment of inflammatory processes were studied. Also, changes in the plasma concentration of protein HC, a recently described protein occurring both as a free protein and as an IgA complex, were studied. The influence of plasmapheresis varied for the different proteins. The plasma levels of IgG, IgA, CRP, alpha 1-antitrypsin, and haptoglobin decreased during plasmapheresis. The simultaneous reduction of the level of protein HC was smaller than that of IgG, IgA and haptoglobin. The plasma concentration of beta 2-microglobulin did not decrease during plasmapheresis. The recovery rates of the proteins were found to be approximately inversely related to their molecular weights.

Amyotrophic lateral sclerosis (ALS) is a progressive motor neurodegeneration resulting in paralysis and death from respiratory failure within 3-5 years. About 20% of familial cases are associated with mutations in the gene for copper/zinc superoxide dismutase ( SOD1 ), which catalyses the dismutation of the superoxide radical to hydrogen peroxide and oxygen. Experimental evidence suggests mutations act by a toxic gain of function but the mechanism is unknown. There are >60 known SOD1 mutations associated with ALS and all are dominant except for one in exon 4, a D90A substitution which is recessive. D90A pedigrees with dominant inheritance have now been reported and this apparent contradiction needs to be explained. We performed a worldwide haplotype study on 28 D90A pedigrees using six highly polymorphic microsatellite markers. We now show that all 20 recessive families share the same founder (alpha = 0.999), regardless of geographical location, whereas several founders exist for the eight dominant families (alpha = 0.385). This finding confirms that D90A can act in a dominant fashion in keeping with all other SOD1 mutations, but that on one occasion, a new instance of this mutation has been recessive. We propose a tightly linked protective factor which modifies the toxic effect of mutant SOD1 in recessive families.

B-cells have previously been shown to be very susceptible to damage induced by superoxide radicals, and protection against such damage has been achieved both in vitro and in vivo with superoxide dismutase. During maturation, db/db mice develop diabetes and accumulation of potentially superoxide radical-producing leucocytes can be demonstrated in the islets during the process. To test for the possibility that superoxide radical-induced damage contributes to the development of diabetes, db/db mice were given daily ip injections of 200 mg/kg polyethylene glycolsubstituted CuZn superoxide dismutase. No effect of the treatment could be demonstrated.

1. 1. Mouse pancreatic α-amylase (EC 3.2.1.1) was purified in a two-step procedure comprising isoelectric focusing and chromatography on DEAE-Sephadex. 2. 2. The enzyme was found to be composed of one polypeptide chain with a molecular weight of 54,000 as determined by equilibrium sedimentation and 58,000 as determined by electrophoresis in polyacrylamide gel containing sodium dodecylsulphate. 3. 3. S° 20w was found to be 4.6 · 10 −13 sec. 4. 4. The amino acid composition is presented and is found to be remarkably similar to that of rat pancreatic α-amylase. 5. 5. The isoelectric point was 7.0 as determined by isoelectric focusing in column. 6. 6. The neutral sugar content was 1.5 %. 7. 7. The specific activity was comparatively low, 211 U/mg amylase.

CuZn superoxide dismutase, Mn superoxide dismutase, catalase, and glutathione peroxidase form the primary enzymic defense against toxic oxygen reduction metabolites in cells. To test the importance...

Lesch-Nyhan patients have high concentrations of hypoxanthine in their tissues and body fluids. Since hypoxanthine is a potential oxygen radical generator these patients might be susceptible to a higher oxidative stress than healthy controls. To test this hypothesis we have measured the concentrations of different oxygen radical scavengers in five patients with the Lesch - Nyhan syndrome.

Since neonatal gramnegative infections seem to increase at the same time as breastfeeding diminishes, the role of breast milk in the newborns defence systems needs evaluation. We have studied the presence of antimicrobial factors in colostrum and breast milk-specific (E.coli antibodies of the classes IgA, IgM and IgG) and unepecific (lactoperoxidase, lactoferrin and complement). 1. Lactoperoxidase, which is present in high concentrations in cows milk, was demonstrated only in low quantities in breast milk. This might be compensated by the high concentrations we found in the newborn's saliva. 2. Specific E. coli O-antibodies of the IgA-claee were demonstrated in breast milk and were probably of local production since they were not present in the serum of the mother. In contrast IgG-and IgM-E.coli antibodies were present in lower concentrations in breast milk than in serum. The IgA-antibodies pass along the GI-tract and appear in faeces with retained agglutinating capacity. 3. The relation between the maternal and infantile E.coli strains is studied. Whether or not the specific E.coli antibodies in breast milk influence the selection of the E.coli strains colonizing the infant's bowel is under investigation.

A procedure was established by which CuZn superoxide dismutase in Chinese hamster cells was inhibited to 95% by diethyldithiocarbamate. The inhibition was retained after the diethyldithiocarbamate, a potent radioprotector, was washed away. Cells thus treated with diethyldithiocarbamate were more radiosensitive than control cells, the ratio in the presence of oxygen between the $D_{0}\text{'}{\rm s}$ of the groups being 1.23. The results are compatible with the proposition that superoxide dismutase contributes to the radioresistance of the cells.

The selenium-dependent glutathione peroxidase activities of three mammalian cell lines, HT29, P31, and N-18, cultured in medium with low serum content, increased about 2-, 5-, and 40-fold, respectively, after supplementation with 100 nM selenite. Catalase, CuZn superoxide dismutase, and Mn superoxide dismutase activities were not generally influenced by selenite supplementation, and there was only a minor nonselenium-dependent glutathione peroxidase activity in the investigated cell lines. Gamma-irradiated control and selenite-supplemented cells showed no changes in the surviving fractions, as estimated by clonogenic survival or [3 H]-thymidine uptake, nor were there any significant differences between the two groups in the induction of DNA strand breaks after γ irradiation under repairing (37°C) or nonrepairing (0°C) conditions. The results suggest that selenium-dependent glutathione peroxidase does not contribute significantly to the radiation resistance of cultured mammalian cells.