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Biomedical subjects

S L Marklund

Publications and source records attributed to S L Marklund.

At least 91 records · Page 5Linked to original sources
PubMed1990

Expression of extracellular superoxide dismutase by human cell lines.

Extracellular superoxide dismutase (EC-SOD) is the major SOD isoenzyme in extracellular fluids, but occurs also in tissues. The sites and characteristics of the synthesis of the enzyme are unknown. The occurrence of EC-SOD in cultures of a large panel of human cell lines was assayed by means of an e.l.i.s.a. Unlike the situation for the intracellular isoenzymes CuZn-SOD and Mn-SOD, expression of EC-SOD occurs in only a few cell types. None of the ten investigated suspension-growing cell lines produced EC-SOD. Among normal diploid anchorage-dependent cell lines, expression was found in all 25 investigated fibroblast cell lines, in the two glia-cell lines, but not in six endothelial-cell lines, two epithelial-cell lines or in two amnion-derived lines. Among neoplastic anchorage-dependent cell lines expression was found in 13 out of 29. EC-SOD was secreted into the culture medium by cell lines expressing the enzyme. The rate of EC-SOD synthesis varied by nearly 100-fold among the fibroblast lines and remained essentially constant in the individual lines during long-term culture. In the nine investigated cases, the secreted EC-SOD was of the high-heparin-affinity C type. It is suggested that tissue EC-SOD is secreted by a few well-dispersed cell types, such as fibroblasts and glia cells, to diffuse subsequently around and reversibly bind to heparan sulphate proteoglycan ligands in the glycocalyx of the surface of most tissue cell types and in the interstitial matrix.

Amnion↗
PubMed1990

Superoxide dismutase as an inhibitor of postischemic microvascular permeability increase in the hamster.

The purpose was to elucidate the involvement of superoxide radical (O2-.) in the postischemic increase in the vascular permeability in the hamster cheek pouch. Cheek pouches of anesthetized hamsters were everted, prepared for intravital microscopy, and superfused with a bicarbonate buffered saline solution. Local ischemia for 30 min was obtained using a cuff placed around the proximal part of the cheek pouch. The vascular permeability in the postcapillary venules was quantified as leakage of intravenously injected fluorescein labeled dextran (FITC-dextran, Mw 150,000), using intravital microscopy and fluorimetry. There was a significant and reversible permeability increase after the reperfusion started. In the first series of experiments, combined intravenous infusion and topical application of human recombinant extracellular superoxide dismutase C (EC-SOD C) reduced the postischemic permeability response by 80%. Bovine CuZn-SOD given in exactly the same way reduced the response by 60%. In the second series of experiments, inactivated EC-SOD C was given to the control animals and active EC-SOD C was given to the treated animals. The topical treatment was excluded. Only active EC-SOD C reduced significantly the postischemic permeability increase when present during the ischemic period. Treatment with mannitol (i.v.) did not alter the postischemic response. Since active EC-SOD C and CuZn-SOD but not inactivated EC-SOD C effectively inhibited the response, we suggest that the superoxide anion is involved in the mediation of the postischemic permeability increase in the hamster.

Animals↗
PubMed1990

Recombinant human extracellular superoxide dismutase reduces concentration of oxygen free radicals in the reperfused rat heart.

STUDY OBJECTIVE: The aim of the study was to determine whether recombinant human extracellular superoxide dismutase type C (rh-EC-SOD C) has antioxidant effects similar to those of the previously studied Cu-Zn superoxide dismutase. DESIGN: Electron paramagnetic resonance spectroscopy was used for quantification of free radical generation during reperfusion of postischaemic isolated rat hearts. The hearts were perfused with Krebs-Henseleit buffer and then subjected to 15 min global ischaemia. N-tert-butyl-alpha-phenylnitrone (PBN) was added as a spin trapping agent to the perfusate. EXPERIMENTAL MATERIAL: 18 hearts from Sprague-Dawley rats (280-400 g) were used. MEASUREMENTS AND MAIN RESULTS: Reperfusion resulted in a burst of radical formation. In presence of rh-EC-SOD C plus catalase, the PBN spin adduct concentration was only 18% (p less than 0.05) of the value in the control group. With bovine Cu-Zn superoxide dismutase plus catalase added to the buffer the corresponding spin adduct formation was 44% of control (NS = v rh-EC-SOD C). CONCLUSIONS: In isolated rat heart, rh-EC-SOD C reduces free radical concentrations at least to the same extent as Cu-Zn superoxide dismutase.

Animals↗
PubMed1990

Limitation of myocardial infarct size by superoxide dismutase as an adjunct to reperfusion after different durations of coronary occlusion in the pig.

Superoxide dismutase (SOD) has been documented to limit myocardial infarct size in the richly collateralized dog heart. This study was designed to explore this concept in a low-collateralized animal model. A blind, randomized, placebo-controlled protocol was used in 65 pentobarbital-anesthetized pigs subjected to closed-chest left anterior descending coronary artery occlusion for 30 (n = 22), 60 (n = 22), and 90 (n = 14) minutes followed by reperfusion up to 24 hours from the start of occlusion. Another seven control pigs were subjected to 24 hours of permanent occlusion. A total dose of 9 mg/kg bovine CuZn SOD was administered as a bolus injection immediately before reperfusion followed by a 1-hour infusion. Infarct size was assessed by tetrazolium staining. Myocardium at risk and collateral flow were determined by using cerium-141-labeled microspheres (15 microns) during the occlusion. After 30 minutes of occlusion, infarct sizes in placebo versus SOD-treated animals were 45.5 +/- 15.7% vs. 23.8 +/- 15.6% of myocardium at risk (p = 0.007). The corresponding values after 60 minutes of occlusion were 78.6 +/- 9.3% vs. 66.9 +/- 14.6% (p = 0.035). SOD administered after 90 minutes of occlusion did not limit infarct size (88.5 +/- 4.8% vs. 92.3 +/- 5.2%). Twenty-four hours of coronary occlusion resulted in infarction of 92.4 +/- 4.2% of myocardium at risk. (All values are mean +/- SD.) Ventricular fibrillation occurred in only nine pigs distributed equally between SOD and placebo. The results indicate that CuZn SOD has the potential to further improve the myocardial salvage established by reperfusion of an ischemic pig heart territory. However, the narrow time window for limiting infarct size in the pig by reperfusion is not much extended by SOD.

Animals↗
PubMed1990

Mercury, selenium, and glutathione peroxidase before and after amalgam removal in man.

In 10 healthy persons all amalgam fillings were replaced with gold inlays. Blood and urinary levels were measured on 10 occasions during a 4-month period before and a 12-month period after amalgam removal. These variables were also measured three times in 10 healthy controls. A strong statistically significant relation was found between plasma mercury values and both the total number of amalgam surfaces (r = 0.71, p = 0.0006) and the total surface area of the fillings (r = 0.73, p = 0.0004). In the immediate postremoval phase plasma mercury rose three- to four-fold, whereas the urinary and erythrocyte mercury rose about 50%. These peak values declined to the preremoval level at about 1 month. Twelve months after the removal the plasma and urinary mercury levels were significantly reduced to 50% and 25%, respectively, of the initial values for the experimental group. Apart from the significantly lower plasma selenium values 5 and 10 days after removal no significant differences were found with regard to plasma selenium or erythrocyte glutathione peroxidase either within or between the experimental and the control groups. A large number of supplementary biochemical analyses did not show any influence on organ functions or any differences between the groups before or after the amalgam removal. Amalgam fillings considerably contributed to the plasma and urinary mercury levels.

Adult↗
PubMed1990

The influence of dental amalgam placement on mercury, selenium, and glutathione peroxidase in man.

Amalgam restorations were inserted in eight healthy persons, previously unprovided with dental restorations, who had several severe carious lesions. The mean number of surfaces restored were 16.1 (range, 11 to 22). The total mean calculated amount of mercury inserted was 2.9 g (range, 1.5 to 4.3 g). Blood and urinary levels were measured on seven occasions during a 4-month period before and a 3-month period after amalgam placement. One and 3 months after placement, the P-mercury mean values were almost equal to the preplacement values (3.3 nmol/l). After placement U-mercury increased continuously; 3 months after placement a statistically significantly higher (p less than 0.05) mean U-mercury value (0.58 nmol/mmol creatinine) was found compared with the mean preplacement value (0.34 nmol/mmol creatinine). No statistically significant correlation was found between the P- and U-mercury concentrations and the total number of amalgam surfaces. Selenium levels in plasma and urine and erythrocyte glutathione peroxidase showed no systematic change of pattern. The results show that the insertion of amalgam fillings contributed to the U-mercury concentration, but apparently even more extensive amalgam therapy and/or longer exposure periods are needed to affect the P-mercury concentration. No negative effects on the P- and U-selenium or the erythrocyte glutathione peroxidase levels could be found during the 3 months immediately after an extensive amalgam placement. The supplementary blood and urine analyses were not influenced by the insertion of amalgam fillings.

Adult↗
PubMed1989

Interactions between human extracellular superoxide dismutase C and sulfated polysaccharides.

The high heparin affinity subtype C of the secretory enzyme extracellular superoxide dismutase (EC-SOD) exists in the body mainly complexed with extracellular sulfated glycosaminoglycans (SGAGs). Addition of sulfated polysaccharides to EC-SOD C resulted in a prompt partial inhibition of the enzymic activity, in most cases amounting to 10-17%, but with the large dextran sulfate 500,000 amounting to 35%. Complex formation between heparin and EC-SOD C could also be observed as increases in apparent molecular weight of the enzyme. The findings suggest that the binding sites for SGAGs on EC-SOD C are localized far from the active site and that EC-SOD in vivo associated with SGAGs should retain the major part of its enzymic activity. Studies with amino acid-specific reagents suggested that both lysine and arginine residues are involved in the binding of SGAGs. In particular, modification of only a few lysine residues/subunit resulted in loss of high SGAG affinity, whereas arginine modification resulted in loss of not only SGAG affinity but also enzymic activity. We propose that this is due to modification of Arg-186, which is homologous to the highly conserved arginine in the entrance to the active site of the copperzinc-SODs.

Animals↗
PubMed1989

Lipid peroxidation and activity of antioxidant enzymes in muscle of the lower leg before and after arterial reconstruction.

Reperfusion syndrome and lipid peroxidation due to toxic effects of free oxygen radicals might be one pathophysiological cause of the oedema that develops during the first week after a femoro-popliteal reconstruction. This paper reports the activity of the protecting antioxidant enzymes superoxide dismutase (SOD), glutathione peroxidase and catalase in gastrocnemic muscle before and after reperfusion of chronically ischaemic legs with comparison to activities in non-ischaemic muscle. Furthermore the susceptibility to lipid peroxidation in the muscle was measured as thiobarbituric acid reactive material (TBAR). The activities of CuZn SOD, Mn SOD and glutathione peroxidase were equal in normally perfused and chronically ischaemic muscle and there was no difference after reperfusion. Muscle catalase activity was low compared to activity of red blood cells and could not be reliably estimated. There was no difference in iron-stimulated lipid peroxidation of ischaemic and non-ischaemic muscle but in reliably estimated. There was no difference in iron-stimulated lipid peroxidation of ischaemic and non-ischaemic muscle but in muscle biopsied 10 min after reperfusion there was a significant increase in production of TBAR indicating an increased susceptibility for lipid peroxidation at this time. The finding is compatible with the occurrence of an oxidant insult on the muscle at reperfusion. Ischaemia--or reperfusion--induced reductions in activity of antioxidant enzymes are however not related to this reaction.

Arteriosclerosis↗
PubMed1989

Mercury, selenium, and glutathione peroxidase in dental personnel.

Eighteen persons, dentists and nurses, with urinary mercury levels higher than the group median value of all dental personnel in the country of Västerbotten were compared with a group consisting of 15 persons with low urinary mercury levels working in the same clinics. A statistically significant difference between the high urinary mercury group and the low urinary mercury group could be seen in the plasma mercury level. In each group a statistically significant relation could be seen between the plasma mercury level and the total number of amalgam surfaces. The two groups did not differ with regard to the levels of plasma selenium and erythrocyte glutathione peroxidase, and no correlation between these two variables and the plasma mercury levels could be found. To evaluate organ functions, a large number of supplementary analyses were performed. These analyses did not indicate any influence on organ functions. Although the persons in the present study were occupationally exposed to mercury, none of the biologic variables analyzed seemed to be affected. Even among dental personnel who handle amalgam professionally the number of amalgam surfaces is a major contributory factor to the P-mercury level.

Adult↗
PubMed1989

Binding of human extracellular-superoxide dismutase C to cultured cell lines and to blood cells.

The high heparin-affinity subtype C of the secretory enzyme extracellular-superoxide dismutase (EC-SOD) was found to bind to cultured mammalian cells, forming an equilibrium between the cells and the medium. To anchorage-dependent cell lines, binding apparently occurred both to the glycocalyx of the cell surfaces and to the sub- and intercellular matrix produced by the cells. Heparan sulfate proteoglycan appeared to be the principal binding substance. The binding capacities of anchorage-dependent cultures were very high, and at maximal binding the amount of EC-SOD C activity associated with the exterior of the cells was several-fold higher than the endogenous intracellular SOD activity. Half-maximal binding occurred at about 8 micrograms/ml EC-SOD C. At low, nonsaturating, physiologic EC-SOD C concentrations, the enzyme concentration in the glycocalyx of cells may be several thousand times higher than in the medium. All 14 investigated anchorage-dependent cell lines, including endothelial cells, bound EC-SOD C avidly. The 10 suspension-growing cell lines were all weaker binders. Blood monomorphonuclear leukocytes and platelets bound little EC-SOD C, whereas no significant binding to neutrophil leukocytes, to erythrocytes and to E. coli could be demonstrated. The findings are compatible with the notion that EC-SOD C in the vasculature forms an equilibrium between plasma and heparan sulfate in the glycocalyx of the endothelium. Furthermore, tissue EC-SOD is probably distributed between heparan sulfate on the surface of most cell types in the organs and in the interstitial matrix. The binding pattern suggests that EC-SOD C has the potential to protect most normal cells in the body and the interstitial matrix, without protecting microorganisms lacking affinity, and without interfering with superoxide radicals produced at the surface of activated neutrophil leukocytes.

Animals↗
PubMed1989

Selenite-induced variation in glutathione peroxidase activity of three mammalian cell lines: no effect on radiation-induced cell killing or DNA strand breakage.

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 [3H]-thymidine uptake, nor were there any significant differences between the two groups in the induction of DNA strand breaks after gamma irradiation under repairing (37 degrees C) or nonrepairing (0 degrees C) conditions. The results suggest that selenium-dependent glutathione peroxidase does not contribute significantly to the radiation resistance of cultured mammalian cells.

Animals↗
PubMed1988

Binding of human extracellular superoxide dismutase C to sulphated glycosaminoglycans.

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.

Affinity Labels↗
PubMed1988

Heparin-, dextran sulfate- and protamine-induced release of extracellular-superoxide dismutase to plasma in pigs.

Intravenous heparin has previously been shown to release the high-heparin-affinity fraction C of extracellular-superoxide dismutase (EC-SOD, EC 1.15.1.1) to plasma in man and other mammals. This paper reports on further studies of the phenomena in the pig. A dose-response curve of the effect of heparin revealed that 1000 IU/kg body weight is needed for maximal release of EC-SOD C. This dose is an order of magnitude larger than that needed for the maximal release to plasma of factors such as lipoprotein lipase, hepatic lipase, and diamine oxidase, which are distributed between plasma and endothelium similarly to EC-SOD C. Thus EC-SOD C appears to have an unusually high affinity for endothelial cell-surface sulfated glycosaminoglycans relative to the affinity for heparin. There was no significant difference in releasing potency between unfractionated heparin and heparin subfractions with high or low affinity for antithrombin III. The heparin structure conferring high-affinity binding to antithrombin III is thus not specifically involved in binding to EC-SOD C. The non-biosynthetic compound dextran sulfate 5000 was an order of magnitude more efficient than heparin. Protamine displayed dual effects. Given alone in high dose it released EC-SOD to plasma, probably due to binding to endothelial cell-surface sulfated glycosaminoglycans displacing fraction C of the enzyme. When given after heparin, in a dose just below that expected to neutralize the heparin, protamine reversed the heparin-induced EC-SOD release.

Animals↗
PubMed1988

Extracellular superoxide dismutase in the vascular system of mammals.

Plasma extracellular superoxide dismutase (EC-SOD) from the pig, cat, rabbit, guinea pig and mouse was found to be heterogeneous with regard to heparin affinity and could be separated into three fractions: A without affinity, B with weak affinity and C with relatively high affinity. Rat plasma EC-SOD was deviant and contained only A and B fractions. There were very large interspecies differences in total plasma EC-SOD activity and in division of the activity between the different fractions. Intravenous injection of heparin resulted in the pig, dog, cat, rabbit, guinea pig and mouse in a prompt increase in the plasma SOD activity. The increase was due to release of EC-SOD C to plasma, most probably from endothelial cell surfaces. In the rat, heparin induced no increase in plasma SOD activity, apparently because of the absence of EC-SOD C in this species. The relative heparin-induced increase in plasma EC-SOD C varied between 2 and 11 in the different species and was distinctly correlated with the heparin affinity of EC-SOD C in the particular species. Apparently the EC-SOD C, present in the vasculature, forms an equilibrium between plasma and endothelium, whereas EC-SOD A and B primarily exist in plasma. The wide diversity of EC-SOD in the vascular system of mammals with regard to total amount, division into fractions and distribution between plasma and endothelium indicates that the pathogenic potential of superoxide radicals in the extracellular space might vary much between species.

Affinity Labels↗