P. De Sole

Superoxide dismutase, glutathione peroxidase and catalase in oxidative hemolysis. A study of Fanconi's anemia erythrocytes

Abstract Superoxide dismutase, glutathione peroxidase and catalase were assayed in the erythrocytes of three patients of Fanconis anemia. Superoxide dismutase was found to be significantly decreased, as previously reported. The enzymes metabolizing H 2 O 2 are normal (glutathione peroxidase in the higher limits of the normal value). The abnormal erythrocytes were found to be as resistant (perhaps more resistant) as normal red blood cells to oxidative hemolysis induced by drugs. Malonyl dialdehyde production was found to be comparable to that of normal erythrocytes. It is concluded that a significant (30–40%) deficiency of superoxide dismutase, when associated to normal values of H 2 O 2 -removing enzymes, does not affect the antioxidative defense capability of erythrocytes, even in conditions of augmented oxidative injury.

Ferritin iron content in haemodialysis patients : Comparison with septic and hemochromatosis patients

OBJECTIVES Hemodialysis (HD) population commonly show high plasma ferritin levels with a poor diagnostic value. The objective of this study is to elucidate the meaning of HD hyperferritinemia through the analysis of its ferritin iron content (FIC). DESIGN AND METHODS FIC (iron atoms/ferritin molecule) was measured by atomic emission spectrometry. Ferritin and FIC values were correlated with iron storage and inflammation markers and the results of HD patients compared to those of septic and hemochromatosis patients. RESULTS 1) In the whole HD population, high ferritin levels were associated to low FIC values; 2) the correlation of ferritin with iron indices and inflammation markers in HD patients was intermediate in between that of septic and hemochromatosis patients; 3) the FIC level of HD patients was lower than that of the other two groups. CONCLUSIONS The high ferritin levels of HD patients are not synonymous with either inflammation or of high levels of iron storage. Their high levels and the low FIC values might be due to the presence inside the ferritin core of oligoelements other than iron.

A two-dimensional electrophoresis preliminary approach to human hepatocarcinoma differentiation induced by PPAR-agonists.

Adopting biochemical and proteomic approaches, we investigated the effect of some PPAR‐agonists, a new class of differentiating agents, on human hepatocellular carcinoma Hep‐G2 cell line. Cancer differentiation was assayed by checking albumin, transferrin and α‐fetoprotein synthesis. Cell metabolism was studied by NMR spectroscopy of cell culture supernatants and by evaluation of mitochondrial respiratory chain enzyme activities. The two dimensional electrophoresis approach was employed to analyze modifications in the expression of cellular proteins linked to cell phenotype differentiation in the attempt to identify potential diagnostic and prognostic biomarkers of hepatocellular carcinoma. Results indicate that PPAR‐agonists are able to act as differentiating inducers in human hepatocellular carcinoma Hep‐G2 cell line as well as to inhibit mitochondrial respiratory chain Complex I, provoking a selective derangement of cellular oxidative metabolism. Lastly, two dimensional electrophoresis showed interesting modifications in the pattern of expression of cellular proteins that confirm biochemical data (increase in albumin and transferrin, decrease of alpha ‐fetoprotein synthesis) and, moreover, emphasize the meaning of these data by the increase of spots indicatively ascribed to HSP70 and catalase.

Iron, zinc and aluminium ferritin content of hemodialysis hyperferritinemic patients: Comparison with other hyperferritinemic clinical conditions and normoferritinemic blood donors

The present study describes the specific content of ferritin iron, zinc and aluminium in four different groups: 1) hemodialysis hyperferritinemic patients; 2) septic patients; 3) iron overloaded patients with hematologic diseases; and 4) blood donors. In all four groups high levels of aluminium and zinc were found in addition to those of iron. However, the sum of the ferritin ions of the control group is significantly higher than that of the other three groups. Furthermore, while ferritin of hemodialysis patients has the same molecular ratio of metal ions as control group (high Al content vs. Fe and Zn), a lower Al/Fe ratio is found both in septic and hematological patients. The results of the present paper might help to explain the high percentage of hyperferritinemia found in hemodialysis patients also in presence of low transferrin saturation and in absence of inflammatory markers. Moreover, the high content of ions other than iron in the ferritin core leads us to believe that ferritin is not only an iron storage protein but rather a regulator of redox active ions.

Impaired reactive oxygen metabolism of phagocytic leukocytes in NIDDM patients. A role for non-enzymatic glycosylation of collagen.

Non-enzymatic glycosylation (NEG) of collagen has been previously shown to significantly influence the reactive oxygen metabolism (ROM) of phagocytic cells in healthy subjects. Considering the role of NEG in the pathophysiology of diabetes, we have further analysed the oxidative metabolism of polymorphonuclear cells (PMNs) and monocytes in 23 patients with non-insulin dependent diabetes mellitus in order to better elucidate a possible pathogenic role of NEG of the extracellular matrix in long-term complications of diabetes. Experiments were performed in triplicate on native-collagen and glycated-collagen coated vials, using a chemiluminescence (CL) assay. Results show that PMNs from diabetic patients display a significant increased basal and zymosan-induced CL activity with respect to controls that are not related to the glycation state of the substrate. Conversely, the CL activity of monocytes induced by zymosan shows a decrease in diabetic patients with respect to healthy volunteers (p < 0.05). Moreover, monocyte CL was reduced by the glycated matrix, both in healthy volunteers and in diabetic subjects (p < 0.05 and p < 0.01, respectively). These data highlight a complex role of phagocytic leukocytes in the pathophysiology of extracellular matrix alterations secondary to NEG that are typically present in clinical conditions such as diabetes or ageing.

Lactic dehydrogenase and glycolytic activity of human platelets in uncomplicated pregnancy.

Lactic dehydrogenase (LDH) of isolated platelets was determined in a group of 103 normally pregnant and 19 unpregnant women. LDH activity is significantly increased in the last trimester of pregnancy while the isoenzyme pattern does not vary in relation to LDH increase. Moreover, glucose consumption and lactic acid production are linearly correlated to platelet LDH activity.

MONOCYTE OXYGENATION ACTIVITIES ARE SIGNIFICANTLY INFLUENCED BY NON-ENZYMATIC GLYCOSYLATION OF COLLAGEN

The pathophysiology of the acquired extracellular matrix alterations, like non-enzymatic glycosylation (NEG) of proteins secondary to diabetes or ageing, is not well characterized, particularly considering its relationship with leukocytes. We have analysed the influence of collagen NEG on the fundamental function of phagocytic cells, i.e. the production of reactive oxygen species (ROS). To this aim, we considered the activity of polymorphonuclear leukocytes and monocytes incubated in the presence of both non-glycated and glycated collagen. ROS production was monitored by chemi-luminescence (CL), a standardized and sensitive assay of phagocytes oxidative metabolism. All experiments were performed in triplicate on collagen-coated and glycated collage-coated vials. Results showed that PMNs ROS metabolism appeared unrelated to the glycation state of the substrate. Conversely, data regarding zymosan-induced CL by monocytes indicated a significant and intriguing decrease in reactive oxygen metabolism, which appeared greatly compromised by the glycation state of the matrix (monocytes in collagen, 197.4 +/- 31.2 vs. monocytes in glycated collagen, 138.0 +/- 20.4; p < 0.001 expressed as counts/cell/60 min). These data highlight the different role of polymophonuclear and monocytic phagocytes in the pathophysiology of the acquired extracellular matrix alteration secondary to non-enzymatic glycosylation (NEG) of proteins.

Lactic dehydrogenase activity of platelet subpopulations during pregnancy.

SummaryWe determined the LDH activity of different platelet subpopulations throughout pregnancy. While no changes in LDH content occurred in large, dense platelets, a significant change was found in small, light platelets.

Platelet heterogeneity based on lactic dehydrogenase activity.

SummaryMean lactic dehydrogenase (LDH) activity of human platelets is about 5 nU/platelet; expressed on a cell volume basis, it is 0.5 nU/fl. Platelet separation by means of centrifugation on a Percoll discontinuous gradient gives different subpopulations with a clear-cut change of LDH at a mean platelet volume (MPV) of about 6 fl. The LDH content of platelets with a MPV >6 fl is constant (about 0.5 nU/fl), while that of small platelets (3<MPV<6 fl) is inversely correlated with the MPV and reaches a value of 1.5–2.0 nU/fl. Because small platelets are granule-depleted, we suggest that platelet LDH activity be considered as an in vivo activation index.