Sándor Túri

Oxidative stress and antioxidant defense mechanism in glomerular diseases

The changes in red blood cell (RBC) lipid peroxidation [measured via the malonyl dialdehyde (MDA) concentration], reduced (GSH), and oxidized glutathione (GSSG) levels, hemoglobin (Hb) oxidation and antioxidant enzyme [catalase (Cat), glutathione peroxidase, and superoxide dismutase (SOD)] activities were studied in 45 pediatric patients with various glomerular diseases [minimal change nephrotic syndrome (MCNS) in relapse or in remission, lupus nephropathy (SLE), poststreptococcal glomerulonephritis (APSGN), IgA nephropathy (IGA gn)], and in 20 adult patients with IGA gn and also in 15 pediatric and 14 adult controls. The in vitro effects of hydrogen peroxide [acetyl phenylhydrazine (APH) test] on the GSH and Hb metabolisms were likewise investigated. There was an increased oxidative stress in MCNS with relapse, IGA gn, SLE gn, and APSGN, which could be detected in the GSH and Hb oxidation and in the lipid peroxidation on the peripheral RBC-s. The RBC SOD and Cat activities were significantly lower in all patients than in the controls. The RBC GSSG level was significantly elevated in all patients, with the exception of MCNS in remission. This stimulated a compensatory GSH production in MCNS with relapse and in IGA gn, but not in SLE or APSGN. The regeneration of GSH from GSSG was reduced in MCNS with relapse, SLE, and IGA gn, but not in APSGN. In remission, the GSH-GSSG redox system normalizes, but in vitro the APH test stimulates an intensive Hb oxidation. In conclusion, there is a correlation between the presence of active glomerular disease and the evidence of oxidative changes in the various parameters measured in peripheral RBCs.

Vitamin E alleviates the oxidative stress of erythropoietin in uremic children on hemodialysis.

Abstract The efficacy of combined therapy with recombinant human erythropoietin (rhEPO) and vitamin E versus rhEPO alone in the treatment of anemia was examined in children (n=10, aged 15.2±3.2 years) on chronic hemodialysis at the restart of rhEPO therapy after a 4-week interval. The results confirmed that rhEPO induced oxidative stress of the red blood cells as observed during the first rhEPO therapy. Vitamin E (15 mg/kg per day per os) was introduced after 2 weeks of rhEPO monotherapy, when the signs of acute oxidative stress appeared. The level of oxidized glutathione (GSSG) increased from 8.9±3.1 to 26.7±5.7 nmol/g hemoglobin (Hb) by that time. After 2 weeks of simultaneous vitamin E treatment, there was a significant difference in GSSG values compared with rhEPO monotherapy (10.1±3.9 vs. 56.7±15.8 nmol/g Hb, P<0.001). A considerable decrease was observed in the previously high ratio of GSSG/reduced glutathione (GSH), an indicator of oxidative stress, and the level of carboxyhemoglobin, indicating hemolysis. A significant increase in Hb and hematocrit (P<0.01) was achieved within 2 weeks of starting the combined therapy, while similar results occurred only at the 8th and 5th weeks without vitamin E. Antioxidant vitamin E supplementation improved the therapeutic effect of rhEPO in patients with chronic renal failure on hemodialysis.

Erythrocyte defense mechanisms against free oxygen radicals in haemodialysed uraemic children

Changes in erythrocyte lipid peroxidation (measured as the concentration of malonyl dialdehyde), glutathione metabolism, antioxidant enzyme activities (glutathione peroxidase, catalase, and superoxide dismutase), the oxidized products of haemoglobin (Hb), and hydrogen peroxide (H2O2)-induced haemolysis were studied in six children with chronic renal failure treated with serial acetate and bicarbonate haemodialysis (HD). Ten age- and sex-matched children acted as controls. Malonyl dialdehyde levels were significantly higher and antioxidant enzyme activities lower in uraemic red blood cells (RBCs) compared with controls (P<0.05). Incubation of RBCs for 1 h with acetylphenylhydrazine induced a decrease in the concentration of reduced glutathione (P<0.001) and an increase in the level of oxidized products of Hb (P<0.001), but only in the uraemic patients. The H2O2 haemolysis test revealed a mild (n=3) to increased (n=3) haemolysis in the uraemic RBCs. Oxidative haemolysis is probably a multifactorial process in uraemic patients, and may be an important risk factor in HD therapy.

Oxidative damage of red blood cells in haemolytic uraemic syndrome

Changes in red blood cell (RBC) lipid peroxidation [measured by malonyl dialdehyde (MDA) concentration], glutathione (GSH) metabolism, antioxidant enzyme activities (catalase, superoxide dismutase, glutathione peroxidase) and haemoglobin (Hb) metabolites (metHb, carboxy Hb) were studied in six children with post-enteropathic (D+) haemolytic uraemic syndrome (HUS) and ten controls. The in vitro effect of hydrogen peroxide [acetyl-phenylhydrazine (APH) test] on GSH and Hb metabolism was also investigated. MDA levels were significantly higher and the antioxidant enzyme activities were lower in HUS patients than in the controls (P<0.01). The oxidised glutathione concentration was significantly higher in the patients than in the control children (26.3±12.6 vs. 10.9±1.8 nmol/g Hb. Percentage values of carboxy Hb and metHb were also higher in HUS (P<0.01). Incubation of RBC with APH induced a more pronounced decrease in the concentration of GSH (P<0.001) and a significant increase (P<0.01) in the level of metHb and carboxy Hb in the HUS patients. This suggests that there is reduced RBC GSH stability in HUS. Utilisation of GSH and antioxidant enzymes leads to increased Hb oxidation and haemolysis. The oxidative damage may have an important role in the pathogenesis of haemolytic anaemia in HUS.

The effect of erythropoietin on the cellular defence mechanism of red blood cells in children with chronic renal failure

The glutathione redox system, haemoglobin (Hb) oxidation, the activity of antioxidant enzymes and the lipid peroxidation product malonyl dialdehyde (MDA) were studied in red blood cells (RBCs) during administration of recombinant human erythropoietin (rhEPO) over 12 weeks in ten children maintained on haemodialysis. A rapid increase in the reticulocyte count was accompanied by a slower rise in total Hb concentration. The mean level of oxidized glutathione (GSSG) increased from 13.2±5.3 nmol/g Hb to 56.7±15.8 nmol/g Hb 4 weeks after the start of rhEPO (P<0.001), followed by a fall to the basal value. Reduced glutathione (GSH) levels showed a smaller though constant elevation during rhEPO therapy (P<0.001). Before rhEPO treatment, incubation of RBCs for 1 h with acetylphenylhydrazine induced a decrease in GSH concentration compared with controls (P<0.001), which became more pronounced in the first few weeks of rhEPO therapy (P<0.001). In addition, the percentage of Hb derivatives (metHb and haemichrome) increased in the first 4 weeks of rhEPO therapy (P<0.001). Although there was no significant difference between the values obtained preEPO and during EPO treatment, MDA levels were continuously higher and superoxide dismutase, catalase and glutathione peroxidase concentrations were lower than in the controls (P<0.001). These results are compatible with oxidative damage to the RBCs in the early period of rhEPO therapy in children with end-stage renal failure. The GSH-GSSG system, as an important cellular defence mechanism of the RBCs, appears to be severely affected.

Long-term follow-up of patients with persistent/recurrent, isolated haematuria: A Hungarian multicentre study

A retrospective multicentre study of 341 children with persistent/recurrent, isolated haematuria is described. The haematuria was isolated for at least 6 months at the beginning of observation. The duration of follow-up was 2–5 years in 201, 5–10 years in 119, 10–15 years in 19, and over 15 years in 2 cases. Of these patients 47.8% became symptom-free. In 18.4% the haematuria remained isolated; in 13.8% it was combined with proteinuria over 250 mg/day more than 2 years later. The occurrence of associated proteinuria increased progressively with time. It was 8.6% between the 3rd and 5th years, and 37.0% after the 5th year. Renal biopsy was performed because of the symptoms of glomerular disease in 47 cases at an average time of 12 months following the appearance of proteinuria. Proteinuria appeared after a 2–5, 5–10, 10–15 and more than 15 years follow-up period in 16, 23, 6, and 2 patients respectively; 14 of them had Alports nephropathy. The percentage of more serious azotaemia was 1.7 (creatinine clearance: 10–50 ml/min per 1.73 m2) and 0.3 (creatinine clearance: < 10 ml/min per 1.73 m2). Mortality was 0.58%. Most of the patients who developed severe azotaemia had persistent microscopic haematuria at the beginning. The prevalence of hypertension was only 1.2%. The time of its appearance was above 5 years in 2 and below 5 years in 2 cases. All these patients had chronic glomerulonephritis. The haematuria was associated with hypercalciuria in 19.9%. In 14.3% of the overall group of patients urolithiasis developed 2–15 years after onset. All of these had hypercalciuria. Our findings suggest that symptoms of isolated haematuria may last for a longterm period and need systematic control. When proteinuria and/or hypertension is associated with haematuria a worse prognosis can be expected.

Mitochondrial mutation as a probable causative factor in familial progressive tubulointerstitial nephritis

Abstract Renal biopsy of two children and a maternal relative, diagnosed with severe progressive tubulointerstitial nephritis, has shown the presence of distorted mitochondria. Mitochondrial DNA from the blood of these patients was analysed. No major deletions were found, but an A to G mutation was detected in position 5656. It is proposed that this mutation might play a causative role in the renal disease of the patients.

The effect of erythropoietin on platelet function in uraemic children on haemodialysis

The effect of 1-year erythropoietin (rHu-EPO) treatment on the bleeding time, platelet aggregation, ATP and thromboxane B2 (TXB2) release, cyclic AMP (cAMP) concentration and platelet surface positive charge were studied in 8 haemodialysed children with chronic uraemia and 8 controls. The pre-dialysis haematocrit (Hct) was 0.21+0.01 before and 0.36+0.01 following 1 year of rHu-EPO therapy. At the end of this period the pre-dialysis bleeding time became normal (P<0.05); this was associated with a significant increase in platelet aggregability (P<0.05), ATP release (P<0.05) and TXB2 production (P<0.01), and with a significant decrease in platelet cAMP concentration (P<0.01). A further increase in platelet aggregation, ATP release and TXB2 production and a decrease in platelet cAMP concentration was observed following bicarbonate haemodialysis (BHD) (P<0.01). There was a significant positive correlation between platelet aggregation and ATP release (r=0.78,P<0.05), as well as platelet aggregation and TXB2 production (r=0.68,P<0.05). A significant negative correlation was found between platelet aggregability and cAMP concentration (r=−0.7,P<0.05). The platelet surface positive charge, which was significantly lower in the patients than in the controls (P<0.01), did not change during rHu-EPO therapy, nevertheless BHD resulted in a significant increase (P<0.05), suggesting the surface charge may influence platelet aggregation. In an in vitro and an in vivo study, rHu-EPO and the higher Hct did not increase platelet aggregation directly. Long-term administration of rHu-EPO stimulated complex functional and biochemical changes in the platelets of uraemic patients, which resulted in an improved aggregability.

Plasma factors influencing PGI2-like activity in patients with IgA nephropathy and Henoch-Schönlein purpura

Plasma factors influencing vascular PGI2-like activity (PSA) were studied in 45 patients with IgA nephropathy, 18 with Henoch-Schönlein purpura, including 8 children with nephrotic syndrome, and 41 controls. The results were compared with the levels of plasma high-density lipoprotein (HDL), low-density lipoprotein (LDL) and fatty acid components of plasma phospholipids. The plasma of 38 of 45 patients with IgA nephropathy and 14 with Henoch-Schönlein purpura showed a diminished ability or no ability to support PSA. Twenty-three patients with IgA nephropathy and 10 with Henoch-Schönlein purpura exhibited an inhibitory activity against PGI2 production. The plasma HDL level was lower, while the LDL level and the LDL/HDL ratio were significantly higher in IgA nephropathy and Henoch-Schönlein purpura cases than in the controls. A high LDL/HDL ratio was associated with a low plasma PSA. The levels of arachidonic acid and its precursor were not lower in the plasma of patients than in the controls. The decreased PGI2 synthesis may play an important role in the pathogenesis of IgA nephropathy and Henoch-Schönlein purpura, but it can not be explained by reduced PG precursors. LDL may have an inhibitory, and HDL a protective effect on PGI2 synthesis.

Laser aggregometer studies, ATP release and thromboxane B2 release and cAMP concentration of the platelets in nephrotic syndrome

Platelet function was studied in 56 children with nephrotic syndrome, 33 were on oral prednisolone (P) treatment (group 1), while 23 were in early (< 6 months) remission (group 2): 12 on P (group 2a) and 11 not on P (group 2b), and there were 18 controls (group 3). The following tests were used: platelet aggregation with collagen in a laser rheoaggregometer; adenosine triphosphate (ATP) release: during aggregation with luciferin-luciferase in a lumiaggregometer; thromboxane B2 (TXB2) release: by radio-immunoassay; platelet cAMP concentration: by binding assay. The changes in plasma cholesterol (C) and triglycerides (TG) were compared with the platelet aggregation results. Patients in group 1 and 2 exhibited significantly higher aggregability, TXB2 release and ATP release in response to collagen than those in group 3 (p < 0.01), but there was no difference between groups 1 and 2 or groups 2a and 2b. Some differences were observed between the histological groups. Patients with IgA and SLE nephropathy displayed higher aggregability than those with minimal change nephrotic syndrome in remission (p < 0.05). The highest level was in membranous nephropathy. The platelet cyclic adenosine monophosphate (cAMP) concentration was significantly lower in groups 1 and 2 than in group 3 (p < 0.001). No differences were observed between groups 1 and 2 or between groups 2a and 2b. Plasma C and TG levels did not show any correlation with the platelet aggregation. The increased platelet aggregation, TXB2 release and ATP release and decreased cAMP concentration in the platelets may play a role in the pathogenesis of nephrotic syndrome which remains altered in early remission. Oral P and plasma lipids did not change the platelet function significantly.