Jaroslav Racek

Influence of chromium-enriched yeast on blood glucose and insulin variables, blood lipids, and markers of oxidative stress in subjects with type 2 diabetes mellitus

The aim of this study was to determine the effect of chromium (Cr)-enriched yeast on blood glucose and insulin variables, blood lipids, and blood markers of oxidative stress in persons with type 2 diabetes mellitus (median duration: 3.0 yr). Thirty-six subjects (9 men, 27 women; mean age: 61.3 yr; mean body mass index: 34.33 kg/m2) were supplemented with 400 μg Cr/d as Cr-enriched yeast (n=19) or placebo (n=17) for 12 wk in a randomized, double-blind study. The most interesting results were obtained by comparison of the change in the placebo group to the change in the Cr group. The Cr group showed a significantly greater increase in serum Cr compared to the placebo group (p<0.05). Supplementation with Cr-enriched yeast was associated with a significant decrease in fasting serum glucose compared to placebo (p<0.01). Blood markers of oxidative stress glutathione peroxidase activity and levels of reduced glutathione were essentially unchanged in the Cr group after 12 wk, but decreased significantly in the placebo group (p<0.05, p<0.01, respectively), Serum HbA1c and glycated protein (fructosamine) were essentially unchanged in the Cr group, whereas HbA1c tended to increase in the placebo group (from 6.94% to 7.11%). Fasting serum insulin decreased in both groups, with a greater tendency in the Cr group (−16.5% vs −9.5%). These data suggest that supplementation of well-controlled type 2 diabetics with Cr-enriched yeast is safe and can result in improvements in blood glucose variables and oxidative stress.

Parameters of oxidative stress in children with Type 1 diabetes mellitus and their relatives

Oxidative stress (OS) plays an important role in the pathogenesis of Type 1 diabetes mellitus (DM). The aim of the study was to compare OS parameters in diabetic children and their first-degree relatives. Fifty diabetic children from the West Bohemian Region were examined as well as their 32 siblings (12 Boys and 20 girls) and 65 of their parents during a period of 6 months. Thirty healthy sex- and age-matched children studied before planned surgeries were normal controls for children, 40 healthy adult volunteers were controls for parents. OS parameters were evaluated in all participants of the study (superoxide dismutase, SOD; glutathione peroxidase, GSHPx; plasma antioxidant capacity, AOC; reduced glutathione, GSH; and malondialdehyde, MDA) and also Type 1 DM-associated antibodies (ICA and GADA). The results in diabetic children showed significantly lower GSHPx and AOC and increased MDA when compared with healthy children. Similar findings were found in their siblings but without statistical significance. It is consequently evident that decreased antioxidative protection and simultaneous free radical (FR) overproduction occur in diabetic children and that there is a similar, but not significant, tendency in their siblings. The findings warrant reducing OS in diabetic children and postponing disease onset in susceptible relatives.

Brain Natriuretic Peptide and N-Terminal proBNP in Chronic Haemodialysis Patients

Background: Brain natriuretic peptide (BNP) and N-terminal proBNP (NT-proBNP) are released into circulation as a result of congestive heart failure (HF). As HF and water overload are frequent complications in haemodialysis (HD) patients, we decided to study the levels of BNP and NT-proBNP and their changes during HD. Methods: BNP and NT-proBNP levels were determined in 94 HD patients before and after a regular 4-h HD. We followed changes in these peptides during HD depending on age, sex, HF (NYHA classification and left ventricular ejection fraction [LVEF]), duration on HD, presence of hypertension, coronary artery disease, type of membrane used for HD [low-flux (LFx) or high-flux (HFx)] and body mass change during HD. Furthermore, patients´ basic medication and creatinine levels and presence of diabetes mellitus were monitored. Results: Respectively,94% and 100% of the patients had pre-dialysis concentrations of BNP and NT-proBNP above the cut-off values for HF. The marker levels correlated significantly both before and after HD (r = 0.903 and 0.888, respectively, p < 0.001). BNP levels significantly decreased (p < 0.0001), whereas NT-proBNP significantly increased (p < 0.0001) during HD on LFx membranes. HD on HFx membranes caused greater decrease of BNP (compared to LFx membranes, p < 0.001), but also a decrease of NT-proBNP (p < 0.001).We did not find any significant differences in marker levels for HF and non-HF patients (NYHA classification). However, both peptides reached higher levels in the group with LVEF ≤50% (p < 0.001 for both peptides). Body mass change during HD negatively correlated only with the change of NT-proBNP (r = –0.27, p < 0.05). In the multiple regression model, the change of both peptides during HD was significantly influenced by membrane type (p = 0.003 for BNP and p = 0.001 for NT-proBNP). NT-proBNP change during HD was further significantly influenced by LVEF (p = 0.012), sex (p = 0.002) and duration on HD (p = 0.006). Conclusions: Both BNP and NT-proBNP levels were significantly increased in HD patients prior to dialysis. The change in concentrations of both peptides during HD is influenced by membrane type. HD probably triggers increased production of both peptides and this increase is emphasized by impaired LVEF. This fact can be clinically observed only on NT-proBNP levels, because BNP levels are biased by significant removal of this protein during HD.

Influence of Antioxidants on the Quality of Stored Blood

Background and objectives: Blood is exposed to oxidation stress and therefore has a high antioxidant capacity (AOC). With the many factors increasing the demands on the AOC, there may be damage to erythrocytes by free radicals. This study was to investigate evidence of erythrocyte damage in stored donor blood and to affect this by premedication of blood donors. Materials and methods: Blood samples of 15 healthy donors were collected in CPDA‐1 solution and analyzed immediately, and then again after 10 days of incubation at 4 °C and 1 day of incubation at 37 °C. Prior to incubation, the following parameters were evaluated: Na+, K+, malondialdehyde (MDA), hemoglobin (Hb), AOC in the supernatant, superoxide dismutase (SOD) in erythrocytes, and glutathione peroxidase (GSHPx) in whole blood. Blood donors of group 1 were not given any drugs or vitamins before blood sampling. The same blood donors were then supplemented with the following daily doses of antioxidants for 10 days before the next blood sampling: 36 mg of β‐carotene, 300 mg of vitamin E, 200 mg of vitamin C, and 40 mg of selenium. Results: The blood from donors of group 2 had a significantly smaller increase in MDA, K+, and Hb, and a smaller decrease in Na+ and AOC in the supernatant compared with that of group 1, while the activity of SOD and GSHPx did not change during blood storage. Conclusions: These results suggest that antioxidants given to blood donors can improve red cell storage parameters by reducing cell damage caused by free radicals.

Parameters of oxidative stress, DNA damage and DNA repair in type 1 and type 2 diabetes mellitus

Objectives: (i) to determine the extent of oxidative stress and DNA damage and repair using a panel of selected markers in patients with type 1 and type 2 diabetes mellitus (T1DM, T2DM), (ii) to find their possible relationships with diabetes compensation and duration, and finally (iii) to test for the effect of functional polymorphisms in the 8-oxoguanin DNA glycosylase (rs1052133), catalase (rs1001179) and superoxide dismutase (rs4880) genes on respective intermediate phenotypes. Methods: A total of 207 subjects (23 children and 44 adults with T1DM, 52 adult patients with T2DM and 88 healthy adult control subjects) were enrolled in the study. The following markers of redox state were determined in participants: erythrocyte superoxide dismutase (Ery-SOD), whole blood glutathione peroxidase (WB-GPx), erythrocyte glutathione (Ery-GSH), plasma total antioxidant capacity (P-tAOC) and plasma malondialdehyde (P-MDA). Furthermore, the extent of DNA damage and repair was ascertained using the following parameters: DNA single strand breaks (DNAssb), DNA repair capacity (DNArc) and DNA repair index (DNRI). Results: Comparison of T1DM vs. T2DM patients revealed significantly higher Ery-GSH content (P < 0.0001) and significantly lower Ery-SOD activity (P = 0.0006) and P-tAOC level (P < 0.0001) in T1DM subjects. T2DM diabetics exhibited a significant increase in DNAssb (P < 0.0001) and significant decrease in both DNArc (P < 0.0001) and DNRI (P < 0.0001) compared with T1DM patients. Patient’s age (irrespective of DM type) significantly correlated with DNAssb (r = 0.48, P < 0.0001), DNArc (r = −0.67, P < 0.0001) and DNRI (r = −0.7, P < 0.0001). Allele frequencies of all studied polymorphisms did not exhibit any significant association with the investigated parameters. Conclusion: We demonstrated significant age- and DM type-related changes of oxidative DNA modification and capacity for its repair in subjects with T1DM and T2DM.

Negative association between plasma levels of adiponectin and polychlorinated biphenyl 153 in obese women under non-energy-restrictive regime

The aim of this study was to reveal whether accumulation of the persistent organic pollutants (POPs), especially polychlorinated biphenyl (2,2′,4,4′,5,5′-hexachlorobiphenyl, PCB 153), affects plasma levels of adiponectin in obese patients. The study was designed as a longitudinal intervention trial with a control group, where 27 obese women (body mass index (BMI)>30 kg/m2; age 21–74 years) were studied before (OB) and after (OB-LCD) a 3-month low-calorie-diet intervention (LCD; 5 MJ daily). As the control group, 9 female volunteers without LCD intervention were used (C; BMI=19–25 kg/m2; age 21–64 years). Plasma levels of PCB 153 were measured by high-resolution gas chromatography with electron capture detection; total adiponectin and insulin plasma levels were quantified by immunoassays; and adiponectin multimeric complexes were quantified by immunoblotting. Plasma levels of total adiponectin, high and medium molecular weight multimers significantly negatively correlated with plasma levels of PCB 153 in OB, but not in C or in OB-LCD, whereas the LCD intervention lowered BMI by 3.3±3.0 kg/m2. Our results may suggest suppression of adiponectin by PCB 153 in obese women under non-energy-restrictive regime, which may contribute to the known association of PCB 153 and other POPs with type 2 diabetes.

Biosensor for lactate determination in biological fluids. I: Construction and properties of the biosensor

The preparation of a biosensor for lactate determination is described. The biosensor is based on an immobilized suspension of the aerobic yeast Hansenula anomala, containing flavocytochrome b2 in high activity. The conditions for yeast cultivation were optimized to gain a sufficiently high activity of this enzyme converting lactate in the cells. The properties of the biosensor are compared with those of a sensor based on immobilized enzyme flavocytochrome b2. The yeast lactate biosensor has a sufficient sensitivity and linearity and short time of response. The precision and accuracy of lactate determination as well as the results of comparisons using an enzyme electrode and the spectrophotometric UV-test, enables this biosensor to be used in routine work. Analysis can be performed in blood plasma or whole blood. The stability of the biosensor makes it possible to work for 4 weeks with one yeast cell pellet.

Increased proinflammatory cytokine production in adipose tissue of obese patients with chronic kidney disease

ZusammenfassungHINTERGRUND: Die Adipositas ist ein Hochrisikofaktor sowohl für die Entwicklung von Gefäßerkrankungen als auch für chronische Niereninsuffizienz (CKD). Ziel dieser Studie ist es, den Einfluss des Fettgewebes auf den Entzündungsstatus bei adipösen Patienten mit CDK zu erfassen. PATIENTEN UND METHODEN: In einer prospektiven Querschnitt-Studie analysierten wir 40 CDK (Stadium 3–4) Patienten mit milder Proteinurie (2,3–3,5 g/Tag): 20 Patienten mit Adipositas (Gruppe 1) und 20 normalgewichtigen Patienten (Gruppe 2) wurde während einer elektiven Abdominaloperation (laparoskopische Cholecystektomie) einmalig Blut abgenommen, sowie Proben des subkutanen und des viszeralen Fettgewebes entnommen. Die Serumkonzentrationen folgender Parameter wurden bestimmt: Asymmetrisches Dimethylarginin (ADMA), Adiponektin (ADPN), C-reaktives Protein (CRP), Interleukin-6-(IL-6), Tumor Nekrose Faktor-α (TNF-α), Pentosidin, Monocyte Chemoattractant Protein-1 (MCP-1). Mit Hilfe von Real-Time-PCR wurde die Expression der Messenger RNA (mRNA) von TNF-α, MCP-1 und der Adiponektin Rezeptoren 1 und 2 sowie des immunkompetenten Zellmarkers CD68 im subkutanen sowie im viszeralen Fettgewebe bestimmt. Das Fettgewebe wurde immunhistochemisch auf CD68 positive Zellen geprüft. Außerdem wurden in beiden Gruppen folgende weitere biochemische Parameter bestimmt: Insulin, HbA1c, Cholesterin, LDL-Cholesterin, und Triglyzeride. ERGEBNISSE: Die Serumkonzentrationen von ADMA, CRP, Pentosidin, Interleukin-6, TNF-α, and MCP-1 waren bei den adipösen CDK-Patienten signifikant höher. Das Adiponektin war signifikant im Vergleich zur Kontrollgruppe erniedrigt. Die subkutane und viszerale mRNA Expression von TNF-α, CD68, Adiponektin Rezeptor-1 and MCP-1 war bei den adipösen CDK Patienten signifikant erhöht. Die mRNA Expressionen waren im viszeralen Fettgewebe signifikant höher als im subkutanen Fettgewebe (p < 0,01 vs. p < 0,05). Die Expressionen der mRNA von Adiponektin, Interleukin-6, und des Adiponektin Rezeptors-2 beider Fettdepots waren nicht unterschiedlich in den beiden Gruppen. Bei den adipösen CDK-Patienten wurde im subkutanen und im viszeralen Fettgewebe eine erhöhte Infiltration mit CD68 positiven immunkompetenten Zellen gefunden. Die Fettstoffwechsel-Parameter waren in der Gruppe 1 gering, aber signifikant (p < 0,02) erhöht. Ausgeprägter waren die Veränderungen in den Triglyzeriden (p < 0,01). Ein ähnlicher Anstieg wurde bei den Insulin und HbA1c Werten der Gruppe 1 beobachtet (p < 0,02). SCHLUSSFOLGERUNGEN: Im Fettgewebe adipöser Patienten mit CKD im Stadium 3–4 wurde eine erhöhte Expression von proinflammatorischen Zytokinen und eine gesteigerte Infiltration mit immunkompetenten Zellen gefunden. Diese hinauf-regulierte Entzündung könnte zur Auslösung eines systemischen proinflammatorischen Zustands bei Patienten mit CDK beitragen und das Fortschreiten der Störung der Nierenfunktion beschleunigen.SummaryBACKGROUND: Obesity is a known high-risk factor for the development of vascular diseases and chronic kidney disease (CKD). In this study we aimed to elucidate the impact of adipose tissue on the inflammatory state in CDK patients with obesity. PATIENTS AND METHODS: A cohort of 40 patients with CKD (stages 3–4) with mild proteinuria (2.3–3.5 g/day) were analyzed in a prospective cross-sectional study: single blood samples and visceral and subcutaneous samples of adipose tissue were taken from 20 patients with obesity and 20 without obesity (control group) during elective abdominal surgery (laparoscopic cholecystectomy). Serum concentrations of asymmetric dimethylarginine (ADMA), adiponectin, C-reactive protein, interleukin-6, tumor necrosis factor-α, pentosidine and monocyte chemoattractant protein-1 were measured. Messenger RNA expression of tumor necrosis factor-α, monocyte chemoattractant protein-1, adiponectin receptors 1 and 2, and immunocompetent cell marker CD68 was measured in subcutaneous and visceral fat samples using real-time PCR. Adipose tissue was examined immunohistochemically for CD68-positive cells. Other biochemical parameters (insulin, glycated hemoglobin, cholesterol, LDL cholesterol, and triglycerides) were assessed in the two groups of patients at the same time. RESULTS: Serum concentrations of ADMA, C-reactive protein, pentosidine, interleukin-6, tumor necrosis factor-α and monocyte chemoattractant protein-1 were significantly higher in obese CKD patients than in the control group; adiponectin was lower in the obese group. Subcutaneous and visceral mRNA expressions of tumor necrosis factor-α, CD68, adiponectin receptor-1, and monocyte chemoattractant protein-1 were significantly increased in the obese patients, whereas expression of adiponectin, interleukin-6, and adiponectin receptor-2 did not significantly differ between the patient groups. In general, mRNA expressions were higher in visceral than in subcutaneous samples (P < 0.01 vs. P < 0.05). Increased infiltration of subcutaneous and visceral adipose tissue by CD68-positive immunocompetent cells was found in the obese CKD group. With respect to lipid metabolism parameters, a small but significant increase in levels was found in the obese patients (P < 0.02). Changes in triglycerides were more marked in this group (P < 0.01) and a similar increase was noted in insulin and HbA1c levels (P < 0.02). CONCLUSION: Increased expression of proinflammatory cytokines and increased infiltration by immunocompetent cells were found in adipose tissue of obese patients with CKD stages 3–4. This upregulated inflammation may contribute to the induction of a systemic proinflammatory state in patients with CKD and could accelerate the progression of renal dysfunction.

The EC4 European Syllabus for Post-Graduate Training in Clinical Chemistry and Laboratory Medicine: version 4 - 2012

Abstract Laboratory medicine’s practitioners across the European community include medical, scientific and pharmacy trained specialists whose contributions to health and healthcare is in the application of diagnostic tests for screening and early detection of disease, differential diagnosis, monitoring, management and treatment of patients, and their prognostic assessment. In submitting a revised common syllabus for post-graduate education and training across the 27 member states an expectation is set for harmonised, high quality, safe practice. In this regard an extended ‘Core knowledge, skills and competencies’ division embracing all laboratory medicine disciplines is described. For the first time the syllabus identifies the competencies required to meet clinical leadership demands for defining, directing and assuring the efficiency and effectiveness of laboratory services as well as expectations in translating knowledge and skills into ability to practice. In a ‘Specialist knowledge’ division, the expectations from the individual disciplines of Clinical Chemistry/Immunology, Haematology/Blood Transfusion, Microbiology/ Virology, Genetics and In Vitro Fertilisation are described. Beyond providing a common platform of knowledge, skills and competency, the syllabus supports the aims of the European Commission in providing safeguards to increasing professional mobility across European borders at a time when demand for highly qualified professionals is increasing and the labour force is declining. It continues to act as a guide for the formulation of national programmes supplemented by the needs of individual country priorities.

The effect of hemodialysis and acetate-free biofiltration on anemia.

The authors monitored, for a period of 12 months, anemia-, nutrition-, and free radical-related parameters and the rHuEPO dose required to maintain target hemoglobin (Hb) in 20 patients with chronic renal failure. Ten patients each were randomized for treatment by either acetate-free biofiltration (AFB) or low-flux hemodialysis (HD). At baseline, Hb levels were 102±2 (AFB) vs. 98±2 g/L (HD) (not significant difference, NS), the rHuEPO dose was 4050±976 vs. 5100±1538 IU/week (NS). Compared with baseline and with HD, lower rHuEPO doses were required during AFB at months 8, 9, 10 and 11, and 12 when they were 2100±510 (AFB) vs. 6000±1153 (HD), p=0.008. Prealbumin, transferrin and cholinesterase levels rose in the AFB group. Kt/V, albumin, transferrin saturation, aluminium, bicarbonate in serum, superoxide dismutase and glutathione peroxidase in erythrocytes, and malondialdehyde and antioxidant capacity in plasma did not differ between the AFB and HD groups. In terms of anemia control, AFB using an AN69 membrane was found to be more advantageous than low-flux HD, AFB improves some nutritional parameters. The compared methods do not differ in their effect on lipid peroxidation and the antioxidant system. (Int J Artif Organs 2000; 23: 173–80)