František Šefrna

Cell-Free Plasma DNA during Peritoneal Dialysis and Hemodialysis and in Patients with Chronic Kidney Disease

The mechanisms of clearance of circulating plasma DNA are not fully understood, and so we aimed to examine it in patients with impaired renal function compared with healthy individuals. We also assessed the effect of peritoneal dialysis and hemodialysis on circulating plasma cell‐free DNA (cfDNA) in our treated patients. Overall, 20 healthy volunteers, 20 patients with chronic kidney disease (CKD), 18 patients undergoing peritoneal dialysis (PD), and 17 patients on hemodialysis (HD; high‐flux polysulfone membrane) were examined. Cell‐free DNA levels were determined using real‐time GADPH gene sequence amplification. The levels of cfDNA in all groups of our patients did not differ significantly from those of healthy volunteers. In HD patients, cfDNA levels were significantly increased compared with those of CKD patients (P < 0.05) and PD‐treated patients (P < 0.01). In PD‐treated patients, cfDNA was detectable in overnight effluent, with its levels correlating inversely with the duration of PD treatment (r=−0.619, Spearmans coefficient, P= 0.008). Factors contributing to these differences may include changes in the quality and quantity of the cell population of the peritoneum, highlighting the need for additional studies clarifying the dynamics of cfDNA during PD. The plasma levels of cfDNA do not seem to be dramatically altered even in CKD patients or those on PD or HD (as long as they are measured prior to the procedure in the latter two). Our data suggest renal elimination is not the main mechanism of circulating cfDNA clearance.

A clinical study to assess the effect of heparin in dialyzer rinsing solutions.

The solution usually recommended for rinsing the blood side, which is an indispensable step in preparing a dialyzer for hemodialysis (HD), contains saline and heparin. The heparin used for rinsing is said to reduce the thrombogenic properties of the dialysis membrane and, hence, also the need for systemic heparinization during the whole procedure. The aim of our study was to establish whether this postulate also applies to polysulphone steam-sterilized dialyzers. To do so, 16 patients on long-term dialysis were randomized into two groups of eight. One group was subsequently treated with polysulphone low-flux dialyzers (F6HPS), the other with polysulphone high-flux dialyzers (F60S). Both groups were examined, in a crossover manner during HD using a dialyzer previously rinsed with 1000 ml of saline plus 2,000 IU of heparin, and during HD using a dialyzer previously rinsed with 500 ml of saline without heparin. Except for the rinsing, HD conditions were completely identical. Blood obtained before HD, and at 15, 60 and 240 min of HD at the dialyzer inlet, was used to determine the activated partial thromboplastin time (to test heparinization control), the thrombin-antithrombin III complex (ELISA, to evaluate coagulation system activation), platelet factor 4 (ELISA, a substance with antiheparin activity), and platelet count. None of the above parameters showed, at any of the collecting intervals, a statistically significant difference between HD with and without heparin with a reduced volume of rinsing solution, or between HD using low- and high-flux dialyzers. It is concluded that heparin used to rinse polysulphone dialyzers before HD has no effect on blood coagulation or on the need for heparin during the procedure.

Fibrinolysis Defect in Long-Term Hemodialysis Patients with Type 2 Diabetes mellitus and Its Relation to Metabolic Disorders

Background/Aims: Patients with chronic renal failure (CRF) secondary to diabetes mellitus show a high incidence of atherosclerosis with its thrombotic complications. Both CRF and type 2 diabetes mellitus (DM2) results in fibrinolysis defects causally related to atherogenesis and thrombogenesis. It is not well known whether or not and, if so, how fibrinolysis is altered in patients with both CRF and DM2. Our study was designed (1) to identify the fibrinolysis defect present in patients with DM2-mediated CRF and treated by long-term hemodialysis (DM2HD), and (2) to establish whether the fibrinolysis defect is related to the metabolic abnormalities observed in CRF or DM2. Methods: Sixteen DM2HD patients and 23 healthy individuals (HI) had their euglobulin clot lysis time (ECLT), and tissue-type plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1) activities (act) and concentrations (ag) assessed before and after standard fibrinolytic stimulus (i.v. administration of 0.4 µg/kg BW 1-deamino-8-D-arginine vasopressin, DDAVP) along with metabolic status markers. Results: DDAVP caused a significant shortening of ECLT, rises in tPA act and ag, and a significant decrease in PAI-1 act. PAI-1 ag declined significantly in HI, but not in DM2HD. A comparison of responses to DDAVP revealed the groups differed significantly in the change in PAI-1 ag. Whereas, in HI, PAI-1 ag decreased by 11.8 ng/ml, no decrease was seen in DM2HD (0.0 ng/ml) (p < 0.0001; medians given; unpaired Wilcoxon’s test). Stepwise regression analysis showed the change in PAI-1 ag was highly group-specific (DM2HD vs. HI, regression coefficient 21.22; partial correlation 0.58; p < 0.0001) and, also dependent on the serum concentrations of apolipoprotein A-I (–32.41; –0.46; p < 0.01) and homocysteine (0.35; 0.36; p < 0.05). Conclusions: Patients with type 2 DM and CRF on long-term hemodialysis have a fibrinolysis defect manifesting itself after standard fibrinolytic stimulus by an insufficient decrease in PAI-1 concentrations. The defect is related to decreased serum levels of apolipoprotein A-I and increased serum levels of homocysteine. The defect might be a factor contributing to accelerated atherosclerosis and thrombotic complications in these patients.

Relationship between Anemia and Adequacy of Continuous Ambulatory Peritoneal Dialysis

Sylvie Opatrná, MD, 1st Department of Internal Medicine, Charles University, Alej Svobody 80, CS-304 60 Plzen (Czech Republic), Tel. +42 19 522564, fax +42 19 531110 Dear Sir, It is generally recognized that anemia is a common symptom of chronic renal failure which causes a number of serious problems to the patient. Some studies claim that, in hemodialysis patients, success in treating anemia depends on the efficacy of dialysis therapy. The importance of adequate hemodialysis for successful control of anemia has recently received authoritative support especially by a study conducted by Ifudu et al. [1]· Regarding the relationship between anemia and efficacy of continuous ambulatory peritoneal dialysis (CAPD), it has been explored to a much lesser extent than was the case for hemodialysis, and, what is more, the results are controversial. While some studies have not demonstrated an association, other authors have, even though in case reports [2-4]. Because of the paucity of unambiguous information, we conducted a study designed to establish whether or not the renal anemia in patients with chronic renal failure is affected by the adequacy of CAPD. We examined 22 patients with a mean age of 51.8 (20-79) years (arithmetic mean and range) treated by CAPD (Twin-Bag System; Baxter, Deerfield, Ill., USA) for 14.8 (1.5-52) months for chronic renal failure caused by chronic tubulointerstitial nephritis in 8 cases, chronic glomerulonephritis in 7, diabetic nephropathy in 5, polycystic kidney disease in 1 ‚ and Fanconi’s syndrome in 1 case. None of the patients received recom-binant human erythropoietin or blood transfusion for renal anemia. At the same time the hematocrit was 30.9% (19.4-43.3). The KT/ V index had not changed for 6.3 (1.5-24) months prior to the study when it was at 2.0 (1.4-2.8) per week. The residual glomerular filtration rate (GFR) was 4.8 (0-9.4) ml/ min. The efficacy of blood purification was assessed by the KT/V index considering both peritoneal and renal urea clearances [KT/ Vurea = (Durea/Purea·VD + Curea)/Vtot] and by weekly creatinine clearance corrected for body surface area (BSA), again considering both peritoneal and renal eliminations (Ccrea/BSA = Dcrea/Pcrea·VD·7 + GFRest). The residual GFR was determined as the arithmetic mean of urea and creatinine clearances [GFRest = (Curea + Ccrea)/2] [5]. Besides the hematocrit, we examined, using standard methods,

Impact of Cardiovascular Risk Factors on Morbidity and Mortality in Czech Middle-Aged Men: Pilsen Longitudinal Study

The impact of biological and life-style characteristics measured during baseline examination on 12-year morbidity and mortality of coronary heart disease (CHD), stroke (STR), and malignancies was investigated in an urban population of 3,540 middle-aged men initially free of clinical disease. The following factors enhanced significantly (at the 5% level) the adjusted relative risk ratios: for total mortality age, smoking, and elevated systolic blood pressure; for CHD age, smoking, elevated systolic blood pressure, serum cholesterol levels, and body mass index, and family history (father or mother). Myocardial infarction was positively associated with age, smoking and elevated serum cholesterol levels. For STR age and elevation of both systolic and diastolic blood pressure were risk factors. The relative risk for all malignancies was enhanced by age and smoking. Regular alcohol consumption was associated with a significantly lower risk for all CHD; however, with only marginal significance for myocardial infarction. Higher education was associated with a significantly lower risk of total mortality, all CHD, and myocardial infarction and a marginally lower risk of STR. A high leisure physical activity was negatively (but not significantly) associated with the risk of all end points.

Tissue Factor, Its Pathway Inhibitor, and Metabolic Disturbances in Long-Term Peritoneal Dialysis

Background/Aim: The tissue factor (TF) plays a key role in triggering the coagulation system in vivo. Our study was designed to determine whether or not the plasma levels of TF and its pathway inhibitor (TF pathway inhibitor; TFPI) in patients with chronic renal failure (CRF) treated by peritoneal dialysis (PD) (1) are pathologically altered; (2) differ between diabetics and nondiabetics, and (3) depend on the metabolic disorders associated with CRF and/or diabetes. Methods: Using ELISA, plasma TF and TFPI levels were measured once in 21 PD patients (10 with diabetes, 11 without diabetes) and in 21 healthy subjects. Results: As compared with healthy subjects (TF 282 pg/ml; TFPI 73 ng/ml), both TF and TFPI levels were significantly higher in PD patients with diabetes (485 pg/ml, p < 0.001, and 106 ng/ml, p < 0.01, respectively) and in PD patients without diabetes (480 pg/ml, p < 0,001, and 121 ng/ml, p < 0.001, respectively). The difference between diabetics and nondiabetics was not significant. In stepwise regression analysis, the TF levels depended on serum creatinine (partial correlation 0.39, p < 0.05), glycemia (0.43, p < 0.01), and insulin (–0.43, p < 0.05), and the TFPI levels depended on creatinine (partial correlation 0.67, p < 0.001), apolipoprotein B (0.46, p < 0.01), and plasma fibrinogen (0.43, p < 0.01). Conclusions: CRF patients on PD show increased plasma TF and TFPI levels. There is no difference between diabetics and nondiabetics. The TF and TFPI levels depend significantly on the renal function, as assessed by serum creatinine, and on some metabolic disorders. Elevated TF and TFPI levels may be related to thrombosis and atherosclerosis in CRF patients on PD.

Cell-Free Plasma DNA during Hemodialysis

Purpose. Cell-free plasma DNA (cfDNA) levels originating predominantly from apoptotic leukocytes were found to rise during hemodialysis (HD) session, and as such are considered a marker of HD membrane biocompatibility. The purpose of our study was to determine the changes of cfDNA during two consecutive high-flux polysulphone HD sessions after a long (HD-L) and short (HD-S) interdialytic interval. Methods. A total of 17 HD patients were examined. Prior to HD and at 30 and 240 min, cfDNA (using real-time PCR) and leukocyte count were determined. Results. No significant difference was found when comparing pre-HD-S with pre-HD-L cfDNA [4893 (1090–28804) vs. 4589 (691–73796) genomic equivalent/mL]. A significant increase in cfDNA at 240 min was seen in HD-L (p < 0.05) but not in HD-S. Leukocyte count correlated with cfDNA levels before HD-S (r = 0.8; p < 0.01); however, no other correlation was seen between routinely measured biochemical markers and pre-HD cfDNA levels or cfDNA changes during HD. The increase in plasma cfDNA during HD did not correlate with dialysis duration, its efficacy, or ultrafiltration. An association between magnitude of diuresis and cfDNA levels in HD-S was found (r = 0.58; p < 0.05). Conclusions. The behavior of cfDNA during HD after long and short interdialytic interval is inconsistent and cannot be explained by changes in laboratory and clinical parameters. The observed relationship of plasma cfDNA levels with diuresis deserves further investigation.

Acid-base balance in peritoneal dialysis patients: a Stewart-Fencl analysis.

Background. Evaluation of acid-base disorders using the Stewart-Fencl principle is based on assessment of independent factors: strong ion difference (SID) and the total concentration of non-volatile weak acids (Atot). This approach allows for a more detailed evaluation of the cause of acid-base imbalance than the conventional bicarbonate-centered approach based on the Henderson-Hasselbalch principle, which is a necessary yet insufficient condition to describe the state of the system. The aim of our study was to assess acid-base disorders in peritoneal dialysis (PD) patients using both of these principles. Methods. A total of 17 patients with chronic renal failure (10 men), aged 60.7 (22–84) years, treated by PD for 25.7 (1–147) months were examined. A control group included 17 healthy volunteers (HV) (8 males), with a mean age of 42.7 (22–77) years and normal renal function. Patients were treated with a solution containing bicarbonate (25 mmol/L) and lactate (15 mmol/L) as buffers; eleven of them used, during the nighttime dwell, a solution with icodextrin buffered by lactate at a concentration of 40 mmol/L. The following equations were employed for calculations of acid-base parameters according to the Stewart-Fencl principle. The first is SID = [Na+] + [K+] + 2[Ca2+] + 2[Mg2+] − [Cl−] − [UA−], where SID is the strong ion difference and [UA−] is the concentration of undetermined anions. For practical calculation of SID, the second equation, SID = [HCO3−] + [Alb−] + [Pi−], was used, where [Alb−] and [Pi−] are the charges carried by albumin and phosphates. The third is Atot, the total concentration of weak non-volatile acids, albumin [Alb] and phosphates [Pi]. Results. The capillary blood pH in PD group was 7.41 (7.27–7.48), [HCO3−] levels 23.7 (17.6–29.5) mmol/L, SID 36.3 (29.5–41.3) mmol/L, sodium-chloride difference 39.0 (31.0–44.0) mmol/L, [Pi] 1.60 (0.83–2.54) mmol/L, and [Alb] 39.7 (28.8–43.4) g/L (median, min-max). Bicarbonate in blood correlated positively with SID (Rho = 0.823; p < 0.001), with the sodium-chloride difference (Rho = 0.649; p < 0.01) and pH (Rho = 0.754; p < 0.001), and negatively with residual renal function (Rho = −0.517; p < 0.05). Moreover, the sodium-chloride difference was also found to correlate with SID (Rho = 0.653; p < 0.01). While the groups of PD and HV patients did not differ in median bicarbonate levels, significantly lower median value of SID were observed in PD patients, 36.3 vs. 39.3 mmol/L (p < 0.01); additionally, PD patients were shown to have significantly lower mean value of serum sodium levels, 138 vs. 141 mmol/L (p < 0.01), and serum chlorides levels, 100 vs. 104 mmol/L (p < 0.001). Despite the higher [UA−] levels in PD patients, 9.1 vs. 5.4 mmol/L (p < 0.001), this parameter was not found to correlate with bicarbonate levels. Conclusions. The results suggest that the decreased bicarbonate in PD patients results from a combination of decreased sodium-chloride difference and mildly increased unmeasured anions.