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Background and objectives Renin-angiotensin system (RAS) inhibitors reduce cardiovascular morbidity and mortality in patients with CKD. We evaluated the cardioprotective effects of the angiotensin-converting enzyme inhibitor ramipril in patients on maintenance hemodialysis. Design, setting, participants, & measurements In this phase 3, prospective, randomized, open-label, blinded end point, parallel, multicenter trial, we recruited patients on maintenance hemodialysis with hypertension and/or left ventricular hypertrophy from 28 Italian centers. Between July 2009 and February 2014, 140 participants were randomized to ramipril (1.25–10 mg/d) and 129 participants were allocated to non-RAS inhibition therapy, both titrated up to the maximally tolerated dose to achieve predefined target BP values. The primary efficacy end point was a composite of cardiovascular death, myocardial infarction, or stroke. Secondary end points included the single components of the primary end point, new-onset or recurrence of atrial fibrillation, hospitalizations for symptomatic fluid overload, thrombosis or stenosis of the arteriovenous fistula, and changes in cardiac mass index. All outcomes were evaluated up to 42 months after randomization. Results At comparable BP control, 23 participants on ramipril (16%) and 24 on non-RAS inhibitor therapy (19%) reached the primary composite end point (hazard ratio, 0.93; 95% confidence interval, 0.52 to 1.64; P=0.80). Ramipril reduced cardiac mass index at 1 year of follow-up (between-group difference in change from baseline: −16.3 g/m2; 95% confidence interval, −29.4 to −3.1), but did not significantly affect the other secondary outcomes. Hypotensive episodes were more frequent in participants allocated to ramipril than controls (41% versus 12%). Twenty participants on ramipril and nine controls developed cancer, including six gastrointestinal malignancies on ramipril (four were fatal), compared with none in controls. Conclusions Ramipril did not reduce the risk of major cardiovascular events in patients on maintenance hemodialysis. Clinical Trial registry name and registration number: ARCADIA, NCT00985322 and European Union Drug Regulating Authorities Clinical Trials Database number 2008–003529–17.

Aims: To evaluate whether angiotensin-converting enzyme (ACE) inhibitor and angiotensin II receptor blocker (ARB) combination therapy is more nephroprotective than ACE inhibitor or ARB monotherapy in people with type 2 diabetes and overt nephropathy. Materials and methods: In this prospective, randomized, open, blind-endpoint phase III trial sponsored by the Italian Drug Agency, 103 consenting patients with type 2 diabetes, aged >40 years, with serum creatinine levels 159 to 309 μmol/L, spot morning urinary albumin–creatinine ratio > 1000 mg/g (or > 500 mg/g in those on ACE inhibitor or ARB therapy at inclusion) were stratified by centre and randomized to 4.5-year treatment with valsartan 320 mg/d (n = 36), benazepril 20 mg/d (n = 34) or halved doses of both medications (n = 33). The primary endpoint was end-stage renal disease (ESRD). Modified intention-to-treat analyses were performed. Results: Recruitment took place between June 2007 and February 2013 at 10 centres in Italy and one in Slovenia. A total of 77 participants completed the study and 26 were prematurely withdrawn. During a median (interquartile range) of 41 (18–54) months, 12 participants on benazepril (35.3%) and nine on combination therapy (27.3%) progressed to ESRD, versus five on valsartan (13.9%). Differences between benazepril (hazard ratio [HR] 3.59, 95% confidence interval [CI] 1.25–10.30; P = 0.018) or combination therapy (HR 3.28, 95% CI 1.07–10.0; P = 0.038) and valsartan were significant, even after adjustment for age, gender and baseline serum creatinine, systolic blood pressure and 24-hour proteinuria (HR 5.16, 95% CI 1.50–17.75, P = 0.009 and HR 4.75, 95% CI 1.01–22.39, P = 0.049, respectively). Adverse events were distributed similarly among the groups. Conclusions: In people with type 2 diabetes with nephropathy, valsartan (320 mg/d) safely postponed ESRD more effectively than benazepril (20 mg/d) or than halved doses of both medications.

Whether convective therapies allow better control of serum phosphate (P) is still undefined, and no data are available concerning on-line haemofiltration (HF). The objectives of the study are to evaluate the effect of convective treatments (CTs) on P levels in comparison with low-flux haemodialysis (HD) and to evaluate the correlates of serum phosphate in a post hoc analysis of a randomized clinical trial.This analysis was performed in the database of a multicentre, open label and randomized controlled study in which 146 chronic HD patients from 27 Italian centres were randomly assigned to HD (70 patients) or CTs: on-line pre-dilution HF (36 patients) or on-line pre-dilution haemodiafiltration (40 patients).CTs did not affect P (P = 0.526), calcium (Ca) (P = 0.849) and parathyroid hormone levels (P = 0.622). P levels were associated with the use of phosphate binders including aluminium-based phosphate binders (P0.001) and sevelamer (P0.001), pre-dialysis bicarbonate levels (P0.001) and pre-dialysis blood K levels (P0.001). On multivariate analysis (generalized linear model), serum P was again largely unassociated with CTs (P = 0.631). Notably, participating centres were by far the strongest independent correlate of serum P, explaining 45.3% of the variance of serum P over the trial and this association was confirmed at multivariate analysis. Bicarbonate (P0.001) and, to a weaker extent, serum K (P = 0.032) were independently related to serum P.In comparison with low-flux HD, CTs did not significantly affect serum P levels. Participating centres were the main source of P variability during the trial followed by treatment with phosphate binders, serum bicarbonate and, to a weak extent, serum potassium levels (ClinicalTrials.gov Identifier: NCT011583309).

Background Predictors of haemoglobin (Hb) levels and resistance to erythropoiesis-stimulating agents (ESAs) in dialysis patients have not yet been clearly defined. Some mainly uncontrolled studies suggest that online haemodiafiltration (HDF) may have a beneficial effect on Hb, whereas no data are available concerning online haemofiltration (HF). The objectives of this study were to evaluate the effects of convective treatments (CTs) on Hb levels and ESA resistance in comparison with low-flux haemodialysis (HD) and to evaluate the predictors of these outcomes. Methods Primary multivariate analysis was made of a pre-specified secondary outcome of a multicentre, open-label, randomized controlled study in which 146 chronic HD patients from 27 Italian centres were randomly assigned to HD (70 patients) or CTs: online pre-dilution HF (36 patients) or online pre-dilution HDF (40 patients). Results CTs did not affect Hb levels (P = 0.596) or ESA resistance (P = 0.984). Hb correlated with polycystic kidney disease (P = 0.001), C-reactive protein (P = 0.025), ferritin (P = 0.018), ESA dose (P < 0.001) and total cholesterol (P = 0.021). The participating centres were the main source of Hb variability (partial eta2 0.313, P < 0.001). ESA resistance directly correlated with serum ferritin (P = 0.030) and beta2 microglobulin (P = 0.065); participating centres were again a major source of variance (partial eta2 0.367, P < 0.001). Transferrin saturation did not predict either outcome variables (P = 0.277 and P = 0.170). Conclusions In comparison with low-flux HD, CTs did not significantly improve Hb levels or ESA resistance. The main sources of variability were participating centres, ESA dose and the underlying disease.

Few data are available regarding the prevalence of burnout among dialysis health care workers. Aims of the present study were to assess and compare burnout levels in a sample of nurses and physicians working in dialysis units, and to investigate their relationships with quality of life, in a cross-sectional observational study.A total of 344 workers from 10 dialysis centres in Northern Italy completed a battery of questionnaires including the Maslach Burnout Inventory, the MOS-36 Item Short Form Health Survey [SF36: physical (PCS) and mental (MCS) component scores] and the 30-item General Health Questionnaire (GHQ30). Data on social and demographic characteristics and working conditions were also collected. General Estimating Equations models were used for the analysis.Overall, burnout scores were lower than the Italian normative sample, with no significant differences between physicians and nurses. However, 30% of nurses had high emotional exhaustion vs 18% of physicians (adjusted OR 2.38, P = 0.003). Emotional exhaustion was also predicted by number of worked hours and months worked in dialysis in the previous 2 years. Depersonalisation was predicted by male gender and bad relationship with coworkers. Having no children and having a permanent hospital position predicted low personal accomplishment. PCS was lower in nurses (50.0 vs 53.3, P0.001), while no significant difference was found for MCS and GHQ30. Lower PCS was associated with emotional exhaustion (P = 0.007) and GHQ305 with depersonalization (P = 0.032).Although burnout is not a general problem in dialysis health care providers, a subgroup of them may be identified, who would benefit from supportive measures to prevent this condition. Nurses appeared more burned-out in the emotional exhaustion scale than physicians.

Background: Previous studies reported significant increases in epoetin dosages when converting hemodialysis patients from subcutaneous (SC) to intravenous (IV) administration. More recent studies that corrected for iron deficiency found a much lower, if any, increase in epoetin dosage and/or decrease in hemoglobin (Hb) level after conversion from SC to IV epoetin administration. Therefore, the matter is still open for debate. Methods: This multicenter observational study evaluated stable hemodialysis patients without iron deficiency who had a stable SC epoetin dosage and Hb level of 10 g/dL or greater (≥100 g/L) at the time of study enrollment. Data for epoetin dosage, anemia, and inflammatory markers were collected retrospectively during the last 6 months of SC epoetin treatment and prospectively for 6 months after conversion to IV administration. The primary efficacy assessment was difference in Hb levels and epoetin dosages between patients administered epoetin SC and IV. Changes in values for iron stores, C-reactive protein, intact parathyroid hormone, and albumin were monitored as control parameters. Results: Data were analyzed for 262 hemodialysis patients from 6 Italian centers. Overall, mean Hb levels were similar with SC and IV epoetin administration (11.49 g/dL [114.9 g/L] and 11.44 g/dL [114.4 g/L]). Mean epoetin dosages also were similar with SC and IV administration (7,185 and 7,270 IU/wk). In patients requiring epoetin dosages of 12,000 IU/wk or greater at study entry, mean dosages tended to decrease after conversion to IV administration. There were no significant changes in control parameters. Conclusion: Conversion from SC to IV epoetin administration did not result in changes in Hb levels or epoetin dosage requirements in iron-replete hemodialysis patients.

Background. Routine water monitoring in a haemodialysis centre revealed high trichloroethylene (TCE) concentrations. The aim of this study is to describe the measures adopted after organic contamination of dialysis water in order to avoid the possibility of patient exposure. We also carried out in vitro experiments to evaluate the accumulation of TCE in various devices normally used in a dialysis water treatment system (DWTS). Methods. In vivo and in vitro blood and water TCE levels were determined by means of solid phase microextraction-gas chromatography/mass spectrometer. Results. High TCE concentrations were found throughout the DWTS; acceptably low levels were obtained only by replacing the activated charcoal, ionic-exchange resins, microfilters and PVC pipes. The adsorption and realising capacities of these devices were tested in vitro, and the elimination curves made it possible to calculate the total percentage of the previously absorbed TCE mass released into the water. Evidence of exposure was confirmed by the relatively high TCE levels in the patient blood samples taken 30 days after the last exposure even if the subjects were asymptomatic. In vivo experiments showed that the blood gain of TCE through the low flux membrane during the course of dialysis was about 77±10.4% of the amount carried by dialysis fluid as calculated on the basis of its partition coefficient value (Kb/w 3.75). Conclusions. This study shows that, when present in dialysis water, the lipophilic TCE contaminant can accumulate in various devices, thus transforming them into possible sources of exposure. This highlights the importance of periodically monitoring dialysis water for organic substances that have a great affinity to the blood compartment, in order to prevent occasional or chronic patient exposure.

Online hemodiafiltration appears to be the most effective technique of renal replacement therapy in many respects. Removal of small and high-molecular weight substances is enhanced. Modern technology ensures a safe, online production of reinfusion fluids. Nonetheless, stringent maintenance rules are required for the production of sterile and nonpyrogenic-dialysate solutions. In this review, we will critically review the state of the art of the clinical effects derived from the use of ultrapure dialysate and the online production of dialysate fluids in high-flux hemodiafiltration.

Regenerated cellulosic membranes are held as bioincompatible due to their high complement - and leukopenia - inducing properties. Adherence of polymorphonuclear neutrophils and monocyte purified from normal human blood to the three membranes were evaluated in an in vitro recirculation circuit in the presence or absence of fresh, autologous plasma after recirculation in an in vitro circuit using minimodules with each of the three membranes. In in vivo studies, 9 patients were treated with conventional haemodialysis for 2 weeks with each membrane and 1 week for wash-out using haemodialysers with the following surface: 1.95 m2for benzyl-cellulose, 1.8 m2for acetate-cellulose and low-flux polysulfone. Measurement of leukopenia, plasma C3a des Arg and elastase-α 1 proteinase inhibitor complex levels as well as urea, creatinine, phosphate and uric acid clearances was performed. Plasma-free neutrophils adhered maximally to acetate-cellulose (65% remaining in the circulation), while there was no significant difference between low-flux polysulfone and benzyl-cellulose (80% circulating neutrophils, at 15 min, p

Introduction Background. Proximal tubular epithelial cells express asurface C3-convertase activity which induces C fixation The development of tubular interstitial injury is aand insertion of the C5b-9 membrane attack complex common finding in the progression of several forms of(MAC) into the cell plasma membrane. The physio- glomerulonephritis. The importance of this phenom-pathological consequences of this phenomenon are enon is underlined by its correlation with renal func-unknown. tional impairment [1–3].Nevertheless, the mechanismsMethods. The effect of C fixation on the production of responsible for the spread of tissue damage from theinflammatory mediators by human proximal tubular glomerular to the tubular compartment are stillepithelial cells in culture was explored. unclear. The well-documented correlation between theResults. Proximal tubular epithelial cells incubated tubular interstitial damage and the severity of pro-with a sublytic amount of normal human serum as a teinuria suggests that proteins abnormally filteredsource of C, but not with heat-inactivated human through the glomerular capillary wall can potentiallyserum, showed a time-dependent calcium influx and a exert toxicity on tubular cells, thereby triggering aconcomitant release of 14C-arachidonic acid (14C-AA). tubular interstitial inflammatory process [1–3]. AmongEicosanoid synthesis following the arachidonic acid the plasma proteins escaping into the urine in nephroticmobilization was studied as prostaglandin E2release.syndrome, complement fractions are potential candid-Mg2+/EGTA, which did not prevent C activation byates for causing tubular injury. In fact, histologicalthe C3-convertase, and p-bromodiphenacyl bromide, astudies have shown that the alternative pathway ofphospholipase A2-inhibitor, inhibited mobilization ofcomplement can be directly activated by the brush14C-AA. These results suggest the activation of anborder of proximal tubules of normal rat [4] or humanextracellular Ca2+-dependent, phospholipase A2.kidney [5]. The presence of a potent C3-convertaseComplement fixation was associated with the synthesisactivity on the surface of proximal tubular cells wasof proinflammatory cytokines such as IL-6 and TNF-confirmed in a recent in vitro study on cultured humana. Experiments with C6-deficient sera indicated thatproximal tubular epithelial cells (PTEC), which dem-the release of 14C-AA and the production of cytokineswere dependent on the insertion of the terminal com- onstrated that the activation of the alternative pathwayponents of complement in the plasma membrane. of complement leads to fixation of the C5b-9 MACIndeed, the reconstitution of normal haemolytic activ- neoantigen on their surface with consequent C5b-ity of C6-deficient sera with purified C6 restored also 9-mediated cytotoxicity [6].the release of 14C-AA and the production of cytokines. The tubular interstitial lesions observed in progress-Conclusions. In vitro complement activation on the ive glomerulonephritis are characterized by the inter-proximal tubular cell surface triggers the generation of stitial infiltrate of mononuclear cells and accumulationproinflammatory mediators, which may potentially of extracellular matrix within the interstitial space.contribute to the pathogenesis of tubulointerstitial In non-immunological experimental disease charac-injury. terized by severe proteinuria, the deposition of comple-ment in the brush-border of proximal tubules isKey words: complement; cytokine; membrane attack accompanied by early monocyte/macrophage transmi-complex; proximal tubule; tubular injury gration from the circulation into the tubulointerstitialspace with subsequent mononuclear cell infiltrationand fibroblast proliferation and differentiation [7]Although several hypotheses were proposed, the

In the late 1960s, ultrafiltration was first used in clinical settings to increase, by convection, the clearance of toxic solutes in patients undergoing dialysis. Unfortunately, the efficiency of convection-based dialysis treatment, or hemofiltration (HF), was limited by the relatively low ultrafiltration coefficient of the dialyzers available at the time. Thus, the exchanged volume was low, and the corresponding clearance of low-molecular-weight solutes was insufficient with respect to the current target value of Kt/V urea. This was probably the cause of the failed improvement in the clinical and metabolic status of patients compared with standard dialysis treatment. In 1977, favorable results of the combination of diffusion and convection demonstrated the potential advantage of hemodiafiltration (HDF) over HF in terms of dialysis clearance. HDF was in fact the only means to obtain significant clearance of high-molecular-weight solutes while maintaining adequate urea clearance, whereas the increase in mean hematocrit in the erythropoietin era limited the exchanged volume in HF, in spite of the improved water permeability of the dialysis membranes. Mixed diffusive and convective clearance is less than the sum of the two parts because of reciprocal interference. Diffusive clearance mainly depends on the membrane permeability and the solute concentration gradient. New, highly permeable dialysis membranes can reach significant clearance of high-molecular-weight solutes such as Beta2 microglobulin (B2m) simply by diffusion, although in clinical settings there is also considerable ''hidden'' convection due to backfiltration. However, convection remains the best way to remove high-molecular-weight solutes, also for this kind of membrane. The ultrafiltration rate and the sieving coefficient account for the amount of convective clearance, as described in detail in the text. To define the treatment dose, the equation of Waniewsky allows the theoretical calculation of the urea clearance in HDF, both in postdilution and predilution mode. Unfortunately, no such equation is available for B2m. With a new mathematical model, well fitting with preliminary measured data although not fully validated, we calculated the relationship between urea and B2m clearance in predilution versus postdilution HDF, also considering the impact of variables such as blood and ultrafiltration flow. In particular, the predilution mode may decrease the urea clearance in comparison to hemodialysis with the same membrane and blood flow. This also applies to B2m clearance in predilution vs postdilution HDF, in spite of a marked increase in the ultrafiltration rate, at least in the more common clinical settings. In conclusion, good knowledge of the physics of solute transport is mandatory for appropriate prescription of HDF, in order to maximize both low- and high-molecular-weight solute clearance.

Alternative pathway activation of complement by cultured human proximal tubular epithelial cells. Human proximal tubular epithelial cells (PTEC) incubated with normal human serum (NHS) were found to fix on their surface C3, properdin, terminal complement components and C5b-9 MAC neoantigen, but not C1q and C4, by immunofluorescence. Complement fixation was abrogated if PTEC were incubated with EDTA-treated NHS or C3-deficient human serum, but not with Mg EGTA-treated NHS or C1q-deficient human serum, showing the prevalent activation of the alternative pathway of complement. This event was followed by marked cytoskeleton alterations with disruption of the actin cortical network, redistribution of actin throughout the cytoplasm and formation of blebs, and by cell cytolysis. In addition, superoxide anion and hydrogen peroxide production and chemiluminescence response were detected in consequence of MAC insertion on PTEC plasma membrane. The dependency on MAC of the observed biological effects of complement fixation on PTEC surface was shown by using sera selectively deficient of terminal components of complement (C6 or C8), and therefore unable to form the C5b-9 MAC, and by restoring the ability to form MAC after addition of purified C6 or C8. The possible pathogenetic relevance of these observations in tubulointerstitial injury occurring in patients with complementuria due to non-selective proteinuria, is discussed.

Lipopolysaccharide (LPS) from gram negative bacteria has been shown to prime human polymorphonuclear neutrophil (PMN) production of platelet activating factor (PAF). PAF is a lipid mediator of inflammation and endotoxic shock and is also involved in leukocyte activation occurring during hemodialysis; PAF induces leukopenia, degranulation of lysosomal granules, and adherence to hemodialysis membranes. Transmembrane passage of LPS has also been shown to occur. To evaluate the relevance of transmembrane passage of LPS on the priming of PMN derived production of PAF, we designed in vitro studies using an experimental circuit equipped with different membranes (Cuprophan, polysulfone, polymethylmethacrylate, polyamide) and recirculation of purified human PMNs. At different time intervals, PMNs were stimulated with FMLP (10 microM) after back-filtration of sterile and LPS contaminated dialysate. The results of these studies suggest that priming of PMN derived production of PAF was related to the percent of backfiltered LPS, and they emphasize the need for careful assessment of microbiologic quality to improve biocompatibility.

Symptomatic intradialytic hypotension is a common complication of hemodialysis (HD). The application of convective therapies to the outpatient setting may improve outcomes, including intradialytic hypotension. In this multicenter, open-label, randomized controlled study, we randomly assigned 146 long-term dialysis patients to HD (n = 70), online predilution hemofiltration (HF; n = 36), or online predilution hemodiafiltration (HDF; n = 40). The primary end point was the frequency of intradialytic symptomatic hypotension (ISH). Compared with the run-in period, the frequency of sessions with ISH during the evaluation period increased for HD (7.1 to 7.9%) and decreased for both HF (9.8 to 8.0%) and HDF (10.6 to 5.2%) (P < 0.001). Mean pre-dialysis systolic BP increased by 4.2 mmHg among those who were assigned to HDF compared with decreases of 0.6 and 1.8 mmHg among those who were assigned to HD and HF, respectively (P = 0.038). Multivariate logistic regression demonstrated significant risk reductions in ISH for both HF (odds ratio 0.69; 95% confidence interval 0.51 to 0.92) and HDF (odds ratio 0.46, 95% confidence interval 0.33 to 0.63). There was a trend toward higher dropout for those who were assigned to HF (P = 0.107). In conclusion, compared with conventional HD, convective therapies (HDF and HF) reduce ISH in long-term dialysis patients.

Exhaled breath condensate (EBC) is composed mainly by water and also contains non-volatile mediators, which are expired in small droplets of airway fluid. Urea has been proposed as a normalization factor for EBC non-volatile biomarkers. Aim of this study was to assess volatility and diffusivity of urea ex vivo and to measure its EBC concentrations in different clinical conditions. Volatility was assessed quantifying EBC concentrations collected at 4 different temperatures, whereas diffusivity was tested by measuring urea concentrations in both plasma and EBC from uraemic patients on intermittent haemodialysis. Urea was also measured in EBC from patients with chronic airway diseases, i.e., chronic obstructive pulmonary disease, asthma, and cystic fibrosis. The concentration of urea but not its absolute amount in EBC increased with condensation temperature. Haemodialysis influenced EBC and plasma urea concentrations in a similar way. The concentrations of urea in chronic airway diseases did not significantly differ from those of controls. Urea is a non-volatile molecule ex vivo and EBC urea depends on its concentrations in plasma. Urea concentrations in EBC are unaffected by three chronic airway diseases. We suggest that there is no need to normalize non-volatile biomarkers in EBC for urea concentrations to account for inter-individual variability. However, in repeated measurements within the same individual, the use of urea either as a normalizing factor or as covariate variable could be proposed to control intra-individual variability.

Chronic inflammation has been repeatedly reported in individuals undergoing hemodialysis. C-reactive protein (CRP) is considered a marker of chronic inflammation, as well as a mediator of the atherosclerotic process. Clinical and epidemiologic studies are based on plasma values obtained with the use of various automated methods. Our aim was to test 3 commercially available methods and compare the values obtained with the use of these tests in a population of individuals undergoing hemodialysis. We compared the following methods: immunoturbidimetry (AU2700 biochemistry analyzer; Olympus, Rungis, France) laser nephelometry (Behring Diagnostics, Marburg, Germany), and nephelometry (Beckman Instruments, Fullerton, Calif. The 3 methods were used in 3 different centers: Montpellier, France; and Pisa and Turin, Italy, respectively. We prepared samples for the estimation of imprecision values (ie, coefficient of variation [CV]) from the plasma of normal patients by adding purified C-reactive protein at concentrations ranging from 2.6 to 180 mg/L for intraassay variation and concentrations of 0, 1, 2, 3, 5, 10, 20, 50, 100, 150, and 180 mg/L for interassay variation. Intraassay imprecision was determined with the use of 10 replicate analyses on the same sample of the same day. We assessed interassay imprecision using the same sample, divided into aliquots and measured on 5 consecutive days. Agreement between methods was assessed on predialysis serum samples collected from patients with stable chronic kidney disease who were undergoing long-term hemodialysis at the 3 different centers (Montpellier,192; Pisa, 56; Turin,98). Serum was separated from the red cells and stored in 3 aliquots at -70 degrees C until it could be analyzed. Samples were thawed only once, circulated among the 3 centers, and each tested with all 3 of the methods. The Beckman method yielded the most precise results, with intraassay CVs ranging from 1 to 2 and interassay CVs ranging from 1 to 4. The Behring method was the least precise, with intraassay and interassay CVs ranging from 12 to 15 and 7 to 16, respectively. The results of the Olympus method fell between those of the other 2 methods. Agreement between the results of the Olympus and Behring methods was satisfactorily. The Beckman and Olympus methods yielded, on average, similar results over the entire range of CRP values. We detected significant disagreement between the Beckman method and the other 2 methods, obtaining results 10 to 100 times lower with the Beckman method. This became evident in terms of kappa-statistics. Our findings emphasize the need for careful assessment of the methods used to detect CRP in serum samples. Failure to do so may ultimately have a negative impact on the real relevance of CRP as a marker and on the role of chronic implication particularly in epidemiologic studies.

Renal proximal tubular epithelial cells (PTEC) are target for LPS during sepsis and renal infections. In the present study, we evaluated whether stimulation of human PTEC by LPS is modulated through the soluble or the membrane form of the LPS receptor CD14. We found that PTEC lacked expression of the membrane form of CD14 and did not release soluble CD14 (sCD14). sCD14 was detected in the urine of normal subjects and it was increased in patients with renal sepsis or with proteinuria. In the presence of sCD14 and LPS binding protein (LBP), PTEC were 10 to 100-fold more sensitive to LPS activation, resulting in cytokine production (IL-6, IL-8 and TNF-alpha) and NO release. We found that sCD14 purified from urine was biologically active on PTEC. Moreover, the presence of sCD14 and LBP was required for cytotoxicity induced by low concentrations of LPS (1-10 ng/ml) in PTEC. Cell death showed the characteristics of both necrosis and apoptosis, as demonstrated by LDH release and by TUNEL and acridine orange staining and caspase-3 activation. Whereas the LPS alone was sufficient to induce necrosis, sCD14 and LBP were required for apoptosis. Our results suggest that sCD14 excreted in urine may participate with endotoxin in the activation and injury of renal proximal tubules. In particular, sCD14 may contribute to the tubulo-interstitial injury in clinical settings characterised by proteinuria and enhanced susceptibility to infections such as in diabetes.

Background Based on the increased hydraulic permeability of the new high permeability polyethersulfone membrane, DIAPES®HF-800, we investigated the kinetics and handling of albumin in high volume on-line hemodiafiltration (HDF). Methods Seven patients on predilutional HDF were studied in two consecutive sessions. Blood flow rate and transmembrane pressure were continuously monitored. Spent dialysate was spilled at 20 ml/h every hour. Albumin was measured in blood and dialysate by immunonephelometry. Albumin and proteins adsorbed onto the dialyzer membrane were eluted after treatment with Triton X. Ultrafiltrates collected at 1 and 2 hours of treatment were pooled from different patients and incubated for 24 hours at 37°C with bovine serum albumin (BSA). Total sulphydryl groups were evaluated using Ellmann's reagent [5, 5'-dithio-bis(2-nitrobenzoic acid)]. Results In all 7 patients, the total loss of albumin was 3.99±1.81 g, ranging between 1.09 and 6.82 g/session. Most albumin loss occurred in the first 60 min of pre-dilutional hemodiafiltration (1.92+0.83 g). There was no correlation between transmembrane pressure, urea clearance and the loss of albumin. Plasma water urea clearance values were stable over the treatment (234±14.3 mI/min). Plasma albumin concentration did not decrease during HDF sessions. Albumin adsorbed onto the dialyzers was 0.7±1.6 mg but the total amount of adsorbed proteins was much higher (130+90 mg). In addition, the ultrafiltrate collected during HDF sessions was able to induce oxidation of bovine serum albumin as measured by total protein sulfhydryl groups: bovine serum albumin incubated in the presence of ultrafiltrate collected at 1 hour had a sulfhydryl loss of 56.3±5.7% (pConclusion The present study shows the high inter- and intra-patient variability of transmembrane passage of albumin in chronically uremic patients undergoing pre-dilutional HDF. Factors involved do not seem to be correlated to transmembrane pressure but rather to an interaction with the polymer surface. Albumin adsorption was minimal and was significantly lower than that of other plasma proteins. Albumin loss during HDF seemed to have no acute impact on plasma albumin. In addition, we demonstrated the presence of prooxidative compounds able to oxidize albumin, of which extracorporeal removal by HDF procedure could be beneficial for HD patients.

Preliminary studies indicate the involvement of interleukin (IL)-12 in experimental renal pathology. In the present study, we evaluated whether cultured glomerular mesangial cells are able to produce IL-12 and whether IL-12 may regulate some of their functions, including the cytoskeletal reorganization, the change in cell shape, and the production of platelet-activating factor (PAF). The results obtained indicate that pro-inflammatory stimuli, such as tumor necrosis factor-alpha and bacterial polysaccharides, induce the expression of IL-12 mRNA and the synthesis of the protein by cultured mesangial cells. Moreover, cultured mesangial cells were shown to bind IL-12 and to express the human low-affinity IL-12 beta1-chain receptor. When challenged with IL-12, mesangial cells produced PAF in a dose- and time-dependent manner and superoxide anions. No production of tumor necrosis factor-alpha and IL-8 was observed. Moreover, we demonstrate that IL-12 induced a delayed and sustained shape change of mesangial cells that reached its maximum between 90 and 120 minutes of incubation. The changes in cell shape occurred concomitantly with cytoskeletal rearrangements and may be consistent with cell contraction. As IL-12-dependent shape change of mesangial cells was concomitant with the synthesis of PAF, which is known to promote mesangial cell contraction, we investigated the role of PAF using two chemically different PAF receptor antagonists. Both antagonists inhibited almost completely the cell shape change induced by IL-12, whereas they were ineffective on angiotensin-II-induced cell shape change. In conclusion, our results suggest that mesangial cells can either produce IL-12 or be stimulated by this cytokine to synthesize PAF and to undergo shape changes compatible with cell contraction.

Today’s patient population is increasingly older. Patients with chronic renal failure therefore start extracorporeal substitutive treatment having congestive heart failure, chronic liver disease, diabetes and so forth. In these patients, however, long-term haemodialytic treatment may add further aggravation on their pre-existing pathological conditions. Oxidative stress and alterations in lipid metabolism are caused by haemodialysis mainly due to (1) bioincompatibility type of reactions such as production of reactive oxygen species by inflammatory cells due to complement-mediated or -independent pathways, and (2) the imbalance between oxidants and antioxidants due to the diffusive loss of hydrophilic vitamins such as ascorbic acid. The events related to the oxidant stress may sustain a state of chronic inflammation. Recent advances suggest that atherosclerosis and proliferation of the smooth muscle are initiated and sustained by inflammatory mechanisms. Therefore, attempts to counterbalance the prooxidant effect of haemodialysis and to reduce the chronic inflammatory state will be presented.

Extracorporeal treatments for acute or chronic replacement of organ function still represent a challenge for today’s technology. Activation of fluid phase (complement, coagulation, fibrinolysis) and cellular systems (leukocytes, platelets) is known to occur in hemodialysis (1). Among other biochemical indications of leukocyte activation as a conseguence of blood-surface interactions such as oxygen radicals, release of granulocyte proteinases, monocyte prostaglandin release and cytokine generation, platelet-activating factor (PAF) has drawn interest in various biocompatibility studies (reviewed in 1–5). PAF is a phospholipid mediator of inflammation with different biologic properties relevant for the development of inflammation and septic shock (7–11). PAF may act at concentrations of 10−12M and requires an ether linkage at the sn-1 position of the glycerol backbone, a short acyl chain, usually an acetyl residue, at the sn-2 position, and the polar head group of choline or ethanolamine at the sn-3 position (7–8). However, it has been shown recently that PAF belongs to a family of structurally related phospholipid molecules of biologic origin that shares many physiologic activities (12). PAF is considered a mediator of cell-to-cell communication that may function both as an intracellular and intracellular messenger. PAF is produced after immunologic or nor immunologic challenge by a variety of cells such as monocytes/macrophages, polymorphonuclear neutrophils, basophils and platelets, that may participate in the development of inflammatory reaction (13). In addition, human endothelial cells were found to produce PAF after stimulation by several inflammatory mediators including thrombin, angiotensin II, vasopressin, leukotriene C4 and D4, histamine, bradykinin, elastase, catheprin G, and plastic (14–18). PAF acts in an autocrine and paracrine way through a specific receptor for which a cDNA has been cloned (19,20). The receptor belongs to the family of “serpentine” receptors containing sever a-helical domains that weave in and out plasma membrane and it interacts with a G protein, which activates a phosphatidylinositol-specific phospholipase C (19,20). PAF receptors exist in all cells that are known targets for PAF. Recently, also human cultured endothelial cells have been shown to express PAF receptors (21). PAF promotes the permeability of the EC monolayer leading to cell retraction and formation of intracellular gaps (22). PAF induces in vitro migration of endothelial cells and promotes in vivo angiogenesis by a heparin-dependent mechanism (21). The aim of the present review is to briefly summarize early evidence for a role of PAF in bioincompatibility events and to highlight recent advances and their possible potential practical application in testing polymer biocompatibility.

Ten patients of a population of 319 on chronic intermittent treatment with Cuprophan dialyzers have undergone surgery for carpal tunnel syndrome (CTS). Since 1971 the dialysis schedule for all patients has been 4 m2/h three times per week. All patients were anuric. The operated patients represent 16% of the total population. In comparison with a dialysis population in other units described in the literature (12), 8 m2/h performed three times per week, with similar dialysis aging and a CTS incidence of 47%, it appears that the duration of the dialysis session per se increases the frequency of CTS and leads to the problem of the consequences of blood membrane interaction.

Studies of drug interactions during Cyclosporin A (CyA) therapy are based mostly on spontaneous reports. Between 1985 and 1987 35 significant clinical manifestations arising from pharmacological interaction were published by three major medical journals. This work is a summary of clinical complications after combined use of CyA and several other drugs.

Postdilution haemofiltration is a treatment indicated for patients who are unable to tolerate standard dialysis. However, its duration is uncomfortably prolonged in cases of low blood flow from the vascular access. In a predilution mode greater clearance rates can be reached even if a greater amount of sterile and pyrogen-free solutions are required. Although an in-line solution-producing system lowers the cost, theoretical calculations indicate that a predilution mode is not advantageous, in terms of the amount of infused solutions, if urea clearances greater than 150 ml/min at 300 ml/min blood flow are required. Theoretically, the combination of predilution and postdilution is the best system to utilise relatively small amounts of sterile solution in order to enhance treatment efficiency at low blood flow. However, the predilution mode may ultimately be preferable because of easier utilisation of the hardware. The results of clinical application of pre-postdilution haemofiltration indicate that at a 400 ml/min blood flow, a urea clearance of more than 220 ml/min can be obtained, and the duration of treatment can be reduced to only 3 h in the majority of patients.

In the present study we evaluated spontaneous and stimulated adherence of human monocytes to regenerated cellulose and polyacrylonitrile (AN69) membranes. Spontaneous adherence at 60 min was significantly higher for regenerated cellulose (28 +/- 2%, P < 0.001) than for AN69 (11 +/- 2) membranes. Stimuli such as bacterial lipopolysaccharide, TNF alpha, interleukin-1 and -6 as well as platelet-activating factor, but not IL-4, significantly enhanced adherence at 60 min to AN69 (28 to 30%). In contrast, adherence was not further inducible in the presence of regenerated cellulose. Both spontaneous and cytokine/bacterial lipopolysaccharide-stimulated adherence were significantly reduced by SDZ-63072, a specific platelet-activating factor receptor antagonist. This difference in sensitivity of monocyte adherence reflects probably the intrinsic ability of regenerated cellulose to provide maximal spontaneous monocyte adhesion. These data suggest that PAF may act as an adherence mediator. This is in line with the ability of regenerated cellulose to directly stimulate monocytes to synthesize platelet-activating factor and with the ability of cytokines and bacterial lipopolysaccharide to stimulate its synthesis. Although AN69 has a low adherence potential, bacterial lipopolysaccharide or cytokines may blunt the biocompatibility of this membrane.