Serum-soluble Fas and serum levels of erythropoietin in chronic kidney disease.

BACKGROUND: Soluble Fas levels (sFas) are increased in the serum of uremic patients and are associated with the presence of anemia and recombinant human EPO (rHuEPO) dosage in dialysis patients. It is possible that sFas levels are associated with an increased need for serum erythropoietin levels (Epo) in chronic kidney disease and dialysis patients in order to maintain hematocrit (Hct) levels. AIMS: To investigate the relationship between serum sFas levels, serum Epo levels and the ratio between Epo levels and Hct in uremic patients. METHODS: We studied 52 predialysis chronic kidney disease patients (CKD; 33 M, 57 +/- 12 years, hematocrit (Hct) = 37 +/- 7%), 29 peritoneal dialysis patients (PD; 12 M, 54 +/- 14 years, Hct = 36 +/- 7%), 29 hemodialysis patients (HD; 19 M, 47 +/- 14 years, Hct = 33 +/- 5%) and 29 healthy volunteers (control group 17 M, 50 +/- 16 years, Hct = 43 +/- 3%). We examined the relationship between Hct and serum levels of Epo, sFas, C-reactive protein, IL-6 and iron status. The ratio of serum Epo divided by Hct (Epo/Hct) was used as an indicator of Epo responsiveness. RESULTS: Compared to normal subjects, the CKD, PD and HD groups presented lower Hct levels and higher serum levels of sFas, Epo, Epo/Hct and IL-6. Serum levels of sFas correlated negatively with albumin (r = -0.24, p = 0.02), IL-6 (r = -0.18, p = 0.04) and Epo/Hct (r = -0.37, p < 0.001). In multivariate analysis, after adjusting for markers of iron store and inflammation, only sFas correlated with Epo/Hct. In the CKD group, there were negative correlations between serum levels of sFas and glomerular filtration rate (GFR) (r = -0.45, p < 0.001) and between Epo/Hct and GFR (r = -0.32; p = 0.02). There was a positive correlation between Epo/Hct and serum levels of sFas in the CKD group (r = 0.31, p = 0.03) and in the HD groups (r = 0.58, p = 0.001). CONCLUSION: Our findings show that serum sFas is associated with higher Epo/Hct ratio, suggesting that sFas may be a marker of Epo hyporesponsiveness in uremia. Further studies are needed to determine whether sFas is just a marker of Epo hyporesponsiveness or is also involved in its pathophysiology.

Physicochemical properties of ferumoxytol, a new intravenous iron preparation.

BACKGROUND: Intravenous iron is a critical component of anaemia management. However, currently available preparations have been associated with the release of free iron, a promoter of bacterial growth and oxidative stress. MATERIALS AND METHODS: We determined the molecular weight, dialysability and capacity for free iron release of ferumoxytol, a semi-synthetic carbohydrate-coated superparamagnetic iron oxide nanoparticle. Ferumoxytol was compared with three intravenous iron preparations in clinical use: iron dextran (low molecular weight), sodium ferric gluconate and iron sucrose. Intravenous iron preparations were also incubated in rat, and pooled human sera (at concentrations of 600 microM and 42 microg mL(-1) respectively) from healthy subjects. RESULTS: The molecular weight of ferumoxytol was 731 kDa. The relative order of molecular weight was as follows: ferumoxytol > iron dextran > iron sucrose > sodium ferric gluconate. The least ultrafilterable iron was observed with ferumoxytol and the most with ferric gluconate. The least dialysable free iron was observed with ferumoxytol and the most with ferric gluconate. Incubation of intravenous iron preparations in rat or pooled human sera demonstrated minimal free iron release with ferumoxytol. The order of catalytic iron release as detected by the bleomycin detectable iron assay was as follows: ferumoxytol < iron dextran < iron sucrose < ferric gluconate. A similar trend was observed for the in vivo serum concentration of free iron in rats. CONCLUSIONS: In vitro observations from these experiments suggest that ferumoxytol has a favourable profile in terms of tendency to release free iron, in comparison with currently available intravenous iron preparations.

The importance of transitions between dialysis and transplantation in the care of end-stage renal disease patients.

Analyses describing outcomes of kidney transplantation usually exclude the survival of wait-listed patients and dialysis patients with failed kidney transplants, and thus reflect only a portion of the typical transplant process. We determined death rates during the continuum of wait-listing, transplantation, and after allograft failure among adult end-stage renal disease patients in the United States between 1995 and 2003. Before transplantation, death rates increased with longer waiting times. Death rates were lowest during the period of allograft function and highest after allograft failure. Patients were at particularly high risk during periods of transition between dialysis and transplantation (death rates during the peri-transplant period and during the re-initiation of dialysis after transplant failure were 8.2/100 patient-years (95% confidence interval (CI) 7.7, 8.8) and 17.9/100 patient-years (95% CI 15.7, 20.3), respectively compared to 6.4/100 patient-years (95% CI 6.25, 6.51) during the period of wait-listing. Diabetic patients and older patients were at increased risk at all time points. The most common known cause of death in all age subgroups was cardiovascular disease. The proportion of death owing to sepsis was greatest after allograft failure (16.8% of all deaths were due to sepsis compared to 14.0% during wait-listing, and 12.7% during the period of allograft function). Consideration of the entire transplant experience as a whole should help to focus patient care on periods of particularly high risk, and emphasizes opportunities to improve outcomes by strategies aimed at preventing death owing to cardiovascular and infectious causes.

CREB transcription factor modulates Bcl2 transcription in response to C5a in HL-60-derived neutrophils.

BACKGROUND: Complement fragment C5a and neutrophils have been implicated in the pathogenesis of renal disease and C5a has also been shown to delay apoptosis of human neutrophils via a transcription-independent pathway. However, transcription-dependent pathways have not been well described. The present study examined whether activation of HL-60-derived neutrophils by C5a modulates the transcription of two members of the Bcl2 family, Bax (pro-apoptotic) and Bcl2 (anti-apoptotic) molecules, and whether the cAMP-response element-binding protein (CREB) transcription factor mediates these effects through the phosphatidylinositol 3-kinase (PI3K)/Akt and extra-cellular signal-regulated kinase (ERK) signalling pathways. MATERIALS AND METHODS: The human promyelocytic leukaemia HL-60 cell line was differentiated into neutrophils using 1.25% DMSO. Differentiated cells were incubated with recombinant human C5a for 30-120 min with, or without, pretreatment with wortmannin or PD98059. The cells were lysed and quantified for gene-specific Bax and Bcl2 mRNA. In separate experiments, cells were incubated with C5a for 5-30 min with, or without, pretreatment with wortmannin, PD98059, or alkaline phosphatase. Cells were then lysed and immunoblotted using antihuman phospho-CREB (Ser133) antibody. Apoptosis was assessed by measuring active caspase-3 in differentiated HL-60 cells. RESULTS: C5a inhibited caspase-3 activation in HL-60-derived neutrophils (P=0.003). C5a significantly increased the expression of Bcl2 mRNA (P=0.028), which was time-dependent, peaking at 30 min, and was abrogated in the presence of either wortmannin or PD98059 (both P=0.028). The C5a had no impact on Bax mRNA expression. The Bax : Bcl2 mRNA ratio markedly decreased at 30 min (P=0.028). Time-dependent effect of C5a on CREB phosphorylation was demonstrable and rapid, peaking at 5 min, and was abrogated by either wortmannin or PD98059 (both P=0.028). Phosphorylation of CREB, but not of Akt and ERK, was inhibited by alkaline phosphatase (P=0.028). The effect of C5a on Bcl2 mRNA expression was abrogated by alkaline phosphatase (P=0.028). The Bax : Bcl2 mRNA ratio markedly increased in the presence of alkaline phosphatase (P=0.046). CONCLUSIONS: This study demonstrates that C5a induces Bcl2 mRNA transcription in HL-60-derived neutrophils, which is mediated in part by CREB through the convergence of the PI3K/Akt and ERK-signalling pathways.

Hepatitis C virus in chronic dialysis patients.

Since the cloning of hepatitis C virus (HCV), numerous serologic and virologic tests for detecting HCV infection have been developed and implemented in clinical practice. As a result, significant advances have been made in the study of HCV infection in patients with end-stage kidney disease. Patients on hemodialysis have a higher incidence and prevalence of HCV infection than the general population. In addition, HCV infection affects adversely survival among patients with end-stage kidney disease. Risk factors for HCV infection in dialysis patients include number of blood transfusions, duration of hemodialysis, mode of dialysis, prevalence of HCV infection in the dialysis unit, previous organ transplantation, intravenous drug use, male gender, older age and nosocomial transmission of HCV in hemodialysis units that can occur due to breakdown in standard infection control practices, physical proximity to an infected patient, cross-infection through dialysis machines, disrupted integrity of dialyzer membrane or dialyzer reprocessing. Suggested strategies to control HCV transmission in hemodialysis units include strict adherence to universal precautions, careful attention to hygiene, sterilization of dialysis machines and routine serologic testing and surveillance for HCV infection. Antiviral therapy with interferon alpha is recommended for selected categories of HCV-infected hemodialysis patients and kidney transplant candidates.

C5a delays apoptosis of human neutrophils via an extracellular signal-regulated kinase and Bad-mediated signalling pathway.

AIMS: We recently demonstrated that complement fragment C5a delays apoptosis of human neutrophils via induction of the phosphatidylinositol-3 kinase (PI 3-K) pathway. In the present study, we examined whether C5a modulates neutrophil survival through the extracellular signal-regulated kinase (ERK) and Bad-mediated signalling pathway. METHODS: Human neutrophils were isolated by percoll gradient and preincubated for 1 h with or without PD98059 (20 microM), a specific ERK inhibitor, followed by incubation with C5a (1 microg mL(-1)) for 24 h. Apoptosis was quantified by flow cytometry, using propidium iodide nuclear staining. Extracellular signal-regulated kinase downstream signalling events were evaluated by measuring the expression of cytosolic total and phosphorylated p44/p42 proteins, and Bad phosphorylation using immunoblot analyses. These time-dependent analyses were performed over a brief exposure to C5a (0-30 min). Modulation of cytosolic caspase-9 and caspase-3 activity was measured by Western blot analyses. RESULTS: C5a inhibited neutrophil apoptosis (P=0.04), which was abrogated in the presence of PD98059 (P=0.04). Time-dependent effect of C5a on p44/p42 phosphorylation was rapid, peaked at 5 min, and was abrogated by the ERK inhibitor (P=0.04). In addition, brief stimulation of neutrophils with C5a induced phosphorylation of Bad, which was inhibited by the ERK inhibitor (P=0.03). Further, C5a suppressed the proteolytic cleavage of caspase-9 and caspase-3, which was reversed by ERK inhibition. Finally, blockade of both the ERK (with PD98059) and PI 3-K (with wortmannin) pathways did not induce additive inhibition of neutrophil apoptosis by C5a. CONCLUSION: This study demonstrates that in addition to the PI 3-K pathway, C5a also inhibits neutrophil apoptosis via an ERK-signalling pathway, resulting in phosphorylation of Bad and blockade of proteolytic cleavage of caspases. The activation of this additional survival-signalling pathway may be another important cellular mechanism that enhances neutrophil survival in inflammatory states.

Quantification of Bax and Bcl2 in polymorphonuclear leukocytes from haemodialysis patients: relation to hydrogen peroxide.

BACKGROUND: Bax and Bcl2 are two apoptosis-related molecules that play an important role in determining cell fate following oxidative injury. In the present study, we explored the relation of hydrogen peroxide (H2O2) generation by polymorphonuclear cells (PMNs) to the cytosolic expression of Bax and Bcl2 proteins and apoptosis in haemodialysis (HD) patients. METHODS: Cytosolic generation of H2O2 by PMNs from control subjects and HD patients was measured by flow cytometry using the dichlorofluorescin diacetate assay. Bax and Bcl2 expression was detected by flow cytometry using FITC-conjugated antibodies. Apoptosis was quantified by flow cytometry using propidium iodide nuclear staining. To examine the effect of H2O2 on Bcl2 and Bax expression, PMNs from control subjects were briefly exposed to H2O2 (0.1-100 microM) for 10 min and then washed and cultured for 6 h, with or without catalase, a H2O2 detoxifying molecule. Bcl2 and Bax expression was determined by Western blot analysis. RESULTS: Basal H2O2 generation by resting PMNs was significantly higher in HD patients compared with control subjects (211 +/- 115 vs. 23 +/- 5 MFI; P=0.002). However, PMNs from HD patients did not undergo accelerated programmed cell death compared with control subjects (58 +/- 7% vs. 46 +/- 5; P=0.14). Polymorphonuclear cells cytosolic Bcl2 was undetected in control subjects but detected in 25% of HD patients, and Bax was more frequently detected in PMNs from HD patients (75% vs. 67%; P=0.04). In the HD patients with detectable cytosolic Bax and Bcl2 proteins, the Bax to Bcl2 ratio inversely correlated with H2O2 levels (P<0.0001). Finally, brief exposure of PMNs to 0.1-100 microM of H2O2 resulted in a marked increase in Bcl2 expression (P=0.001), which was prevented by catalase (P=0.05). There was no apparent effect on Bax expression. CONCLUSIONS: This study demonstrates that in HD patients, high-resting cytosolic H2O2 production by PMNs is not associated with accelerated in vitro apoptosis, and that the Bax/Bcl2 system may counter-balance the deleterious effects of reactive oxygen species in human PMNs.

Transmission of viral hepatitis by kidney transplantation: donor evaluation and transplant policies (Part 1: hepatitis B virus).

This two-part article discusses serologic testing of prospective donors for viral hepatitis B and C, as part of the comprehensive donor evaluation, and reviews the current policies and practices aimed at preventing donor-to-recipient transmission of hepatitis B and C viruses (HBV, HBC). This first part of the review discusses HBV. Organs procured from HBV-infected donors can transmit the virus to their recipients. Because infections with HBV have been associated with increased morbidity and mortality among renal transplant recipients, it is important to prevent HBV transmission with renal transplantation. Routine serologic evaluation of prospective organ donors for markers of HBV infection includes testing for hepatitis B surface antigen (HBsAg), anti-hepatitis B surface antigen antibody (HBsAb), and antibody to hepatitis B core antigen (anti-HBc). The risk of HBV transmission with kidney transplantation is a function of the serologic status of both donor and recipient. Knowledge of this risk is essential for the rational use of kidney allografts. HBsAg-positive donors are at high risk of transmitting HBV infection to their organ recipients, particularly if these donors are concurrently positive for hepatitis B e antigen (HBeAg). Kidneys from donors with isolated presence of HBsAb are unlikely to transmit HBV infection to their recipients. The risk of HBV transmission with the use of kidneys from IgG anti-HBc-positive, HBsAg-negative donors is low. Kidneys from donors negative for both HBcAg and anti-HBc are at low-to-negligible or no risk of transmitting HBV to their recipients. Under certain conditions, kidneys from HBV-infected donors can be safely used and thus prevent unnecessary discarding of organs. Kidneys from HBsAg-positive donors, who are negative for HBeAg, carry no risk or only minimal risk of transmitting HBV infection to their recipients if these recipients are immune to HBV or HBsAg-positive. However, the safety of these policies deserves further evaluation.

Impact of iron dextran on polymorphonuclear cell function among hemodialysis patients.

BACKGROUND: Polymorphonuclear cell (PMN) dysfunction and the increased use of parenteral iron may be important contributory factors to bacterial infections among patients with end-stage renal disease (ESRD) on maintenance hemodialysis (HD). We compared the in vitro impact of a commonly used parenteral iron preparation, iron dextran, on PMN function and viability between a group of HD patients with normal iron indices and healthy subjects. METHODS: Eleven patients with ESRD on HD and 10 healthy subjects were studied. PMN harvested from heparinized blood were incubated with iron dextran (0 - 20 mM) in culture medium (RPMI) for 24 hours at 37 degrees C with 5% CO2 following which function and viability were assessed by flow cytometry using appropriate fluorescent labels. RESULTS: Unstimulated, S. aureus and N-formyl-methionyl-leucyl-phenylalanine (fMLP)-stimulated hydrogen peroxide (H2O2) production was significantly higher in PMN unexposed to iron dextran from HD patients compared to those from healthy subjects. Iron dextran had no impact on unstimulated PMN H2O2 production in either group. In the healthy group, the only significant change occurred with 4-beta-phorbol 12-beta-myristate 13-alpha-acetate (PMA) stimulation, where cells exposed to 0.2 and 2.0 mM iron dextran produced less H2O2 relative to PMN unexposed to iron dextran (p < 0.05). In the HD group, all concentrations of iron dextran significantly attenuated H2O2 production stimulated by S. aureus, fMLP and PMA compared to PMN unexposed to iron dextran. Although PMN phagocytosis decreased with exposure to increasing concentration of iron dextran in both healthy subjects and HD patients, these changes did not achieve statistical significance. No significant changes in PMN viability or apoptosis were seen in either group after exposure to iron dextran. CONCLUSIONS: These results indicate that iron dextran, a standard parenteral iron preparation, attenuates PMN function in HD patients with normal iron indices at clinically relevant concentrations. Further studies are required to evaluate and compare the impact of newer preparations of parenteral iron, such as iron sucrose and ferric gluconate, on PMN function.

Effect of biocompatibility of hemodialysis membranes on mortality in acute renal failure: a meta-analysis.

BACKGROUND: The effect of biocompatibility of hemodialysis membranes on mortality in acute renal failure (ARF) has been a subject of intense debate, with some, but not all studies reporting a lower risk of death among patients with ARF dialyzed with biocompatible membranes (BCM) compared to bioincompatible membranes (BICM). OBJECTIVES: We performed a meta-analysis of group data extracted from previously published studies of controlled clinical trials to assess the impact of BCM on the mortality among patients with ARF who required intermittent hemodialysis (IHD). METHODS: BCM and BICM were defined as synthetic and cellulose-derived membranes (cuprophan and cellulose acetate), respectively. All controlled clinical trials comparing the effect of BCM to BICM on clinical outcomes in the setting of ARF were included. Original articles as well as abstracts were included. Data in Tables, Figures, and text were independently extracted by 2 of the authors. Risk ratios (RR) for mortality were combined using the random-effects model. RESULTS: Seven studies with a total of 722 patients met the inclusion criteria. One hundred seventy-two (45%) of 384 patients died in the BCM group, compared with 156 (46%) of 338 patients in the BICM group. The RRs for mortality ranged from 0.56-1.28. Overall, the pooled RR for mortality was 0.92 (95% CI = 0.76-1.13) in favor of the BCM group. However, the test for heterogeneity in RR among studies was significant (chi2 = 8.6, p < 0.05). One study accounted for this significance, and once removed from the model, the RR for mortality was 0.94 (95% CI = 0.79-1.12), and the test for heterogeneity among studies lost its significance. Subgroup analyses comparing BCM to cuprophan membranes revealed that the RR for mortality was 0.82 (95% CI = 0.62 - 1.08) in favor of the BCM group, whereas in the subgroup of studies comparing BCM to cellulose acetate, the RR for mortality was 1.11 (95% CI = 0.87-1.44) in favor of the BCM group. CONCLUSION: This metaanalysis demonstrates that the use of BCM does not significantly affect mortality among patients with ARF who require IHD. However, subgroup analyses suggest that cellulose acetate membranes may offer a survival advantage when compared with synthetic membranes, which, in turn, may be more beneficial than cuprophan membranes. Available evidence does not permit a recommendation for or against the use of BCM in ARF. Large trials and pooled analyses of individual patient-level data will be required to assess sources of variability among studies and non-fatal outcomes of ARF.