Egidio Lusvarghi

Oxidative Metabolism of Polymorphonuclear Leukocytes and Serum Opsonic Activity in Chronic Renal Failure

Luminol-amplified chemiluminescence was used to study the oxidative metabolism of polymorphonuclear leukocytes (PMN), in resting state and in response to opsonized zymosan, in 65 patients with different degrees of chronic renal failure (CRF) or on regular dialysis treatment (RDT). Every patient was compared on the same day with a normal subject. Furthermore, the serum opsonic activity was evaluated, cross-matching zymosan opsonized by serum from CRF-RDT patients and normals with PMN from CRF-RDT patients and normals. PMN resting chemiluminescence showed a progressive increase inversely related to the glomerular filtration rate, and it remained high in patients on RDT. Zymosan-activated chemiluminescence indicated a deficit in phagocytosis for PMN of patients with a glomerular filtration rate lower than 10 ml/min, persisting in RDT patients. The serum opsonic activity was always significantly lower in CRF and in RDT patients than in the control group; this defect was already present in patients with mild renal impairment. Our findings suggest that PMN from CRF or RDT patients have an increased reactive oxygen metabolite production in the resting state that may cause cell and tissue damage; the opsonization impairment and the decreased PMN phagocytic activity contribute to increased vulnerability to infection in these patients.

No Direct Evidence of Increased Lipid Peroxidation in Hemodialysis Patients

Lipid peroxidation, as measured by the thiobarbituric acid test, has been reported to have increased in hemodialysis (HD) patients, even though the test has low specificity in vivo. Conjugated diene fatty acid (CDFA) hydroperoxides are formed during lipid peroxidation, but not all conjugated dienes (CD) detected in humans originate from lipid peroxidation: octadeca-9,11-dienoic acid, a nonhydroperoxide CD derivative of linoleic acid (CDLA), has a dietary origin. We evaluated CDFA hydroperoxides, CDLA and linoleic acid, using high-performance liquid chromatography, in lipids extracted from plasma, adipose tissue and RBC membranes obtained from 25 patients treated with HD, 16 patients treated with hemodiafiltration (HDF) and 29 controls. No differences in the levels of CDFA hydroperoxides and linoleic acid were seen in any of the groups. Concentrations of CDLA were found to be significantly high in the adipose tissue and low in the RBC membranes of HD patients. HDF-treated patients showed the same results as HD patients. No direct evidence of increased lipid peroxidation was found in HD patients. This does not exclude the possibility that lipid peroxidation is increased and escapes direct detection due to the bodys homeostatic control eliminating the increased production of hydroperoxides. Both HD- and HDF-treated patients showed a significant change in CDLA concentrations, either in the adipose tissue, or in the RBC membranes. These dietary CD may be mistaken for markers of lipid peroxidation by conventional methodologies.

Conjugated Diene Fatty Acids in Patients with Chronic Renal Failure: Evidence of Increased Lipid Peroxidation?

Conjugated diene fatty acids (CDFA) were evaluated by second derivative spectrophotometry in the plasma and adipose tissue of 42 chronic renal failure (CFR) patients in conservative treatment, 40 patients treated by hemodialysis (HD) with cuprophane, cellulose acetate or hemophan, 29 treated by hemodiafiltration (HDF) with polysulfone, polyacrylonitrile or polyamide, and 28 healthy controls. Plasma CDFA were also evaluated at the beginning, at 30 min and at the end of the dialytic session. CDFA were unchanged in CRF patients with creatinine clearance (Ccr) > 10 ml/min respect to the controls, CRF patients with Ccr < 10 ml/min showed a higher level of CDFA both in plasma and adipose tissue (p < 0.02). HD patients showed values similar to those of the control group. The lowest level of CDFA was found in HDF patients (p < 0.01 for plasma, p < 0.05 for adipose tissue versus both control and any other group). A significant relationship between plasma and adipose tissue CDFA was found in all groups. In the group of CRF patients with Ccr < 10 ml/min, females exhibited a higher level of CDFA both in plasma and adipose tissue. No significant change was found during dialytic session, independently from the membrane used. CDFA are not only primary products of lipid peroxidation, but also have a dietary origin, primarily from dairy products. Taking into account the reduced dietary intake, the increase in end-stage CRF may be due to an enhanced oxidative stress and/or to abnormalities in CDFA metabolism. Uremic patients, particularly in the predialytic stage, should be considered at risk for increased oxidative stress. HDF treatment better corrects the abnormality compared to conventional HD.

Very late activation-3 integrin is the dominant β1-integrin on the glomerular capillary wall : an immunofluorescence study in nephrotic syndrome

The expression of alpha 2; alpha 3; alpha 5; alpha 6-subunits of the beta 1 [very late activation (VLA)] integrin family was studied in kidney specimens using an immunofluorescent technique. 6 specimens from normal kidney were compared with 10 specimens from patients affected by various glomerulopathies [minimal change nephropathy (MCN), membranous nephropathy (MN) and systemic lupus erythematosus nephritis (SLEN)]. On normal glomeruli, alpha 3 was the dominant integrin, being mainly present on podocytes and showing a linear fluorescent pattern codistributed with laminin. In MCN and SLEN, alpha 3 presented a normal pattern. In MN, alpha 3 revealed a trabecular picture on thickened glomerular basement membranes. Moreover, in stage-III MN, a segmental loss of alpha 3-integrin was detected. In our opinion, VLA-3 may offer an interesting approach to the study of the relationships between podocytes and their substrate.

Beta-1 Integrins in the Normal Human Glomerular Capillary Wall: An Immunoelectron Microscopy Study

The localization of the alpha 2, alpha 3 and alpha 6 subunits of the beta 1 integrin family on different cells of the glomerular capillary wall and on juxta-capillary mesangium was investigated using an immunoelectron microscopic technique on freshly harvested normal human glomeruli. Alpha 2 beta 1, alpha 3 beta 1 and alpha 6 beta 1 were weakly expressed on both luminal and abluminal surfaces of glomerular endothelial cells; alpha 2 beta 1 and alpha 3 beta 1 were also found on the mesangium of the juxta-capillary areas. Alpha 3 beta 1 was regularly present in great density on the basal and lateral surface of podocyte foot processes, confirming alpha 3 beta 1 as the unique beta 1 integrin on glomerular epithelial cells. None of these integrins was strictly polarized along the glomerular basement membrane, thus suggesting, in agreement with recent literature, that these molecules perform other biological functions in addition to adhesivity.

On line filtration of dialysate: structural and functional features of an asymmetric polysulfone hollow fiber ultrafilter (Diaclean).

The endotoxin transfer across dialysis membranes has been investigated using specific in vitro circuits. Backdiffusion and backfiltration have been analyzed and most dialysis membranes have shown to be permeable to LAL positive substances. Synthetic membranes however display the better capacity of retention of these products despite their higher porosity and permeability. For such reason synthetic polysulfone ultrafilters are used as pyrogen filters to obtain ultrapure dialysate. We have investigated the characteristics of a polysulfone ultrafilter named Diaclean and manufactured by Amicon Ireland. The capacity of endotoxin retention has been investigated both in filtration and backfiltration modes on new and used ultrafilters. The capacity of endotoxin adsorption was investigated as well. Used ultrafilters appeared to maintain the retention capacity and the adsorption capacity up to 4 months of use. Only slight differences were noted from the baseline values (p = n.s.). The best adsorption capacity is always displayed by the outer layer of the membrane suggesting its best utilization in back filtration mode with tangential flow. No morphological changes were observed in the used membrane analyzed by scanning electron microscopy.

Polymorphonuclear Oxygen Free Radical Production and Complement Activation Induced by Dialysis Membranes as Assayed in an Experimental Model

Activation of polymorphonuclear leukocytes with subsequent production of reactive oxygen metabolites has been reported to occur during hemodialysis related to a membrane bioincompatibility. We used an experimental dialysis model to evaluate, by chemiluminescence, the production of reactive oxygen metabolites and, by C3a, complement activation induced by cuprophan, cellulose acetate, hemophan, polysulfone, polyacrylonitrile, polymethylmethacrylate or polyvinyl chloride blood lines alone. No differences were obtained in the system, at time 30 min compared to initial values, as far as zymosan-activated chemiluminescence is concerned; resting chemiluminescence increased markedly with cellulose acetate (+71%), cuprophan (+49%), polymethylmethacrylate (+22%), hemophan (+21%) but had no variation with polysulfone, polyacrylonitrile and blood line. The time course of C3a levels up to 120 min showed a marked rise with cuprophan and cellulose acetate, a moderate increase with hemophan, polysulfone and blood line, and a decrease with polymethylmethacrylate and polyacrylonitrile. The results obtained documented a different behavior of the production of reactive oxygen metabolites compared to complement activation and support the hypothesis that the production of reactive oxygen metabolites by polymorphonuclear leukocytes is stimulated not only by complement activation but also by a direct dialysis membrane interaction.

Behavior of Antibody Profile against Hepatitis C Virus in Patients on Maintenance Hemodialysis

The prevalence of anti-hepatitis C virus (HCV) in dialysis setting is still a nonstandard datum. In particular, it is not known of the phenomenon is stable or increasing or decreasing, even in a given geographical area. We studied the behavior of anti-HCV prevalence during a 12-month follow-up in 415 hemodialysis patients treated at a single institution and belonging to a limited geographical area with standard HCV endemic. Point prevalence of anti-HCV has shown a tendency to growth linked in part of the incidence of infection, in part to new positivities in patients already on dialysis treatment. More than 50% of the new HCV-positive patients, had no history of classical parenteral transmission of the virus. These findings suggest that HCV infection is a phenomenon on the increase in dialysis units and that dialysis treatment emerges as an independent risk factor in contracting infection.

Assessment of Occlusion of the Vascular Access in Patients on Chronic Hemodialysis: Comparison of Physical Examination with Continuous-Wave Doppler Ultrasound

Background: Dialysis access occlusion is the most common cause of hospitalization and a frequent indirect cause of mortality in patients on chronic hemodialysis. The clinical assessment of an arteriovenous shunt is presently the most widely adopted method for the diagnosis of vascular access occlusion in hemodialysis patients, but no studies have yet investigated objectively its sensitivity and positive predictive value (PPV). Continuous-wave (CW) Doppler ultrasound is a simple, inexpensive, and noninvasive technique for the assessment of arterial blood flow. We have carried out a prospective evaluation of the PPV of CW Doppler for the diagnosis of vascular access occlusion in hemodialysis patients and compared it with clinical investigation. Methods: Fourty-one hemodialysis patients with clinical diagnosis of occlusion of their fistula were studied, and in 23 of them the diagnosis of occlusion was objectively validated. Results: CW Doppler in the patients in whom occlusion was objectively validated showed PPV of 86 and 83% under basal conditions and after fistula compression, respectively, with sensitivities of 95 and 100%, respectively. Clinical diagnosis, under the same conditions, showed a PPV of 83% and a sensitivity of 100%. Conclusions: CW Doppler and clinical examination have a similar high sensitivity for the diagnosis of occlusion of the dialysis access; thus, there is no need to use routinely Doppler CW examination, unless objective documentation is required.

Leaching and corrosion of polyvinyl chloride (PVC) tubes in a dialysis water distribution system.

PVC tubes, from the dialysis water distribution system of a 14 and a 7 year old plant, were analysed by scanner electron microscopy (SEM) and X-ray microanalysis (EDAX). Results showed that the leaching of material occurs depending on time of use. The phenomenon does not modify the chemical composition of the material. Results suggest that PVC alterations, induced by time, could support bacterial proliferation thus reducing the quality of dialysate.